oe ee ete ee nm betas «& teh ae ‘ ete eee ee ee Tee Aes eeaaeenens ak arr naen C Sreee Nreae . Rw Me a cso Te Rete ee ‘o vin i aii 7 wie wes ce Pete wesw” no £ 7 ii Fh) Te ui ' Ave Tay : — : ra ad : 7 s ; | ry , it . . NA i ‘ + AUGUST 1954 PROCEEDINGS NATIONAL SHELLFISHERIES {39/3 Volume 45 wa the Sia Ope @4 4 f ~ - PROCEEDINGS of the NATIONAL SHELLFISHERIES ASSOCIATION Official Publication of the National Shellfisheries Association; an Annual Journal devoted to Shelifisheries Biology Volume 45 August, 1954 Published for the National Shellfisheries Association by the Fish and Wildlife Service, U. S. Department of the Interior Washington, 1955 hae vanes ae “ay hs byl te ak , , ‘= oa / ) if ae ' eos Ps: an a 2 2 a’ ir - ea) oe en cite or ii phe ‘ag ti) Shale Bd eta iin % ‘Ale Ga ' a anae Wade : soni in a ae a2 oT “Tv ae) TABLE OF CONTENTS 2 1 ih GEIL ESTEE EN Gis’ Bilteralc ekattra oidicl do1 6 avcrac gland are amtonanonerera staat cua.ovm obs, .6 a Aratdipumiond lavarenarate) 0, (6 Brief History of the National Shellfisheries ASS OGHELb AGIA Wale plore & 6,0 4/010, 0.0 0: 0.414 0 e146 Ala loramlaiaiotoia bras Cree RANGES, BEAVIENc xo 3 ANNUAL CONVENTION: OEt teeromena toma LESS asus nid « asatataiateln ie one eRe aia ana, dha eas a dioarale ater RES Olli OMS eearavalere io aoa ofalera Hn olola ona pid eto alalaonatelatotaen aaa cuakdton a: audratcka: akasa Oe Cherie) EC PHSUMerrS) REPELS as.nsa04 44046 2d Aw RRR OCR CEM CA ee ea wen one e mee OC & Opening Remarks of the President of the Nat-= ional Shelifisheries Assotiatione.csssade¢oanase. As Bs CHESTNUT... 7 Report of the President of the Oyster Growers and Dealers Association of North America.....J. RICHARDS NELSON... 9 Annual Report of the Director of the Oyster Institute and Secretary-Treasurer of the * Oyster Growers and Dealers Association of Merch AMETIER actacccadcasccamdeacsanceionacase, DAVID He WALEACH x, oie CONVENTION SYMPOSTUM ON VARIOUS ASPECTS OF OYSTER SETTING: DyBLER SGtbing .accauiomecacuadiaodd anatase eeaadis RICHARDS NELGON, = 16 How to Increase Production of Seed Oysters in COonneetiewt. «cilos OOOO NE OOS LI Ote GOMCEO BOGE EO a ling TMOG SVAIIGIHMa cg eAls) © Observations of the Behavior and Distribu= tion Of Oyster, LArVae s 26446 cscaaaes Aso mad occa THURLOW C.) NEPOONnceS, < Various Aspects of Oyster Setting in Mary= MEHL cu Ue UMC O COON EEC ORCC Ee GCeeRondesaacene (Cle INWANCARSy 1a Waite eZS) Setting of Oysters: in Virginiaasacsscssdenasecacaae JAY D. ANDREWS...38 The General Pattern of Oyster Setting in SOuth C&rVPl AGA. 66 <5,3220 cama dma tame als centers) G. ROBERT LUNA ¢ «647 Oyster Setting on the Gulf G@ensteinca chee sees 2. SEWER H. HOPKINS...52 OTHER CONVENTION PAPERS ; Distribution of Oyster Larvae and Spat in Relation to Some Environmental Factors abiél teh AEAUGb srk Jays cibhehis fc mv ericeCON CUD 6 OU ORO EM 2 6 J. H. MANNING and H. H. WHALEY...56 Food Requirements of Some Bivalve Larvae...«oeoeee> Vo Le. LOOSANOFF, H. C. DAVIS and P. E. CHANLEY... 66 Possible Causes of Growth Variations in Cuber TS WEE a ata arate kata atauateratelal satan Mer ataata at aia tnt x 2atala\ aloe Hae AOA ICH geet AE Selective Setting of Oyster Larvae on $Neenbtaiehhsdl (OM welsh Parco pene Go ka Ce ee aad! JNs ISUHMLINS As Leh The Tidal Spat Trap, a New Method for Col- leebine Séed Claris c.ccaaaaascasesametsaaeaacaae COUN Ba GLUON ..c 106 Recent Advances in the Studies of the Struc- ture and Formation of the Shell of Crass- ostrea VAPginicasscercccccecccecsssceccoseces PAUL S. GALTSOFF...116 > On the Rate of Water Propulsion by the Bay SEGIMIGD cdacau dasa secede dan en dagen aecneananaccn« We AS MCHMPMAN eee Growth Studies in Venus mercenaria....ccosccscoscsesh. He. GUSTAFSON...1L40 A Fungus Disease in Bivalve Larvae...ccsesceacoseeeeH. C. DAVIS and V. L. LOOSANOFF...151 Notes on Fungus Parasites of Bivalve Mol- lusks in Chesapeake Bay....ccsacccsccsescccscoesdAY D..ANDREWS...157 Studies of Pathogenesis of Dermocystidium MALL oat apslehale Alaa aola spac amieleinaas ae a a ales ca ela giavey oe) (Mins RAW etc J. G. MACKIN...164 Studies on the Effect of Infection by Dermo- cystidium marinum on Ciliary Action in Oysters (Crassostrea virginiens) «.scscaaagacae Ja Gaz MACKIN and S. M. RAY...168 A Haplosporidian Hyperparasite of Oysters......2.. J+ Gs» MACKIN and HAROLD LOESCH...182 Effects of Two Parasites on the Growth of OYSLETS oc. ck saeco tonnwecokegaesn850e605 Ra WINSTON MENZEL and SEWELL H. HOPKINS...184 The Functional Morphology of the Aliment- ary Canal of Asterias forbesi and the Predation of Bivalve Mollusks.........+.++FREDERICK.A. ALDRICH...187 Seasonal Vertical Movements of Oyster Drills (Urosalpinx cinerea)....ccseesee+eeeee+s MELBOURNE R. CARRIKER...190 Preliminary Experiments in the Use of Ground Controlled Aerial Photography in Inter- tidal Hydrographic Surveybesacnacasasecacccaacas ROBERT L. DOWs. 6199 The Use of Equipment and Techniques in Applied Shellfish Management ...0..020e+e+e2.+sDANA EK. WALLACE... Report on Certain Phases of the Chincoteague BaVarhnVieS Gl Sab ONS aa « aiacss ms ousiovs erent enneatce Mlle bro iW «= nln TING 2cace Computation of Oyster Yields in Virginia.........J3. L. MCHUGH and J. D. ANDREWS... Shellfish Sanitation as Related to the Export and: import Trade: i Canads. accadaesecuniosdvnae dia Re MENZATHS. 2. The Sanitary Aspect of Importation of Shell- fish into the United States..... L a i @ oe ; = | | FOOD REQUIREMENTS OF SOME BIVALVE LARVAE V. L. Loosanoff, H. C. Davis, and P. E. Chanley U..So Fish and Wildlife Service, Milford, Connecticut Almost at the very beginning of our work with bivalve larvae we began to suspect that the various species differed in their feed- ing behavior and had widely different food requirements. With more experience in culturing larvae and more knowledge of their behavior we concluded that, in general, the larvae of all the species that we have grown thus far, and they number 16, can be roughly divided into two groups. The larvae of the first group, probably best represented by oysters of the genus Crassostrea, which includes our Atlantic oyster, C. virginica, and the Japamese uyster, C. gigas, are able to utilize only a few of the many food forms which are fed to them (Davis, 1953). The second group, represented by larvae of the clam, Venus mercenaria, or the common mussel, Mytilus edulis, seems, on the contrary, to thrive on most micro-organisms provided they are small enough to be ingested (Loosanoff and Davis, 1950). The difference in the food requirements of these two groups was clearly demonstrated several years ago both in laboratory experi- ments and in outdoor tanks (Loosanoff, 1950). In these experiments the larvae of the clam, Venus mercenaria, and the oyster, Crassostrea virginica, were kept together in the same jars or tanks, and were, therefore, under identical conditions. They received the same food consisting of mixed nannoplankton in which the cells of small green algae predominated. The larvae of V. mercenaria grew rapidly and metamorphosed in about 13 days, while those of C. virginica showed virtually no growth after attaining the straight hinge stage and, eventually, died without any appreciable increase in size. The experiment was varied by changing the relative numbers of larvae of the two different species and by offering different quantities of food. Nevertheless, in all these experiments the larvae of the clam grew rapidly and metamorphosed, but the oyster larvae showed no growth. Apparently the latter were unable to utilize the food forms that were abundant in the surrounding water and which were, obviously, assimilated by the clam larvae. * Another circumstance which attracted our attention during the early stages of our work with larvae was that their survival and rate of growth depended, to a large extent, upon the quantity of food cells present in the water. On many occasions the larval cultures were killed by what appeared to be overfeeding. On others, the rate of growth was abnormally slow, prebably because of an insufficient quantity of food. Obviously, to succeed in growing larvae with consistent results their quantitative and qualitative food requirements had to be ascertained. abba The food requirements may vary with the species of larvae, their age, or with the species of the food organisms themselves. Because of our limited personnel, we could not undertake studies of the food require- ments of all the species of larvae with which we worked. Instead we con- fined our work to a few selected forms, which we believe to be representa- tive of much larger groups of bivalves. In this report we shall describe some of our experiments on clam larvae that we think are of general inter-= est. We may begin by repeating, what has already been reported by us in various articles, that contrary to the generally accepted view advanced by Blegvad (1915) we found no evidence that organic detritus is utilized by larvae of clams (Loosanoff, Millerfand Smith, 1951) or oysters (Davis, 1953). On the other hand, while clam larvae seem to be able to grow on sulfur bacteria, oyster larvae can utilize neither sulfur bacteria nor any other of the 13 species of marine bacteria isolated for us from the sea water of Milford Harbor by Dr. Burkholder formerly of Yale University. These results are rather interesting because they con- tradict another old and generally accepted, although unverified, point of view, that marine bacteria constitute an important part of the oyster diet. As may be remembered, Davis (1953) showed that young oyster larvae cannot utilize green algae, such as Chlorella, while larger larvae of the same species, after reaching the size of about 125 u, are able to do so. We had already made many observations indicating that larvae of several species, such as Venus mercenaria, Mya arenaria, Mytilus edulis, and even the European and Olympia oysters, can be grown on a diet con- sisting chiefly of Chlorella (Loosanoff and Marak, 1951). Davis' experi- ments, however, definitely proved that bivalve larvae are able to utilize certain green algae. This conclusion was supported by another series of experiments in which clam larvae were grown on a unialgal culture of Chlorella, reaching metamorphesis in about 12 days. Since this alga was isolated from the sea water of Milford Harbor, it probably constitutes part of the normal diet of the larvae of this region. Further experi- ments showed that clam larvae also grew well, reaching metamorphosis, on pure cultures of any one of the following three flagellates: Chla- mydomonas sp-, Chromulina pleiades, or Isochrysis galbana (Davis and Loosanoff, 1953). These studies demonstrated two important points, namely, that clam larvae can live and grow on a very restricted diet, actually con- sisting of a single species of algae, such as Chlorella, or flagellate, and that unlike oyster larvae they can utilize green algae during all stages of development. Our conclusions, obviously, differ from those of some European workers (Cole, 1936) who maintain that bivalve larvae do not possess certain enzymes needed for the digestion of cellulose of which the cell walls of algae, such as Chlorella, are made and, therefore, cannot survive on such a diet. a Ga =67= The next problem was to determine the effect of different con- centrations of certain food organisms upon the rate of growth and sur-= vival of larvae. From some of our early experiments we already knew that if fed approximately 200,000 to 300,000 cells of smail Chlorella, measuring only about 3 u in diameter, per cc. of water, the larvae would live and metamorphose. However, we did not know whether the optimum concentration of food celis would be dbove or below this range. To answer this question a series of cultures of larvae of Ve mercenaria, each containing approximately 6.5 individuals per CCo, were given different quantities of food. The principle foods tried were a mixed culture of plankton consisting largely of small Chlorella, measuring only about 3 u in size, and a unialgal culture of large Chlorella, the cells of which averaged about 8 u in diameter. As was shown by the increase in size and survival of the larvae, the optimum concentration of food depended upon the size of the food cells (Loosanoff, Davis, and Chanley, 1953). When large Chlorella was used the optimum concentration was approximately 50,000 cells per cc. (Fig. 1). However, approximately 400,000 cells per cc. of smaller cells were needed to maintain the growth of the larvae at an equal rate. The difference between the length-frequency distribution of the larvae in cultures that were given the optimm concentration of food and those that were somewhat overfed is illustrated in Figure 2, which shows that the larvae given approximately 400,000 cells of small Chlorella per cc. were, at the end of the tenth day, considerably larger than those that were fed approximately 750,000 cells. As can be concluded from the similarity of the two curves, the data also suggest that the food value of 400,000 small Chlorelia closely approaches that of 50,000 cells of the larger form. Were we to assume that the cells of Chlorella of both spe- cies were perfect spheres, the largest measuring 8 u and the smaller about 4 u in diameter, we would find that regardless whether we fed 50,000 cells of the large form or 400,000 of the small, the total volume of cells in_one cc. of water containing larvae would be approximately 13.40 x 10°3mm3 . Carrying this comparison further we may calculate that 750,000 cells of the smail form have a total volume of 25.13 x 10°3mm ay which closely approaches the volume of 26.81 x 10°3mm3. of 100,000 large cells. As can be seen from Figure 1, the larvae fed these quantities of the two foods grew at approximately the same rate, and both cultures showed signs of overfeeding. It would be interesting to determine whether the addition of a similar volume of other food forms per ec. of water would enable larvae to grow at the same rate. The larvae were usually killed when the concentration of food cells became too heavy. This concentration depended again upon the size and kind of cells. For example, a concentration of about 300,000 cells of large Chlorella per tc. of water killed approximately 90 per cent of the larvae within a few days and those that survived grew very slowly or not at all, as did the larvae in the unfed control. The volume of 300,000 =68= LEGEND SMALL CHLORELLA ---- LARGE » 400,000 ‘y 50,000 LENGTH IN MICRONS _# 300,000 UNFED CONTROL Fig. 1. Mean length of larvae of hard clam, Venus mercenaria, of different ages in cultures kept at the same population density of 6.5 larvae per cc. but given different numbers of cells of small or large Chlorella per cc. of water. 69 LEGEND ---- 50,000 LARGE CHLORELLA — 400,000 SMALL 5 “= 750,000 PERCENT LENGTH IN MICRONS Fig. 2. Length-frequency distribution (expressed in per cent) of ten day old larvae in cultures kept at the same population density of 6.5 larvae per cc. but given different numbers of cells of small and large Chlorella. 10 cells of large Chlorella is approximately 80.43 x 10°3mm3, When the concentrations of large Chlorella were increased to approximately -=500,000 or 700,000 celis per ce. total mortality occurred sometimes o~ within 24 hours. However, when fed the much smailer form of Chlorella the larvae survived and grew comparatively well in concentrations as high as 750,000 cells per ec. (Fig. 1). The larvae surviving in heavily fed cultures usually displayed some anatomical abnormalities which often made the larvae unable to ingest the food. Perhaps these abnormalities were to some extent res#- sponsible for the survival of such larvae under conditions which were lethal to normal ones. In discussing the effects of certain concentrations of food organisms upon the growth and survival of larvae it should be remem- bered that the number of celis given constitutes only one approach for estimating the value of the food form. It has been pointed out by other workers (Burlew, 1953) and it has also been noticed by us on numerous occasions, that the food value of such forms as Chlorella may show con- siderable variations according to the age of the culture, its density of population, and, of course, the media in which it is grown. While trying to standardize as many factors as possible in our feeding ex- periments we still have not tried to feed the larvae with Chlorella cultures of one age only. Comparable to the effects noted in adult oysters by Loosanoff and Engle (1947) we found that clem larvae may be killed either by the cells alone or by the filtrate only of the food culture or, of course, by a combination of the two (Loosanoff, Davis, and Chanley, 1953a) Dense concentrations of the food affect the larvae both mechanically, by the cells interfering with the swimming and feeding mechanism, and chemically, by the accumulated metabolic products of the cells which are toxic to the larvae. For example, in an experiment in which the larvae were kept in millipore filtered sea water, the larvae grew well in a culture receiving 100,000 large Chloreila ceils per cc., plus the media in which the Chlorella was grown, even though this concentration was somewhat above the optimm (Fig. 3, A). However, both the larvae receiving cells alone, filtered off by use of a millipore filter and resuspended in sea water, at the rate of 1,000,000 cells per cc. (Fig. 3, D) and the larvae receiving the filtrate only from the above cells (Fig. 3, E) soon died. The results indicate that the filtrate containing metabolites of the cells may be more detrimental than the heavy concentration of the cells themselves. This toxicity of the so called external metabolites and their ecological effect have long been recognized by aquatic biologists (Lucas, 1947). In this experiment the larval culture receiving filtrate from relatively small quantities of Chlorella grew somewhat better than the unfed control, especially during the first few days of life of the etal LEGEND A~- COMPLETE CULTURE-BASIS 100,000 CELLS PER C.C. B - FILTRATE ONLY =i ; pe ANSE C - UNFED CONTROL D-CELLS ONLY -BASIS 1,000,000 CELLS PER C.C. E- FILTRATE ONLY- “ ss i ae te LENGTH IN MICRONS ~ 90% DEAD Fig. 3. Effects of whole culture of Chlorella, of cells alone, and of the filtrate only at two concentrations upon growth of larvae of Venus mercenaria. larval culture (Fig. 3, B, C). Additional experiments will be needed to determine whether this growth was due to the suspended matter present in the water or to the ability of the larvae to utilize some of the dissolved substances in the filtrate. In observing the behavior of clam larvae we noticed that they showed both mechanical, or quantitative, and chemical, or qualitative, selectivity in feeding. By regulating the amount of food ingested the larvae kept in heavier-than-optimum, but still nonkilling concentrations of’ food cells, often contained less food in their stomachs than the larvae in lighter food concentrations. This suggests that larvae are not merely mechanical feeders but possess a mechanism by means of which they can control the food in- take. Nevertheless, if kept in a heavy concentration of food cells for a long time, the larvae lose this regulating ability and soon become choked with food cells. However, if these larvae were removed to clear sea water, they would, if not too seriously injured, expel the excess ingested food and continue to develop and grow normally if properly fed. -73- The ability of larvae to select food was observed when they were given a mixture of several food organisms but showed definite preference for one of them. As an example, given a mixture of Porphyridium and Chiamydomonas the larvae ingested the much larger cells of Chlamydomonas while rejecting the small celis of Porphyridium. Another problem was to find the optimum concentrations of larvae that should be maintained in the cultures. In the early stages of our work it appeared that if cultures are properly attended and fed, the majority of the larvae even in rather dense cultures will survive and, although displaying a slow rate of growth will, never- theless, reach metamorphosis. We now know that to some extent optimum, as well as maximum, concentrations of larvae depend upon the species, but such hardy varieties as V. mercenaria can be grown having as many as 50 larvae per cc. of water. Aithough this is a much heavier con- centration than those advocated by other investigators, who usually emphasize the danger of overcrowding, we have frequently reared larvae to metamorphosis in such densities and, on many occasions we have successfully grown even denser cultures. We believe that, as far as earrying the larvae of some species to metamorphosis is concerned, the danger of overcrowding may not be as acute as believed. This opinion is shared by our colleague, Melbourne R. Carriker, of the University of North,Carolina, who has also succeeded in growing cul- tures of V. mercenaria at the above mentioned concentrations (personal communication). To verify our impression, and at the same time learn more about the effect of crowding upon larvae a quantitative experiment was de= Signed to determine the survival and rate of growth of larvae in different concentrations (Loosanoff, Davis, and Chanley, 1953b). Larvae of V. mercenaria were grown in concentrations of about 6255 Sy 26 and 52 early straight hinge larvae per cc., with each culture re= ceiving the same quantity of food, namely, 100,000 ceils of large Chlorella per cc. The larvae in all cultures showed a low rate of mortality and grew to metamorphosis. However, the mean rate of growth of the different cultures had an inverse relation to the population density (Fig. 4). For example, on the tenth day the mean length of the larvae in the series of cultures containing 6, 635,13, 26, and 52 individuals per ce. was 162, 156, 151 and 144 u respectively. On the 12th day the difference between the two lightest cultures was even more pronounced. {tt is assumed that the slower growth in the more crowded cule tures was caused by more frequent collisions between the larvae, which interfered with the feeding, by the deleterious effects of the greater concentration of the excretory products of the larvae accumulating in the water and, at ieast during the later stages, by competition for food. The effects of crowding were furthér demonstrated by the experi- ments in which the cultures containing either 6.5 or 32.5 larvae per cc. oe LEGEND A- 6.5 LARVAE PER C.C. B- 13.0 " ri " C - 26.0 D - §2.0 IN MICRONS LENGTH Fig. 4. Mean length of larvae of Venus mercenaria of different ages in cultures kept at different popu- lation densities but receiving the same quantity of food per cc. of water (100,000 cells of large Chlorella). 75 were given what are considered optimum quantities of two kinds of food cells. Some cultures received 50,000 cells of the large Chlorella, while the others received 400,000 cells of the smaller form (Pires 5). The cultures containing 6.5 larvae per cc. grew quite rapidly on both kinds of food. The more crowded cultures, containing 32.5 larvae per cco, grew well between the second and fourth days suggesting that there was sufficient food for this number of larvae at this size, but grew much more slowly during later stages. At the end of 12 days they averaged only about 147 u, as compared with 189 and 185 u for the two lightly populated cultures. The discrepancy in the sizes of the larvae of the cultures widely differing in population density is further emphasized by com- paring the length-frequency distribution of the larvae. We may take as an example the cultures shown in Figure 5 that received 50,000 cells of large Chlorella but differed in population densities. By plotting the length-frequency distribution of the larvae of these cultures at the end of the tenth day one can easily see that while the modal class of the lightly populated culture was approximately 170 u, that of the denser culture was only about 145 u (Fig. 6). Furthermore, while the maximum size of the larvae in the first culture was at that time al- ready 200 u, the largest larvae in the more crowded culture measured only 17>u. it was also noticed that the range of lengths of the larvae was much greater in the lightly populated than in the denser cultures. Earlier Davis (1953) came to the same conclusion in his studies of oyster larvae, C. virginica. These experiments showed that, as we thought previously, clam iarvae will grow and reach metamorpho= sis even if heavily overcrowded (Loosanoff, Miller and Smith, 1951). However, present observations indicate that, contrary to our earlier opinion, the larvae in such overcrowded cultures will reach the setting stage later than those in less populated cultures. As a result of our experiments on crowding of larvae, the ques= tion arose whether the growth of larvae in overpopulated cultures could be maintained at the same rate as in lightly populated ones by increas- ing the quantity of food proportionally to the increase in population. Accordingly, experiments were conducted in which the increase in the larval population was accompanied by a proportional increase in the number of food cells. The cultures containing approximately 6.5 clam larvae per cc. were given 100,000 large Chlorella cells per cc. per day; the cultures containing 13 larvae received 200,000 cells; 26 larvae re- ceived 400,000 cells and, finally, the cultures with 52 larvae were given 800,000 cells per cc. per day (Fig. 7). At the end of the 14th day the larvae in the least crowded culture averaged aimost 190 U in length and many of them were already metamorphosing. In the next least crowded culture, containing 13 larvae per ec. of water and receiving 200,000 cells of large Chlorella per cc., the larvae averaged only slightly over 150 u, even though the same num- ber of food cells per larva was available as in, the first culture. A majority of the larvae given 400,000 cells of Chlorella per cc. died within four days. Those fed 800,000 cells per cc. died within 48 hours , =76 LEGEND oo SMALL CHLORELLA ~--- LARGE - » 6.5 LARVAE PER C.C. 6.5 LARVAE PER C.C. 32.5 LARVAE PER C.C. --a--# 93215 LARVAE PER C.C. 4 it LENGTH IN MICRONS Fig. 5. Mean length of larvae of Venus mercenaria of different ages in cultures kept at two different population densities but given the same quantities of food. The foods were 400,000 small or 50,000 large Chlorella cells per cc. iM LEGEND _ 50,000 LARGE CHLORELLA PER CC 6.5 LARVAE PER G.C. 50,000 LARGE CHLORELLA PER C.C. 32.5 LARVAE PER CC 20 PERCENT 3 Tr) 120 130 140 150 160 170 180 190 200 LENGTH IN MICRONS Fig. 6. Length-frequency distribution (expressed in per cent) of ten day old larvae of Venus mercenaria in cultures maintained at different population densities but fed the same quantities of food. 78 LENGTH JIN MICRONS LEGEND A- 6.5 LARVAE FED 100,000 CELLS B - 13.0 i "200,000 , C - 26.0 "400,000 D - 52.0 "800,000 100 % DEAD 100 % DEAD Fig. 7. Effects of increase in food con- centrations (large Chlorella) made in proportion to increase in density of larval population upon growth and survival of larvae. 79 the majority succumbing within the first 24-hour period of heavy feed- ing. This and similar experiments have demonstrated that an increase in the larval population beyond certain limits cannot be compensated, as far as rate of growth and survival are concerned, by a proportional increase in the quantity of food made available. On some occasions, recently developed carly straight hinge clam larvae were kept in cotton filtered sea water without receiving additional food for as long as 14 days. The growth of these larvae was; as a rule, considerably siowed down or entirely arrested, but the larvae themselves remained normal in appearance and continued to swim. If after such a long period of virtual starvation the larvae were given food, they began to grow and eventually reached metamorphosis. Obviously, clam larvae can endure a near starvation diet better than overfeeding. As may be concluded from this short report, the studies of elam larvae have already considerably added to our understanding of the behavior of these organisms, placing us in a position where we can now decide what, from a practical point of view, should be the most advantageous ratios in our cultures between the numbers of lar- vae and numbers of food celis of certain kinds. Future experiments should he directed towards making such ratios even more accurate by introducing a progressing correlation factor that would compensate for the increase in the size of the larvae during the life of a cule ture o We hope the information on the effects of temperature on the rate of growth of larvae (Loosanoff, Miller and Smith, 1951), together with the present data concerning the optimum ratio of food to larvae, will soon give us definite formulae indicating the most advantageous combinations of these factors for growing the largest number of clam set in the shortest time in small volumes of water. Even now, utilize ing our present limited experience, we are able to grow to metamorphosis approximately 1,000,000 small clams per month per five-gailon jar the bottom area of which is only about one square foot (Fig. 8). Our present methods allow us to grow clams during the entire year. There- fore, we can roughly estimate that a single jar can be used to grow approximately 12,000,000 clams per year provided we can keep mortality at a low level. Although we still do not know of any efficient method for con- trolling fungi, such as Sirolpidium, which at times causes heavy more tality among our larvae, we are optimistic enough to think that we may easily prevent and control many bacterial and, perhaps, virus diseases which we suspect to be prevalent among the larvae. Our optimism is founded on the results of some of our preliminary experiments in which larvae were kept in water to which certain quantities of sulpha drugs or such antibiotics as procaine penicillin G or chloromycetin were =8Q= Fig. 8. Battery of jars for rearing bivalve larvae. 81 added. Mortality of the larvae in cultures so treated was usually lower than in the controls. Further studies in this field will be conducted at our laboratory. Clam sets obtained by the use of our methods should be free of the admixture of juveniles of other bivalves, which in the early stages look almost the same as small V. mercenaria, and which later on will ‘have to be removed, a slow and expensive process. The clams will also be free of external enemies, such as crabs, or borers, the larvae of which may set in nature simultaneously with clams. Finally, the set will originate from known stock or, if necessary, even from known pairs of parents, as has often been done at our laboratory. From then on, if we learn how to control diseases of larvae and juvenile clams, the breeding of clams of desired qualities, such as rapid growth, high glycogen content, and resistance to certain diseases, will become commercially feasible. We wish to express our appreciation to our colleague, Charles A. Nomejko, for participating in some aspects of these experiments and for making the illustrations for this article, and to Miss Rita Riccio for her generous help in editing and preparing the manuscript. =Obs LITERATURE CITED Blegvad, H. 1915. Food and conditions of nourishment among the come- munities of invertebrate animals found on or in the sea bottom in Danish waters. Rept. Danish Biol. Sta. to Board Agric. 22: 4i=78. Burlew, Jo So (Editor). 1953. Algal culture from laboratory to pilot plant. Carnegie Inst. Wash. 1-357. Cole, H. A. 1936. Experiments in the breeding of oysters (Ostrea edulis) in tanks; with special reference to the food of the larva and spat. Fish. Invest. Series II, 15: 1-28. Davis, H. C. 1953. On food and feeding of larvae of the American oyster, C. virginica. Biol. Bull. 104: 334-350. Davis, H. C., and V. L. Loosanoff. 1953. Utilization of different food organisms by clam larvae. Anat. Rec. 117: 646. Loosanoff, V. L. 1950. Variations in intensity of setting of oysters in Long Island Sound. Atlantic Fisherman 30: 15-16, 47. Loosanoff, V. L., and J. B. Engle. 1947. Effect of different con- centrations of micro-organisms on the feeding of oysters (0. virginica). Fishery Bull. 42, Fish and Wildlife Service 51: aL=3i(« Loosanoff, V. L., and H. C. Davis. 1950. Conditioning Ve mercenaria for spawning in winter and breeding its larvae in the labor- atory. Biol. Bull. 98: 60-65. Loosanoff, V. L., and R. R. Marak. 1951. Cuituring lamellibranch larvae. Anat. Rec. lll: 129-130. Loosanoff, V. L., W. S. Miller, and P. B. Smith. 1951. Growth and setting of larvae of Venus mercenaria in relation to temperature. Jr. Mar. Res. 103 59-01. Loosanoff, V. Lo, H. C. Davis, and P. E. Chanley. 1953a. Behavior of clam larvae in different concentrations of food organisms. Anat. Rec. 117: 586-587. Loosanoff, V. L.; H. C. Davis, and P. E. Chanley. 1953b. Effect of overcrowding on rate of growth of clam larvae. Anat. Rec. 117: 645-646. Lucas, C. Ee 1947. The ecological effects of external metabolites. Biol. Reve 22: 2702295. =036 POSSIBLE CAUSES OF GROWTH VARIATIONS IN CLAM LARVAE P. Eo. Chanley U. S. Fish and Wildlife Service, Milford, Connecticut Ta recent years advances in larval research have greatly in- creased the possibility of raising the hard clam, Venus mercenaria, under hatchery conditions. As the commercial cultivation of clams becomes increasingly feasible, numerous questions are raised about all its phases. Perhaps the most frequent questions cencern the causes fordifferences in the rate of growth of larvae. These causes may be those brought about by environmental differences and those that are inherited. The effect of variation of environmental factors on the rate of growth of larvae has been studied to some extent. Thus, the effects of variations in temperature, density of population, and the type and quantity of food have been reported (Loosanoff, Miller and Smith, 1951; Carriker, 1952; Loosanoff, Davis and Charley, 1953a; Loosanoff, Davis and Chanley, 1953b; Loosanoff and Davis, 1953). However, very little is known about inherited variations. Loosanoff, Davis and Chanley (1953c) reported that, in general, there was no relationship between the age or size of the parent and either the size of the larvae or the viability of the spawn. They also reported that variations in both the viability of the spawn and the size of the larvae were common. Individual differences in larval size, within the same culture, have also been observed on many occasions (Loosanoff and Davis, 1950; Marak, 1951; Davis, 1953). Since environmental conditions for all larvae, within the same culture, should be virtually identicai, some of these variations must be caused by inherited differences. This study was designed to evaluate some of the effects of in= herited factors on the growth and survival of clam larvae. It should be emphasized, however, that this report presents the results of pre-= liminary studies and, therefore, the conclusions are tentative and may need to be revised in the future. ; In the first experiment we compared the larvae from several combinations of parents to determine the influence of each parent on the rate of growth of the larvae. Three female and three male Venus mercenaria were induced to spawn in the laboratory by increasing the water temperature and adding miilipore filtered sperm water. The eggs of one female ( 3) were then divided into three groups and each group was fertilized by the sperm of a different male. This resulted in the erosses = 3x1, 3x2, and 3x 3. Sperm from 1 was also used to fertilize the eggs of the other two females ( 1 and 2), giving the crosses 1x land 2x i. Triplicate cultures from each cross were then started, using approximately equal numbers of eggs, in 18 liter earthenware culture jars. These jars were kept in a common water bath to minimize the temperature differences between the cuitures. During =8h.a the course of the experiment the temperature ranged from 19.0° to 22.1, but at no time did the maximm temperature difference between the jars exceed 0.7°C. All cultures were fed equal quantities of Chlorella daily. Water in the jars was changed and larval samples taken at two day in= tervals. On the second day, aiter fertilization, a count was made to determine the percentage of eggs that had developed te normal straight hinge larvae and the population in each culture was then adjusted to approximately 180,000 to 200,000 larvae. The percentage of survival and the number of cultures continued for each cross are shown in Table I. Several factors were probably responsible for the differences in the percentages of eggs that developed normally into straight hinge larvae. The length of time between the discharge of spawn and fertili- zation may be cited as an example. We have found that the percentage of developing eggs decreased rapidly as the time interval between spawn-= ing and fertilization increased. This was apparently caused by the loss of viability of the sperm since another sample of the same eggs showed a high percentage of development if fresh sperm were used. Since the first male spawned more than two hours earlier than the last female, it is possible that the differences in survival in some of these crosses were partially due to the differences in the age of the sperm at ferti- lization. it is also possible that mechanical damage may result from screening or transferring the eggs after cleavage has begun. This may be responsible for some of the abnormalities and lower rates of survival in this experiment, since some of the eggs had undergone cell division before all the cultures could be started. These factors cannet aecount for all the observed differences, however, for in the 1x 1 cross the sperm was fresh and the same sperm, used later in the day, initiated good development in the eggs of 2 and 3. Moreover,the eggs in the 1x il cross were handled soon after fertilization and the cultures were started before cleavage had begun. Nevertheless, such a small percentage of the eggs developed to normal straight hinge larvae that the cultures of this cress were discarded. Since this female spawned very lightly while the other clams spawned more heavily, it seems reasonable to suspect that the condition of the egzs was responsible for their poor development. Possibly the eggs were immature. Davis (1949) reported the release of immature eggs by oysters and Locsanoff and Davis (1950) stated that, "Probably some clams were compelled by the strong temperature stimula- tion to abort the eggs even if the eggs were not fully ripe." on the cther hand, clams can retain mature gametes for extended periods of time without the loss of viability. Loosancff and Davis (1951) found that normal larvae could be raised even when the gametes were not released until several months after the end of the normal spawning period. Table I. Number of fertilized eggs used in each cross, percentage developing to normal straight hinge larvae, and number of cultures continued for growth studies. TOTAY, EES TOTAL PERCENTAGE NUMBER CROSS Cultured Normal Larvae Of Of Cultures At 2 Days Normal Larvae Continued iL se i 753,000 == less than 10 (0) eee ae 1,638,000 672 ,000 45 3 ce ames 1,500,000 gh2 ,000 63 3 See 1,500,000 360, 760 ek 2 3 en $3 1,500,000 180,000 12 aL =O6= When two females were crossed with the same male, 2x 1 and 3 x 1, there was a significant difference in the size of the larvae at two days (Fig. 1). Significance was determined by the use of the "+" test. The "t" value was 13.014, consequently the probability was much less than .001. However, from the second day to the con= clusion of the experiment, this difference in larval size remained constant and consequently the growth rates were nearly identical. Since no similar difference in larval size at two days was recorded when one female was crossed with three males and since the rate of growth, after the second day, was nearly equal in ali crosses of this experiment, it seems unlikely that any inherited difference in rate of growth was responsible for the difference in size of the 2x 1 and 3x 1 larvae at two days. We may conclude, therefore, that significant differences in the size of the larvae at two days are caused primarily by some factor, such as the size of the egg. Such a concludion is, however, tentative since at present we have only limited data on the correlation of egg size with the larval size at two days. There was a considerable range in size within each culture although all the larvae within a single culture were from the same parents. This size range increased with the age of the culture, as can be seen by comparing the length-frequency distribution of larvae at two days with that of the same larvae at ten days (Fig. 2). Apparent- ly, then, certain individuals grew consistently faster or slower than average, although the environment must have been virtually identical for all larvae within a culture. Although the effect of possible en- vironmental differences, within a culture, cannot be entirely eliminated, the range in length, observed in these crosses, must have been at least in part due to inherited differences. It would seem, then, that even among sibling larvae, there is a wide range of inherited differences that affects the rate of growth. In @ repeat experiment stripped sperm were used so that all sperm would be approximately the same age at fertilization. Ail eggs were placed in jars before cleavage had begun, to avoid damaging the early embryos by screening. Temperatures ranged from 22.8° to 24.3°C. with a maximm difference of 0.4°C. between jars at any given time. On the second day, populations were reduced to about 150,000 larvae per jar. Otherwise, the procedure employed was the same as in the first experiment. In the stripping process, the body fluids of the maie are un- avoidably mixed with the sperm. It is not known how these fluids affect the gametes or the developing zygotes. Some workers (Just, 1939) have demonstrated that, in other species, the body fluids interfere with the normal development of the eggs. This danger was probably slight in our experiment, since the water containing the eggs and sperm was greatly diluted when the eggs were put in culture jars. These fluids o87~ LEGEND o—- 22 BY | o-=--0 9 3, BY’ o || MICRONS DAYS Fig. 1. Growth rates of larvae from two different females crossed with the same male. Each point repre- sents the mean length of 100 larvae from each of tripli- cate cultures. 88 LEGEND —- $2 Bro! o---9 $3 BY) 10 DAYS PERCENT 100 110 120 130 140 160 160 170 180 190 200 MICRONS Fig. 2. Length-frequency distribution of the same two crosses at two and ten days showing the increase in range with the age of the larvae. 89 should not, in any event, have had any effect on the larvae after the water was changed on the second day. The numbers of larvae were not equal in all cultures for the first two days. There is no reason to believe that this affected the percentage of eggs developing to the normal straight hinge stage since the heaviest concentration was no higher than in many other experiments in which the development was normal. Nor is there any reason to be= lieve that this affected their growth during the first two days since there was no appreciable difference in the size of the straight hinge larvae with the possible exception of the larvae from the Bx A cross. The larvae of this cross were somewhat smaller but so few developed that they were discarded. Triplicate cultures of the A x A and Ax C crosses, duplicate cultures of the A x B cross, and a single culture of the C x A cross were continued for growth rate studies. The differences in the percentage of eggs that developed normally must be primarily due to the condition of the eggs in this experiment. The larvae from the C x A cross grew more slowly than those from the Ax Across (Fig. 3), and by the sixth day the difference was statistically significant, that is, the 95% confidence limits did not overlap. Since the growth rates of these crosses were different throughout the experiment, the difference appears to have been caused by factors inherited from the female parent. The differences in rates of growth, when one female was crossed with three males, were not nearly as pronounced. However, the difference between the mean lengths of the larvae from the Ax A and the Ax B crosses gradually increased and by the tenth day the 95% confidence limits determined statistical signis= ficance. (Fig. 4), This difference must have been the result of in- herited influences of the male parent. The mean lengths of the larvae from the Ax C cross are not significantly different from either the mean lengths of the Ax Aor the Ax B larvae. I would like to express my appreciation to Dr. V. L. Loosanoff and Mr. H. C. Davis for their invaluable assistance in all phases of the work. I am also indebted to Miss R. S. Riccio and Mr. C. A. Nomejko for their assistance in the preparation of the manuscript. SUMMARY 1. Significantly different rates of growth were found between larvae from the same female crossed with two different males and also between larvae from two separate females crossed with the same male. It is tentatively concluded that inherited differences from either of the parents may be responsible for these differences in the rate of growth. 290= LEGEND —- 2A BYOA e----- 96 BYOA MICRONS DAYS Fig. 3. Growth rates of larvae from two different females crossed with the same male. Points on the ZA x @A line are mean lengths of 100 larvae from each of triplicate cultures. Points on the $C x #A line are mean lengths of 100 larvae from a single culture. Shaded areas are the 95 per cent confidence limits. 91 LEGEND —- 3A BYWA o----- $A BY WB *~$a BY MICRONS DAYS Fig. 4. Growth rates of larvae from a single female crossed with three males. Each point on the ZA x #A and SA x A&C lines represents the mean length of 100 larvae from each of triplicate cultures. Each point on the gA x@B line represents the mean length of 100 larvae from each of duplicate cultures. 92 2. Significant differences in the mean lengths of larvae at two days were not correlated with subsequent differences in growth rate. It is tentatively concluded that these early differences are caused by physiological differences in the eggs. 3. The range of length of larvae, from a singie pair of parents, increased as the larvae grew larger and older. Apparently, then, sibling larvae have widely different rates of growth. At least part of this difference is believed to be a result of inherited dif- ferences. 4, The ability of sperm to fertilize eggs decreases as the time between the discharge of sperm and fertilization increases. Eggs remain fertilizable for a longer period of time. =93= LITERATURE CITED Carriker, M. R. 1952. Preliminary studies on the field culture, behavior, and trapping of the larvae of the hard clam, Venus (Mercenaria) mercenaria L. Nat. Shellfisheries Assoc. Conv. Add. 1952: 70-73. Davis, H. C. 1949, On the culture of oyster larvae in the labore atory. Nat. Shelifisheries Assoc. Add. 1949: 33-38. Davis, H. C. 1953. On food and feeding of larvae of the American oyster, C. virginica. Biol. Bull. 104: 334-350. Just, Ho Ee 1939. Basic methods for experiments on eggs of marine animals. 89 pp. Philadelphia: P. Blakiston's Sons & Co., Abrateye Loosanoff, V. Lo, and H. C. Davis. 1950. Conditioning V. mercenaria for spawning in winter and breeding its larvae in the laboratory. Biol. Bull. 98: 60-65. Loosanoff, V. L.,; and H. C. Davis. 1951. Delaying spawning of lamellibranchs by low temperature. Jr. Mar. Res. 10: 197-202. Loosanoff, V. L., and H. C. Davis. 1953. Utilization of different food organisms by clam larvae. Anat. Rec. 117: 6h6. Loosanoff, V. L., H. C. Davis, and P. E. Chanley. 1953a. Behavior of clam larvae in different concentrations of food organisms. Anat. Recs 117: 586-587. Loosanoff,; V. L., He C. Davis, and P. E. Chanley. 1953b. Effect of overcrowding on rate of growth of clam larvae. Anat. Rec. 117: 645-646. Loosanoff, V. L., H. C. Davis, and P. E. Chanley. 1953c. No req= lationship found between age of oyster and quality of spawn. Atlantic Fisherman 34: 22-23. Loosanoff, V. La, W. S. Miller, and P. B. Smith. 1951. Growth and setting of larvae of Venus mercenaria in relation to temper- ature. Jr. Mar. Res. 10: 59-01. Marak, R. Ro 1951. Variations in sizes and rates of growth of lamellibranch larvae of the same parents. Nat. Shellfisheries Assoc. Cony. Add. 1951: 45. xe) ea i. i ae 0 af ig ed merry > cae Lai a ap ka | | a) i tna & 2 a By tle <=" ; wee r Y io fies xe ea) > ee ook, hive ; y C. ped a rw = en ot 2p Agree Da . i *4 ey ps Se aioe oma hy 7 , ee, ae hes aac Pe cuales ro ‘qud es; Zhe 4% * ath r ey pcten) tihiae wee) oat ‘ dels fe earsn! aby Baths _ a “Wait, 7 “fim ' witeiee 20) daiwadea eet rash, “SM coh ose sanreniyh oD cebtaur Dig. .f22t iret aa — ii wo hye i has MA « «@eraP) aal4 J : = 7 ei a® ee itt aj mocymdes = akeel: -ysctei ado) be a ) ‘ ~ [whens Pa. 19 -noe hye Cigaes Sew wate Ve.29 a: : te - | i ‘ poe sacl : OM oye WJ 9 2a Bete minh! | i ==” p oe, dented xe” =eet ary hs ) ae |? LE oa. ntana ot ETAL gurl 30 let epee an] Tl ~ we OM =2EC0L . «Qiinnd) «2.58 ame We (ttf maa “tet erS fu. ae @ st dig 6 ROl iste 8G weet @2 dl tls eh a! zicunes a's 10 26585 eel Vikas sadn SELECTIVE SETTING OF OYSTER LARVAE ON ARTIFICIAL CULTCH Philip A. Butler U. S. Fish and Wildlife Service, Pensacola, Florida The consistently high oyster spatfall in the Gulf area sets this region apart from many other areas where oysters are cultured. Since knowledge of the causes of success or failure of a set is of great importance to the industry, we initiated studies on this pro-= blem at the Pensacola Laboratory in 1949. We made systematic counts of the time and frequency of spatfall in conjunction with continuous records of hydrographic conditions, This paper presents some of the general conclusions derived from a study of five years of data, es= pecially those relating to larvel behavior at the setting stage. It was necessary to select materials and methods for this study which would permit comparable results over a period of years and not be cumbersome. The collection of spat on oyster shells while having many desirable features has also some obvicus objections, especially the difficulty of computing the exposure area. For this reason, several flat artificial materials were tested with the re= sults shown in Figure 1. We sanded the surface of the Plexiglas so its texture was comparable to the other materials used. The fact that spat avoided the white Plexiglas is obviously not due to a color reaction, since the rates on black are nearly as low whereas the rate on white oyster shell is very high. We examined only the white surfaces of the oyster shells and carefully computed the area of the exposed surfaces, so that these data could be expressed as rates per square centimeter. The cement board, which received the highest set, is that commonly used for building purposes. It is easily handled and costs approximately one cent per 4 inch square plate. Such a plate provides one hundred square centimeters of exe posed surface. We also tested smooth and rough surfaces and found over a long period thet each receives the same amount.of set. Since the smooth surface of the cement board is so much casier to examine, it is the only one we used routinely. In the five years of observations, we have never had a routine plate, that is one exposed for 7 days, without at least cne sedentary animal form. We have had concentrations of oysters as high as 30 per square centimeter; that is roughly comparable te a set of 700,000 per bushel of shell. Barnacle rates have at times been over 100 per square centimeter, In general, when the concentration of a species reaches O.5) pex or more, counting 10% of the surface provides an accurate index of the total set. When oyster concentrations were below this figure, we examined from one to four full plates each week. We wash the plates under a strong jet of water before examination so that any debris or animais not firmly attached will be removed. =95- OYSTER SPATFALL SELECTION OF SUBSTRATE PLEXIGLAS - WHITE [+] PLEXIGLAS - BLACK FARIONS IME Die GIEANS!S (Sine saactal OYSTER SHELL CEMENT BOARD 46 % Fig. 1. Percentage occurrence of spat on plates of different materials exposed for 7 days. Plates were exposed simultaneously and adjacent to each other. Data include approximately 3,000 oysters. 96 Figure 2 shows the rack containing cement board plates which is exposed each week throughout the year. In special studies, other racks of similar nature carried plates at different angles. The racks hang halfway between surface and bottom in three meters of water on one of our docks. Figure 3 shows a horizontal rack carrying different colored plates about i meters below the water surface. This photograph indi- cates the usual clarity of the water during the oyster spawning season. One of the first things noted in our examinations was the pre-= ponderance of spat cn the upper surface of exposed plates. Figure 4 shows the relative numbers of spat setting on upper and lower surfaces of plates exposed throughout the setting season for four years. I have included here similar data on barnacles because of their probable cor-= relation with the data on oysters. This figure also shows the daily incidence of oyster spat on upper and lower surfaces of plates exposed for limited periods. Dark plates were exposed from 7:30 P.M. until 3:30 A.M. on consecutive moon- less nights, while light plates were in full sun from 7:30 A.M. until 3:30 P.M. on the intervening days. These data are somewhat limited and the experiment will be repeated next summer, but it appears that the majority of setting in this area occurs in daylight hours and on upper surfaces. Since the weekly plates are exposed at mid water level, we deter- mined the relative frequency of set at different levels in our area. Figure 5 shows the percentage of the total set which occurs at one foot intervals below mean low water. The first series, held on a fixed rack, shows approximately one per cent of the set occurring at the surface and nearly 80 per cent confined to the 4 = 7 foot levels. We repeated this work using a floating rack which fluctuated with the tide and obtained very similar results. The slight variations in numbers setting on the lower plates in the second series are of no real significance. In the third series, we extended the rack so the last plate was a scant inch above the kottom. This experiment showed a very different picture as to the location of the oyster set. Nearly 50 per cent of the set was on the bottom plate. The disparity between our data and reports in the literature led us to repeat some of the experiments first reported by A. E. Hopkins in his work on the Olympia oyster in 1935. Figure 6 shows the incidence of set on plates held at different angles in the water. Tnese plates were in separate racks but exposed simultaneously for { day periods. Again, as on our other plates, we foynd the majority, 78 per cent, setting on upper surfaces (135° plus 180°) and only 18 per cent on lower surfaces. These data were difficult to reconcile with Hopkins’ results which are shown on the third line of the table. More than 86 per cent of the Olympic oyster set occurs on under surfaces ard less than one-half per cent on the upper surface. =9[= Fig. 2. Weighted wood rack with four inch square asbestos cement board plates. Each plate surface provides 100 cm© area for attach- ment of sedentary organisms. 98 transparent, and Photograph of rack carrying white, meters below water surface. Fig. 3. black plates 13 99 OYSTER SPATFALL SELECTION OF SURFACE 4- YEAR AVERAGE DAILY INCIDENCE % 3644 DARK ON. Smee 6424 LIGHT Oe) BARNACLE Fig. 4. Percentage of oysters and barnacles setting on upper and under surfaces of cement board plates throughout spawning season for four years. Data include approximately 70,000 oysters and twice as many barnacles. Daily incidence of setting shows percentage of oysters found on upper and under plates exposed for 8 hour periods of continuous darkness or light. 100 OYSTER . SPATFAEE At’ .DIFFERENT . EEVEES % 10 20 10 20 10 30 50 RIGID FLOATING RIGID 1g0) 84 78 43, | 35 oO 16 22 Sy Tf 96.5 Fig. 5. Percentage of oysters setting on both surfaces of horizontal plates held in rack at one foot intervals. Racks were fastened to wharf (rigid) or suspended from float which fluctuated with the tide. Ratios show percentage of oysters setting on upper (180°) and under surfaces. Last ratio in- dicates plate at 9 foot level. 101 OYSTER SPATFAEL ANGLE OF INCIDENCE-% 90° 135° ee eee 5 Sir 1g0° ce 8 oe 8645 90 | 135 | 180 | " ee sie | SUSPENDED 8 ieee er Pe or | oo ON BOTTOM 96.6 ON BOTTOM 86 3.4 SUSPENDED 70.5 15.5 Fig. 6. Percentage of oysters setting on cement board plates held at several angles. Data include approximately 6,000 oysters. Tabulation compares results obtained at Pensacola (first two series) with results obtained by other investigators. 102 .We find the answer to these apparently conflicting results by referring again briefly to Figure 5. As noted earlier, 50 per cent of the set in the third series occurs on the bottom plate, but more signi« ficantly, 96 per cent of these are on the upper surface of the bottom . plate. On all of the remaining plates, the majority of the set occurs on the under surfaces. The ratio of top to bottom set for each series is shown in the lower part of the figure. The last ratio is that of the bottom plate of the third series. If we examine Figure 6 again, we find reasonably good agreement between my plates and Hopkins' plates which were located on the bottom. The final line in the tabie indicates the data obtained by a third in- vestigator using frosted glass plates suspended above the bottom. These data inciude all bivalves but were primarily oysters. The results are in good agreement with Hopkins' but are almost the exact opposite of my own data shown on the first line of the table. This was especially in- teresting to me, since Dr. Pomerat did this work at Pensacola in 190. We repeated his experiment as closely as possible, using the same locations and, perhaps, some of the identical glass plates which he used, but are unable to confirm his odservations. Since our results have been consistent over the past three seasons, I conclude that some special hydrographic feature was present during the two weeks that he conducted his experiment, Possibly in 1940 excessive turbidity caused a rapid sedimentation of his plates preventing bivalves from setting on the upper surfaces . Figure 7 shows one of the special crate collectors used to com= pare spatfall here with other areas where crate collectors have been tested. Each cubicle of the crate measures approximately 2 x 2 x 4 inches. We find this type of cultch expecially efficient in sttracting oyster spat, as many other investigators have reported. I have tabulated the actual numbers of spat counted in a single test. The uniformity of these num- bers, depending on the angle of surface, is extremely interesting. Var- ious organisms covered only about 10 per cent of the surfaces of the plates and hence there was very little competition for attachment space. The fact that each section in the crate caught nearly identical numbers of oysters indicates to mé an unexpectedly uniform dispersion of larvae in the plankton. The percentages of oysters found on the different sur= faces of all of these collectors agree with results obtained on plates_ held singly and in series. From these and other experiments I gain a general picture of the behavior of oyster larvae at the setting stage in the Pensacola area. The greatest deterrent to setting is a layer of sediment, just as in other areas investigated. A deposit of silt only as deep as the larvae, about 1/75 of an inch, will prevent their attachment on what is normally a preferred surface and cause them to set on a surface they would ordinar= ily avoid. The larvae have no particular attraction for black or white =103= OYSTER SPATFALL = CRATE COLLECTOR NUMBER ON ONE CRATE PERCENTAGE BY ANGLE OF SURFACE EXPERIMENTAL — CONTROL A = 320 Oo B= a9) b= 19 o° 6 20 C - 255 C= 287 90° 16 10 Ol = S32 ad- 31 180° 78 70 T = 326 t= 354 Fig. 7. Photograph of cement board "crate collector" and tabulation of setting on inner surfaces of the ten cubicles. Lettered diagram designates surface positions; T - t indicates sum of all surfaces in each vertical row of cubicles. Collectors were suspended at mid depth in three meters of water for seven day periods. Data include approxi- mately 5,000 oysters. 104 backgrounds either in the dark or daylight hours, and whén above the bottom have little preference to the lighted or unlighted side of a plate. I find no evidence that phototaxis plays a part in their selection of cultch. In one limited series of experiments, the larvae set primarily during the daylight hours. They appear to set equally on the rising and falling tides as well as during slack water periods. This may be a special condition of the Gulf environment, where there is only one tide each day and the change in water level is slight. We can predict the location of the spat, other conditions being comparable, almost entirely on the basis of the larvae's struggle against the laws of gravity. The majority are located and set on the bottom and their numbers decrease as the distance from the bottom increases. As the larvae swim upwards through the water they are just as likely to hit and set on an under surface as, when they fall back down through the water, they may hit and set on an upper surface. I believe there is no relation bwtween the swimming position of the larvae, i.e. with its foot upwards, and the surface on which the larva attaches. There are two factors of prime importance in determining the utilization of a surface as cultch. Sediment physically interferes with setting on uppér surfaces; this factor increases in importance near the bottom where currents that might wash silt away are relatively weak. The second factor of importance is the occurrence of barnacles. Barnacies May set more quickly than oysters on newly exposed cultch and the sweeping action of their appendages in collecting food repels the larvae. On upper surfaces, this sweeping action does not interfere too much with larvae which drift down between the barnacles and set. On vertical and under surfaces, however, when an oyster larva comes into contact with this field of activity it closes and fall® away from the surface. Relative- ly few barnacles can seriously interfere with the setting rates on vertical and under surfaces. in all of our work we have found that deviations from the expected i:1 ratio of spat on upper and under sur-= faces can be correlated with the numbers of barnacies on the under side. Barnacles have a marked preference for setting on under surfaces, and this leads to the predominance of the oyster sét on upper surfaces in this area. The importance of barnacles is obscured or overshadowed on bottom cultch where siltation is of paramount importance. In summarizing this work, it appears that in this area the move- ment of mature larvae is governed almost entirely by the laws of chance and gravity. The only selectivity larvae show in setting resuits from their avoiding silt or other organisms and they will be found on the first clean surface they can reach regardiess of its position in the water. it should be noted that these data refer to the incidence of setting and not spat survival. In some areas, factors causing early mortalities operate at certain levels or positions. This condition may produce an erroneous picture of the original location of the spatfali. =105= ’ - r ; e 4 ry Saye eg OV herve. Dy 54) SY og «nchfite ix aor ‘ae Foe , he. i. i es, 1 By ae js . Vs iy THE TIDAL SPAT TRAP, A NEW METHOD FOR COLLECTING SEED CLAMS John B. Glude U. S. Fish and Wildlife Service, Boothbay Harbor, Maine The Fish and Wildlife Service was directed by Congress in 1948 to determine the causes of the decline of soft and hard-shell clams along the Atlantic Coast, and to develop methods for increasing pro- duction. Our initial approach was to accept, for the time being, the statements that a scarcity existed and to attempt the most logical method for increasing production, viz., clam farming. Clam farming is a well established practice in a number of parts of the world and it seemed reasonable that methods which had been used in other places might be applied to the New England Coast. For many years the Japanese have utilized the tidal flats in Tokyo Bay for the production of clams as a farming venture (Glude, 1947). This industry is based on the collection of seed clams from the flats at the mouths of several rivers and the transplanting of these clams to private grounds. After a growing period of about one year the clams are dug and sent to market. Another commercial clam farm exists in Puget Sound, Washington, where one concern has title to approximately 10 miies of beach. Re= production appears to be adequate on these flats and the entire farm- ing procedure depends on natural seeding of the flats. In this case clam farming approximates the management of forestry land where it is simply a matter of harvesting a crop as it comes along and then leav- ing the area until a new crop has reached commercial size. The third area where commercial clam farming is practiced is the eastern shore of Virginia. (Tiller, Glude, Stringer, 1952). Here small hard-shell clams are placed on private beds, protected from pre- datory fish, and marketed when they have attained sufficient growth and when economic conditions are favorable. In each of these clam farming ventures the primary requirement is a source of seed. There- fore, we have devoted a considerable amount of our effort to a search for a source of juvenile clams. We recognize three principal methods of obtaining seed clams. 1) Collection of juveniles which have naturally set in large numbers , 2) Artificial propagation, 3) Collecting spat or juvenile clams by some device. -106- Natural setting appears to be the most practical method for the collection of soft-shell clams. We have found one place in Maine where each year soft-shell clams, 1/2 to 1 inch in length are found in concentrations of 1,000 to 1,200 per square foot over a consider- able area of sandy beach. We have developed methods for collecting these small clams by sorting them from the sand using a hydraulic seed rake (Glude, Spear, Wallace, 1952). Young hard-sheil clams, on the other hand, are seldom found in heavy concentrations. Ordinarily our bottom samples in Rhode Island contain one to 20 per square foot and it is difficult to work out mechanical methods for removing clams when they occur in these low concentrations. Only once in Rhode Island have we found hard clams in concentrations which might be usable as a seed source. This in- stance occurred in Greenwich Bay in the summer of 1951 when some of our samples contained as many as 600 Venus per square foot. This, however, appeared to be a very unusual instance and could not be de- pended upon as a source of seed quahaugs for a commercial farming venture. The best natural source of juvenile hard shell clams which we have seen is in Casco Bay, Maine. The Maine Department of Sea and Shore Fisheries is now transplanting these overcrowded and stunted clams to deeper waters where they will grow faster and where they will be protected from freezing (Dow and Wallace, 1951). ‘These clams set during recent years when temperatures have been high. A cold eycle might prevent Venus from reproducing in Maine. Therefore, we consider natural setting as a sporadic source of seed which should be utilized when available, but which can not be depended on each year. Artificial propagation has long been the goal of both oyster and clam biologists. Reliable methods for propagating Venus have now been developed by Dr. Loosanoff and have been applied in several laboratories. Hard-shell clams, fortunately, are among the easier bivalves to rear and this method may some day be usable on a com- mercial scale. It is entirely possible to rear sgoft-shell clams in a hatchery, but since they are so often available from natural setting it would probably be more economical to use this source of seed rather than artificial prepagation. The third method of collecting seed is that of employing some device or procedure to induce setting or of collecting the young juve- nile clams. In the oyster industry this would include spreading shells on the bottom to catch the set. In Japan, it might consist of hanging fibre mats out in the water to which the young larvae of some of the clams would attach. We have tried a great number of methods of in- ducing setting, some of which have been successful and most of which have failed. A timely suggestion from Roger Munsey, who was then Pala ye President of the Massachusetts Shellfish Officers’ Association, led us to design an automatic filtering device which we have named the Tidal Spat Trap. b The Tidal Spat Trap is simply a box which fills and empties with the tide and a system of check valves to force the outgoing water through a sand filter. The bivalve larvae are carried into the box by the incoming tide and are kept there by a tray of sand which filters the water on its way out. The outlet is located about eight inches above the filter so the larvae and spat can remain in water even though the tide falls below this level. Figure 1 is a diagram of the operating principle of this device. -108= SNOILVOILSAANI NV 19 WANA AGL Mom i Ve ad aids M0 ad SC) dO00A5 =—M0414 AVYL YALTIS GNVS ‘der, yedg TeptL ayy go wemetd “TT “Std as 1 SCM ONE bel ee gVal IVdsS= iV Giet 109 Results, 1953 The first Tidal Spat Trap was installed at Boothbay Harbor, Maine in May, 1953, to test its operation. Table I srows the number of bivalves caught in the sand filter during the first month. The trap caught over 10,000 bivaives per square foot of sand filter in the first 24 days. Of these, nearly a thousand were soft-shell clams. The ratio of mussels to soft-shell clams was about what we expected from plankton samples. Table I: Catch of Bivalves per Square Foot of Sand Filter. Mark I Tidal Spat Trap, Boothbay Harbor, Maine 1953 Soft Total Date No Days Clams Bivalves May 26 Trap installed. June } 8 =o= 2,000 June 12 16 648 6,372 June 18 22 952 10,639 June 18 Sand filter changed. June 19 1 73 LU7t On June 22 we transferred the spat trap from Boothbay Harbor to Robinhood Cove and attached it to the side of an oid sailing ship which had been beached there. This location proved to be less suitable than the wharf at the Boothbay Harbor Laboratory because of the large amount of silt in the water which tended to plug the sand filter. In spite of the plugging by silt, the trap continued to catch bivalve lar= vae in fairly good numbers as long as they were present in the water. The sand filters were taken from the spat trap at intervals and placed in running water at the laboratory. Periodically, the number of Juvenile clams and musseis in these trays were checked, We found that nearly ail of the mussels died after they were brought to the laboratory; whereas, many of the clams survived. By Octoher the soft clams present aoe in the tanks ranged from 2 to 9 millimeters in length which we con- sidered very good growth in comparison with naturally set juvenile clams. The spat trap was also tried as a plankton collector by re- placing the sand filter with a plankton net tray made from No. 18 triple extra heavy plankton netting. This proved to be quite success- ful in straining bivalve larvae from the water until the plankton net plugged. We found that the net would filter successfully for about two days which would represent four tides or about 700 gallons of water. It appears that the spat trap might be useful in some places as an intermittent plankton sampler since it filters all of the water which is taken during each incoming tide. -lll= Results, 1954 During the winter, two additional spat traps were designed ard built. The Mark Ii, or Maine model, is 2 feet square and 10 feet tall, to accommodate tidal ranges of 8 te 10 feet. The Mark III, or Rhode Island model, is 5 feet high and has a cross section of 2 fe feet. This trap is designed for tidal ranges of 4 to 5 feet. in June, i954, the three spat traps were put into operation. The Mark I, or original model, was piaced at Loves Cove on Southport Island, Maine which is th® area where we are now conducting most of our plankton studies concerning soft-shell clams. The Merk IZ spat trap was installed in Orr's Cove, Maine, which contains a large popu- lation of hard-sheli clams and is noted for its heavy sets. This cove was chosen as 8 typical area in Maine where hard=-shell clams are found. The Mark 137 spat trap was installed at Wickford, Rhode Island, on June 2, 1954, and placed in operation the following day. Mechanical difficulties with the check valves prevented its successful operation until June 10. Following this time the check valves and filters operat- ed successfully and Mya, Venus, and Mytilus larvae began to appear in the samples which were taken. Table Ii shows the catches of larvae and spat by the Leves Cove spat trap during June and July of 1954. Few larvae were present in the water during the period frem Jume 10 to July 2 and this accounts for the low numbers of muss¢is and clams that were obtained by the trap dur- ing this pericad. Following that time the number of clam larvae in the plankton increased which resulted in fairly good catches by the trap during the following week. Highest catch recorded to date was for the period from July 9 te 12 when over 400 soft clams were caught by each Square Toot of sand filter in the trap. in & commercial operation of a spat collecting Gevice such as this, the most productive period woula be during times like that from July 9 to 12 when large mumbers of mature lervae are present in the water. Parse Table II: Catch of Bivalves per Square Foot of Sand Filter. Mark I Tidal Spat Trap, Loves Cove, Maine, 1954. Total Date No Days Soft Clams Bivalves June 10 Trap installed. cially; 2 22 93 162 July 7 27 139 789 July 9 29 464 1,995 July 9 Sand filter changed. July 12 3 418 627 The best catch to date of the Mark II trap at Orr's Cove was during the period of July 15 to 19. In these four days each square foot of filter caught 1,044 Venus larvae and spat. The total catch of bivalves was 2,396 per square foot. The location of this trap appears to be very favorable since a strong current flows past it during the flooding tide. The Mark III spat trap at Wickford, Rhode Island, has twice the filter area of the Mark T and Mark II traps. Therefore catches of spat per square foot of filter are expected to be lower. Best results to date were obtained during the week before July 19 when each square foot of filter caught a total of 5,296 larvae of which 656 were Venus. At this time plankton counts averaged 172 Venus per hundred gallons of water. From July 19-22 the Wickford trap caught only 48 Venus per square foot of filter, but plankton samples averaged only 6 per hundred gallons during this period. It appears that the spat trap catches follow closely the abundance of larvae in the water. aliz= Conelusions 1. The principles involved in the design of the Tidal Spat Traps are sound. 2. Mechanical difficulties have been overcome. The only moving parts of the spat trap are the two check valves which have been found to operate satisfactorily. The sand filter retains larvae and spat and allows the water to pass through. No outside power source is required. 3. The results depend on the location, time, and presence of advanced or mature larvae. The location in Orr's Cove appears to be satisfactory; whereas, the location of the Mark I trap in Robinhood Cove last year was unsatisfactory because of the silt and sediment in the water. The Loves Cove spat trap this year may not be in the best location possible since it is in a cove where there is little circulat- ion of water. The maximum catches indicate that this device may have commercial potentialities for hard-shell clams. 4. Methods for holding the seed until it grows to a size when it can resist predators are now needed. This is the next project. We have tried holding the spat in the laboratory at Boothbay Harbor but have found that we would have insufficient room although the growth and survival appear to be satisfactory. Perhaps screened or fenced plots could be developed where the seed could be planted until it had reached a sufficient size. It may also be possible to find certain areas which are free from predators. Growth plots could be established in such areas and the seed transplanted to their final grounds after they had reached a large enough size to resist predators. 5-« We can visualize several applications of the Tidal Spat Trap when the details of its operation are perfected. It could be used in Rhode Island to obtain seed which could be planted along the Maine Coast during cold cycles when there was no natural reproduction in Maine. The spat trapsmight also supply seed for private hard clam farms. Biologists could use this as a research tool for evaluating the success of setting or for determining the amount of plankton in the water. 6. This description has been presented as a report on an applied research project which is now in progress. A complete evaluation of the Tidal Spat Trap will be given when the present tests are completed. male Re Literature Cited Dow, R. L., and D. BH. Waliace. 1951. A method for reducing winter mortalities of quahkogs (Venus mercenaria) in Maine waters. Maine Dept. Sea & Shore Fish. Res. Buil. No. 4: 32 pp. Glude, J. B. 1947. Observations on the Japanest clam fisheries. Wash. Sta. Dept. Fish. Ann. Bull. No. 47; 34=43. Glude, J., H. Spear, and D. Wallace, 1952. The hydraulic clam rake, a new method for gathering seed clams. Nat. Shell- fisheries Assoc. Conv. Add. 1952: 163-166. Tiller, Ro. Ha; d« B. Glude, and La D. Stringer. i952. Hard clam fishery of the Atlantic coast. Comm. Fish. Rev. 14(10): 1-25. “115 RECENT ADVANCES IN THE STUDIES OF THE STRUCTURE AND FORMATION OF THE SHELL OF CRASSOSTREA VIRGINICA Paul §. Galtsoff Fish and Wildlife Service, Woods Hole, Massachusetts Knowledge of the structure and formation of an oyster shell has not advanced far enough to enable us to give a complete story of this complex and extremely interesting problem. Like many other marine in- vertebrates, oysters extract calcium salts from sea water and deposit it in their shells. In this way millions of tons of calcium salts are taken annually from sea water and deposited in the living bodies of the animals. After their death they accumulate on the bottom. The calcareous material is partially redissolved, though the process is slow, depending on local conditions, chemical composition of shell’ material, and the amount of carbon dioxide in sea water. Oysters play an important role in the calcium cycle since their shells constitute about 90 percent of their total weight. Deposition and accumulation of calcareous sediments are the problems of great interest to oceanographers and geologists. The marine biologist, however, is concerned primarily with the biochemical reactions and morphogenetical processes which control the formation of shells. In recent years this problem has attracted the attention of a number of investigators who materially contributed to the understand- ing of the physiology cf calcification and shell formation in mollusks. Microscopic studies of the sections of an oyster shell show the presence of three distinct elements: (a) a very thin and poorly developed horny layer called periostracum. Its presence in the oyster ean be detected on a cross section of the mantle edge, or by examining the edge of the mantle of a living mollusk; (b) = prismatic layer con-= sisting of a series of calcite crystals arranged in a honeycomb pattern (Fig. 1). In Crassostrea virginica this layer is well developed only in the right valve; and Cel a calcite-ostracum layer which comprises the greatest part of the sheli material. It has a foliated structure made of flat sheets of calcite arranged either horizontally or at an angle to the surface of the shell. In many specimens of Crassostrea virginica, C. gigas, and Ostrea edulis, the structure of the shell is complicated by the pre- sence of irregular masses of soft and porous material (known as chalky deposits) embedded between the layers of hard material. They consist of loosely packed minute crystals of calcite. In exceptional eases, the chalky material covers the entire inner surface of the shell but it may be completely absent. About 50 percent of New England oysters in my coliection were free of chalky deposits. In the other half the deposits occupied only a part of the shell surface. -116= Prismatic layer at the edge of a growing Whole mount x 250. Ialfeas kg shell of Crassostrea virginica. Woods Hole, Mass. al Calcium salts of shell can be easily dissolved in dilute mineral acids or in chelating agents such as sodium versenate. The insoluble residue appears in the form of thin, homogenous sheets of organic material kept together like pages of & book. This sub- stance, discovered in 1855 by Fremy, is known as conchiolin. It is a scleroprotein, the structural formula of whick has not yet been determined, The elementary composition of conchiolin of Ostrea edulis, according to Schlossberger (1856), is as follows: H, 6.5%; C, 50.7%; N, 16,7%. Wetzel (1900) found that conchiolin contains 0.75 percent of sulfur and Halliburton (quoted from Schmidt, 1928) gives the follow- ing formula: C29» Hig» Ng» O77 The calcareous material of mollusk shells is laid either as aragonite or calcite. The shelis of edible oysters consist exclusively of calcite; on the other hand, the nacreous layer of the pearl oyster consists of aragonite. Taking advantage of the fact that both calcite and aragonite are present in the two distinct layers of shell of the fan oyster (Pinna) and of the pearl oyster (Pinctada), the French in- vestigators (Roche, Ranson, Eysseric=-Lafon, 1951) attempted to find whether theré is a difference in the organic material of the two layers of the same species, They found a distinct difference in the percentage of the two amino acids obtained from the conchiolin of the prismatic layer (calcite) and of the nacreous part (aragonite) of these mollusks. In the first one the content of tyrosine (11.6=17.0%) and of glycocolle (25~36%) was much higher than in the latter (tyrosine 2.8-6.0%; glycocolle 14.9-20 «8%) » Conechiolin from the sheils of Ostrea edulis was found to contain 3.2 percent tyrosine and 15.7 percent glycocolle. The content of other amino acids found in the shell of ©. edulis, namely, arginine, lysine, leucine, valine and methionine varied between O45 and 3.55 percent, These findings are interesting, but at present pro- vide no clue concerning the role of various amino acids in determining the minerological type of shell structure. To the naked eye and under the ordinary microscope the con- chiolin appears as amorphous, viscous, and transparent material, which hardens shortly after being deposited. By using electron microscope technique, Florkin of Liege University and his associates (Gregoire, Duchateau, and Florkin, 1950) have shown that the conchiolin of gastro- pods and lamellibranchs consist of a material laid as a fine network with many meshes of irregular shape and variable dimensions. This can be clearly seen in the photograph of conchiolin of abalone (Haliotis tuberculata) at magnification of 100,000 times. (Fig. 2). It is ob= vious that the organic membranes of molluscan sheils have rather com= plex structures which can be seen only with very high magnification. It is too early yet to speculate regarding the significance of this structure and its role in calcification. The question of the amount of conchiolin in the oyster shell was studied by several investigators. As early as 1817 Brandes and ~118- Fig. 2. Conchiolin sheet of the shell of abalone (Haliotis tuberculata) photographed after decalcification and ultra-sonic treatment. [Elec- tron microscope; phosphotungtic acid x 100,000. Courtesy of Dr. Ch. Gregoire, University of Liege, Belgium 1. 119 Bucholz estimated that the entire oyster shell consists of about 98.6 percent CaCO3 and 0.5 percent organic material. Schlossberger (1856) found 6.3 percent of organic matter in the prismatic layer of the oyster shell, but only from 0.8 to 2.2 percent in the calcite ostracum. According to Douville (1936), the albuminoid content of oyster shell is 4.8 percent. Determinations made for Korringa (1951) by Dr. A. Grijns gave similar results. It is interesting to note that in the latter tests the conchiolin content of the prismatic layer varied from 3.4 to 4.5 percent egainst the 0.5-0.6 percent in the calcite ostracum. My own observations made this year on Crassostrea virginica from Cape Cod, Narragansett Bay, and Long Island Sound show that in the 34 samples studied, the organic content of the calcite ostracum layer varied from 0.46 to 1.1 percent. There was no significant difference between the organic content of chalky deposits or the hard portions of calcite. Duplicate and triplicate tests of the hard portions of shell of the same specimen show that the conchiolin content is fairly constant, the difference not exceeding 0.2 percent. Among the diff- erent specimens, the conchiolin content of the entire shell, varied from 0.3 to 1.1 percent. The presence of chalky deposits and their structure was sub- ject to many speculations. Orton and Amirthalingam(1927) advanced a theory that chalky deposits occur in places where the mantle epi- thelium looses contacts with the shell. This theory received support from Ranson (1939-41) who, without making any additional study, stated that chalky deposits are formed wherever there is a local detachment of the mantle from the shell. Korringa finds no difference in the percentage of chalky deposits in the oysters placed with the cupped valves uppermost and in those which are kept in their normal position with cupped valves undermost. He assumes that the topmost mantle epithelium of the exnalant chamber “is certainly liable to sagging”. My observations on Crassostrea virginica show that the mantle epi- thelium adheres rather strongly to the inner sides of the valves and does not sag unless the mantle is pushed down by force. To determine whether local detachment of the mantle from the inner surface of the shell expedites the formation of shalky deposits, I made the following experiments: Small pieces of thin plastic, about 1 em.©, were bent in the shape of shallow cups and introduced between the mantle and the shell. in five oysters the concave side of the cup was facing the mantle, in another five the position of the cup was re- versed, i-.e., its concave side faced the shell. Oysters were kept for 55 days in running sea water in the laboratory. During this time they fed actively and there was considerable shell growth along the margin of the valves. After removal from the shells, the plastic cups were found to be covered with hard calcite deposit. There was not the slightest indication of the formation of chalky material. On the =-120- other hand, conspicous chaiky areas were formed along the marginal area of shell growth in places where the two valves were in close contact with each other (Fig. 3). It is clear from these observations that chalky deposits may be constructed by the mantle at any place on the surface of the shell and that the detachment of the mantle epithelium from the inner surface of the valve is not a factor in their formation. Korringe (1951) advances a theory that "chalky material is used by the oyster as a measure of economy and as a ‘eheap padding' in smoothing out the shell's interior and in creating the right shell shape to maintain its efficiency of function. The more the oyster shell attains a cupped shape, the more layers of 'chalky' material are deposited beyond the muscle to maintain the proper shell dimensions." He emphasizes the significance of the presence of chalky deposits be- ; yond the muscle scar and for reasons which he fails to explain disre- gards their presence in the other areas. The theory goes far beyond the experimental evidence and its teleological approach does not con- tribute to the understanding of the problem. Since so much emphasis was placed on the significance of chalky deposits in the area beyond the muscle scar, it was of interest to determine whether there is a definite tendency in the formation of chalky deposits in different parts of the shell. For this purpose the surface of the valve was arbitrarily divided into the four quad- rants shown in Figure }. Examination of several hundred oyster shells of Crassostrea virginica from various parts of the Atlantic Coast show- ed that chalky deposits were absent in 53.1 per cent of right valves and 47.9 per cent of the left valves. To have some idea of the extent of chalky deposits present, the following categories were established: i. from 1 to 25% of the area of a quadrant covered by chalky deposits; TI. from 26 to 50%; III. from 51 to 75%; and IV. from 76 to 100%. The results, given in Table I, show that the majority of oysters. in which chalky deposits of any size were found had less than 25 per eent of the total shell area covered with then. Table I. Occurrence and Extent of Areas of Chaiky Deposits in Crassostrea virginica (percentages); Exclusive of Oysters without Chalky Deposits Percent of shell Zi surface occupied by chalky deposits Left valve 4739 25.9 HEINE: 9.8 2.8 Right valve 52.1! 24.9 12,0 8.4 v5 ~l21- Fig. 3. Chalky deposits (white areas) at the edge of a rapidly growing shell of Crassostrea virginica and near the muscle scar. Woods Hole. 122 ALT CTR Fig. 4. Four quadrants of the shell area used in a study of the occurrence of chalky deposit in the values of adult Crassostrea virginica. 125 Chalky deposits of any size were almost uniformly distributed, 48 per cent occurring on the dorsal half (above the muscle scar) and 52 per cent on the ventral half of the shell. The percentage of Occurrence among the four quadrants (A, B, C, D) was also rather uni- form as can be seen from Table II. Table II. Occurrence of Chalky Deposits in the Four Quadrants of the Shell (percentages ) Areas A B C D Left valve 22,2 25.5 29.8 22.5 Right valve 25.2 23.0 22.9 28.8 Although there seems to be an equal chance for a chalky deposit of small or medium size to be found in any of the four quadrants of the shell, large deposits covering more than three quarters of the area of a quadrant are more frequently found in the dorsal half of the valves (quadrants A and B, Table III.) Table ITI. Frequency of Occurrence of Large Areas of Chalky Deposits (Category IV) within the Four Quadrants of the Valves (percentages ) Mir Sian (or Gta i a mies A B C D Percentage occurrence 33.3 Wien 916.8 8.6 (both valves) Studies of the distribution of chalky deposits in Crassostrea virginica do not support Korringa's suggestion that they are purposely formed by the oyster in order to maintain a constant distance between the shells in the posterior area opposite the exhalant chamber. It is well known that the valves of oysters differ in size and shape, the left valve usually being cupped and heavier than the flat and lighter right valve. Normally the oyster occupies a position with the right (flat) valve uppermost. The majority of Atlantic oysters, when placed on their left valves with the dorsal end away from the observer, show a curvature to the left. This indicates more rapid growth of the shell along the dorso-ventral axis, with a slight shift of the direction of growth to the left. In very young oysters which frequently appear to be round aos this tendency is less pronounced. Examining my collection of sheils, I found some oysters in which the growth along the dorso=-ventral axis was almost equal to the growth along the anterio-posterior direction. The width of these specimens was almost equal to their length and sometimes even exceeded it (Fig. 5). The growth of shell in any direction is an expression of a metabolic gradient along the given axis. The latter is not definitely fixed in the oyster but changes in response to environment. As a rule, oysters living on soft bottom or in very crowded condition on natural reefs tend to form long narrow shells while their growth in width is suppressed. There are, however, other, apparently intrinsic, factors which cause the change in the direction of growth probably not associated with environmental condi- tions. In several specimens collected in various parts of the coast the direction of growth shifted from left to right. This can be clear- ly seen in Figure 6 showing the two left shells of the oysters, one normal and the other with a distinct shift to the right. The "right handed" oysters were found in Texas, Chesapeake Bay, Narragansett Bay, and New Hampshire. There was no indication of the presence of any mechanical obstruction that might have interfered with normal growth. In every other respect the "right handed" oysters were normal and had the typically cupped left valves with well developed grooved beaks. The "right handed" specimens are comparable to the so-called inverted shells in which the typically left valve structures appear on the right side and vice versa. Inversion was found in several pelecypods (Lamy, 1930). In case of Crassostrea virginica, the inversion of growth does not affect the morphological structures but is expressed only in the shifting of the axis of growth. There is a difference in the physiology of the right and left valves of the oysters. The rate of calcification is significantly higher on the left side than on the right one. This can be easily noticed by examining the newly formed shells. The material secreted by the left mantle is thicker and heavier than the material deposited during the same time by the right mantle. For these observations, con- dueted since last April, I used adult oysters which had no recently deposited shell areas along the edges of the valves. They were placed in laboratory tanks supplied with running sea water. About two months later the pieces of the newly deposited shell on eaca valve were care= fully removed from the shell. After measuring their areas with plani- meter, they were dried at 55°C. and weighed. The results are summarized in Table IV. =125- Large Crassostrea virginica, from Hadley Harbor, Naushon Island near Woods Hole. IBaWee Dc 126 peasgttt* yes 1 <7 pod 2," oy or ginica from Maine (on the Two left valves of Crassostrea vir Pig. 6. left) and from Texas (on the right). Wat Table IV. Areas of New Growth and Rate of Deposition of Shell Material in Milligrams per Day per Square Centimeter during April -- June, 1954, Woods Hole, Mass. Area of Weight Ratio Number of rew shell er L‘R. Valve days under Oysters em= em.© (mg.) Ratio (mg.) _ observation Five-year old Narragansett Bay Left valve 5.80 156 2.8 55 days Right valve 5.16 59.3 aye 2.6 Adult Narragansett Bay Left valve ail: 123 £8 68 days Right valve et 19.9 0.3 6.2 Adult Narragansett Bay Left valve opal 74.2 1.09 68 days Right valve 8.8 P55 6.37 2.9 Two-year old New Hampshire Left valve 3.68 163.8 2.98 55 days Right valve 4.20 52.0 0x95 6 Old New Hampshire Left valve 6.83 alee 123 55 days Right valve Tees: 33.0 0.6 2 -128- In every case the amount cof calcified material deposited over a wiit of area was considerably greater on the left valve thar on the right one, the difference varying from 2.2 to 6.2 times. The ratio of calcification along the inner portion of the mantle may be studied by inserting small pieces of plastic or other material of known area and weight between the mantle and shell. In interpret- ing the results cne should consider, however, that the presence of a foreign body may produce pathological conditions which upset the nor- mal rate of calcification, The results obtained with this method show great variability. Observations made with 16 adult oysters at Woods Hole during the period of August 9-20, 1953, show that in 15 specimens the daily rate of shell depositimp=: square centimeter varied from 0.4 to 2.1 milligrams. One oyster deposited 14.2 milligrams in two days or 7.1 mg. per day. Data obtained this spring with the Narragansett Bay oysters kept in laboratory tanks for 68 days during the period of April-June show the variation from 0.1 to 0.79 mg. per day per square centimeter. im one oyster the rate of deposit under a plastic cup was 16.4 mg. per day. In some oysters the presence of plastic material produced pathological conditions which resulted in the formation of leathery capsules similar to the blisters frequently found in the area of the adductor muscle. These experiments are being continued using various techniques designed to minimize the effect of local stimulation of shell secreting tissues, French physiologist Monnip2ult (1939) apparently was the first one to point out the role played by phosphatase as a factor in the acceleration of shell formation. This enzyme igs probably an agent in the transfer of calcium. Later on, Bevelander and Benjer (1948) ap- parently unaware of Monnigault's work arrived at the same conclusion. Freeman and Wilber (1948) have demonstfated the presence of another enzyme, carbonic anhydrase, in the mantle tissues snd body fluids of some pelecypods and gastropods. While this enzyme may be important in she]1 formation in some species, its negligible activity in other mollusks suggests, however, that shell may be deposited in its absence. Physiological and histological studies on growth and regenera-= tion of shell of the oyster disclose the great complexity of the pro- cess. The role piayed by different parts of the mantle epithelimm in secreting conchiolin and in depositing calcium carbonate in the form of well arranged prisms of the prismatic layer or as densely packed lamellae of the calcite presents a stimulating problem of research. Observations made in my laboratory on isclated pieces of mantle and on the mantle edge after the removal of a portion of the shell show that clear, viscous, sometimes stringly conchiolin is secreted from the periostracal groove located between the outer (secretory) and the middle (sensory) folds of the mantle. ‘The sensory fold is capable of great extensibility and seems to serve as a temporary support for the viscous “19. econchiolin until the latter hardens. This conclusion is confirmed by the examination of the sections of mantle which clearly show that con- chiolin produced at the mantle edge originates in the periostracal groove. The results are in agreement with the observations made by other zoologists on the shells of QO. edulis and Anodonta. Mucus secreted by the mantle of the oyster contains a large number of blood cells, Their role in calcification is not yet under- stood. The newly deposited material contains minute granules which sometimes give a positive reaction for calcium with alizarin and other reagents used for identification of this metal. Unfortunately, none of the available color reactions for the detection of calcium are dependable for they often give negative results. Because of this technical difficulty, it has been impossible to trace the origin of the tiny granules known as calcosphaerites (Fig. 7) which appear in the conchiolin shortly after it is secreted by the mantle. The growth of these crystals and the formation of large crystalline units which eventually form the prismatic layer (Fig. 1) can be seen on the black and white enlargements of Kodachrome photographs made with the polariz- ing microscope (Figs. 8 and 9). In polarized light these crystals of calcite present a picture of great brilliance and beauty. The conditions under which the calcium carbonate forms the prisms of the prismatic layer or is deposited as a foliated structure of the calcite ostracum are not known. The problem presents an oppor- tunity for further research of calcification. In conclusion, I want to remark briefly on the source of calcium used in the formation of the shell of Crassostrea. in 1938, Robertson and Prentice pointed out that sea water is the source of calcium for building the tubes of an annelid worm, Pomotoceros trigueter. I in- dicated at that time (Galtsoff, 1938) that calcium salts required for the building of the shells of Ostrea (Crassostrea) virginica are pro- bably taken directly from the sea water. Experimental work by Jodrey (1953) using Ca). clearly shows that pieces of oyster mantle separated from other tissués are able to deposit calcium taken directly from sea water. She also shows that the rate of calcium turnover in the edges of the mantle is about twice as rapid as in the interior of the mantle. The use of radioactive isotopes gives a new tool for solving many pre- sent mysteries connected with the formation, maintenance, and growth of shells of oysters and other marine invertebrates. =130= Fig. 8. Calcite crystals found in conchiol- in 24-48 hours after it had been secreted. Black and white enlargement of a Kodachrome photograph taken in polarized light with the magnification Cnt »¢ 20> 132 Fig. 9. Large calcite crystals at the begin- ning of the formation of the prismatic layer. Black and white enlargement of a Kodachrome photograph taken in polarized light with the magnification of X 250. 133 Literature Cited Bevelander, G., and P. Benzer. 1948. Calcification in marine mollusks. Biol. Bull. 94: 176-183. Douville, H, 1936. le test des lamellibranches: sa formation dans l'Ostrea edulis. Compt. Rend. Acad. Se. Paris 203: 965-968. Freeman, J. A., and K. M. Wilber, 1948. Carbonic anhydrase in molluscs. Biol. Bull, 94: 55-59. Fremy, E. 1855. Recherches chimiques sur les os. Amn. de Chimie et Physique, 3rd. Ser. 43: 47-107. Galtsoff, P. S. 1938. Seurces of calcium for shell of Ostrea virginica. Nature 141: 922. Gregoire, Ch., Ch. Duchatesu, and M. Florkin. 1950. Structure, étudiee au microscope electronique, de nacres decalcifiees de mollusques. (Gasteropodes, Lamellibranches et Cephalo- podes). Arch. Intern. Physiol. 58: 117-120. Jodrey, Louise H. 1953. Studies on shell formation. TIT. Measure-= ment of calcium deposition in shell and calcium eee in -wantle tissue using the mantle-shell preparation and Ca Bull. 104: 398-07. Biol. Korringa, P. 1951. On the nature and function of "chalky" deposits in the shell of Ostrea edulis Linnaeus. Proc. Calif. Acad. Sei., 4th. Ser. 27(5): 133-158. Lamy, E. 1917. Coquilles senéstres chez les Lamellibranches. Bull. Mus. Nat. d'Histoire Naturelle 22: 489-93. Mannigault, P. 1939. Researches on the calcareous materials in mollusks; phosphatase and histochemical precipitation of cal- cium. Ann. Inst. Oceanogr. 18: 331-426. Orton, J. H., and C. Amirthalingam. 1927. Notes on shell-depositions in oysters. Jr. Mar. Biol. Assoc. U.K., N.S« Lt: 935-953. Ranson, Ga 1939-41. Les huitres et le calcaire. I. Formation et structure des "chambres crayeuses". Introduction a la revision du genre Pycnodonta F. de W. Bull. Mus. Nat. Hist. Nat. Paris (2) 11: 467-472, 12: 426-432, 13: 49-66. Roche, Jean, G. Ranson, and M. Eysseric-Lafon. 1951. Sur la com- position des scléroproteines des coquilles des mollusques (conchiolines). Comptes Rendus d. seances de la Soc. de Biol. 145: 1474-1477. Sih Schlossberger, J. 1856. Zur naheren Kenntniss der Muschelschalen, des Byssus und der Chitinfrage. Annalen Chem. u. Pharm. Diz 99120. Schmidt, W. J. 1928. Perlmutter und Perlen nebst einem Anhang uber Phauenstein. Die Rohstoffe der Tierreichs, hearusgeg. von F. Pax und W. Arndt 2: 122-160. Wetzel, G. 1900. Die organischen Substanzen der Schalen von Mytilus und Pinna. Hoppe-Seyler's Zeitschr. f. Physiol. Chem. 29: ~ 366-410. =135« ON THE RATE OF WATER PROPULSION BY THE BAY SCALLOP Walter A. Chipman Fish and Wildlife Service, Beaufort, North Carolina Studies of the movement of sea water through the mantle cavity by lamellibranch molluscs and the efficiency of the gills in removing particulate matter from this flow are of considerable interest to bio- logists concerned with investigations of the feeding activities of these bivalves. Only through a more complete understanding of the factors controlling the rate of water propulsion, the retention of particles by the gills, and the ingestion of filtered material can we approach problems in the nutritional physiology of these animals. The rate of water propulsion of a number of species of lamelli- branchs has been reported by investigators using either direct measure- ment techniques or indirect methods based on the reduction of the num- ber of particles in a suspension in which the animals have been placed. Considerable work has been done on the factors affecting the rate of pumping of oysters using direct measurements of the rate of flow. In many lamellibranchs direct measurements cannot be made and indirect methods have been widely used. Investigators have followed changes brought about by their experimental animals in suspensions of various inert materials and of living plankton cells. For the most part, the observations have been limited to those employing rather dense sus- pensions and to those in which the changes were quite great. With the availability of methods of labeling plankton cells with radioactive isotopes and the extremely accurate technique of measuring changes in cell numbers based on the detection of small amounts of radioactivity contained in such cells, it is advantageous to employ these methods in studies of the feeding and filtration rates of filter feeding invertebrates. At the Beaufort laboratory we have employed various species of plankton containing different radioactive elements in studies of the foods and feeding processes of various shellfish. The work which I wish to report today deals with use of radioactive plankton in the measurement of the rate of water pro- pulsion by the bay scallop, Pecten irradians Lamarck. Methods Observations were made on single scallops immersed in suspensions of plankton cells in sea water. The suspensions were always very ade- quately stirred. The volume of suspension used was varied for the size of the scallop immersed so as to allow satisfactory measurements of the decrease in suspended plankton cells within convenient measuring in- tervals and during satisfactory experimental times. -136- The plankton species to be used were grown in such manner ag to give rather dense cultures of rapidly dividing cells containing the desired amount of radioactivity. The cell population was measured using a haemocytometer and the radicactivity of a known number measured. By dilution of the culture, a suspension of the desired concentration was prepared. Knowing the radioactivity per cell, changes in the num- ber of cells in the suspension were easily and very accurately follow- ed by radioactivity measurements of small aliquots of the suspension. Tests were made to make certain that the radioactivity was due to the isotope within the cell and not to its presence in the water. In earlier work the cells of the aliquot were destroyed by acid and the radioactivity of the liquid measured. We now find it more convenient to filter the cells onto a millipore filter and to ascertain the radioactivity of the cells of the aliquot directly on the filter. Results On being immersed in an experimental phytoplankton suspension, the scallops opened almost immediately and soon started to filter the water through their gills. This resulted in a lowering of the cell content of the suspension. A semilogarithmic plot of the cell con- centrations measured at frequent intervals throughout the period of observation was made to visualize these changes. The rate of clear- ing varied, but followed the same pattern in nearly all of the tests. If one assumes that the conditions of the experimental arrange- ment were satisfactory, the straight line decrease in the logarithm of the cell concentration with time fits the mathematical equation for a constant rate of water filtration with complete removal of the sus- pended phytoplankton from the water filtered by the scallop. During this time of complete removal, the observed rate of decrease in cell concentration may represent the rate of water filtration of the scallop. Changes in the rate of clearing of the suspension can be interpreted as resulting from changes in the amount of water filtered. As the observations were continued, the decrease in phytoplankton became less and less. This resulted in a flattening of the curves. Actually, the number of cells in suspension gradually increased in instances where the ovservations were prolonged. There are a number of factors concerned with determining the rate of reduction of particles in suspensions in which the scallops were immersed. Different ones of these may affect the rate at differ- ent times, or more than one may be acting at one time. The flattening of the curves of our observations, therefore, may not have resulted from a change in the rate of water Tiltration. -137- The decrease in rate of removal was apparently related to time in the suspension. it was not related to cell concentration. Series of observations were made starting the scallops in concentrations at which there was a previous leveling off in the rate of removal. In each series there was rapid removal followed by a less rapid rate re- gardless of the starting cell population. Incomplete mixing of the filtered water with the unfiltered would result in a decrease in the observed rate of phytoplankton re- moval that would get less and less as the cbhservations were continued. Changes in the amount of stirring, however, did not alter the shape of the curves in our experiments. Although it is realized that it is virtually impossible to get the mixing required to fully meet the requirement of the mathematical expression giving a straight line de- crease, we believe that recirculation of filtered water through the scallop before mixing was not wholly responsible for the flattening of the curves. The decrease in the rate of removal refleéted in the flattening of the curves could be interpreted as a change in the rate of the water propulsion of the scallop. However, such a change was not apparent. The scallops were seen to be creating a considerable current of water even when there was no appreciable decrease in suspended plankton. It seems not unlikely that the changes in the rate of clearing of the suspension was a result of a changed efficiency in the filtering by the scallop, together with a return of cells previously entrapped in the mucus of the gills. If one assumes that the scallop was filtering the suspension efficiently with complete removal of the suspended particles, it is possible to calculate the rate of water propulsion by the application of the formula given by Jorgensen in 1943. ate (log conco - log conc; )M leevte Tat Undoubtedly the rates obtained are not absolute. They can only repre- sent an approximation since the original assumption made is not likely to be entirely correct. Escapement of plankton through the gills, if occurring as a fixed percentage of the concentration in the suspension, would not change the nature of the curve but would alter the slope. The measured pumping rate may actually be lower than the true rate. Individual scallops in observations made at different times differed in their rate of filtration, but under like conditions of test the rates were reasonably uniform, The average rates _were the same for experiments using either Nitzschia or Chlamydomonas cells =136= for the suspended material. They did not appear to be related to concentration. As the scallops increased in size, their rate of water filtration increased. In June or early July the small scallops averaged about three liters per hour. Later during the summer they filtered increasing amounts as they grew in size. By fall they were pumping an average of nearly 15 liters per hour. The maximum rate observed was 25.4 liters per hour for a scallop measuring 65 milli- meters in length. The average rate of water filtration per gram of tissue was greater for the small scallops than it was for the larger. Summary In conelusion I should say that the use of radioactive plankton cells in studies of the feeding activities of shellfish is particularly valuable. In using such techniques to measure the rate of water filtra- tion by the bay scallop, we have found that the scallop has a high rate of water filtration, probably correlated with its active mode of life. The work reported indicates that the filtering activities and feeding of lamellibranch molluscs have many phases not yet fully understood, but the application of radioisotope techniques offers opportunity to investigate a number of these more easily. -139- GROWTH STUDIES IN THE QUAHOG VENUS MERCENARTA Alton H. Gustafson Bowdoin College, Brunswick, Maine, and the Maine Department of Sea and Shore Fisheries Introduction Venus mercenaria, the quahog or hard-shell clam, is a bivalve molluse of considerable commercial importance widely distributed along the Atlantic coast. It grows from between the tide levels to depths of at least fifty feet. Brunswick, Maine, is the center of commercial digging operations in the state. Here the receding tides expose great areas known as mud flats. Most of the digging is done between the mid tide and low tide levels. Recognizing the need for accurate information about the ecolo- gical factors responsible for the occurrence, distribution, and growth of the organism, the Maine Department of Sea and Shore Fisheries is engaged in a series of studies designed to yield data of value both scientifically and for the promotion of the fishery. The program in- cludes investigations of (a) the early life stages: reproduction, planktonic existence, and setting habits, (b) the fate of the heavy set of 1952 in certain areas, (c) the growth of natural populations, (d) comparative growth studies of populations of known sizes planted under differing conditions, and (e) the factors which presumably in- fluence growth. In spite of the abundance, availability, and commercial im- portance both actual and potential, surprisingly few studies have been made of either the ecology or the growth of the organism. Pub- lished data by Belding (1912), Chestnut (1952), Haskin (1949), Kellogg (1903), and Pratt (1953) and unpublished data by Carriker and Kerswill have yielded information of great value. However, they should be re- garded as but the first steps in acquiring the data necessary for either a real understanding of the organism or developing a sound management program for the fishery. In Maine, studies by Dow and Wallace (1951) on winter mortalities and some aspects of growth have pointed up the scarcity of our knowledge of the factors of importance in growth and survival. The material here presented is in the nature of a preliminary and progress report. it deais chiefly with a comparison of growth of populations of several sizes planted under differing conditions in several localities, an analysis of the annual increment, and some comparisons with conditions reported in other geographic areas. -140- Methods From time to time huge sets of Venus have occurred in several of our coves and we thus have been given an opportunity to obtain specimens of almost any size in any quantity for experimental pur- poses. The specimens are gathered, measured to the nearest milli- meter in length, given an identifying mark with a durable ink, and planted in chosen areas legally closed to digging. They have been planted in bottoms of different types, at various tide levels, and in several concentrations. This report deals with a small fraction of the data accumulated in 1952-53 from plantings of over 13,000 specimens. Comparison of Growth at Simmons Reservation and Avery Cove. At Simmons' Reservation 200 individuals of each of the following sizes were planted in June, 1952, and harvested exactly one year later, and replanted for further study. Table I shows the pertinent data: Table I Original length Actual increase % increase alley) 1760 100 27 16513 (eas SHE 14.85 t@) Dt 12.37 22 Wi 6.52 8.5 Graph I shows that there is a fairly uniform decrease in the percentage of increment as the initial size increases. From the graph one may make an estimate of the probable growth increment of specimens of any size. For example, 30 millimeter specimens might be expected to show a 60 per cent increase in size in a year or to add 18 millimeters in the year. Table II and Graph II show the results of a similar planting at Avery Cove. Unfortunately, vandals removed the larger sizes so we cannot complete the curve. it would have been interesting to be able to determine whether or not the curve changed in a manner similar to that shown for Simmons. It is evident that growth is not as great-in Avery as at Simmons. -141- INCREASE PER CENT 100 90 80 70 (100) GRAPH | VENUS MERCENARIA GROWTH IN SIMMONS' RESERVATION, FREEPORT, MAINE, (72.5) JUNE, 1952—JUNE, 1953 10 20 30 40 50 60 70 80 ORIGINAL LENGTH IN MM. 142 PER CENT INCREASE 80 on. GRAPH 2 VENUS MERCENARIA GROWTH IN AVERY COVE, HARP MAINE. oe RPSWELL, MAIN JUNE, 1952- JUNE, 1953 10 20 30 40 ORIGINAL LENGTH IN MM. 143 Table II - Initial length Actual increase Percentage increase eee 18 1346 1305 28 i, Wal 52.5 38 11.97 31.5 Here, also, we may use the curve to determine the probability of size increase for specimens of intermediate sizes. Specimens originally 30 millimeters in length would be expected to increase 48 per cent or 14.4 millimeters in the year. Curves plotted for several other coves resemble the curve from Simmons' Reservation. Comparison of Specimens of Several Sizes Grown in Different Local Areas. Table III compares a number of plantings of different sizes in different areas. The fact that different plantings differ in respect to their productivity is not surprising. Not only have we many ana- logues in many groups of plants and animals but data from other studies on quahogs indicates the same things. These investigations do give us data on a wider range of sizes than previously known. A comparison may permit us to narrow down the factors responsible for the differences and lead us to a knowledge which may be put to practical use. It will be noticed that in a general way the annual percentage increment decreases with an increase in the original size as was the ease at both Avery and Simmons and-indicates that this situation is quite general. The actual size increase will be useful in comparing data from other geographic areas. The data from Simmons has been underlined to emphasize the great- er growth here than in any other plantings no matter what the original size of the specimens. -144- Table r eit One Year Growth Study = 1952-53 Location Original Year's Percent size growth increase (length in mn.) Shelldrake 16 16.06 100 Simmons 17 sil 100 Avery 18 13.61 75.5 Stetsons Mills 19 13.83 72.8 Dam 20 10.75 oh. Simmons 21-25 16.58 72.5 Shelldrake 26 15.76 60.5 Simmons 27 Toads Avery 28 ew al 52.5 Webbs 2630 8.63 31 Stetsons Mills 29 14.52 51 Dam 30 12.5 AT Shelldrake 36 13333 34 Simmons 37 14.85 40 Avery 38 IAL i 31.5 Diamond 36-0 1HGNere 29.4 Stetsons Mills 39 1636 29 Dam ho 10.14 25 Avery yp-45 Oe St 2h Simmons 56-57 12.37 21.9 Diamond Salt SEE 16 Simmons 76-80 6.52 S55 Stetsons Mills 79 4.55 Breil Comparison with other Geographic Areas. Without discussing the situation in detail those familiar with growth data in Venus may recognize that the annual increment in our beds in general is considerably higher than that reported by Kerswill for Prince Edward Island and equal to or surpasses that published by Kellogg (1903) and Belding (1912) for Massachusetts. =Ly5— It may be of interest to make a closer comparison of our data and that shown in the recent report by Pratt (1953) from Rhode Island for a second comparison will also be made in comparing seasonal data. Pratt planted specimens ranging in size from 20-50 with an average size of 30 millimeters. Unfortunately, the paper does not give a detailed analysis of the results by sizes. We can, however, compare our 30 millimeter size with his average size. His specimens planted in sand show a mean annual increment of 16.1 millimeters which is 24 per cent higher than the mean of 12.9 for those planted in mud. Possibly it should be mentioned in passing that Kerswill found no difference in growth in the two types of bottom. All our specimens were planted in mud. The only planting we had of specimens exactly 30 millimeters in length was at Dam Cove and here the increase was 12.5 millimeters or roughly the same as Pratt's mud specimens. Our 28 and 29 millimeter specimens planted at Stetsons Mills and at Avery Cove showed increases of 14.52 and 14.71 millimeters respectively. We have already called attention to the probable increase in 30 millimeter specimens at Avery Cove and Simmons. The 14.4 increase at Avery represents a 48 per cent increase and the 18 millimeter in- crease at Simmons represents a 60 per cent increase. Pratt's figures translated to percentage is 43 per cent. One may call attention to the fact that the increase in mud at Simmons is actually greater than the increase in sand in Rhode Island. These comparisons indicate that the Brunswick area is a very favorable one for the growth of quahogs. It seems probable that Simmons' Reservation is one of the most favorable areas for growth yet known along the entire Atlantic Coast although we hasten to add that we have not made a complete check with the figures from New Jersey. The data from the New Jersey experiments is given in terms of weight rather than length and we have not yet had time to convert all the New Jersey figures to length. Seasonal growth analysis. Tables IV and V which follow summarize certain observations made on additional beds planted at Simmons' Reservation and Avery Cove. No other study has followed seasonal growth so closely for an entire year. The Simmons bed was planted on June 12, 1952, with specimens whose initial size was 21-25 millimeters. It was made up of twelve sections, one to be removed each month for measuring and replanting. Each section consisted of five square feet. Two square feet were planted in a concentration of ten per square foot, two with a con- centration of twenty per square foot, and the fifth contained fifty -146- specimens. At the end of the year 91 per cent of those planted had been recovered. As nearly as could be determined the concentrations seemed to have no marked effect and have been ignored in the tables. After a few months it was evident that specimens of this size move about somewhat, as has been shown by Chestnut (1952), and this inter- fered with attempts at measuring sharp concentrations. Table IV shows the date of removal, the duration of each sectional planting in days, the number removed, and the growth data. The fourth column shows the actual increase in length at the end of each interval (approximately a month). The final figure in the column thus represents the increase for the year. The fifth column shows the percentage increase in a cumulative series. The final figure in the column shows that specimens of this size had increased 72.47 per cent in the year. The sixth column shows the increment in millimeters for each monthly interval and the final column translates this into the percentage of the annual increment. It is evident that growth ceased by the first of December and probably resumed in late March or early April. “4 Table IV Seasonal Growth Simmons' Reservation, Freeport, Maine Planted: June 12, 1952 Original length: 21-25 mm. Removed Duration No. Gain %Gain Gain in %of year's (days) (mm. ) (gain) interval growth (Original length) 7/9/52 27 107 2.5 alee (al; 2.5 1532 8/7/52 56 107 677. 30.00 4.23 25.51 9/5/52 8 100 9.20 40.00 2.43 14.05 10/2/52 111 110 12.45 52.10 3425 19.60 10/31/52 140 108 13.79 57.33 1.34 8.08 12/1/52 wae 110 14.58 66.40 «79 4.76 1/20/53 221 104 14.08 60.21 - - 2/13/53 2h5 104 14.37 63.30 - - 3/13/53 273 107 14.47 62.29 ~ - 4/12/53 303 101 15.17 67.78 259 3.56 5/16/53 337 72 15471 69.18 54 3.26 6/13/53 366, 160), 16.58 72.47 .87 5.25 Similar data is shown in Table V for a bed planted in the same manner at Avery's Cove. Here the initial size of the specimen was -147- 41-45 millimeters. The actual increment was 10.31 millimeters which is, of course, considerably less than that attained by the population from Simmons' Reservation. This is a percentage increase of 2). Specimens of this original size would have grown 30 per cent in Simmons' Reservation, if we may judge by consulting the curve for Sinmmons'. Table V Seasonal Growth Avery Cove, Harpswell, Maine Planted: June 18, 1952 Original length: 41-45 mm. Removed Duration No. Gain % Gain Gain in % of year's (days ) (mm. ) (gain) interval growth (Original length) 7/10/52 22 10 IRE 2.97 i IST 8/6/52 te) 108 3.67 8.34 2.39 23.81 9/3/52 TT 105 6.01 13.36 2.34 22.70 10/1/52 104 98 eG 20.40 2.36 22.70 10/30/52 133 105 8.87 21,20 -50 4.85 11/29/52 163 110 8.81 20.37 = - 1/20/53 215 103 8.46 20.00 - - 2/12/53 239 101 8.47 19.88 = ~ 3/20/53 275 4g 8.03 18.90 = = April - - - - ~ - May - = ~ ~ - - 6/19/53 366 11g 10.37 24.00 1.44 13.96 Comparisons with other Geegraphic Regions by Season of the Year. Table VI shows a comparison of the seasonal growth of quahogs in Rhode Island taken from Pratt's paper with the closest correspond- ing dates taken from my plantings at Simmons' Reservation and Avery Cove. The Rhode Island specimens planted in sand showed their greatest period of growth early in the year and continued growing for a longer period in the fall than those planted in mud. The peak of growth for specimens planted in the mud was between mid June and mid July when nearly half the year's growth was added. The percentages for Avery and Simmons' are quite consistent. In both cases growth was slow until mid July. The period of greatest growth was between mid July and mid September. Growth continued much more actively in the fall than in the ease of the Rhode Island specimens. ~148- Perhaps it should be added that our results were consistent with those obtained by Belding in Massachusetts and Kerswill in Prince Edward Island. Table VI Analysis of Seasonal Growth Percentage of Total Year's Growth Rhode Island Maine Sand Mud Avery Simmons Noy. G4June 16 35 °«—C Ds COR ee, Lee June 18-July 16 20 47 15 12 June 12-July 9 July 16-Sept. 14 27 22 39.5 46 July 9-Sept. 3 Sept. l4-Nov. 27 16 2 32.5 27.5 Sept. 3-Dec. 1 Summary 1. Specific data has been accumulated for the monthly and yearly growth of quahogs planted in Maine waters both in terms of actual increase and percentage increase for populations of planted specimens of a variety of sizes grown in several localities. 2. Curves drawn from this data seem to enable us to predict the pro- bable size increments of specimens of other sizes. 3. The percentage of increase diminishes with the increase in size of the specimens. 4. The actual monthly and annual increments differ in different localities. 5. A comparison with other geographic areas indicates that growth in Maine waters equals or exceeds growth rates in other areas. 6. The seasonal growth rate is consistent with that in Massachusetts and in Prince Edward Island. The major portion of the growth occurs considerably later in the season in these areas than it does in Rhode Island. =149- Literature Cited Belding, D. L. 1912. A report upon the quahaug and oyster fisher- ies of Massachusetts. Boston: Wright and Potter Printing Company, 134 pp. Chestnut, A. F. 1952. Growth rates and movement of hard clams, Venus mercenaria. Proc. Gulf & Carib. Fish. Inst. 4th annual session; 49-59. Dow, R. L. and D. EK. Wallace. 1951. A method of reducing winter mortalities of quahogs (Venus mercenaria) in Maine waters. Maine Dept. Sea & Shore Fish. Bull. 4:3-32. Haskin, H. H. 1949. Growth studies on the quahaug, Venus mercenaria. Nat. Shellfisheries Assoc. Conv. Add. 1949 ~ 67-75. Kellogg, J. L. 1903. Feeding habits and growth of Venus mercenaria. Ns. ¥. Sta. Mus. 10 (71): 3-28. Pratt, D.« M. 1953. Abundance and growth of Venus mercenaria and Callocardia morrhuana in relation to the character of the bottom sediments. Sears Found. : Jr. Mar. Res. 12:60-74. -150- ? ee angie: i= eee be prem ay a Atay “piece sit ek wien: eT a * aiita’ Une 7 ie r Saar ; Ah co a es i _ 7 -- _ = - pio ae ee ee eo) | ee —— - COPY + : ~ : a 7 & 7 a _ = . : nr vets yo a @ aac F ua oral” % v on - _ Saran wet +) SR 11) Oe a 7 es rite st 7 ye - ‘ iia op Ve erm etahre bina! ate oy har ae < “ete Ve Wee? Gre Mes Slane ; : aon an) A FUNGUS DISEASE IN BIVALVE LARVAE H. C. Davis and V. L. Loosanoff U. S. Fish and Wildlife Service, Milford, Connecticut The method for growing bivalve larvae as now practiced at the U. S. Fish and Wildlife Service Laboratory, Miiford, Connecticut, has offered many new possibiiities for studying the behavior and physiological and ecological requirements of these organisms (Loosa- noff, 1945; Loosanoff and Davis, 1950). Using this method larvae of 16 species of bivaives have been successfully cultured through meta- morphosis. Of these, however, the larvae of the common clam, Venus mercenaria, and of the American oyster, Crassostrea virginica, have received the most attention (Loosanoff, Miller, and Smith, 1951; Davis, 1953). Because of their hardiness and simpler dietary requirements clam larvae are especially suitable for experimental work and their cultures have been reared through metamorphosis as a matter of routine. On several occasions, however, some cultures of an experimental series would show a heavy mortality unrelated to the experimental treatment. In the spring of 1953, while examining such a culture of clam larvae, we noticed an organism, which was tentatively identified as a fungus, in many of the dead or dying larvae. We later observed the same or a related fungus in larvae of C. virginica, Venus mortoni, and the hybrids of V. mortoni x V. mercenaria and V. mercenaria x V. mortoni . In both clam and oyster larvae the fungus may develop as a thallus of limited extent occupying a relatively small part of the larva so that its form and structure can readily be seen (Fig. l, A, B, C). In contrast, in many of the specimens of invaded clam larvae there is a much more extensive growth, the fungus occupying a large proportion of the space within the larval shell with its lobes or segments so densely crowded together in a radiate arrangement that the details of thailus organization cannot readily be discerned (Fig. 1, D). It has not yet been determined whether this difference in thallus character indicates that the simpler, less extensive growth represents a younger phase which may later develop into the more ela- borate one or signifies that two different species of fungi are con- cerned. With either type of thallus as the fungus develops, the seg- ments mature into sporangia, eacn with an exit tube extending to the exterior (Fig. 1, D). The zoospores, which develop fully and become motile within the sporangium, emerge through these tubes into the surrounding water. In some cases these tubes appear to become excessive- ly = wlleani and extend for some distance beyond the larval shell (Fig. i Bi) =151- Fig. 1. Larvae infested with fungus. A and B, young oyster larvae; C, young clam larva; D, old clam larva; E, young clam larva. Larvae A, B, C, and E are stained with Neutral Red; larva D, with Cotton Blue. Magnification 190 times. 152 No sexual stage of the fungus, nor any resting spores or other resistant bodies have yet been observed. The fungus is transmitted by the zoospores which emerge through the tip of the exist tube. These zoospores are released in an intermittent stream and are frequently seen for a short time in a swarm near the edge of the shells of the larva from which they have escaped, but soon disperse throughout the water. Theoretically densely crowded larval cultures should present a better opportunity for development of fungus epidemics than would field conditions be- cause the larvae are so crowded that it must be almost imoossible for them to avoid taking in some of the zoospores released into the water by infected larvae. The presence of fungus in our larval cultures usually was of endemic nature, i.e., there were a few infected larvae in many of the cultures. Only in a few instances did the disease acquire epidemic proportions involving the majority of the larvae and, within two to four days, killing almost the entire population. During the course of an epidemic many larvae were observed in various stages of disintegration with the fungus quite apparent inside the shell. Usually several species of flagellates and ci- liates invaded the dead or dying larvae to feed upon their tissues leaving, within a short time, empty shells with practically no trace of the fungus. Rapid disappearance of the tissues made it very easy, in certain instances, to overlook the fungus which caused the mor- tality. The zoospores are very small and early hyphae are not readily visible. It may be significant that even in those cultures in which mortality was especially severe there were usually some larvae that lived through the epidemic without contracting the dis- ease. The presence of these individuals raises the question as to whether some larvae are immune to the fungus. Larvae of all ages, from the very early free swimming stage to those ready to metamorphose, have been found parasitized by the fungus. Also on several occasions we have observed juvenile clams, just after metamorphosis, that were heavily infested and releasing fungus zoospores. At present we do not know the optimum salinity or temperature for the growth of the fungus nor the limits it can withstand. The larval cultures in which the fungus has been observed have all been in our normal sea water which has a salinity of about 27.0 parts per thousand, and have been kept at constant temperatures between 19.0 and 27.0°C. Wor do we know whether some infected larvae may survive the infection, recover from it and proceed with normal development. In a few instances we have observed the fungus in larvae that were still capable of swimming about. It seems, however, that the majority of the larvae drop to the bottom, stop growing, and die soon after they become infected. -153- Dr. Weston and Mr. Martin cf the Biological Laboratories, Harvard University have tentatively identified the fungus as Sirolpidium sp. among the lower members of the biflagellate series of Phycomycetes. Although its thallus is similar to Lagenidium calinectes, which is known to cause similar destructive epidemics among the larvae of the blue crab in Chesapeake Bay (Couch, 1942), the maturation and release of zoospores are completely different and affiliate our fungus most closely with the genus Sirolpidium. The two known species within the genus Sirolpidium, however, while both parasitic and both marine, have only been reported on such algae as Bryopsis and Ceramium and have never been found on animal hosts. The indications are, therefore, that these fungi attacking clam and oyster larvae represent one or perhaps even two new species. We have found that if living larvae are placed in a weak solu- tion of Neutral Red in sea water, the fungus acquires a distinct reddish color and becomes easily distinguishabie. The method is promising not only as a practical way to demonstrate the presence of the fungus, but also as a help in detecting its first appearance. By "tagging" the infected larvae it may permit us to determine how long fungus infected larvae can survive, whether they grow, and answer other pertinent questions arising in the course of an epidemic. Although we have not yet been able to obtain and test the new fungi-static antibiotic oligomycin, isolated by Dr. Elizabeth McCoy et al. at the University of Wisconsin, we have attempted to find other methods for controlling this fungus. Since it remains endemic in many of our cultures, or may disappear after killing only a small percentage of the larvae, it is difficult to be certain that any given treatment is effective in reducing the death rate from fungus. However, as the motile zoospore stage is presumably the most susceptible to poisons, we have concluded that a treatment is ineffective if motile zoospores are found following the treatment. By this criterion the fungicidal stains Fast Green and Malachite Green were found to be ineffective both when the larvae were dipped in the stain for periods up to ten minutes in concentrations as great as one part of stain to 15,000 parts of sea water and when the stain was added to the culture jars in concentrations up to one part per million. Bursoline, a fungicide developed for the Pacific Theater of World War II, was also found to be ineffective. Copper sulfate was ineffective in killing the zoospores even when added to cultures in concentrations of one part per 100,000. Many clam larvae survived six days of treatment in this high concentration of copper sulfate although they remained relatively inactive during this period. In contrast, oyster larvae were killed by concentrations of copper sulfate as low as one part per million and the culture jars in which copper sulfate had been used required several washings before they lost their toxicity for oyster larvae. -154- Ultraviolet irradiation of larval cultures was ineffective in doses up to four hours per day. Fluorescent sunlamp irradiation up to seven hours per day did not rid the cultures of the fungus but continuous irradiation killed the clam larvae and appeared to steri- lize a culture killing everything including bacteria. We believe this is the first report of a fungus parasite in bivalve larvae, and only a few parasitic fungi have been reported in marine animals. This discovery may indicate, therefore, that para- Sites and diseases of bivalve larvae may be more destructive in nature than heretofore thought. It is also possible that this fungus may be responsible for the heavy mortalities sometimes observed in nature among clam and oyster set, for it has killed almost every young clam in one of our laboratory trays that contained several thousand recently metamorphosed clams. Moreover, there is the possi- bility that such an epidemic among bivalve larvae in nature could upset the balance of events in the food chain and thus affect other species. If we can develop a method for control of this fungus under laboratory conditions, the artificial cultivation of certain bivalves may soon be commercially feasible. We wish to express our thanks to our colleague, Mr. C. A. Nome jko, for making the photographs for this article and to Miss Rita Riccio for her assistance in editing and preparing the manu- SCript.« -155- Literature Cited Couch, J. N. 1942. A new fungus on crab eggs. Jr. Elisha Mitchell Sei. Soc. 58: 158-162. Davis, H. C. 1953. On food and feeding of larvae of the American oyster, C. virginica. Biol. Bull. 104: 334-350. Loosanoff, V. L. 1945. Precocious gonad development in oysters in- duced in mid winter by high temperature. Science 102: 124-125. Loosanoff, V. L. and H. C. Davis. 1950. Conditioning V. mercenaria for spawning in winter and breeding its larvae in the laboratory. - Biol. Bull. 98: 60-65. Loosanoff, V. L., W. S. Miller, and P. B. Smith. 1951. Growth and setting of larvae of Venus mercenaria in relation to temperature. Jr. Mar. Res. 10: 59-81. -156- NOTES ON FUNGUS PARASITES OF BIVALVE MOLLUSKS IN CHESAPEAKE BAY Jay D. Andrews Virginia Fisheries Laboratory, Gloucester Point Note 1. Discovery of Fungus Infections in Numerous Bivalve Species. My hobby is collecting the mollusks of Chesapeake Bay. Having placed a few specimens in museums, and having made a check list (no new species yet) with appended distribution records, I found my hobby less stimulating than my research. But then my research had taken a turn which opened up new and inviting fields of discovery. First came a devastating mortality of oysters in the Rappahannock River, for which no explanation has been found. Then Mackin et @l. (1950) discovered the fungus disease of oysters, Dermocystidium marinum. But not until Ray (1952) developed the thioglycollate culture technique for easy detection of the fungus did we seriously begin to study oyster mortalities and their causes in Virginia (Hewa t+ and Andrews, 1954). For some time Ray and Mackin searched among the invertebrate associates of oysters for alternate hosts, only to find that infection was easily accomplished directly from one oyster to another through water-borne spores (J. G. Mackin, Personal Communication). Since other bivalve mollusks would not be suspected as alternate hosts for an oyster disease, little effort was made to check them. With this background, we at the Virginia Fisheries Laboratory were surprised in August, 1953, to find the meat of a dead clam, Venus mercenaria, infected with a D. marinum-like fungus. During the fall and winter of 1953-54, eee 1 species of bivalve mollusks collected near Gloucester Point, Virginia, were found infected with similar fungi (Table I). None of the fungus parasites has been identified except the one causing a mycosis in oysters. How many species of fungi are in- volved? Can spores from one host species infect individuals of other species? And of most immediate importance, how many bivalve species will serve as host to the oyster parasite? Very early it was noticed that infections in some bivalve mollusks differed from infections in oysters in two ways: (aoe several host species 100 per cent infections have been found for groups of 25 animals. Infections in live oysters have never exceeded 80 per cent. (2) Nearly all infections of bivalve mollusks other than oysters have been "light" whereas most groups of oysters with a high percentage of infection show some “moderate" and "heavy" infections indicating Contribution from the Virginia Fisheries Laboratory, No. 54. -15'7= that the disease is becoming worse in some individuals. The high incidence and low intensity of fungus parasites in certain bivalve mollusks may indicate greater tolerance by the hosts and a lower level of lethality than exists in oysters. Morphological differences have been noted in the cultured para- sites of various bivalve mollusks but these are not understood at present. Now hobby has become inextricably entangled with research. Happily, knowledge of bivalve species and their distribution is an asset to studies of fungus parasites of mollusks. Table I Occurrence of Dermocystidium-like Fungi in Bivalve Mollusks Species in which fungus has been found: Mercenaria mercenaria Linné Mya arenaria Linn Macoma balthica Linné Macoma phenax Dall Macoma tenta Say Mulinia lateralis Say Anomia simplex Orbigny Tagelus plebeius Solander Anadara transversa Say Laevicardium mortoni Conrad Ensis minor Dall Lyonsia hyalina Conrad Hard-shell Clam Soft-shell Clam Little Round Clam Tenta Macoma Dwarf Surf Clam Jingle Shell Stout Tagelus Transverse Ark Morton's Cockle Razor Clam Glassy Lyonsia Species in which fungus has not been found: Solemya velum Say Bankia gouldi Bartsch Volsella demissa Dillwyn Brachidontes recurvus Rafinesque Atlantic Awning Shell Gould's Shipworm Ribbed Mussel Hooked Mussel Note 2. The Disappearance of Fungus Infections in Late Winter and Spring. According to the thioglycollate test (Ray, 1952), D. marinum almost disappears from live oysters in Chesapeake Bay during late winter and spring (March, April, and early May). Based on samples of 25 oysters, the disease apparently disappeared completely by March in oysters that had been 80 per cent infected in November. Despite the apparent absence of the disease in late spring, oysters which have once had infections develop earlier and greater mortali- ties the following summer than oysters transplanted from areas where infections never occur. Also, oysters once infected, but testing negative in late spring, will develop the disease in areas where the fungus is not present. This suggests that latent infections, not detected by the thioglycollate method, are present in these oysters throughout the winter and spring. Apparently Chesapeake winters may not be quite long and severe enough to eliminate in- fections from all oysters. The possible rele of other bivalves as sources of infective material for the oyster disease must not be overlooked. Sketchy records suggest that fungus infections in the other bivalves also disappear in late winter except in Macoma balthica and Anadara trans= versa. Note 3. Racial Differences in Susceptibility to D. marinun. Dermocystidium is a fascinating disease: it resembles a human disease called Blastomycosis in that nearly all organs and tissues are attacked. This makes it easy to study: almost any piece of a dead or live oyster can be cultured with reasonable ex=- pectation of making a correct diagnosis of infection. Dermocystidium is a deadly disease! We are continually as- tounded at its scope. From 80 to 85 per cent of all our dead oysters from trays show serious infections of the fungus. Only young oysters under one year of age escape the disease. Excluding predation and adverse physical conditions such as too much silting, the disease appears to be the dominant cause of oyster deaths in lower Chesapeake Bay and the lower areas of the major tributaries in Virginia. Worst of all for the oysterman, there is as yet little evidence of resistance to the disease. Six year old oysters in trays at Gloucester Point are still dying at about the same rate and with the same degree of fungus infection as they did three years ago. -159= One ray of hope came out of last summer's experience with tray grown oysters originating outside Chesapeake Bay. It was noticed that one year old oysters from Seaside of Virginia were dying at a rate comparable to that of older Chesapeake Bay oysters. Deaths among yearlings are usually very light. Furthermore, a tray of two year old oysters from South Carolina was not following the pattern for oysters of that age: mortality was low and fungus infected oysters were few and late in appearing. Thioglycollate tests of live yearling oysters from South Caro- lina, Chesapeake Bay, and Seaside of Virginia, all grown in trays at Gloucester Point, revealed that the Seaside oysters were indeed more heavily infected than the others (Table II). Among two year olds, South Carolina oysters had fewer and lighter infections than Chesa- peake Bay oysters. A further comparison of various age oysters from the three sources (Table III) shows that both yearling and three to four year old oysters from Seaside had much higher mortalities than Chesapeake Bay oysters; and South Carolina two year olds had much lower mortality than native oysters. Regardless of age, nearly all groups from which a considerable number of gapers were recovered showed about 90 per cent infection of the gapers with D. marinum. Only the South Carolina two year olds deviated from this pattern. Pending the outcome of studies now in progress, it appears that Seaside oysters are more susceptible to the fungus than Chesapeake Bay oysters and South Carolina oysters are more resistant. The fungus is present in South Carolina waters but apparently absent from Seaside and Chincoteague Bay. For commercial oystermen, does this explain repeated failures of Seaside oysters in Chesapeake Bay? Does it signal danger for Sea- side if the fungus is introduced and water conditions are favorable? Would it be wise for oystermen from Seaside and farther north to know the areas in Chesapeake Bay that are infected and the seasons when the disease is rampant? These are questions that we cannot answer, but the disease is already serious in lower Chesapeake Bay and could be- come a problem in more northerly waters. -160- Table IT Effects of Source (Race?) and Age on Susceptibility of Oysters to D. marinum Incidence in live oysters--September 1953 History Source Number Percentage Weighted tested infected incidence** Yearlings* South Carolina (Tray 28) 50 10 0.16 Chesapeake Bay (Tray 33) 50 0 0.00 Seaside of Virginia (Tray 15) 25 64, 0.88 Two-year South Carolina old (Tray 4) 25 20 0.20 oysters Chesapeake Bay (Tray 11) 37 35 0.78 * All moved as spat to Gloucester Point in fall of 1952. ** Weighted incidence combines intensity and incidence of infection by assigning artificial values of O for negative, 1 for light, 3 for moderate, and 5 for heavy infections. To get weighted incidence the sum of all values is divided by the number of oysters tested. These ratings can be compared directly with the six categories assigned Antegers from 0 to 5 by Mackin (1951). Our ratings (ten in all) have been grouped into 4 categories. = Gil= Table Tae Effects of Source (Race?) and Age on Susceptibility of Oysters to D. marinum Incidence in gapers--June to October 1953 History Source Percentage No gapers Percentage Weighted mortality tested infected incidence Yearlings Seaside of 30.1 31 Sy eal 2A Virginia South Carolina (1952) (Tray 1) i(oal 4 25.0 0.25 Chesapeake Bay (1952) (Trays 11 & 12) By 1 100.0 5.00 Two year South Carolina old (Tray 4) 9.3 28 50.0 1.43 oysters Chesapeake Bay (Tray 11) 24.5 79 91.0 4.20 (Tray 12) algal) 33 90.9 4.12 Three-four Seaside year old (Tray 5) 46.5 h7 93.7 4.21 oysters Chesapeake Bay (Tray 7) 31.4 66 90.0 4.27 (Tray 8) 27.0 51 95.5 3.98 ~162- Literature Cited Hewatt, W. G. and J. D. Andrews. 1954. Oyster mortality studies in Virginia. I. Mortalities of oysters in trays at Gloucester Point, York River. Texas Jr. Sci. 6 (2): 121-133. Mackin, J. G. 1951. Incidence of infection of oysters by Dermo- cystidium in the Barataria Bay area of Louisiana. Conv. Add. Nat. Shellfisheries Assoc. 1951. Mackin, J. G., H. M. Owen, and A. Collier. 1950. Preliminary note on the occurrence of a new protistan parasite, Dermocystidium marinum n. sp. in Crassostrea virginica (Gmelin). Science 111: 325-329. Ray, S. M. 1952. A culture technique for the diagnosis of infections with Dermocystidium marinum Mackin, Owen, and Collier, in oysters. Science 116:360. -163- STUDIES OF PATHOGENESIS OF DERMOCYSTIDIUM MARINUM S. M. Ray Rice Institute, Houston, Texas J. G. Mackin Marine Laboratory, Agricultural and Mechanical College of Texas, Galveston A considerable series of studies have been carried out by the authors, some as collaborations and some independently, which have aimed at measuring, as accurately as possible, the effect of in- fections of Dermocystidimm marinum, a fungous parasite, on their host oyster, Crassostrea virginica. This paper aims to summarize the re- sults of these studies. The detailed descriptions of the various ex- periments will be published elsewhere. Methods In all cases oysters used in experimentation were disease free prior to experimental infection, as shown by check of representative samples by the thioglycollate method (Ray, 1952). All oysters dying in the course of the studies were checked and graded for intensity of infection by the same method, as were also all survivors. The in- tensities were graded according to the definitions as presented by Ray, (1953). Infections were induced in the experimental oysters by exposing them to fungous cells obtained by making minces of the meats of dis- eased oysters using a Waring Blendor for that purpose. Two techni- ques were used in exposing the oysters to infection. In the first of these, the tissue minces of the diseased oysters were mixed with the water of the experimental aquaria. This method is referred to as the "feeding" method because it is assumed that in most cases the in- fective cells are ingested by the experimental oysters and entry into the oyster tissue is via the gut epithelium route. In the other method, the tissue mince is injected into the shell cavity of the oyster through a small hole, bored through the shell. Oysters so "injected" are held out of water for a period of 15 to 24 hours until penetration of the infective fungous cells has been accomplished, and it is assumed that the cells penetrate the mantle, gill and general body epithelium. In some of the experiments, aerated sea water in closed aquaria was used, the water being changed at about weekly in- tervals. In the remainder or the studies, a system of filtering run- ning sea water was used, the filters of glass wool preventing in- fection of the controls. In all cases control oysters were treated as were the experi- mental oysters, with the exception that the minces of oyster meats to -16)- which they were subjected were made up of oysters which were free of D. marinum infection. Some of the oysters used in making up the control minces were gapers and their tissues had been invaded by saprophytic bacteria. Data The data derived from these studies are incorporated in Table I. Discussion The data presented in Table 1 hardly need discussion. They show that D. marinum can destroy oysters, and under the conditions of the experiment destruction of all infected oysters will result if the experiments are continued over a long enough period. High water tem- perature is one of the conditions of experiment since all were carried out in summer or late spring. Data have been presented in previous years (Mackin, 1951) to show that under field conditions nearly 90 per cent of gapers re- covered were in stages of acute fungous disease. Unpublished data have shown that uninfected oysters placed in endemic areas of dis- ease become infected about as rapidly as do those in aquaria and develop high mortality of approximately the same order as do those experimental oysters held under aquarium conditions. The dat® pre- sented in 1951 have been vastly supplemented since that time and there can be no doubt that heavy intensities of infection are the rule on planted beds throughout the heavy mortality areas of the oyster producing bottoms of Louisiana. The experimental data pre- sented here corroborate the inferences derived from the field data that D. marinum infection of oysters is highly lethal when the dis- ease develops to acute stages, which it does in a very high percen- tage of cases both under field and laboratory conditions. -165- -99I- ; *quaa gad 2G pue ‘og ‘96 ‘9g ‘gg sem etienbe sATS 34 UT AYTTeYIOMW 944 faZapso UL °4Ua0 Jad 2G peyosert suo stud UT AYTTeIA0m 3944 UsyM paddoys sea Apnys 944 pue ‘yseTaeey ayy JO a#esop 944 FO OOT/T ATUO Butatacer 4seot au. ‘STTeo unuUT Tem °q JO aBesop SATJOSJUT JUaTATIIp © paataoer untzenbe yoeg *Yyoee siteysko Tejueuttedxea Gg yytmM ‘etzenbe oaTy (T) "i L Ve OT 92 €€ 2G untzenbe tTos,u00 ut Ts €6 Z6 €3 96 96 96 etaenbe *zedxe ut AATTe¢fow 4ueo. Iag iE z 9 (S 9 8 €[ umtrenbe ToryU00 uy TOT 82 €% G2 €?% WA v2 etiende *zedxe ut syzeep jo *oN Sd o€ Gz o€ €z ve G¢ wumntsenbe TosqU09 uy (1) 92T o€ SZ O€ Ue Ge G6@ Btaenbe *stedxe uy pesn szeqysfo jo requny uotzyooefuy uotTyoefuy Burpesy Jutpseq uotyzosfuy uotyoofuy Burpeag uoTyooesur jo LeuuEW YS-0£-9 €S-8T-6 ES-8T-6 «= ES“-TT-6 €S-St=-8 «= €S-9T-L €S-TI-8 pepue *dke a72q WG-6T"S = ESGMETH8 ES-9T—L «= EG=-9THL ES-9T-L = ES TT-9 ES—TT“9 MoT 799 JUT Teyuemtazsedxs fo o4eq 0) 0) ) fe) 0) 0) o*dxe erojeq psyoesuT °ON Gz 69 OL OL OL 86 86 °dxe etoyeq *mexe °oN eeqT ‘keg *umoy °ey ffeq ey fkeq eq Sfeg *um09 *uu09 ekeTeyeyoqyy*pIOFTIN YSTJpey ystspeyY yYstspey ‘psoFTIW ‘PIOITIW sieysho Jo UTSTIO 3 8 9 0 8c on Apnyzs I 9TqeL Literature Cited Mackin, J. G. 1951s Incidence of infection of oysters by Dermo- eystidium in the Barataria Bay area of Louisiana. Nat. Shellfisheries Assoc. Conv. Add. 195%: £2=35. Ray, S. M. 1952. A technique for the diagnosis of infection with Dermocystidium marinum in oysters. Nat. Shellfisheries Assoc. Conv. Add, 1952: 9-13. Ray, S. Mo 1953. Studies on the occurrence of Dermocystidium marinum in young oysters. Nat. Shellfisheries Assoc. Conv. Add. 1953: 80-88. -167- @ iv 1 fe sb Pex Fees HE TH ALTE, ae ee oe 3 sci tS08) i es bE se. Pataki ee ia ot ee : he rie r . Sy é . & af if f ( '.. ‘ “a” ; ie ts -_ > 7 | _ ; =e _ of a eat Siew a a Bod , y % EB = Se ee 5 Be HS ae | ! 2 = a e ’ Pig = i = ong an ‘a . rf , - 1 i ov 7. & Ve : 2 - — - ; b. i }'* » es At * ef ; - ty 7 ne dy ~ .* ' ¥ ‘ iF ; 7 a x u ; ; 3 Pd ; 1) — > Ps" S is 22 a Ag 2 4 eee ! oe 0 J ry iy a; 7s =m, OF , i ‘ - > hf F : i ; - . 0% } a sas 3 | “s " w a Bs an we i ~ Fd 5 = : > : é ' | STUDIES ON THE EIFECT OF INFECTION BY DERMOCCYSTIDIUM MARTNUM ON CILIARY ACTION IN OYSTERS (CRASSOSTREA VIRGINICA) Jo Ge Mackin Marine Laboratory, Agricultural and Mechanical College of Texas, Galveston S. M. Ray Rice Institute, Houston, Texas introduction Analyses of the effect on oysters of infection with Dermocystid- ium marinum have been made over a period of several years. "It has been shown (1) that there is a definite association of the disease with dead and dying oysters in the field (Andrews and Hewatt, 1953; Mackin, 1953), (2) that extreme damage is done to tissues in the course of the disease (Mackin, 1958), (3) that heavy losses in weight of tissue are incurred resulting from infection (Ray, Mackin, and Boswell, 1953), (4) that the disease is water borne, and that transmission may be direct (Mackin 1951b; Ray, 1954), and (5) that infections are highly lethal (Ray and Mackin, 1954; Mackin, Ray, and Boswell, 1954). Continuing the analysis of effect of D. marinum on oysters, several studies of a physiological type designed to measure the effect of infection on ciliary action were set up. The data resulting from these studies are presented here. Methods EK. L. Lund invented a very efficient method cf measuring ciliary activity and feeding in oysters. The basis of his method is measure= ment of the amounts of feecal and pseudofaecal matter deposited by oysters. This is the material extracted from the surrounding water by ciliary action, and when faecal matter is measured it becomes a measure- ment of feeding activity. These activities ere physiologically basic and irreversible depression or cessation of ciliary activity may be taken as an indication of extreme weakness. The apparatus designed and made by Lund to eaten faecal and pseudofaecal matter consists of a long lucite aquarium (Fig. 1) which is divided into three indeperdent longitudinal troughs. Each trough is further subdivided into six compartments. Depending on the size of the oyster each compartment may be subdivided by baffles into 3 or 6 in- dividual supports in such manner that the hinge of the shell is down, and faecal matter collects on one side, pseudofaecal matter on the other. Median dividers separate the two sides of an individual cell. Figure 2 shows oysters in position in one compartment of the Lund aquarium. -168- ~ ct mall | pee , 3 S. 4 Fig. 1. A view of the experimental apparatus used for Experiments Number 1, 2, and 3. Above is a constant head tank provided with an overflow which regulated the pressure on the water flow. Three tubes lead from the bottom of the constant head tank to the three longitudinal troughs of the lucite tank. The level of water in the troughs was regulated by means of overflow tubes which are not visible in the photograph. Measurement of the amounts of faecal and pseudofaecal matter was done with the graduates in the fore- ground. A settling period of 24 hours in the graduates was allowed before readings were made. Fig. 2. The positions of the oysters in the troughs are shown here. They rest, hinge down, on the rounded lucite supports and are separated from each other by the slanted partitions. Faecal matter collects on the dorsal side of the median partition; and pseudofaecal matter, on the ventral side. The ventral sides of the oysters are toward the observer in this photograph. 170 An equal flow of sea water was maintained through each of the three troughs, the water entering at one end and leaving the other. A constant head tank was used to maintain constant pressure and the tapered tips of the inlet tubes were matched for size to regulate the amount of water flow. The turbidity of the water varied during the period of the experiments but control and experimental troughs varied together. The inlet tubes were cleaned once or twice daily and checks were kept on the volume of the water flow to see that it did not vary materially between control and experimental oysters, or from time to time. The deposited pseudofaecal and faecal matter was pipetted at intervals into 100 milliliter graduates, allowed to settle for a period of exactly 24 hours, and the amounts recorded. In some cases records for individual oysters were kept, in others the records were kept by compartments. Experimental oysters were matched for size with control oysters. In Experiment 1, each experimental oyster was matched with a corres- ponding control oyster by shell length. In Experiments 2 and 3 the oysters were matched by weights. In Experiment 2 the control oysters totaled slightly less than the weight of the experimental oysters which was unavoidable considering the methods of selecting the con- trols. Further methods peculiar to each study will be discussed in the individual sections. Experiment Number 1 Experimental oysters used in this study were taken from a planted bed in Louisiana owned by Mr. Emmet Eymard. He had reported a considerable mortality, and a study of his oysters showed that about 67 per cent were infected with D. marinum and the intensity was relatively high. The oysters were checked for intensity by the thioglycollate method (Ray, 1952). Control oysters were taken from the Chene Fleur station and were matched for size with the experi- mental oysters. A check of a sample of the controls for disease showed that they were free of disease at the beginning of the study but certainly picked up some infection during the study. All control and experimental oysters were carefully cleaned of silt before placing them in the Lund aquarium. There were 18 oysters in each group. A flow of 1000 cc. per minute of sea water was introduced into the upper end of both the experimental and control troughs and the oysters were placed in position and the study begun on April 26, 1952. On May 5, 1952, the study wes brought to a close and the faecal and pseudofaecal matter produced by control and experimental oysters was measured for volume, and the amounts recorded for each compartment of three oysters separately. Saal Resuits The data from Experiment Number 1 are contained in Table I. Table I Amounts of Faecal and Pseudofaccal Matter in cts Produced by Control (Uninfected) and Experimental (Mostly Infected) Oysters in Experiment No. 1 Compartment 1 2 3 rf 5 6 Total Control Oysters 106 178 198 Veep" eis 107, 1008 Experimental Oysters 166 130 112 ey) 29 614 A peculiar pattern was established by the control oysters, so far as a comparison of the amounts produced in the various compart-= ments is concerned. Water entered the number one compartment and thus the amount of suspended silt must have been highest in this one. It decreased gyucessively through the trough from there to the end compartment, number 6. However, the efficiency of the oysters in removing the materials apparently increased with the decrease of the amount of suspended silt, for the amount precipitated by ciliary action increased through the first five compartments. Apparently the small amount of silt remaining became a limiting factor when the sixth compartment was reached, for only half as much was precipitated as was the case in compartment 5. The pattern of precipitation of silt for the experimental oysters was rather irregular. Oysters of compartment number 1 pro= duced the greatest amount, those of compartment number 5 the next largest, and those of number four the least. No experimentai compartment attained the average amount pro-= duced by the controls. In the case of the experimental oysters, the erratic variations in amount of faecal and pseudofaecal material produced in the different compartments probably represented variations in disease intensity. According to the sample checked for incidence of the population, one third was not infected, and another third was lightly infected. Thus Se it is possible that uninfected oysters were placed in compartments 1 and 5 (which compare favorably with the controls), while in com- partments 4 and 6 (in which deposits were negligible in amount) there were two or three oysters in advanced stages of disease. The total amount of combined pseudofaecal and faecal material produced by the diseased oysters in the experimental aquarium was only 61 per cent of the amount produced by the control oysters, show- ing a considerable ciliary depression caused by the infection. Experiment Number 2 Oysters used in this study were raised from spat at the Chene Fleur field station in Louisiana. At the time of this study they were just over two years old. Each oyster was weighed and they were then matched in pairs for equal weight. In all, 22 pairs were thus matched for weight. One of each pair was artificially infected with D. marinum using the injection method (Mackin, Ray, and Boswell, 1954). Since this involved injection of infected tissue mince into the shell cavity, the other oyster of each pair was similarly injected with a mince of uninfected oyster tissue. Since the oysters used were not free of disease to start with, it was hoped by this method to obtain a considerable number of heavily infected oysters to compare with a group of similar size which were uninfected or with light infections. When placed in the Lund tank, they were so arranged that the artifi- cially infected group which could be depended upon to develop heavy infections was placed in one trough and that group from which it was expected to get controls was placed in another parallel trough. Only 18 pairs of oysters were used in initiating the study, since it was assumed that there would be mortalities. The four extra pairs were used to replace the early mortalities. When an oyster died it was removed along with the one matched to it by weight, and another matched pair was used for replacement. The volume of pseudofaecal and faecal matter produced by each oyster was recorded along with the number of days of production. At the end of the study, or at the time of death each oyster was classed for intensity of infection by the thioglycollate culture method. In this manner data on ciliary action and feeding of a number of oysters in various stages of disease, as weil as some uninfected controls, were obtained. These data were reduced to pseudofaecal and faecal production per oyster per day. Results The data are presented in Table If. Those oysters dying or removed in one day were eliminated because the data were not considered eae Table II Basic Data from Experiment No. 2 Pseudo= Oyse- Infec- Faecal faecal Pseudo-= Oys- ter tion produc- produc-~ Faeces faeces ter wt. inten= tion in tion in per per Total No. pms . Days sity ec. ee. Total day day per day Averag 1 72.5 2 H 3.20 4.0 TsO» “Leb@ 2.00 3.50 2 68.5 8 H 2.0 45 Geb, Ones 0.56 0.81 3 66.0 5 H 2G 2.5 LoS - Oak 0.50 0.90 4 59.3 6 H 2.0 4.0 6.0 6533 0.67 1.00 5 (ey: 13 H 5.0 6.0 Tio G35 0.46 0.85 1.48 6 15-5 i H 8.0 12.0) ‘BOs0y) ATE 172 2.86 Te 76.5 15 H 6.0 9.5 i525 7 @2ho 0.63 1.03 8 759 14 H 6.0 70m 1cOnm OLS 0.50 0.93 9 70.0 15 M Ti 15.0 2650 6.73 1.00 hers 10 68.5 15 M 8.0 Tis> “LBs = Oeh3 Gary 1.30 11 78.6 15 M 9.0 1Gs55 AGEs “G260 0.70 1,30 12 Ts 15 M 8.0 Tt.) Ses. C253 Gaty 1.30 1.39 13 7505 15 M 9.5 Tae. | R7s0. 6263 0.50 13 14 75-0 15 M 10.5 130% 23-5 G<76 0.87 1.56 15 73=5 13 if 19.5 SNO- baee | elaso 2.62 1d 16 TH.5 15 18 12.5 1955 3258) 6.83 1.30 2.13 17 76.5 15 Tih 9.5 95 — 1920 G263 0.63 127 18 aves L Te) Te) S26... G67 0.67 13,33 2207 19 67.8 8 iT 365 6.0 9.5 O.4k 0.75 1.19 20 68.0 1 i 16.5 T7eGe 3565 > 1210 Tey 2.36 21 5.5 ii VL 12.5 20.0 425 1.79 4.30 6.10 22 1.5 6 VL 10.0 20.0 36:@ 1.67 EWE 5.00 23 TH.S 15 VL 14.0 Liae SSkebu | 6353 egane7 2.10 3.58 an 77-5 15 VL 7aO 19,05 biycer weeks 0.67 Heals) 25 73-5 15 N 14.0 3350) 47.0) “Gage 2.20 3013 26 72.0 2 N 5.0 FeO LOO, V2250 2.50 5.00 27 70.0 15 N 15.0 26,0 3560 1.60 1.433 2333 3.40 28 65.5 5 N TIO 17.0 2660. 2,20 3-40 5.60 29 Tose 15 N 1245 1555) £820 ‘0x63 1.03 1.86 30 59.7 6 N 726 B30" Lae ey 1.33 2.50 -174- significant. This left a total of 30 oysters, of which 8 were heavily infected, 6 were moderately infected, 6 were lightly infected, 4 very lightly infected, and 6 were negative for disease. The data on pseudo- faecal and faecal production for individual oysters and for each in- fection class are contained in the table. The production for differ- ent classes of acutely infected oysters (moderate and heavy infections) are not significantly different. Those lightly infected are inter- mediate in capacity, and those very lightly infected and those negative do not differ significantly from each other. But when those acutely diseased are compared with those negative for D. marinum and very lightly diseased, the contrast is very striking, for those in stages of advanced disease produced only about 41 per cent as much pseudo- faecal and faecal matter as did those not diseased. The data are further broken down in the graph Figure 3. Experiment Number 3 Experiment Number 3 was set up on August 14, 1953, and the study was terminated at the end of a 21 day period. The oysters used in Experiment Number 3 were obtained from Milford, Connecticut, through the courtesy of Dr. Victor Loosanoff, Director of the Milford Labora- tory of the U. S. Fish and Wildlife Service, and Mr. Joseph Uzmann, of the same Laboratory. A considerable sample of these oysters was checked by the thioglycollate method to determine whether there were any infected ones among them. No infected oysters were found. Experi- mental oysters were infected with Dermocystidium by the injection method, the injections taking place on August 13, 1953. Ali oysters were weighed and matched in triplets; one of each three was a control and the other two were used in experimental groups A and B. Each of the two experimental groups contained nine oysters and the control group contained nine oysters. The data show that the injection method was successful in producing immediate infection in all experimental oysters. Control oysters picked up a considerable natural infection from the water flow, but no control oyster reached a heavy infection stage in the period of the study and none died. On the other hand 67 per cent of the experimental oysters (both groups) died prior to the end of the study (21 days), and on the following day two more died, making a total of 78 per cent. Excepting as noted, Experiment Number 3 followed the procedures for Experiment Number 2, receiving the same water flow. Ali three divisions of the Lund trough were used, the controls in the right side, Experiment A in the middle, and Experiment B on the left. In assessing the study the following points should be kept in mind. (1) All oysters started out equal as regards infection. (2) Experimental oysters received a massive initial infection by injection of infective elements into the mantle cavity, and were all infected on -175= PRODUCTION IN cc PER DAY EXPERIMENT 45 4r — - ——— - - — —————_—__—— — FAECAL PRODUCTION UDOFAE PRODUCTION U FAECA D SEUDOF, AL PRODUCTION | | 3.5 |- N = NEGATIVE L — VERY LIGHT L = LIGHT | M — MODERATE TO 30 MODERATELY HEAVY | | H — HEAVY | | 25 20 | Si ‘ale | | | | | 5 & | | N VL E M H N VL L M H N VL L M H INTENSITY OF INFECTION Fig. 3. Experiment No. 2. Graphic illustration of the production of faecal and pseudofaecal matter by infection classes of the oysters used in this study. The intensity of infection in each class is indi- cated at the bottom of the column. The left hand scale indicates the amount of material precipitated in cubic centimeters. 176 Oyster No. Table IIT Basic Data from Experiment No. 3 Period: August 14 to September 4, 1953 Faecal Pseudo- Intensity Faecal Pseudofaecal matter faecal Total of matter matter per per per Days infection in cc. shal (era Total day day day Experiment A if Pal H 22.0 45.5 67.5 1.20 Di eoeee 2 21 H LTe5 26.0 43.5 0.83 12 S207 3 Pa H Tae ME) 35.20 On52 114 1.66 4 19 H 1385 26.0 BAG Ola val Sie 2208 5 5 M 20 {20 OL, ..C.60 LAG 2.00 6 25L H 10.0 31.0 HO On48 147 1.95 if 15 H Tao TO) 18.0 G46 Gn73) 1,20 8 6 H 255 5.0 a Onste 0.83 1.25 fe) 16 H 8.5 12.5 2140) 01553 Os7o. dest Experiment B 10 alk H 19.5 34.5 > BO 1.15 S102 1 Salar Tuy 21 H 18.0 a0 55.0 0.86 1.76 2x60 12 18 H 13.0 24.0 2740) Oa 72 1a33e 2a 13 15 H 9.5 16.5 26.0 0.64 ied @ 71273 14 15 H 9.0 16.5 25.5 0.60 TaD On Leo 15 17 H 8.0 19.0 2720) O47 Lele 1259 16 16 H 8.5 19.5 28.0 0.53 1222 1 a7 gy. 12 H 5.5 12.0 7.5 Ozt6 1.00 1.46 18 eal H Ta) 1470 23;0 0.33 0.76 1.09 Control Group 19 al MH 29.0 69.0 68.0) 1338 3n27. e666 20 21 M 23.5 83.0 166.5 1ede 3.95 5.06 al ei N 17-5 51.5 69.60 0.83 Shh 3528 22 21 M 14.5 28.5 43.0 0.69 1.36 2.04 23 eit MH 13 20 2580 38.0 0.62 12d Or ter 2k 21 MH 15.0 34.5 419.5 0.72 1,64 2235 25 21 MH sme) 22.5 33.5 0.52 1.07 1.59 26 eae L 15.0 20.5 3545, Gere 0.98 1.69 ela i IM 13.36 21.5 34.65 6.62 1,02 1,64 mies Averages of expe1 imental and con- trol oys ters 1.90 2.68 the first day. This infection was added te by natural infection from the water stream. (3) The infection proceeded rapidly in the ex- perimental oysters, ranging probably from light to moderate in the first week, from moderate to moderately heavy or heavy in the second week, and attained a heavy level in all oysters by the end of the third week. Because of this progressive development of intensity from zero to heavy, faecal and pseudofaecal production should show an inverse level of production, becoming less from week to week. (4) Control oysters also, with one exception, became infected from the natural water stream. (5) Level of infection in the control group was never as high as in the experimental group, but more than half had attained a level of acute disease (but not heavy) by the end of the third week. (6) Progressive development of disease in the control oysters should be reflected in decreased faecal and pseudofaecal matter, at least by the end of the third week. (7) The decrease should not be great in comparison to that shown by the experimental oysters, because of the difference in level of intensity of disease. Results Table III contains the basic data derived from Experiment Number 3, in terms of total production of faecal and pseudofaecal matter. Experimental groups A and B were nearly equal in production of pseudofaecal and faecal matter, and these two were much below the level of the controls. Assuming that the level of the controls re- presents 100 per cent, the two experimental groups produced only approximately 71 per cent. These data are presented graphically in Figure 4. The data showing weekly production of faecal and pseudofaecal matter are also presented in Figure }. In computing the values on a weekly basis, al] oysters producing for a part or all of each week were included in the calculations. In the case of those producing for only a part of the week, the produetion figures were increased proportionately to represent a full week. As shown by the graph, ciliary and feeding action was not only considerably less in the experimental oysters, but it decreased progressively for the three weeks of the study as infection intensity increased. The controls showed a different pattern. Ciliary activity and feeding showed a distinct increase in the second week over the first. This probably was due to acclimatization to the aquarium conditions. The third week showed a drastic slowing down of ciliary action as the acute disease stage was reached in some control oysters. =-178- EXPERIMENT 53 H TOTAL FAECAL AND PSEUDOFAECAL | MATTER PRODUCED PER OYSTER PER DAY cc OF PRECIPITATED MATTER nN T cc OF FAECAL MATTER SS T eles FAECAL MATTER PRODUCED PER OYSTER PSEUDOFAECAL MATTER PRODUCED PER PER WEEK FOR THE THREE WEEKS OF OYSTER PER WEEK FOR THE THREE WEEKS EXP 53. EXPERIMENTS A & 8 COMBINED OF EXP 53-EXPERIMENTS A & B COMBINED EXPERIMENT — [i 4|8 contro. - [| | 415 42 49 46 | =e) J CONTROL EXPER.A EXPER. B WEEKS WEEKS Fig. 4. Experiment No. 3. Graphic representation of the effect of Dermocystidium marinum on ciliary action and feeding of oysters. On the left is shown a comparison of the total production of pseudofaecal and faecal material in controls and experimental oysters. In the center is a comparison of faecal production per oyster per week for the three weeks of the experimentation. On the right is a similar graph for pseudofaecal production. The latter two not only compare the controls with the experimental oysters, but show the decrease in ciliary action as disease intensity rises. 179 cc OF PSEUDOFAECAL MATTER Discussion The three studies described here show that physiological ab- normality accompanies the development of Dermocystidium disease in oysters. Retardation of ciliary activity of the gills not only affects feeding, but also produces derangement of respiratory function. It is interesting that even light infections by Dermo- ecystidium produce a measureable decrease in ciliary action, and it may well be that this depression sets up the vicious cycle of weakening which in turn results in miliary spread of disease, pro- ducing even more severe respiratory derangement. In this connection it may be noted that it is not thought that decreased feeding materially speeds death of an oyster from Dermo- cystidium disease. The disease works far too rapidly for that to occur under summer conditions. Cessation of feeding by acutely diseased oysters is to be considered as a symptom, and as evidence of physiological abnormality due to disease, but our studies have shown that, all other things being normal, it takes a very long time to starve an oyster to death. -150- Literature Cited Andrews, J. D., and W. G. Hewatt. 1953. Incidence of Dermocystidium ' marinum Mackin, Owen, and Collier, a fungus disease of oysters in Virginia. Nat. Shellfisheries Assoc. Conv. Add. 1953: 79 (abstract). Mackin, J. G. 195la. Incidence of infection of oysters by Dermo- cystidium in the Barataria Bay area of Louisiana. Nat. Shellfisheries Assoc. Cony. Add. 1951 3 22-35. Mackin, J. G. 195lb. Histopathology of infection of Crassostrea virginica (Gmelin) by Dermocystidium marinum Mackin, Owen, and Collier. Bull. Mar. Sci. Gulf & Carib. 1(1): 72-87. Mackin, J. G., S. M. Ray, and J. L. Boswell. 1954. Studies on the transmission and pathogenicity of Dermocystidium marinum IT.) im press 2 Ray, S. M. 1952. A culture technique for the diagnosis of in- fections with Dermocystidium marinum Mackin, Owen, and Colliery, in oysters. Science 110 (3014): 360-361. Ray, S. M. 1954. Experimental studies on the transmission and pathogenicity of Dermocystidium marinum, a fungous parasite of oysters. Jr. Parasit. LO(2): 2356 Ray, S. M., and J. G. Mackin. 1954. Studies on the transmission and pathogenicity of Dermocystidium marinum I. In press. Ray, S. M., J. G. Mackin, and J. L. Boswell. 1953. Quantitative measurement of the effect on oysters of disease caused by Dermocystidium marinum. Bull. Mar. Sci. Gulf & Carib. 3(1): -33. -181- A HAPLOSPORIDIAN HYPERPARASITE OF OYSTERS do G Mackin Marine Laboratory, Texas Agricultural and Mechanical College, Galveston Harold Loesch Department of Conservation, Bayou La Batre, Alabama Introduction Oystermen operating in Mobile Bay, on Kings’ Bayou and Buoy Reefs and the south part of Whitehouse Reef, complained of excessive mortali- ties of oysters (Crassostrea virginica) in the late summer and fall of 1953. They described oysters as being blackish in color on the vis- cera and mantle, but did not definitely connect this characteristic with the mortality. t appears that their examination was of live survivors and they probably did not examine gapers. In an effort to determine what was the cause of the reported mortality, several samples of oysters were sent to the Grand Isle Laboratory of the Texas A.& M. Research Foundation for study. A check showed that the samples had a comparatively high intensity of infection with Dermocystidium marinum which accounted for the reported mortality. However, there was no correlation with the blackening which had been described. A thorough search revealed only one oyster with such characteristic. The sides of the viscera of this one were streak- ed with brownish black areas which extended down into the mantle and palps. This oyster had only a light infection with Dermocystidiun, and a few Nematopsis spores were found. Sections through the visceral region of this oyster were made and the slides were variously stained with Giemsa, Heidenhain's iron haematoxylin, and Delafield's haematoxylin and eosin. These slides showed that the oyster was heavily infected with Bucephalus, and the sporocysts and cercariae were in turn parasitized with a Haplosporidian gsporozoan. The sporocysts had been destroyed by the sporozoan. and only remnants of host tissue and the cuticular walls remained. ii some areas the Bucephalus cuticula had broken down liberating the spores into the oyster host tissue. Intense cellular reaction was evident in these areas, and a careful search showed tht the hyperpara- sites had been carried through vascular channels and the remnants of the gonadal ducts of the oyster host and were generally distributed through the tissue. They were undergoing development, but the develop- ment was obviously abnormal. Spores and amoebula stages could be seen in leucocytes, and in some epithelia of the oyster host. Generally speaking, however, the oyster tissues seemed normal, excepting for the local cellular reactions already referred to. -182- Development of the Haplosporidian All stages of development were easily observed in the Bucephalus sporocysts. Free amcebulae were seen scattered in the cavity of the sporocysts or occasionally in the epithelia of the host worm where such tissues were not destroyed. Most of the stages of development were found free but may originally have been intracellular. Since most tissues of the sporecyst host were destroyed prior to section-= ing, the developmental stages appeared to be free in 2 cavity bounded only by the cuticula of the worm. A direct development takes place resulting in multinucleate plasmodia with variable numbers of nuclei up to more than fifty. Mature spores may form in the sporocysts in any stage from about 12 nuclei up. In the leucocytes of the host amoebulae may form capsules without going through divisions of the nucleus, which results in formation of single spores. These latter are cytologically as they are when they escape from the spore capsule which escape is accomplished by rupture of the spore case. There is no "lid" to the spore. Vacuoles form in the cytoplasm as the amoe- bulae grow and with successive steps in development of the plasmodial syncytium. When the definitive number of nuclei is attained, islets of cytoplasm form around the nuclei, followed by deposition of the spore wall. The spores themselves are variously shaped according to the pressure of surrounding tissues, but are usually ovate, about 3 LO) Sabie ko. Wt. The apparent absence of a polar filament places this sporozoan in the order Haplosporidia Caullery and Mesnil. Certain phases of the development are so obviously parallel to those of the Micro- sporidia that it is thought best to indicate that this placement in no way indicates a relationship with the aberrant Icthyosporidium or Bertramia which have been referred to the haplosporidian wastebasket. The parasite of sporocysts of Trematodes in Donax trunculus (Europe) is a parallel case of hyperparasitism (Caullery and Chappellier, 1906), and there are several other similar hyperparasites among the Micro- sporidia and the Haplosporidia. The Eye arasite of Trematodes in Donax trunculus (Anurosporidium pelseneeri) is not the same as the form described “here, since the spore in that species is described as spherical and has a smali operculum. investigation has shown that Donax variabilis from the Texas coast has a Haplosporidian parasite of Trematode sporocysts, but preliminary study indicates that this one is neither Aeuroeperid iia pelseneeri nor the hyperparasite reported in this paper. However, it is thought best to reserve final judgement in this matter, pending study of more material. Literature Cited Caullery, M., and A. Chappellier. 1906. Anurosporidium pelseneeri no. g-, No sp., Haplosporidie infectant les sporocystes d'un Trematode parasite de Donax trunculus. C. R. Soc. Biol. Paris 60: 325-328. -183- EFFECTS OF TWO PARASITES ON THE GROWTH OF OYSTERS R. Winston Menzel and Sewell H. Hopkins Department of Biology, A. & M. College of Texas College Station This report is based on an experiment conducted by my colleague, Dr. Menzel, at Bay Sainte Elaine oil field in Terrebonne Parish, Louisiana. On May 1, 1948, Sea Rac trays containing a number of }} by 3 inch strips of roofing tin were placed in the bay. The tin strips were held in place, in a vertical position. On June 26 all but one of the numerous spat were cleaned off of each side of each strip, and each was marked by a number on the metal. The height and length of each spat was measured each month. Between June 26 and October 26 some of the original spat fell off and were lost, and these were re- placed by younger spat which had set on the tin strips. October 28 was the end of the setting season, so no replacements were made after that date. On December 20, 1948, all of the young oysters were re- moved from the plates, and each was marked by a number painted on its shell. On January 23, 1949, and once each month thereafter, each oyster was weighed individually, three dimensions (length, height, and thickness) were measured, and the total volume of all oysters in each tray was determined. A record was made of the average length, volume, and weight of all of the oysters in one tray from May 1948 to the end of the experiment in May 1950. These average measurements are very nice for making pretty graphs, but averages do not tell everything. We were more interested in the differences between the individuals and the reasons for those differences. Incidentally, the fact that a few oysters which had been added to replace lost spat were one, two, or three months younger than the original ones did not make any difference in the long run. Within a year these younger oysters were as large as the oldest ones, or even larger. Now, we had noticed in other experiments, as well as this one, that oysters which grew very slowly usually ended by dying while the faster growers survived. Looking back over the records of an oyster which died, we usually found that it had stopped growing or had even lost in length and weight for several weeks or months before it died. The ones that died had grown very little or none at all during the last four or five months, while the survivors had continued to grow, on the average, at an undiminished rate. Only one tray was kept to May 1950. The 23 survivors in this tray were fixed and sectioned. Dr. Mackin, who had never seen the -18)- individual growth records, was asked to examine each siide for Dermo- cystidium marinum. On the basis of his notes, we divided the oysters into three groups: uninfected, (8 oysters), lightly infected (9 oysters), and heavily infected (3 oysters). There was also one oyster which was negative for Dermocystidium but had a well developed infection of the trematode Bucepnalus cuculus. The parasite had completely des- troyed the gonad and replaced it by a mass of sporocysts. The Bucephalus infected oyster had shown unusually rapid growth. The oysters with light Dermocystidium infections had grown at the same rate as the uninfected ones until the last four or five months, and then lagged slightly behind. The heavily infected group (3 oysters) had been growing at a slower rate for several months. A comparison of the increase in length of the same four groups during the same 16-month period shows that the Bucephalus infected oyster had stopped gaining in length, and the ones with heavy Dermo- cystidium infections had not gained for five months. We had observed many times before that when an oyster is affected by some adverse factor, the gain in length quickly comes to a stop and the oyster may even lose several millimeters of the bill because the mantle is drawn back farther inside the shell, but it may continue for some time to gain in thickness and consequently in weight. At the end of the experiment, May 9, 1950, the largest individual with heavy Dermocystidium infection weighed 141 grams and was 83 milli- meters long, while the smallest one in the uninfected group weighed 127 grams and was ‘(7 millimeters long, so there was some overlapping in final sizes. However, the largest oyster in the group heavily in- fected with Dermocystidium had gained only 22 grams in weight and 2 millimeters in length in the last five months, while the smallest and slowest growing oyster in the uninfected group had gained 38 grams and 7 millimeters. in growth rate there was no overlapping. The data have been analyzed statistically. The analysis shows that the differences between the uninfected, the lightly infected, and the heavily infected oysters are real, and that there is less than one chance in one hundred that these differences are due to acci- dent or coincidence. The single Bucephalus infected oyster was not included in the statistical study. We have no information on parasitism in the oysters which died during this experiment, but Dr. Mackin has examined those which died in many other experiments. In a growth experiment conducted by Dr. W. G. Hewatt during the same period (in 1949) all of the 25 oysters which died proved to have heavy infections of Dermocystidium; nearly all of these individuals had stopped growing some weeks before death, while the uninfected survivors had continued to grow at a rapid rate. ~185- We conclude that an infection by Dermocystidium usually causes a slowing and eventually a complete stopping of growth, long before the oyster finally dies. Oysters may stop growing several months before death. Inclusion of parasitized oysters in a growth experiment may change the shape of the growth curve, so a study of parasitism should be made a part of every growth study. We have growth data on only one Bucephalus infected oyster. $0 far as we know, this is the only one on which anyone has any growth data. However, it is interesting that our Bucephalus infected oyster showed exceptionally rapid growth until the last two months of the experiment, for there are theoretical reasons for believing that this parasite may stimulate growth. It is well known to parasitologists, since the work of Miriam Rothschild in the 'thirties, that snails castrated by larval trematodes have rapid growth and prolonged life, resulting in giantism. The same may be true of oysters. So long as the sporocysts are confined to the gonad, Bucephalus infected oysters have large fat meats, even in summer when other oysters are poor. We know from personal experience that these caponized oysters are ex= ceptionally good eating. However, the sporocysts later spread to other tissues and seem to cause considerable damage. It may have been the effect of such a late stage infection that finally halted the rapid growth of our pet. -186- = ye cy - os bade eas gre, *- ov ey 7 hy} Maban an rans af) oe a era - at woe nin: ase oe ‘esries Ai Ae eel oT oe: eee Gea is a9es ‘pont bd Oradea Sik Lae fe Pye we #5 tik iteteige we l:5-3 14] eres Stam fy : int : ee: ioua.. Ba ata jie LPO Cam, 4 n fj OP b vpn ne et Leung ph: q =) eat: T wilt legions 26: Lats Whe iap SPs mg. 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THE FUNCTIONAL MORPHOLOGY OF THE ALIMENTARY CANAL OF ASTERTAS FORREST AND THE PREDATION OF BIVALVE) MOLLUSKS (A Summaz’y) Frederick A, Aldrich Academy of Natural Sciences, Philadelphia, Pennsylvania Introduction The importance of the common sea star Asterias forhesi (Desor) of the middle Atlantic coast as a predator of commercially important bivalves has long been recognized. Very little work has been done on the functional morphology and digestive physiology of this animal. The present report summarizes work on the functional morphology of the alimentary canal of the sea star conducted as a part of a more comprehensive study which will appear in print elsewhere. The author is indebted to the encouragement and direction of Dr. M. R. Carriker, now of the University of North Carolina, and to Dr. Victor L. Loosanoff, through whose cooperation much of the work was performed at the Milford Laboratory of the U. S. Fish and Wild- Tite SErvAee. General Aspects of the Alimentary Canal The alimentary canal of A. forbesi is suspended by mesenteries from the dorsal body wall in the oral-aboral axis of the central disk at the base of the antimeres or rays. The mouth at the oral terminus is ringed by a peristomial membrane bordered by large per- ioral spines bearing numerous pedicellariae. A short esophagus con= nects the mouth with the stomachal portion of the canal. The stomach is divided by a superficial constriction into two portions: an oral portion, the much convoluted and lobulated cardiac stomach, and an aboral pyriform portion, the pyloric stomach. From the mid intraradial surfaces of the pyloric stomach five ducts arise, each of which soon bifureates upon entering the antimeres, and then ramifies repeatedly into the multilobulated paired pyloric diverticula. The pyloric stomach leads aborad beyond the radial diverticular ducts and abruptly tapers to the very short and slender intestine. The latter terminates at the anus, when the anus is present. Two short botryoidal rectal caeca project interradially from the walls of the in- testine. =187- Eversion of Cardiac Stomach In the course of feeding on oysters and other bivalves too large to be ingested the sea star everts the cardiac portion of its stomach through the mouth to cover the meat of the pelecypod prey. This phenomenon of extracorporeal digestion is characteristic of sea stars whose tube feet bear suckers (Schiemenz, 1896). It is the lobes of the cardiac stomach, or those portions which fill the pro- ximal ends of the antimeres, which are everted. Cuenot (1887) des- eribes the mechanism of eversion as one of pressures transmitted through the coelomic fluid following the contraction of the body walls. MacBride (1906) echoes this explanation of the mechanics of eversion. However, no experimental data has been offered to cor- roborate or deny this hypothesis. In the present study sea stars, up to 14 centimeters in dia- meter, were injected with known quantities of sea water after being relaxed with MgSO,. The cardiac lobes were everted upon the injection of 10.5 to 14.0 milliliters of water. The injection of 28 milliliters of sea water into an unrelaxed animal failed to produce eversion. Following extracorporeal feeding the cardiac stomach is with- drawn into the body of the sea star by muscular action. There are five pairs of cardiac retractor muscles, with origins on the ambula- eral ridge of each antimere, at about ossicle 33-35, or at a point one third of the length of the ridge. At the point of insertion of each of these muscles to the wall of the cardiac stomach the muscles bear hook-like calcareous structures, here named the "procardial restrictors." The hooks are 2.8 millimeters in length on a retractor muscle measuring 34.4 millimeters. The procardial restrictors of the paired cardiac muscles of each antimere are connected by a thin cal- careous bar, forming an H-shaped configuration, thus providing a common insertion for the paired muscles. When the cardiac stomach is everted the procardial restrictors are locked on the circumoral ring of the endoskeleton, thus limiting the amount of tissue which is everted. it is difficult to visualize the withdrawal of the cardiac lobes by constriction of the retractor muscles alone. The area under the pro- cardial restrictcrs is pulled into the proximal portion of each anti- mere and may initiate the return of the stomach through the mouth. It is suggested that for the stomach to resume its normal position within the sea star the major component of tne forces accomplishing this would be directed aborally. It is clear that the cardiac retractor muscles cannot supply these forces because of their sites of insertion. The mesenteries, previously mentioned, which suspend the stomach from the dorsal body wall are in a position to exert this force. It would appear that as the lobes are everted the mesenteries are greatly extended. Their site of attachment on the aboral walls of the stomach come to lie inside the ring of everted stomach tissue. Upon contraction the mesen- teries resume their normal position and in so doing draw the lobular por- tions of the stomach into the coelom. -188- Literature Cited Cuenot, L. 1887. Contribution a l'etude anatomique des asterides. Arch. Zool. Exp. et gen. (2) 5: (supplement). MacBride, E. W. 1906. Echinodermata. Cambridge Natural History. London. MacMillan and Company, Ltd. 1: 425-623. Schiemenz, P. 1896. Wie offen die Seestern Austern? Mittheilungen des Deutschen Seefischereiverseins 11-13: 102-118. -189- LAY he (as aa be SEASONAL VERTICAL MOVEMENTS OF OYSTER DRILLS (UROSALPINX CINEREA) Melbourne R. Carriker * Rutgers University, New Brunswick, New Jersey Introduction A number of preliminary observations (Adams , 1947; Cole, 1942; Engle, 1935-36; Federigni, 1931; Galtsoff, Prytherch, and Engle, 1937; Gibbs, personal communication; Mistakidis, 1951; Orton, 1930; Stauber, 1943) suggest that as water temperatures drop in the fall oyster drills, Urosalpinx cinerea (Say), at least in the northern areas of their geo- graphic range, exhibit certain migratory movements. Drills inhabiting subtidal surfaces crawl downward onto the bottom, and some of these bury in the sediment; some drills in intertidal areas also burrow in the bottom, and others migrate into deeper water. The reverse migra- tion is said to take place as water temperatures rise in the spring. Federighi (1931) in North Carolina noted that drills retained in running sea water aquaria maintained at temperatures prevailing outdoors became inactive below 50°F and remained attached to the substratum or lay passively on the bottom. A temporary rise in temperature above 50°F anytime during the winter stimulated slight creeping, and the activity increased as the temperature rose. Galtsoff, Prytherch, and Engle (1937), working in the laboratory in the northeastern states in water temperatures ranging from 26 to hoor noticed that locomotion in drills completely ceased below 35.6°R; the drills either hibernated on the surface or buried in the bottom attached to partially buried hard objects, with the tip of the siphon projecting slightly above the bottom. They discovered no evidence that drills seek and congregate exclusively in cavities of empty shells for protection during the cold weather. In Canada (1947) Adams observed that hibernating drills attached to oysters and shells became covered with a layer of silt which hid their typicai form and made them very difficult to detect. Engle (1935-36) and Stauber (1943) both noticed in Delaware Bay that not all drills bury in the bottom since some were found deeply wedged in crevices formed by clusters of oysters. Stauber (1943) reports that drills migrating off intertidal reefs in the fall were discovered later more or less compietely buried in the hottom around the edges of the reef usually clinging to shell and with siphons up and presumably in contact with the water. In the winter Engle (1935-36) noticed in Delaware Bay that the drill dredge collected drills on hard as well as on soft bottom, and that numerous drills were collected with a drill dredge equipped with a scraper bar, indicating that many of the snails were present on the surface or not far below it. *Present address: Department of Zoology, University of North Carolina Chapel Hill, North Carolina =190- The precise depth to which drilis burrow, the proportion that pass the winter thus buried, and details on the behavior of burrow- ing have not been recorded. Such information is of considerable interest to marine ecologists and to oyster farmers who may wish to eradicate drills from infested oyster bottoms during the winter. This report presents additional information on the wintering over behavior of the oyster drill obtained in a series of laboratory and field studies carried out during the winter of 1953-54. Grateful acknowledgment is made to J. Richards Nelson and to the J. & J. W. Elsworth Co. for support of the project. Laboratory Methods The laboratory studies were conducted in the Rutgers University Vivarium where it was possible to stimulate the prevailing outdoor temperatures and to carefully observe the activities of drills during the period October 21 to March 17. A glass covered aquarium, with a bottom area of two square feet, located within the Vivarium, was connected by one half inch diameter rubber and glass tubing to a glass covered hard rubber bucket placed outside of the Vivarium. The bucket was covered with aluminum foil to reduce warming from insolation. The bottom of the aquarium was filled to a depth of about three inches with three different types of sediment in equal parts: clean fine sand, coarse sand and gravel, and fine black mud. Sediments were overlaid with a layer about an inch thick of mussel shells, living mussels, and a few young oysters. some of the shells were pressed deeply into the sediments. The 10 gallons of sea water (salinity range during the observations was 29.6 to 33.5 o/oo)in the system were continuously circulated at the rate of about a gallon every two minutes by means of an air lift pump. Sea water was changed every two months. Hydrogen ion concentration, determined weekly, fluctuated between 7.5 and 7.6 during the winter. Temperature of the water in the aquarium was recorded once or twice daily. Minimum water temperatures, 34.6°F, were encountered in December and with minor upward fluctuations lasted about three days. Unfortunately no subfreezing temperatures occurred. Two hundred oyster drills ranging in height from 17 to 34 millimeters were distributed uniformly over the bottom of the aquarium and examined almost twice daily. Hach day drilis which had crawled onto the sides of the aquarium during the previous day were returned to the bottom, and a count of these was employed as a partial index of the gross activity of the drills during that period. A careful check was also maintained of the number and position of drills among the shells on the bottom, and of the number and behavior of drills which buried in the sediments along the sides of the aquarium. These were clearly visible through the glass on which the vertical position and outline of the foot was marked and dated with a wax pencil. At the end of the experiment before the drills became active in the spring their vertical stratification in the sediments was determined by successive strneening of one inch layers. ake nee Laboratory Results In the fall at temperatures between 55 and 70°F as many as 42 drills climbed onto the sides of the aquarium per day. During the first cold period when water temperatures dropped to 41°F movement onto the sides ceased altogether, but only after a lag of about 7 days during which water temperatures rose to a maximum of 61°F. Thereafter temperatures in the range of 52 to 70°F brought as many as 11 of the hardier drills onto the glass per day. From the middle of December to the middle of March temperatures below 41°F occurred with sufficient frequency to almost completely inhibit the movement of drills away from tue bottom. During the coldest part of the winter, December 17=19, when water temperatures fluctuated between 34.8 and 39°F, the disposition of the drills did not change appre- ciably, except that drills seemed to move a little deeper among the shells and were less evident. During the winter about 50 drills remained visible among shelis on the bottom. As the winter passed a fine layer of detritus accumulated snugly about them and made them difficult to detect, particularly those aggregated in the cavi- ties of upturned shells (Fig. 1). The remaining 150 drills stationed themselves under shell or in the sediment. Drills on the surface of the bottom displayed considerable activity and irritability even during low temperatures. At 34. 8°F some of the drills crawled slightly, with tentacles extended. These were slowly retracted when touehed,. Drills turned on their backs at this temperature had very slowly righted themselves 8 hours later by which time the temperature had risen to 39°F. At temperatures between 34 and 39°F most attached drills were easily dislodged. It was surprising to find that as early as the end of October at water temperatures between 55 and 70°F a number of drills started crawling into the sediment along the glass of the aquarium. Since no snail was ever seen actually burrowing, the exact method of pene- tration is unknown. But, because of the danger of blocking of the siphon with sediment and because no partially buried drill was ever seen with its siphon pointed into the bottom, it is possible that crawling into the bottom is performed backwards. in every observed case under these conditions buried drills remained partly or completely attached to hard surfaces and moved no deeper than the fleshy tip of the siphon which projected slightly above the surface of the bottom as a tiny yellow-brown bud. Thus the tip of the spire of the largest drills, which measured 34 mm. in height, were buried about 1 1/4 inch below the surface of the bettom. During warmer periods the yellow ventral surface of the foot of buried drills approached the shape of a circle, and remained closely appressed to the site of attachment (Fig. 2). During periods below 41°F the sides of the foot curled inward, leaving a furrow down the middle, and attachment was thereby considerably weakened (Fig. 3). Im a few cases partial retraction of «19D» Fig. 1. As the winter advances a fine layer of detritus accumulates snugly about drills, Urosalpinx cinerea (Say), remaining on the sur- face of the bottom and makes them difficult to detect, particularly those aggregated in the cavities of upturned bivalve shells. Magnifi- cation approximately natural size. 193 Fig. 2. The foot of a buried drill, Urosalpinx cinerea (Say), closely appressed to aquarium glass during temperatures above 41°F, and photographed through the glass from outside the aquarium. The siphon tip of the drill extends upward to the sur- face of the sediment but is not visible in this photograph. Magnification approximately 2X. 194 Figay se Lhe foot worsaspuried dri | Urosalpinx cinerea (Say), loosely appressed to aquarium glass during the lowest winter temperatures. At these temperatures the sides of the foot curl inward leaving a furrow down the middle, and attachment is thereby consider- ably weakened. Slight compaction of the sedi- ment (mud with some fine sand) is visible about the periphery of the foot. Magnification ap- proximately 5X. 195 the foot was observed. Flow of water through the respiratory chamoer of the drill, although probably much reduced, undoubtedly continues‘during hiber= nation. Examination of buried drills suggests that water is drawn into the gill chamber through the siphon, which always appears to remain in contact with the water, and is ejected from the right side of the snail into the sediment to diffuse upward out of the bottom. This explanation is supported by examination of drills buried in soft mud. The mud immediately around and above the drills appeared oxidized and brown in color, clearly islanded from the sur- rounding reduced black mud. Although a few drills buried in the sand and gravel; the majority were found in the mud; in all cases burrow- ing seemed to occur over hard surfaces such as shell and glass. The depth to which different drills buried varied from a few millimeters to complete submergence. Drills also buried in deep depressions in the bottom, with siphon tips stili in contact with the water, and . thus were stationed some distance below the average level of the bottom. Individually marked buried drills exhibited great variation in movement, although this movement was quite localized and in general did not exceed one half inch; it continued throughout the winter at temperatures approximately above 35.6°F. Many drills moved deeper, if partially buried, or horizontally, or upward, or sought other hibernating sites, or did not bury again; a few remained stationary for periods varying from a few to 56 days inspite of intervening warmer periods. Field Methods and Results The protected waters of Home Pond, Gardiners Island, New York, made possible the use of outdoor cages to determine the disposition of drilis in the bottom during the winter. On October 1, four cages approximately one cubic foot in size and open at one end, ef one quarter inch galvanized wire, were pressed open side downward four to five in- ches into a variety of sand and mud bottoms in siowly moving tidal water. Hach cage contained a layer about one to two inches thick of oysters and shell and 100 drills 16 to 26 millimeters in height, and was covered by approximately six inches of water at low tide. Boat traffic and ice demolished three of the cages. The fourth cage, on firm fine sand, was dismantled during low water on November 18. About one half inch of mud had accumulated in the cage. Ninety-nine of the 100 drilis, 91 alive, were recovered and all were buried in the mud cupped in empty shells or clinging to the underside of shells or to the sides of the cage, all withir an inch of the sur- face of the mud. ; ~196- An abundant population of large drills up to 34 millimeters in height on the shoaler bottom of the Shark River Inlet, New Jersey, afforded a good opportunity to observe the disposition of drills on native subtidal bottoms in swift tidal currents umer natural winter conditions unobstructed by enclosures of any kind. Collections and observations were made on warm days at low water on December 6 and February 20. A small shovel with the handle bent at right angles to the blade was used to scoop up relatively undisturbed strata of the bottom. After macroscopic examination these samples were washed on one quarter inch mesh screen. Where during the previous summer drills were abundant on intertidal rockwork and on intertidal mussel covered bottom, no driils were found even at depths of six inches in the sediment. They did occur, and in quantities comparable to those collected intertidaily during the summer, on bottom below the low water line of a -0.4 tide, especially at depths exceeding one foot. When present drills were not visible through clear water on the bottom because of siltation. In layers of the bottom brought up carefully in the scoop they were found clinging to the under and upper surfaces of shell, and buried partly or to the siphon tip in sediment which had accumulated in upturned shells. This disposition was simi- lar to that observed in the aquarium studies. Summary and Conclusions These studies confirm the observations of earlier investigators and demonstrate that at least 75 per cent of the drills bury partly or completely in the bottom during the colder months of the year in the New York-New Jersey area. Those that bury completely move no deeper than the siphon tip. Since drills also bury in depressions of varying depth, so long as they remain in contact with the water, they may occur some distance below the average level of the bottom and thus escape the scrapers, teeth, and suction of drill dredges. Considerable variation in the degree of torpor occurs among different drills at temperatures above 35°F. Complete inactivity probably does not occur except at temperatures below 35°F. Adherenge of the drill to the substratum is noticeably weaker at lower temperatures, parti- cularly below 41°F. This should faciliate drill dredging in the winter time; on the other hand the characteristic of many drills to crawl under partly buried shell in the level bottom and in depressions in-= creases the difficulty of drill eradication during the winter. =197= Literature Cited Adams, J. R. 1947. The oyster drill in Canada. Fish. Res. Bd. Canada, Atlantic Coast Stas., Atlantic Biol. Sta., Note No. 99. Cole, H.« A. 1942. The American whelk tingle Urosalpinx cinerea (Say), on British oyster beds. Jr. Mar. Biol. Assoc. U. K. 25: 477-508. Engle, J. B. 1935-36, Preliminary report of the U. S. Bureau of Fisheries oyster drill control project in New Jersey, 1933-36. Unpublished manuscript, U. S. Bur. Fish., Washington, D. C. Federighi, H. 1931. Studies on the oyster drill (Urosalpinx cinerea Say). Bull. U. S. Bur. Fish. 47: 83-115. Galtsoff, P. S., H. F. Prytherch, and J. B. Engle. 1937. Natural history and methods of controlling the common oyster drills (Urosalpinx cinerea Say and Eupleura caudata Saye Ux S. Bur. Fish. Cir. No. 25: 1-24. Mistakidis, M. N. 1951. Quantitative studies of the bottom fauna of Essex oyster grounds. Ministry of Agr. & Fish., Fishery Investigations Ser. II, 17(6): 1-47. Orton, J. H. 1930. On the oyster drill in the Essex estuaries. Essex Nat. 22(6): 298-306. Stauber, L. A. 1943. Ecological studies on the oyster drill, Uro- salpinx cinerea, in Delaware Bay, with notes on the associated drill, Eupleura caudata, and with practical consideration of control methods. Unpublished manuscript, Oyster Res. Lab., N. Js Agr. Exp. Sta., Bivalve, N. J. -198- 2 os -_ nl BmURNARNY AER “sat ws ae Ula fa fA: : sie aad hice ° : pene chp a Bos ae and Ae > 2 a we es 4 ho thst f & i bt R eet nied ade 26. piace’ ' Sua ety : ik « bth 1. 4 dw ine ian ee ed Spel , ha y r : aero om c rs | PRELIMINARY EXPERIMENTS IN THE USE OF GROUND CONTROLLED AERIAL PHOTOGRAPHY IN INTERTIDAL HYDROGRAPHIC SURVEYS Robert L. Dow Department of Sea and Shore Fisheries, Augusta, Maine Purpose The purpose of these experiments was to obtain base map data for hydrographic, hydrogeological, geological, biological, or com- bined geological-biological surveys of intertidal areas occugded by commercially exploited shellfish. In this paper the meaning of the term hydrography is limited to that branch of surveying which em- braces the determination of the contour of the bottom. Characteristically, hydrographic charts, topographic quad- rangles and even shoreline survey manuscripts developed from high altitude verticals lack the detailed accuracy essential to base map requirements for intertidal surveys. Discussion Earlier bage maps had been obtained solely from ground sur- veys, a method which is both time consuming and expensive. Results in some areas are of questionable accuracy because of hydrogeological changes which could occur during the course of a several weeks survey. Casual observation between 1946 and 1949 of Western Beach, Scarboro, Maine, the area used in these experiments, had indicated that surface conformation and elevation were drastically modified from time to time. Two marked subareas were photographed at irregular intervals sbetween July 1949 and May 1950. These photographs demon- strated that changes in the conformation and direction of ripples took place during the period covered. Changes in ripple direction were associated with changes in prevailing winds. During the summer the ripple axis ran northwest southeast at nearly right angles to the prevailing winds from the southwest (Fig. 1). By mid winter the axis was nearly southwest-northeast (an estimated turn of 80 degrees) while the prevailing wind direction was northwest (Fig. 2). Intermediate photographs indicated that this change in ripple direction was generally gradual, although some severe storms appeared to accelerate the process. rN As a result of these periodic photographic records it appeared desirable to measure quantitatively the amount of change in surface elevation taking place on both a periodic and a seasonal basis. -199= Encroachment of a shoreward migrating sandbar during the summer was destroying large numbers of soft-shell clams (Fig. 3). By sampling both sides of the bar and the bar itself it was found that there was no survival beyond the forward edge of the bar. The following March the bar was completely leveled by a severe storm (Fig. 4). It was theorized that by determining quantitatively some of these changes which were going on it might be possible by engineer modification to reduce the nearly 100 per cent mortalities occurring annually in this area. Western Beach has prolific sets of soft-shell ciams each year, but very few survive to commercial size. Only occasionally are conditions sufficiently favorable, so that there is a limited commercial fishery. Predator mortalities (Polinices) were reported for one year to be nearly 28 per cent (Dow and Wallace, 1950). ° Other mortalities have been attributed largely to hydrogeological changes (Fig. 5). Green crab damage appears to be Hegligible. Under the state's combined geological-biological program approximately 33 man days were spent in a partial ground survey of Western Beach. On the basis of a fortnight's work it was estimated that at least 100 man days of field work over a six to eight week period would be necessary to provide the minimum data. During this preliminary ground survey, changes in surface conformation and elevation from wind and water erosion and deposition were observed (Fig. 6). It was obvious, then, that a ground survey at best would provide only a composite of daily conditions for the period. To compensate for this deficiency an experiment utilizing ground controlled aerial photography was designed. It was proposed to obtain all field data on an instantaneous basis, either during one half=tide cycle or during consecutive dayiight half-tide cycles. Procedure Preliminary experiments were carried out in May, 1952, using a light ground based plane and a.four by five Speed Graphic camera. In addition to the limitations of noncontrolled verticals, weather conditions --- freezing at 5,000 feet, rain at sea level and gusty onshore winds -=- did not permit accurately interpretable results. More carefully organized experiments were carried on between October 9 and 11, 1952. A Fairchild aerial camera mounted in the belly of a multipassenger pontoon plane was used. Weather conditions, except for 10-12 mile per hour southsouthwest winds, were excellent. As in the previous experiments, ground control was exercised by a survey party using a plane table. -201- Fig. 3 202 204 A closed traverse (plane table and telescopic alidade) including all air-ground control panels was commenced October 9, 1952, during the low water period prior to the taking of aerial photographs. The traverse was resumed and closed during the daylight low water period of October 11, 1952. h The traverse consisted of 8 legs and 8 occupied stations. Initial Station "A", also used for photographic control arid water elevation readings, was reoccupied to close the traverse. The length of traverse was 4,860 feet with a horizontal closure error of minus four feet. Within the traversed area the vertical range was 4.44 feet and the vertical closure error was 0.09 feet. The lateral closure error was not measurable. Simultaneous with the two aerial photographs taken on October 9, vertical readings were made of the water elevation near Station "A". Control was effected by radio communication. When the first photograph was taken at 1233 the elevation of the water was minus 5.35 feet (Fig. 7). The second photograph was taken at 1345; at this time the water elevation was minus 2.74 feet (Fig. 8). The first photograph was taken at three hours and thirty two minutes from zero low water; the second at four hours and forty four minutes from zero low water. Zero low water was taken from tide tables for Portland, Maine. Plotting Control points on the ground were inciuded in the ground survey. Since these same points were visible in the aerial photographs, addi- tional detail, including tide lines, could be accurately plotted on the base map (Fig. 9. For purposes of illustration normal computation procedure ---- data assembled on base ground survey map --- is reversed). Tide lines which serve as continuous contour lines can be plotted directly from photographs without the inaccuracies normally associated with the interpolation of contours between critical elevations in con- ventional topographic surveys. There are several methods by which base map data on the photo- graphs can be combined with data from the ground survey: (1) by pro- jection of radii through critical points of control net; (2) by plotting location of critical points in relation to control net and transferring these data to base map, and (3) by superimposing detaiis of photograph on base map and plotting additional data in relation to controls. Results Results of ground control are shown in Table If. =205- ie 4 Se Dee ; ee fe vies Wes ee Sane le Table I. Control Data (Station "A" - assumed elevation minus 3 feet MHW) Station Stadia Elevation (feet) Panel #1 402 minus 3.63 Searboro River Beacon 389 eo Ferry Rock 165 plus 16.6 Panel #2 810 minus 5.09 Panel #4 870 Big 8i Panel #5 520 P76 Panel #6 380 eee) Panel #7 B45 ae 1k 62 Stake 609 * 5a Qh Station "A" 420 a Aor Length of traverse: (1) Stadia - 4,860.0 feet (2) Plotted - 4,864.0 feet Horizontal closure error - minus 4 feet Per cent error horizontal closure - 0.082 Range of vertical traverse - 4.44 feet Vertical closure error - minus 0.09 feet Per cent error vertical closure - 2.03 Time and elevation: October 9, 1952 lies tsiea Ube Tide Time (from zero low water) Elevation 1233 40332 =5.35 feet 1345 FOKKY -2.74 feet =20i- Conclusions l. Satisfactory base maps can be obtained by means of ground controlled vertical aerial photographs. 2. This is the only method presently available which permits nearly instantaneous measurements. 3. The amount of field work entailed by this type of survey is approximately 10 per cent of that required for conventional ground survey methods. Literature Cited Dow, R. L., and D. E. Wallace. 1950. The story of the Maine clam. Bull. Dept. Sea & Shore Fish., Maine, Sec. 7:22. -208- THE USE OF EQUIPMENT AND TECHNIQUES IN APPLIED SHELLFISH MANAGEMENT Dana E. Wallace Department of Sea and Shore Fisheries, Augusta, Maine Maine's hard and soft-shell clam fishery is dependent upon natural sets, their survival and growth to commercial size, and, in the soft-shell industry, the efficiency with which the clams are har- vested. The following summaries cover projects of the Department of Sea and Shore Fisheries as well as cooperative work with the Fish and Wild- life Service Clam Investigations, and the application of results as carried on with shellfish producing communities. Soft-shell Clam Seed (Mya) In the soft-shell clam industry our early work was concerned with developing ways of using soft-shell clam seed in an effort to find out if it was economically feasible to transplant from heavy concentrations to commercially depleted areas. At the present time transplanting of Mya is economically unsound. Projects to learn more about predators and their control are continuing and will be reported another year by Fish and Wildlife Service Clam Investigations. Digging Breakage and Mortalities of Soft-shell Clams We find that the present methods of digging in intertidal areas are extremely wasteful for present and future production of soft-shell clams. This fact has to be considered in any management plans for our soft-shell clam industry. It appears that less than one half of the available or potentially available commercial clam population in Maine ever reaches the consumer. Our work with the commercial diggers in sampling their catch and the flats they have dug indicate that appro- ximately 20 per cent of the clams are broken in digging and can be considered as lost to the industry. Federal Clam Investigations found that less than one per cent survival when broken in the flats and approximately one half of the unbroken clams remaining in the flats each time the flats are dug died because of depth or position. All of this digging information means that we must try to cut down the frequency of digging of the flats and work out a flexible rotation of areas based upon surveys of area, population, size dis- tribution, growth, and the presence or absence of predators. It is likewise always important in cooperative planning with communities to determine the best time to harvest the clams, considering the biological factors as well as the economics of the fishery and area. -209- Quaheg. Research Unlike the soft-sheli clam fishery, we have found that it is definitely economically feasible to transplant quahogs, Venus mercen- aria. Quahogs of approximately over one half inch in size are relative- ly immune from green crab damage. They establish themselves in the flats quite readily and growth in our areas is exceptionally good as will be reported by Dr. Gustafson in his paper. Gathering Seed and Establishing Ciam Farms A pumping arrangement is being used to gather seed clams. A dredge consisting of a two inch suction hose connected to a Homelite eentrifugal pump ratea at 10,000 gailons per hour picks up the water. The unit is portable and weighs only 88 pounds. The water is played through jets down to the flats and the entire dredge is dragged along at approximately 5 to 10 feet a minute. Tiny seed clams in concentra- tions of 200 to 300 per square foot are washed out of the flats. The stream of water picks up the seed clams, the silt and sand is screened out through a one quarter inch mesh screen used on the drag, and jets of water keep the clams moving into the dredge and then back into a detachable mesh bag. Very little breakage occurs. The level of the dredge is set with the shoes on the side. Clam Breakage and Mortalities Caused by Digging Soft-shell Clams (Mya) On silty clay bottom breakage is quite high. Comments on clam breakage in Maine are part of a paper published by our Department by Dow, Wallace, and Taxiarchis. The degree of breakage of clams varies with the concentration and the range of sizes of clams, as well as with the amount of growth and the thinness of the shells. Even in sediments in which a high percentage of breakage occurs, careful and experienced diggers have kept the breakage below 10 per cent. In our studies along the coast, five of the 47 diggers had less than 10 per cent breakage. Two of these dug under good flat conditions while three dug under flat conditions ranging from fair to very poor. The breakage varies both geographically and seasonally. Digging with a hoe produces about 19 per cent breakage, and with a shovel breakage may be cut down to approximately 9 per cent. With a shovel a much larger piece of the flats is turned over, increasing efficiency as well as lessening breakage of clams. Dredging Quahogs With the Venus M, bearing an eight inch discharge hose and mounted on a berge, we were able to get 256 bushels on one day whereas -210- the carrying capacity of the barge alone was about 110 bushels. These quahogs averaged slightly over an inch in size and were compacted in the flats anywhere from 200 to 400 per square foot. One such area covered three to four acres. Already approximately 4,200 bushels have been transplanted from this area with 3,000 bushels being moved by the Venus M. At least 10,000 bushels have yet to be moved. Quahogs are being taken into conservation areas and spread out by sifting them over the sides of the boats or shoveling them as we do from the barge when planting. In one area quahogs are being left to grow to chowder size. Quahogs are screened over one half inch mesh hard- ware cloth and most of the water is discharged through scuppers on the side of the craft. The rate of discharge of quahogs varied from one to six bushels per minute; however, over an entire tidal operation of approximately four hours, it averaged out to about a bushel a minute. This is in the order of approximately 2,800 seed quahogs each minute that are pumped from the flats. The six inch suction hose passes to the centrifugal pump where the design of the impeller allows the tiny seed quahogs to go through tle pump unharmed and out the discharge hose. -2ll- REPORT ON CERTAIN PHASES OF THE CHINCOTEAGUE BAY INVESTIGATIONS Me Es Wa Sieling Department of Research and Education, Solomons, Maryland This is a report on progress made on an ecological survey directed toward, among other things, the re-establishment of the oyster industry in the Maryland part of the Chincoteague Bay area. It will discuss primarily problems encountered in planting shells for the purpose of establishing potential seed areas. This survey was begun in 1951 and a brief progress report was made to this group in 1952. Since that time progress has been made along several lines. The area studied runs from Fenwick Island, Delaware, to Chincoteague Inlet, Virginia, and includes Asawoman Bay, Isle of Wight Bay, Sinepuxent Bay, and Chincoteague Bay. This is a contin- uous body of water and is served from the ocean by two inlets located at Ocean City, Maryland, and at Chincoteague, Virginia. No actual barriers exist between the different bays. The physical aspects and the history of the area were given in the 1952 report and will not be repeated here. The hydrography of the area has been studied during the course of the survey and the data will be published elsewhere. However, it might be said very briefly here that the temperature shows a very slight vertical gradient throughout the area except at the inlets where it may be as great as Se, Throughout most of the area the gradient is not more than 1°c. The horizontal gradient is usually much greater and during the summer varies from the inlets where the ocean water is cooler to the center of the bay where the water is warmer. During the winter this pattern is reversed and the water is warmer toward the inlets and colder in the center of the bay where the shallow water cools more rapidly. Salinities also show a slight vertical pradient but a marked horizontal gradient from the center of the bay to the inlets. In the summer salinities are higher in the center of the bay due to the small land drainage area and the high evaporation in the shallow water. Salinities decrease toward the inlets where the tidal surge brings nearly pure ocean water into the inlets. During the winter and spring this patter is reversed and higher salinities occur in the inlets and drop in the central part of the bay due to the lower evaporation rate and increased rainfall during that part of the year. There is no great tidal movement except at the inlets, the tidal amplitude there normally being about three feet, and about one foot in the center of the bay. Currents through- out the bay are of no great magnitude but may have some influence in distributing shellfish larvae throughout the area. uD1D= During the course of the investigation about 12,000 bushels of shells have been planted annually throughout the area in small test plantings. These plantings, averaging about 2,000 busnels each, have been placed in locations which were selected on the basis of findings from our test shells and on advice from oyster plantérs in the local areas. In most parts of the area, however, fouling of the shells has been a major factor in the success or failure of the plantings. Consequently, considerable attention has been given to determining the period of attachment of the common fouling organisms which cause trouble on the sheil plantings and which were unknown for this area. One of these organisms in particular, the serpulid worm (Hydroides), has been more trouble than the others. Its setting period nearly coincides with that of oysters in the area. The worms can cover a shell with a coating of their calcareous tubes in about two weeks. After this occurs there is no hope of a young oyster ever attaching to the shell since this coating becomes thicker as the colony of worms becomes older. These animals set in greater numbers in certain sections of the bay than they do in other parts. Anomia, barnacles, bryozoans, and Crepidula are some of the other major foul- ing organisms present. Serpulid worms have a definite pattern of setting in the bay which appears to be tied in with the temperature gradient of the water. They set with much less intensity near the inlets and their setting starts there later than it does in the central part of the area. In the central part of the region they begin to set about the middle of May in most years and continue setting with increasing intensity until about the middle of August and then begin to decline slightly. There is then a period of much less setting intensity until about the last week in September when there is again a slight increase in setting and then a complete cessation of setting which occurs about the first week in October. Near the inlets the initial set occurs about the middle of June and continues with a slight dip in intensity in August, and an upturn at the end of September which terminates about the middle of October. Setting is also much more intense in areas where there is little tidal movement, thus suggesting that there is also a con- nection between intensity of setting and water movement. For example, the set in areas where there is very slight tidal movement was as high as 1,077 worms per 10 shells in 14 days exposure, whereas the set near the inlets was in no case over 300 worms per 10 shells for that same period of time. Throughout the central part of Chincoteague Bay the set of these worms through the months of July and August was con- siderably greater than 300 worms per 10 shells at all statiors. Shells planted on the bottom in the central portion of the area were covered with worm tubes in three weeks so that it became impossible for the oyster larvae to attach. Planting cultch at a time to avoid this set of worms is also impossible in this central part of the bay as the oyster set usually occurs at the height of the set of the serpulids. 2213= Barnacles are not a serious fouling organism in most of the area as their setting season is more sharply defined and cultch may be planted at such a time as to avoid a heavy set of this organism. In several sections of the area they do set in great numbers in late May and early June but this does not interfere with oyster setting as shells can be planted after that time. Near the iniets the set of barnacles is usually later and lighter and extends over a longer period of time than it does in the central part of the bay. Also, oyster drills which are numerous in the area kiil great numbers of these before the oysters begin to set. Bryozoa are not too serious a pest in the Chincoteague Bay region as they set sparsely. There are several different species which are common but none is as abundant as in Chesapeake Bay, and they do not form the thick crust on shells commonly observed in that area. Crepidula is not a serious fouling organism in the area as its setting period is rather long, but the number of individuals is small, so that it has not created a problem as yet. These occur more fre- quently near the inlets than in the central part of the bay. Anomia is a much more serious fouling agent in some parts of the area as it sets at the same time as the oyster larvae and it grows much faster, thus overgrowing young oysters. At certain times Anomia may completely cover a shell and any other organisms thereon in about a month after attachment. They are a very serious problem in the best potential seed area which has been located to date. Other fouling organisms are not dangerous and in most cases do not set in large enough numbers to be of any significance in a shell planting program. The part of the program which has had as its aim the rehabil- itation of the oyster industry in the Maryland part of the urea has been conducted with the goal of producing seed locally for the planters. Small plantings of shells made in several different parts of the bay, usually about 2,0CO bushels each, have given varied results. Some of these plantings were made with plantings of brood stock oysters in the immediate area and others were made near private plantings of oysters to take advantage of thespawn from them. Very little success was ex- perienced where the plantings had small lots of brood stock oysters in the midst of them. It is believed that too few oysters were used in the plantings, since where the cultch was near large private plantings of oysters the set of oysters was moderately successful. Two different methods of planting shells have been used in the bay: near the inlet at Ocean City the shells have been planted on the tidal flats in windrows and in the deeper parts of the bay the shells -21)- were broadcast on the bottom. The plantings made near the Ocean City inlet were planted in long windrows on the tidal flats so that they were exposed on each low tide. These have been moderately successful, having a set averaging about 300 spat per bushel at the end of the setting season. However, the bottom is mostly sand and due to a heavy storm during the fall, approximately half of the shells were covered with sand. Some which were planted some little distance from the inlet were not covered but received a very light set so that their value is slight. These plantings were made over the last two years. This year a different region near the inlet which has a muddy bottom and does not completely ebb out on low tide was planted. It is hoped that there the bottom will not shift and cover the shells. Fouling in the Ocean City area on the flats is not serious but it is not known how it will be on these shells which will not be exposed each day. Test shells during the past two years have shown some fouling, particularly by Bryozoa. Survival of spat is very good in that sectian and it is hoped that a seed area similar to that in the Virginia Seaside may be established there. In the deeper parts of the bay where the shells are broadcast on the bottom varying degrees of success have been experienced. Several factors have contributed to the failures, the main one being the heavy fouling already mentioned, and the other being the lack of oyster brood stock in many parts of the area. In those areas where there was no brood stock about 200 bushels of mature oysters were planted in the spring in the spot selected to be planted with shells. This was done early in order that they would become acclimated before the onset of the spawning season. Late in June the shells were planted on and around these oysters. A light set resulted on those shells which were among and very close to the brood stock, but none on those which were 50@ feet away from the oysters. There were no other oysters in any direction from these plantings for several miles and in this one area the tidal movement was very slight, so that there was no other source of spawn. Fouling by serpulid worms in this region was particularly heavy so that three weeks after the shells were planted they were completely covered with the worm tubes. It appears that here the 200 bushels of brood stock were not adequate to give a set and also that larvae did not move very far away from the parent oysters. Test shells placed in a pattern around this planting failed to pick up any spat even a -quarter of a mile away from the brood stock oysters. Other plantings placed near large commercial oyster beds re- ceived moderate sets. Some of these shells, however, were covered with fouling organisms within a very short time and had practically no surviving spat. One planting, however, located just a short distance above the Virginia line, received a good commercial set and at the end of the season gave a spat count of 900 per bushel. This planting came through the winter and still had a count of 700 spat per bushel this spring. This is the only area which gave a really good 275 = set and its location takes advantage of the great number of oysters which are planted in Virginia. The tide which comes in through the Chincoteague Inlet carries larvae from the Virginia oysters into Maryland where they set. In addition there are several large beds of oysters in Maryland not too far from the shell planting. This is an area which does not get a heavy set of serpulid worms but does receive a heavy set of Anomia. The set of these coincides with that of oysters and so they cannot be avoided when planting shells. They also grow faster than oysters and so may overgrow them and cause considerable mortality among the young spat. Further they cover a large percentage of the shell surface and so reduce the efficiency of the shells. If there were intertidal areas in that part of the bay we would have much less fouling on the shells and consequently better sets. However, in Maryland there are no such areas as are found in Virginia, where the set is good and the foul- ing is negligible. It is believed that once a good population of oysters is established in the area, sets will be better and the seed program could be extended. Many very small areas now give good sets but are too small to be of practical use. The two common oyster drills or screwborers are very numeroug in the bay and cause considerable loss to the planters killing great numbers of spat at an early age. This of course is a big factor in the survival of a set which may occur and it is hoped that a program of drill control can be initiated in the near future. The future of the industry in the area depends upon the control of drills and a source of good seed oysters. -216- COMPUTATION OF OYSTER YIELDS IN VIRGINIA * J. L. McHugh and Jj. D. Andrews Virginia Fisheries Laboratory, Gloucester Point, Virginia Drs. Andrews and Hewatt have been holding oysters in trays sus= pended from the Virginia Fisheries Laboratory pier at Gloucester Point, Virginia, for the past four years. The primary objective has been to study mortality rates, but other information has been gathered from time to time, particularly on the growth rate. During the course of these investigations we have been impressed by the yields that have been ob- tained, for it has not been uncommon to realize three bushels of market- sized oysters for each original bushel of seed placed in the trays. Reduced to the simplest terms, the yield of market oysters from planted seed is determined by the interaction of growth and mortality. This has been pointed out by Hopkins and Menzel (1952), who have outlined methods by which planters can determine growth and mortality rates from which they can calculate the net yield. Owen (1953) has described the relationship between growth, mortality, and yield at given locations in Louisiana waters, using figures obtained from experimental plants of seed. Thus, our work is not original in the sense that it represents a new approach. It is originai, however, to the extent that it concerns the Chesapeake Bay region, and that it utilizes the methods of computation applied to fish populations by Ricker (1945, 1948) and others. Hewatt and Andrews (1954) have presented extensive data on oyster mortalities in trays at Gloucester Point, Virginia, and information on oyster growth is accumulating. Both items of information are available in some detail, for mortality records were made daily in summer and at intervals of 10 days to two weeks in winter, and growth measurements have been made at intervals of two weeks to one month. Oystermeri usually report that planted grounds in Chesapeake Bay yield about one bushel of market oysters for each bushel of seed planted. The crop is harvested two to four years after planting, depending on the characteristics of the particular piece oi ground, usually determined through past experience or by occasional sampling, and based on the size of the oysters. It is relatively simple to calculate the mortality that occurs between planting and harvesting. A bushel of seed oysters from Wreck Shoal in the James River may contain as many as 3,000 oysters of various sizes. If he counts a sample of seed, the planter will ignore the small spat, for he knows that these tiny oysters will not survive the planting operations, or if they do, will fall prey to oyster drills and other enemies shortly after, and hence carnot contrioute to the harvest. The * Contributions from the Virginia Fisheries Laboratory, No. 55. =o 7= planter, therefore, will conclude that the viable seed in each bushel number perhaps 1,000 or 1,200 at the most. The market oysters that he harvests in an average period of three years will run akout 300 to each bushel. Therefore, when the yield is 1:1, about 900 of the original 1,200 oysters, or 75> per cent of the number planted, will have been lost. The true mortality, based on all the oysters in the original planting, is of the order of 90 per cent, but the lower figure is more realistic from the oysterman's point of view. On first thought, it might seem that a mortality of 75 per cent in three years is equivalent to a death rate of 25 per cent per year. Percentages cannot be summed or divided so simply, however, and actually the annual rate is considerably higher. t+ can be demonstrated simply that an annual death rate of 37 per cent will produce a total mortality of 75 per cent in three years, by applying this annual rate to a group of 100 oysters, as follows: Original number = 100 Subtract 37 per cent Shih Survivors = 63 (End of first year) Subtract 37 per cent mean Survivors = ho (End of second year) Subtract 37 per cent ples Survivors = 25 (End of third year) Total survival rate = 25 per cent Total mortality rate = 75 per cent Mathematically, the conversion of short period observations on mortality or growth rates to annual rates is somewhat complicated. For- tunately these calculations have been made and recorded systematically in tables (Ricker, 1948) from which mortality rates on a percentage basis can be converted to instantaneous rates, which can be summed- directly. The Rate of Growth in Length Growth rates were measured on oysters held in trays at the Vir- ginia Fisheries Laboratory pier. The most extensive data were available on the rate of growth in length, hence length was used in setting up the basic growth curves (Fig. 1). The curves in Figure 1 were obtained by grouping data from various trays of oysters according to their average -218- Length in millimeters Length in /aches 10} —- Ralicsiee — (2) —L i —— — Oo Apr May June July Aug Sept Oct Nov Dec Jan feb Mar Fig. 1. Average growth rates of oysters in trays at Gloucester Point, Virginia. The data were grouped into broad categories based on the average length in April, which marks approximately the beginning of the year's growth. The points on which the successive curves were based are indicated alternately as black and open circles for ease in recog- nition. 219 length at the beginning of April, the approximate time at which the year's growth commences. The decision to group was dicvated by two considerations, namely, that the data were not sufficient to permit grouping according to specific lengths, and that the average process is much more practical from the oysterman's point of view. From the curves in Figure 1, the lengths at the end of each month were recorded. Figure 2 was then constructed, after the method described by Walford (1946), by which the lengths at a particular date are plotted against the lengths a given time interval later, in this case at intervals of one month. Smooth lines were drawn through each set of points. By reading off lengths from these curves, or by inter- polating between them, the growth in length of oysters in trays at Gloucester Point, starting with any given original size, can be re- constructed easily. The Rate of Growth in Weight The available data on growth in weight at Gloucester Point, though less extensive than the length records, are adequate to con- struct a graph of the length-weight relationship. Plotted on logarith- mic coordinates the resulting points assume a linear relationship, which can be represented by a line fitted by the method of least squares, as in Figure 3. Weights corresponding to the lengths read off Figure 2 were plotted as in Figure 4, which represents the best available average estimate of the growth in weight of Wreck Shoal seed transferred to trays at Gloucester Point. The lower curve in this figure illustrates the growth rate of the small oysters (mostly less than one inch in length) that do not survive planting operations in Chesapeake Bay. The upper curve re= presents the growth of the larger seed oysters (those recognized as seed by the planters). The Instantaneous Rate of Growth The instantanecus growth rate can be computed from the following: expression (Ricker, 1945): eK 214b where e = 2.71839 the base of the natural logarithms, k © the instantan= eous growth rate, and b = the fraction by which the surviying oysters have increased in weight during the period in question. For the present purpose, however, the computations can be present= ed more simply by the method outiined by Ricker and Foerster (1948), as =220= Length in inches on April / / 2 3 Hr race zs 100 + Length in inches x Mmonrrs saver Length in millimeters x months tater bf eke vil etl (2) _— = - i [ere (4) 0/0 20 30 40 50 60 70 80 90 100 110 Length in millimeters on April / Fig. 2. Growth curves for oysters held in trays at Gloucester Point, Virginia, trans- formed according to the method of Walford (1946). 221 Average weight im grams Average sengrh in inches Uf O4 zZ 3 456 350 I (cede [cal PSs) /0.0 250 8.0 200 6.0 /50 50 + 4.0 100 80 30 Ce) 20 40 os 22 +10 20 Os 10 0.2 Ss f 0.04 / Ea 10 20 30 40 60 80 100 /50 200 300 Average /ength in millimeters Fig. 3. The relationship between length and weight in oysters held in trays at Gloucester Point, Va. i? ouwrices Average we/ght vy N U +/. S 4 NEw, == N 4 = S Neos = v he : eal 5S NY | < I | | 104 i el =| SS } Ounces = 2 g g i} | (2) | f Se Ale ar : d t | | | % | | | OS Wea \ | | N | I 1 el \ 0.2 =(}| = 1 \ \ ese A Be ae ee fh ONDSFMAMSSASONDSFHMAMSSASONDSFMAMSSASO~ Months Fig. 4. Seasonal patterns of growth in weight of oysters held in trays at Gloucester Point. The lower curve represents the growth of the current year spat in seed oysters from Wreck Shoal in the James River, which do not survive when transplant- ed to Chesapeake Bay. The upper curve represents the thick- shelled larger oysters in Wreck Shoal seed, that are recognized as seed by the planters. 223 illustrated in Teble I. The instantaneous growth rates k were computed by dividing the values in the previcus column by 0.4343, the logarithm of Cc The Mortality Rate As demonstrated by Hewatt and Andrews (1954), the mortality of oysters in trays at Clovcester Point igs concentrated for the most part in the summer months (July to Octeber inclusive). From the original data on which their report was based, the monthly mortality rates have been computed, that is, the percentage of the oysters alive at the be- ginning of each month that died during that month (Fig. 5). The Instantaneous Rate of Mortality The instantaneous mortality rate can be computed, as was the instantaneous growth rate, from a similar formula (Ricker, 1945): e “4 ale-a where e = 2.7183, g = the instantaneous natural mortality rate in trays at Gloucester Point, and a = the fraction of the original number of oysters that died during the period under consideration (usually a signifies the annual rate). Here again it is simpler to use Ricker's (1948) table to read off the corresponding values directly, according to the value of a. The instantaneous rates listed in Tables IT and III were obtained by this method. Computation of Yields The instantaneous rates of growth and mortality were combined, as in Tables Ii and ITI, to calculate the net increase in total mass of oysters (k - q). The corresponding changes in biomass (total volumes of oysters) were read from column 12 (if positive) or from cgqlumn 2 (if negative) in Ricker's (1948) appendix table. Assigning the arbi- trary value 1CO to the original volume of oysters planted, the relative biomass at the end of each month was computed. The absolute volume of oysters in 100 bushels of seed was then calculated, and this value was substituted for the original arbitrary value of 100. The subsequent -: absolute biomass at the end of each month after planting was derived by ~* simple proportion. =22h- *peseq sem (S6T) SMOIpuy pue 44eMOH JO Jeded 944 YOTUM UO e4ep TeUTSTIO 344 WOrTs peTtdwmog -sazoumms SATSS9OONS 99144 YSnoIm} SUTMOTTOF pue ‘pees Teoyug YootM UT $ia3sko JaBsIeT 24 UTM BuTousmMOD *4UTOg JaqysaonoTyD 12 sfkerq UT PTey staysho jo AqtTeyIom fo uszseyyzed ATYQUOM aU, °S “STW GNOSVSIWYWALTINOSYV ST WYWAILSANOSVSSWYWAILSONO 1 ] ! ] | l G00 NN | ZOO 8 x00 N 900 Y s00 Oro XY zo #10 % es 7OR 225 Table II presents these computations as applied to the current year spat in 100 bushels of Wreck Shoal seed. The increase in biomass ig very large, reaching 168 times the original volume at the end of 35 months, when this group reached its greatest computed yield. The original volume of these oysters is relatively small, however, being only one-half bushel for each 100 bushels of seed. Consequently, this tremendous increase in biomass produces a maximum of only 84 bushels of market oysters for each 100 bushels of seed. It must be pointed out that these figures are only approximations, for the original estimate of 0.5 bushels in each 100 bushels of seed was derived from the average length of these oysters. It is probable that this represents an over- estimate rather than a low figure, thus the maximum volume may be too high. Nevertheless, this group of small seed oysters probably con- tributes significantly so the higher yields obtained by planters in the upper estuaries, where driils are not a problem. In Table III the same computations are applied to the group of larger oysters in Wreck Shoal seed, recognized as useful seed by the planters. Here, although the maximum volume, reached in 22 months, is only about five times that of the original planting, the oysters when planted make up about half the entire volume of seed. Thus, a yield of about 2.5 bushels is possible from the larger oysters in each original bushel of seed. Another consideration must be introduced in combining these two sets of figures to derive the total yield. At planting, few, if any, of the oysters are of market size, and our growth studies have shown that some of the survivors may never reach the arbitrary length of three inches or over that we have used to designate market oysters. Allowance has been made for the size factor in computing the yields in Table IV. It will be noted that two maxima in the yield of market- sized oysters are reached, the first, of about 2.8 bushels for one, in 22 months after planting, and the second, of about 2.9 bushels for one, in 34 months. The slightly larger value for the second maximum pro- bably is not significant, and the greater total volume of oysters in existence at 22 months (see column 4) would almost certainly contain significant numbers smaller than three inches worth shucking so as to boost the computed yield. Applications to Oyster Planting The yields discussed above are illustrated graphically in Figure 6. Cbviously, it is not wise to apply results obtained from tray cul- ture directly to practical oystering problems, at least without attempt- ing to determine how these rates of growth and mortality compare with those on planted grounds. Considering first the growth rate, it is fairly certain that the seasonal variations observed in trays are similar in order of time, -226- Yield in bushels per /00 bushels of seed °OMBY PHAM IIA SONDJ FMAMIIASOND/FMAMIIASOND Fig. 6. The yield, in bushels of live oysters, in successive months after planting in trays at Gloucester Point, Virginia, from Wreck Shoal seed. The upper curve represents all oysters, including the current year spat. It must be noted that the original yield at planting is only about one-half the actual bulk of the seed, because each bushel of seed oysters contains about half a bushel of shell, to which the oysters are attached, and fouling organisms. The three lower curves represent respectively the yield of oysters larger than three inches in length from the entire volume of seed, from the larger seed oysters, and from the current year spat. 227 if not in magnitude, to growth cn planted ground. Jt can be assumed alse, that because the trays are exposed to a more effective cire n culation of water, and because there is relatively less silt to be rejected, oysters in trays will grow faster thar those on the bottom. This appears to be supported by the available data, and we hope to gotain better data soon. The seasonal pattern of mortality rates in trays at Gloucester Point appears to bear a close relationship to the seasonal eyele of water temperature (Hewatt and Andrews, 1954). Therefore, for the sources of mortality ca which tray oysters are subject, it would not appear unrsasonable to assume that a similar mortality pattern would apply on planted bottom, with respess to time though not necessarily in magnitude. Oysters om the hottom, however, are subject to death from other important causes, the depredations of drills or screwborers being perhaps the principal factor. Unpublished observations of cyster drill activity in the vicinity of Gloucester Point show that the activity of these predators is closely associated with the temperature cyele, and these obseryations carn reasonably be extended to cover the activities of other predators, all of which are cold blooded and thus quite sensi- tiye to temperature variations. Thus it seems safe tc assume that the average seasonal pattern of mortality on the bottom is similar to the pactern observed in trays. Hewatt and Andrews (1954) report annual mortalities of about 25 per cent for oysters in trays at Gloucester Foint, but the calculations made earlier in the present paper suggest that the annual mortality on the bottom is of the order of 37 per cent. The ansual mortality rate associated with bottom factors, therefore, is about 16 per cent. This Suggests that the fungus Dermocystidium marinum is a more serious source of mortality than the oyster drill, at least insofar as the larger seed oysters are concerned. There seems to ke good reason to believe that growth and mors tality oa planted bottom differ from the same rates in trays chiefly in magnitude rather than im seasonal pattern. Thus, curyes showing the yield on plamted ground at various levels ef growth and mortality can be constructed by adjusting by appropriate faztors the rates deter-= mined from tray culture. Such a series of curves, based on a growth ‘rate three-quarters as great as the rabe im trays, is illustrated in Figure ‘7. It is worth noting that the maximum yield, unless mortality is exceptioually low, is reached about a year and a half after planting. If these oysters were not harvested umtil fall, a mere three or four months after the maximum yield was reached, the yield would have failen consideranly, and although the spring growth of the follewing year would eause the yield to increase again, it would never apparently reach the former -Lleyel.. Families of ecuryes, based on various rates of growth and mortality, can be constructed readily. The oysterman can determine the rates char-= acteristic of his grounds by methods described by Hopkins and Menzel 22286 Mortality rates: Z| \ 8 Growth rate O75: ! ' ' ' i] i eee) ee Oe ees / ee OO OOO 6 1 ~~ 8 40 Yield in bushels per /00 Lushe/s of seed Oo Bs i] ONDSFMAMSSASONDISJFMAMISASONDSFMAMSSA SOND Fig. 7. Hypothetical yield curves, based on growth rate three-quarters as rapid (0.75k) as the rate in trays at Gloucester Point, Virginia, and at mortality rates equal to one-half, one, one and one-half, and two times (0.5q, q, 1.5q, and 2q) the rate in trays. The figures within the arrows represent the number of oysters per bushel. 229 (1952) and by selecting the appropriate curve, can determine when to haryess for the greatest yield in busheis of oysters. The Bushel Count at would be of little benefit to the planter if he were to haryest his oysters at the point of maximum yield, only to find that the size was too small for economical shucking. The oysterman's eriterion of size is the count per bushel, and it is useful to know the relavionship between the average length, or average weight of oysters, and the bushei count. By actual measure of oysters of various Sizes, grown im trays at Gloucester Point, we haye found that the relationships between both length and weight with yield, can be expressed as straight lines on logarithmic coordinates, as with the length-weight relationship in Figure 4. Furthermore, the ree= lationship of weight to yield can be expressed roughly by an even simpler expression: ne (3 x 104) W where m is the number of oysters per bushel, and w is the average weight or the oysters in grams. in Figure 7 the counts per bushel at the points of inflection ave given as numbers within arrows at the appropriate positions. is Further investigation Necessary? several factors important to the oysterman have been ignored in the preceding sections. Perhaps the most important is the question: “Ave the oysters in prime condition at the time of maximum yield, and is the shucking ratio high?" The planter is perhaps better able than the biologist to answer this question. The producer wiil also be interested in the demand and the price, for he may find it necessary often to hold his crop past the point of meximum yield whether he wishes to or not. Some practical considerations such as the labor supply, will tend to foree him to spread his operations over as many months as possible; others, such as the necessity te obtain high yields, favor a concentration of effort. VTechrological developments that would eliminate such conflicting pres- sures, such as the discovery of mechanical shucking methods and the development of quick freezing processes, seem to offer the best hope for solution of these preblems. . 2230-6 Much more accurate information is necessary on the growth and mortality rates characteristic of planted bottom. It is hoped to get this information in two ways, by examining representative samples from planted grounds in various areas of the Bay and estuaries, and by experimental plantings of marked oysters. We hope also that some planters will be stimulated by these findings to examine our figures carefully. If our argument appears reasonable, we would urge them to experiment by harvesting at various time intervals. It goes with- out saying that for maximum results, such experimentation should be planned carefully and should be accompanied by careful and systematic recording. The Virginia Fisheries Laboratory will be willing and anxious to cooperate in such experiments. a231 = Hewatt, Literature Cited Willis G., and Jay D. Andrews. 1954. Oyster mortality studies in Virginia. I. Mortalities of oysters in trays at Gloucester Point, York River. Texas Jr. Sei. 6(2): 121-133. Hopkins, Sewell H., and R. W. Menzel. 1952. How to decide best time to harvest oyster crops. Atlantic Fisherman 33(9) Oct. 1952: 15, 36+37. Cwen, H. Malcolm. 1953. Growth and mortality of oysters in Louisiana. Ricker, Ricker, Ricker, Bull. Mar. Sci. Gulf & Carib. 3(1): 44-54. William E. 1945. A method of estimating minimum size limits for obtaining the maximum yield. Copeia 1945(2): 84-9h. William E. 1948. Methods of estimating vital statistics of fish populations. Indiana Univ. Publ., Science Ser. 15. Bloomington, Indiana, v 1Ol pp. William E., and R. E. Foerster. 1948. Computation of fish production. Bull. Bingham Ocean. Coll. 11(4): 173-211. Walford, Lionel A. 1946. A new graphic method of describing the growth of animals. Biol. Bull. 90(2):141-147. =p2o5 TABLE I Computation of seasonal growth rates for Wreck Shoal seed oysters transferred to trays at Gloucester Point, The lengths are inserted for reference, and are not used in the computations, One year of age or older Current Year Spat poet oe Logio Logio . fo) Lengt weight Diff- k Length weight Diff- k month inmm, , erence in™M™M. in gms, erence in October 12 -0,43 48 1,22 0,30 0.69 0.13 0.30 November 15 -0.13 54 1535 0.21 0.48 0.11 0,25 December; 18 +0,.08 59 1,46 | 0,06 0,14 0,03 0,07 January | 19 0,14 60 1.49 0.05 Onti2 0.00 0,00 February 20 0.19 60 1,49 0,00 0.00 0.00 0.00 March 20 0.19 60 1,49 0,05 0.12 0,00 0,00 ‘April 21 0.24 60 1,49 0.11 0.25 0,08 0,18 May 23 0,35 64 1,57 0.38 0.88 0,08 0,18 June 32 0.73 69 1,65 Os2t 0, 62 0.08 0,18 July 40 1,00 74 1.73 0.19 0,44 0.05 0,12 August 47 1,19 78 ens 0.13 0.30 0.06 0,14 September 53 loose 81 1, 84 0,11 0,25 0,03 0,07 October 58 1, 43 84 1, 87 0,11 Ona5 0.03 0,07 November 63 1,54 86 1.90 0.08 0.18 0,03 0,07 December 67 1,62 89 1,93 0,02 0.05 0.01 0,02 January 68 1, 64 : 90 1,94 0.01 0.02 0,03 0.07 February 69 165 91 1,97 0,00 0.00 < 0, 00 0,00 March: 70 1,65 "92 1,97 0.00 0.00 0,00 0.00 -233- Beginning of month April May June July August September October | November | December January February | March April May June July - agest September October November December 88 94 Current Year S TABLE I (continued) at One year of age or older Length °810 | pitt weight k in mm, invgmiss Czence 92 1,97 0.03 0,07 94 z.00 0,05 0,12 97 2,05 0,03 0.07 100 2,08 0.03 0.07 102 2,11 0,01 0,02 103 2,12 0.01 0,02 104 2.13 0.00 0,00 105 2,13 0.00 0.00 105 2.13 0.00 0.00 106 2,13 0,00 0,00 106 2,13 0.00 0,00 106 2,13 0.00 0.00 106 2,13 0,00 0,00 106 2,13 0,03 0.07 107 2,16 0.01 0.02 108 2,17 0,02 0.05 109 2.19 0,00 0.00 109 2,19 0,00 0.00 110 2.19 0.00 0,00 110 2,19 0,00 0.00 110 TABLE II Computation of relative biomass, and absolute biomass per original bushel of seed oysters, for the current-year spat in Wreck Shoal seed, beige: Absolute Beginning Change . biomass of k q k-4q in Bro mae 2 ee ptoONbEe Biomass of seed month October 100 0,5 0,69 0.01 0,68 +0,97 November 197 1.0 0.48 0,00 0.48 +0.61 December 317 1,6 0.14 0,00 0,14 +0,15 January 365 1.8 0,12 0.01 Oelei +0,12 February 409 2.0 0,00 0.00 0,00 0,00 March 409 2.0 | 0.12 0,01 0.11 +0, 12 April 457 2.3 On25 0.00 0,25 +0, 28 May 585 2.9 0. 88 0.01 0,87 +1.39 June 1,398 7.0 0.62 0.02 0.60 +0. 82 July 2, 544 12,7 0,44 0.03 0,41 +0,51 August 3, 841 19,2 0,30 0,08 0,22 +0,25 September 4,801 24,0 0.25 0.08 0,17 +0,18 October 5,665 28.3 0,25 0.03 0.22 +0,25 November 7,081 35.4 0,18 0.01 0.17 +0,18 December 8, 356 41.8 0,05 0.00 0.05 +0,05 January 8,774 43.9 0.02 0.00 0,02 +0,02 February 8, 943 44,7 0,00 0,00 0,00 0,00 March 8, 948 44,7 0.00 0.00 0.00 0.00 April 8, 948 44,7 0,18 0.00 0,18 +0,19 TABLE II (continued) Absolute Beginning Change biomass of k q k-q He Biomass) |) en g0.bu: month Biomass of seed May 10, 648 0.14 0.01 0.13 +0,14 June 12,139 60.7 0.12 0.02 0,10 +0, 10 July 13, 353 66, 8 0.07 0,06 0.01 +0,01 August 13, 486 67.4 0,07 0,12 -0.05 -0.05 September 12,812 64,1 0,07 0.10 -0, 03 -0.03 October 12, 428 62,1 0,09 0,04 0,05 +0,05 November 13,049 65.2 0.07 0.01 0,06 +0,06 December 13, 832 69,2 0.02 0.00 0.02 +0, 02 January 14,109 70.5 0,02 0.01 0.01 +0,01 February 14, 250 lene 0,02 0.00 0.02 +0,02 March 14, 535 Uitte U 0.02 0,00 0.02 +0,02 April 14, 826 74,1 0,02 0,00 0,02 +0.02 May 15,122 T5a6 0.07 0.01 0.06 +0, 06 June 16,029 80.1 0,07 0,02 0.05 +0,05 July 16, 830 84, 2 0.02 0,04 -0,02 -0.02 August 16, 493 82, 5 0.02 0,12 -0,10 -0.10 September 14, 844 74,2 0,02 0.16 -0.14 -0,. 13 October 12,914 64, 6 0.02 0,05 -0,03 -0,03 November 12,526 62,6 0.02 0,02 0.00 0,00 December 12, 526 62.6 0,00 -236- TABLE III Computation of relative biomass, and absolute biomass per original bushel of seed oysters, for the yearling and older oysters in Wreck Shoal seed, Absolute Beginning Change Nominee of | k q k-q in Biomass periculnn: month Biomass of seed Cctober 100 50 0.30 0.01 +0.29 +0, 34 November 134 67 0.25 0.00 +0,25 +0,28 December 172 86 0.07 0,00 +0,07 +0,07 January 184 92 0.00 0,01 -0,01 -0.01 February 182 91 0,00 0,00 0. 00 0,00 March 182 91 0,00 0,01 -0,01 -0,01 April 180 90 0.18 0.00 +0, 18 +0,19 May 214 107 0,18 0.01 +0,17 +0,18 June 252 126 0,18 0,02 +0, 16 +0,17 July 295 148 0.12 0,03 +0,09 +0,09 August $22 161 0,14 0.08 +0,06 +0, 06 September 34] 170 0,07 0.08 -0,.01 -0,01 October 338 169 0.07 0.03 +0, 04 +0,04 November 352 176 0.07 0,01 + 0,06 +0,06 December 373 186 0,02 0.00 +0,02 +0,02 January 380 190 0.07 0,00 +0,07 +0,07 February 407 204 0.00 0.00 0.00 0.00 March 407 204 0.00 0.00 0.00 0.00 April 407 204 0.07 0,00 +0,07 +0,07 TABLE III (continued) Recina; Change Absolute eginning ; of k q k-q in Biomass biomass month Biowinee per 100 bu, of seed May 0.12 0.01 +0,11 +0,12 June 487 244 0,07 0,02 +0,05 +0,05 July 511 256 0,07 0,06 +0.01 +0.01 August 516 258 0.02 0,12 -0,10 -0.10 September 464 232 0.02 0.10 -0. 08 -0,08 October 427 214 0,00 0.05 -0.05 -0,05 November . 406 203 0,00 0,01 -0.01 -0.01 December 402 201 0,00 0.00 0.00 0.00 January 402 201 0.00 0.01 -0,01 -0,01 February ‘ 398 199 0.00 0,00 0.00 0,00 March 398 199 0.00 0,00 0.00 0,00 April 398 199 0,00 0.00 0.00 0. 00 May 398 199 0,07 0,01 +0.06 +0,06 June 422 211 0,02 0,02 0.00 0,00 July 422 211 0,05 0.04 +0,01 +0.01 August 426 213 0.00 O22 -0.12 -0,11 September 375 188 0.00 0,16 -0, 16 -0,15 October 315 158 0.00 0.05 -0.05 ~0.05 : November 299 150 0.00 0,02 -0.02 -0.02 December 293 146 TABLE IV Total biomass resulting from the planting of 100 bushels of Wreck Shoal seed intrays at Gloucester Point, Absolute biomass aan Beginning per 100 bu. of seed market oysters Bushels of market of per 100 bu.of seed oysters por 100 bu, month of sced Spat Young Total October OS 50 50 0 12 0 + 6-= 6 November | 1,0 67 68 0 18 One eli2ye= sled December 1.6 86 88 0 19 0+ 162+ 16 January Ves 92 94 0 20 ORS Se Sees February 20 91 93 0 20 a Ue Ss alts March 2.0 91 93 0 20 Uy ap hc UGS) April ons 90 92 0 20 ese iG May 2.9 107 110 1 23 (eS A es 7A‘) June Caw 126 133 3 31 Qa 39 a oF July ras 147.5 160 5 38 1+ 56= 57 August 19,2 161 180 8 41 Zt 166" =" 168 September | 24,0 170.5 194 14 43 B) ee Wiehe w/e) October 28.3 169 197 19 53 5+ 90 = 95 November 35.4 176 211 21 61 7 + 107 = 114 December | 41,8 186.5 228 25 75 10 + 140 = 150 January 43.9 190 234 26 84 11 + 160 = 171 February 44,7 Z03%.5 248 27 85 12 + Wse= 85 March 44,7 20315 248 28 85 Ns} ab NYS SS HS April 44,7 2035 248 29 86 13 + 175 = 188 May BS}, C2 outa rat(\ 31 87 16 + 189 = 205 June 60.7 243.5 304 36 90 (yf, ts fasl®) ep Asi July 66,8 255.5 gia, 40 91 27 + 232 = 259 August 67.4 258 525 55 95 37 + 245 = 282 September | 64,1 232 296 60 95 38 + 220 = 258 October O21 ANS 5S) 276 70 95 43 + 203 = 246 November | 65,2 203 268 US) 96 49 + 195 = 244 December 69.2 201 270 80 96 55 + 193 = 248 January OD 201 2c 83 97 Be) Ge) fie) Sy als February le 2 199 270 84 98 Xt) ap GIG) eS AGI March UU 199 ne 84 98 61 + 195 = 256 April 74,1 199 273 64 + 195 = 259 May U5 199 275 66 + 195 = 261 June 80,1 211 291 72 + 209 = 281 July 84,2 211 295 77 + 209 = 286 August 82,5 ils! 296 75 + 213 = 288 September | 74,2 188 262 69 + 188 = 257 October 64.6 158 223 61 + 158 = 219 Novernber 62.6 150 213 59 + 150 = 209 December 62,6 146 209 59 + 146 = 205 =239= mae = a shared Sank alee od bdbm Po.od 00) NG. : gacuY sae > eee ' ‘ ® — a cr . o4@ ses Pere e~se > | we : = = Le gn tas m2 ee —= = # hates “ts = #4) 4 1. . ° P te = t ” ety aa = sé $ ‘$ ne ¢ @ os; z eet oe * i se . tae °. * &5 ir? . } wy m s y Ba a @ s 4° = PHS «<2 eae y + # = of b- \ ray sae we éLissuastes Te. 2 os == 455% ~ » > — eb ee a4 = ae +o © | i, - > = ~~ = - SHELLFISH SANITATION AS KELATED TO THE EXPORT AND UMPOry TRADE TN CANADA J, Ro Menzies Department of National Health and Welfare, ttawa, Canada When shelifish sanitation in North America ie considered, it is not surprising that the past thirty years must be revieved. Wo account of shelifish sanitation in North America would be complete without re- ference to the events of late 1924 and early 1925 which are clearly etched on the memories of both producers and health officials. The typhoid fever cases then attributed to the eating of shellfish were undoubtedly responsible for most of the control procedures which have since been developed. It was generally agreed that control methods mist be developed which would restore public confidence in shellfish as a food. The sale of shellfish in Canada wae seriously affected by the publicity given to the occurrence of typhoid fever in the United States. Since most of the oysters eaten in Canada were imported from your country some measure of protection was needed and regulations were passed on July 3, 1925, under authority of the Fish Inspection Act which re- quired that "Each consigument of oysters imported into Canada, whether in the shell or in bulk, shall be accompanied by a certificate by 2 competent authority, that will be satisfactory to the Department of Health, that will show that the oysters contained therein are a safe food product". This wes generally interpreted to mean a license or certificate issued by appropriate state agencies and endorsed by the Public Health Service of the United States Treasury Department. This regulation is still in force. There does uot appear te have been similar legislation invoked in the United States at that time in respect to Canadian shellfish but the States of Massachusetts and New York, which then received the bulk of our then very limited supply, were rightfully coxcerced with the control exercised over producing areas and production methods in Canada. Thus the Federal Department of Health found it necessary to concern it- self with these matters and began to issue certificates to exporters who were shipping to the United States. From a consideration of the correspondence at that time it is evident that the Federal Hesith Department of Canada accepted the responsibility of isswing export certificates rather reluctantly and opposed, for a time, the proposal that zhelifish other than oysters should be considered. Thie reluctance is not surprising since in- vestigations required sending personnel from Ottawa to carry out field work, there being no field offices in the vicinity of the producing areas. This phase of the control programme contimed for some ten years o2hQn at which time detaiied Baateary surveys of individual producing areas were started. Unly ome public health engineer was available for shell- fish control work ani only a part ef his time was assigned to this work. In the meantime there had been developed an export trade in scallop meat, which led to a demand by the States concerned for regulation of the industry and cert ification by the Federal Department of Health. A detailed study was required of production metheds and equipment on which was based legislation establishing minimm sani- tary requiremerts. Aise during the thirties seme interest was shown in the export of soft-shell clams. The first application for an export eertificate for shucked clams was received in 1938. The trend toward the export of shucked clams was accelerated by Canadian legislation forbidding the export of softesheii clams in the shell from New runswick in 1945 ani Neva Scotia in 1946. Other Pecters which had a definite effect on the export of shellfish from Canada were (a) the outbreak of a disease among oysters in 1915 which spread to ether areas in the twenties and thirties and destroyed a high percentage of the population, and (b) the discovery of toxicity in ciams and mussels in the Bay of Fundy area. The former reduced our control problem considersble for a time but the same canngt be said of the latter. Patal poisonings, believed te have been caused by the consumptios of mussels, were reported in Nova Seotia in 1936, These occurred on ae Bay of Fundy shore and led to action by the Provineiai government forbidding the use of mussels in the general area. ‘urther studies were them undertaken for the purpcse of identifying all the areas subject t6 toxicity amd it was found that some of the most bree tive sec etic ens im the Province of New Srunswick, berdering the Bay of Pumiy, were also affected. Arrangements were made with the Department of Fisheries and the Fisheries Research Board of Canada for the regular collec a Of samples from which extracts were sent %6 the Laboratery ef Ayzlene of the Federal Department of Health for testing. Mice were injeeted with an acid-aqueous solution of the extract and the degree of tos psonngesiesd determined by the time required tS praduce ¢ death, As a result of this study, which is still being pursued, the danger season, ENE dangerous areas and the dangerous species af sheilfish have been identified. Ts fee be poiwmted out that it required several years of care-= ful investigations before a thorough understanding of this problem was gained. During the early years of the study those exporters holding certificates covering toxic areas had their certificates cancelled during periods when toxic conditions prevailed. ‘This was too drastic sinee it prevented those exporters from taking clams from non-toxic areas. The sl li procedwre is to close the toxic areas by legis- lation which is exforeed by the Department of Fisheries. Certificate holders are advised of the closures as they oceur and also of the release of the areas when toxicity falls to permissible levels. eT It was later found that toxicity was also a problem in the Province of Quebec on both the south and north shores of the St. Lawrence River. This, until recently, was a local problem since shellfish were not exported from the province of Quebec witil early this year. A comprehensive programme of sampling is sow underway in Quebec and as a result toxic conditions have been found along the Gaspe Coast of thet Province adjacent to the Gulf of st. Lawrence. It should also be noted that toxicity is widespread on the Pacific Coast of Canada and presents, in some respects, a more difficult administrative problem since it persists throughout the year and becsuse it has not been possible to observe any definite pattern in relation to the severity of the occurrence. This is contrary to our experience in the Bay of Fundy area where toxicities are usually low or nonexistent in most areas except in the summer and early autumn. Prier t¢ 1954, the limits for toxicity in shellfish exported from Canada was less than 200 mouse units per 100 grams of meat, this being the lowest detectable amount with the test being used. This year, by agreement with the United States authorities, the permissible limit was established as 400 mouse units per 100 grams of meat. Reverting now to the sanitary controls established on our Atlantic Coast reference has been made to detailed sanitary surveys of producing areas by a public health engineer. It was agreed that the Federal Department of Health would be responsible for this phase of the control pregramme some twenty years ago and this practice has since been followed. For several years the sanitary survey was earried out first and a decision was then made as to whether a bacteriological investigation was needed. In recent years an effort has been made to have both studies made together but this has pre- sented some difficulties because of limited laboratory facilities. In carrying cub these investigations and in submitting re= commendations fer closures because of pollution, the Federal Department of Health acts in an advisory capacity to the Federal Department of Fisheries which has the legislative authority to control fisheries. The actual closures are made by regulations under the Fisheries Act, 1932. This act is basic to the contrel of the whole fisheries in=- dustry in Canada sinte control of fisheries was as signed to the Federal Government by the British ‘Worth America Act under which Canada was founded, Authority to administer the Fisheries Act has since been delegated to some of the provimees but is administered by the Federal Department of Fisheries in the Provinces of Newfoundiand, Prince. Edward Island, Nova Seotia, ani New Brunswick which, until this year, have produced the shelifish exported from our Atlantie coast to the United States. Reference should be made to an advisory group which was first ee in 1941, was allowed to lapse during World War IT, and was evived in 1945. It is known as the Interdepartmental Shellfish Committee and includes representatives of the Federal Departments of Health and of Fisheries and the Fisheries Research Board of Canada, This come mittee meets annually to diseuss all phases of the shellfish industry including the review of data collected during sanitary and bacterio-= logical surveys. Prior to the meeting, officers of the Department ef Health review the field work of the previous year and submit to the cémmittee recommended closures required by excessive pollution. The Interdepartmental Shelifish Committee considers these recommenda- tious, usually in subcommittee, when the Department of Eisheries is afforded an oppertunity to present its views. Recommended closures endorsed by the full committee are then transmitted to the Department of Fisheries for legislative action. A similar committee deals with control problems oe the Pacific Coast of Canada. In recent years representatives of the U. S. Public Heaith Service have attended committee meetings. The advice and information supplied by them has been invaluable. It will be apparent that there is considerable delay involved in handling closures related to poliution between the time of carrying out the field work and legislative action. Consequently it is some- times considered expedient to deal with these matters more quickly and senior officers of the Departments of Health and Fisheries then agree on the action to be taken, necessary legislation is enacted and the open portions of the areas are released for production. Why are the problems related to closures still of importance after some twenty years of fairly detailed investigations? Two conditions affect the situation. The first, of course, is the gradual development of the export industry from a few cysters toa several species ef sheilfish which are taken from an increasing number of producing areas. The second condition is lack of personnel and equipment to carry out field investigations. At the present time the fieid staff of the Federal Department of Health consists of four professional engineers who are respoesibile for all sanitary surveys in our four eastern provinces and only a part of their time can be assigned to shelifish control work. ‘The Leboratory of Hygiene assigms a mobile lsboratery with a bacteriologist and a technician for a few months Gach year. During the early years of the export trade, fishing was centered around the more prodvetive areas, More recently, with an imerease in the number of closures and the increasing demand for shell- steck, every potential producing area is being explored. If even a small shelifish bed is fowad there is an immediate demand for permission te fish it. Within a short time data should be available regarding sanitary conditions in all our Atlantic coastal waters. Fishing for export is not permitted in areas which have not been surveyed. At the same time, ef course, changing conditions frequently require re-surveys which, in turn, result in changes of the closure limits. Some success has been achieved in having existing sources of pollution corrected. =D? Brief reference has been made to an informal agreement existing between the U. S. Public Health Service and the Canadian Department of Health in regard to certification of shellfish producers. This agree= ment has been very helpful in the shelifish control programme in Canada. This mutual agreement was formalized on April 30, 1948, and included the following points: i. Approval by both countries of a manual of recommended practice for sanitary control. 2. Kath country agreed to report to the other the degree of compliance. 3. Both countries agreed to facilitate inspections required by the other. 4, The agreement may be terminated by either country on thirty days notice. The Department cf Health agreed to use the United States manual as a @uide in assessing satisfactory compliance with accepted standards. Completion of the revised manual now being prepared by the U. S. Pubiic Health Service is awaited with much interest. Our views were requested and have been considered. The list of shellfish shippers published in Washington has been used continuously in cur control of imported shelifish. In issuing export certificates to Canadian producers this is the procedure. in the Provinces of Newfoundland, Prince Edward Island, Nova Scotia, and New Brunswick, the exporter completes an application in triplicate which is signed by the Fisheries Officer in the area. The application lists the areas from which it is proposed to take shell- fish, the varieties to be handled, and whether they are to be exported as sheilstock or shucked} a statement is inciudecd in which the appli- cant agrees "to conform to all regulations, rulings or requirements in the taking, handiing, packing and shipping of for export purposes", with the word "shucking" added if shucked shellfish are to exported. The application is then sent to the district offices of the Department of Fisheries and of the Public Heaith Engineering Divi- sion of the Federal Heaith Department where senior officers recommend the application when they are satisfied that conditions in the pro- ducing areas and shucking plants (for shucked stock) are satisfactory. The application is then sent to Ottawa where it is signed for the Deputy Ministers of Health and Fisheries. Only then is an export certificate issued. This is done by the Chief, Public Health Engineering Division of the Federal Health Department. (Administrative control of the shell- fish industry in Canada, in respect to the Health Department's interest was assigned to the Public Health Engineering Division in 1943.) A copy of the certificate is sent to Washington and the name and certifi- cate number of the exporter is then included on the next list of shell- fish shippers which is published twice monthly in Washiugton. eehhe The control procedure which has been outlined has dealt with our four eastern preyinces. OGuly brief reference need be made te crontrel procedures in the Provinces ef Quebec and British Columbia since, in those provimeesy it is similar te that followed in the United States. Controls are established by the prevince which issues certificates to Seaeee producers. The Federal Department of Health, -wnen it is satisfied that the control programme meets its requirements, transmits to Rabhinpees as list of the certificates. This action implies endorse= oe ef the provingial comtrol programme and may be discentinued when- ver it is felt that adequate precautions are uct being observed. Similerly whem any producer who receives an export certificate directly from the Fetersal Department ef Health fails te live up to his signed agreement, suspension or caucellation of his certificate serves as an effective control. It is very seldom that such drastic measures must be taken. Im March, 1950, tentative limits of bacterial contamination were established im relation to shellfish imported into Canada. Three classes were established, (1) asceptable, (2) acceptable on condition, and (3) rejectable. Reports of analyses are sent to Washington. In the “seceptable on confiition" category competent authorities examine pro- cessing and handiing methods to determine if possibie the cause of the high bacterial eontent. While data is limited, this arrangement seems to have been effective in securing a better quality product. This has been an incemplete review of Canadian snelifish control but will perhaps serve to indicate that the shelifish industry is not a simple one to deal with from the point of view of the government official. Im fact there are prehably more administrative headaches associated therewith than with any other phase of our work and yet it has its fascinations too. New -preblems require consideration and additional knowledge in ey fields is essention before maximum use can be made of our shelifish without endangering public health. The wesenee of sbundant shelistock in polluted areas, from the point of £ the health department, is probably the most serious threat to adequate control aud is no doubt recognized as such by the industry itself. Therefore some satisfactory mears must be fowsd te purify and utilize polluted shellsteck in the iia of both the industry, the regulatory athorities, and the consumer. it is a challenge to our research workers aol shevld have the acti ive support of the industry. pheuleé polluted shellfish by intent or accident reach the consumer we may again experience a recurrence of disease similar im character and results to that of thirty years ago. Every effort must be made to prevent such an oceurrence. This deseription of our control programme has emphasized the role of the Federal Department of Health. It showld be noted, however, that a close liaison exists with the Federal Department of Fisheries. Its officers enforce the closures required by pollution and toxicity. Without the wholehearted support of that department 1+ would be impossible to maintain an acceptable degree of compliance with sanitary requirements. THE SANITARY ASPECTS CF IMPORTATION OF SHELLFISH TiVO THE UNITED STATES Richard S. Green U. S. Department of Health, Education, and Welfare, Washington, D. C. It is a real privilege to report to this group on the importation of shellfish from abroad. This.is one of the most complex shellfish sanitation problems facing health and food control officials and the incustry itself. T am especially pleased that Mr. Menzies ef Canada could be with us today. His excellent discussion of the development and operation of the agreement on shellfish sanitation between the Canadian Department of National Health and Welfare and the Public Health Service has presented a fine point of departure for my talk. I feel rather sure, also, that some of the implications of recent developments will be of direct in- terest and mutual concern to him and his Canadian colleagues, and so it is fine to have this opportunity for us to exchange views. Some of you may remember that last year I had an opportunity to outline for you in broad terms the impact of several recent technical and administrative developments on shellfish sanitation control in the United States. On that occasion, mention was made cf the question of foreign shellfish imports, but neither time nor the state of develop- ments then existing permitted more than a brief statement of this partic- ular problem. During the past year, there have been many significant developments, and a very considerable amount of time has been devoted to deliberations on this problem. We believe that it is especially im- portant for members of the Oyster Institute and related groups to be brought up-to-date. Since all of you are familiar with the program of shellfish sanitation control of the Public Health Service, it will be unnecessary for me to give you the details of our technique of cooperating with the States on the endersement of State operations. You know that the listing of certified dealers in our routine compilation, designed for use in consumer areas, is the backbone of this system of voluntary control. It was the acceptance of this concept of the certification system, and an understanding of the health department surveiliance involved in it, which brought about the reactions that followed the recent growth of shipments of shellfish from abroad. So many health officials in the United States require oysters and clams to be from certified dealers that shipments from foreign countries other than Canada have not easily been sold, even though admitted legally to the country under the terms of the Food, Drug, and Cosmetic Act administered by the Food and Drug Administration. That agency is responsible for permitting or denying entry to food imports. Faced with these difficulties, representatives of the foreign countries concerned and the United States importers have ~2h6~ asked the Public Health Service how their shellfish can b & manner similar t6 domestic and Canadian shellfish. At some State and local health departments have askea the Se they should do about foreign shellfish which have appeared on the market. Thus, the Public Health Service, which has no legal! jurisdiction, but which has guided the domestic contrel program for many years, has been eaught in the middie, so to speak, without being able to furnish answers that are satisractory. I co not heve the final answers to thes= problems to give you today, but I can tell you what we are facing and some of the reasoning we have used. Since this program was developed, and still functions, in cooperation with the States and industry, the Public Health Service dbes not intend to proceed in the direction of major adjustments without first discussing these matters with these groups. As indicated above, responsibility for permitting or denying entry of such shipments when presented at perts of entry, under the terms of the Food, Drug, and Cosmetic Act, lies with the Food and Drug Administration, a sister agency of the Public Health Service in the Department of Health, Education, and Welfare. Whenever the Food and Drug Administratien finds, from the examination of samples or otherwise, that such shipments of shellfish have been produced under unsanitary conditions, or are otherwise adulterated or misbranded, their entry is refused admission, it is emphasized that refusal of admission to entry of food is based on evidence of adulteration, or misbranding and that, in the absence of such evidence, entry must be permitted. Few will deny that, in the case of shellfish (oysters, clams, and mussels), a mich greater degree of protection is afforded the con- sumer if sanitation controls can be based upon a sure knowledge of conditions surrquoding the growing, harvesting, packing, and shipping of the shellfish, instead of on an objective examination of the final packages as received in the market. This principle of "control at source" has been the basis of the Public Health Service and the State domestic shellfish programs, as all of you who have seen State inspectors at work will attest. The effectiveness of this program may be judged by the low incidence of shellfish borne enteric disease, in spite of the fact that we have had to be concerned at all times over such problems as king sure that no shellfish are used from the mere than 400 areas on our coast line that are legally wlosed because cf pollution. Objective examination of samples collected from fereign shell- Tish shipments at the time of entry do not always give satisfactory evidence of sanitary conditions under which the shellfish were produced and packed. It is, therefore, difficult to decide which shipments should be admitted anc which denied entry, particularly when bacteriological findings do not clearly show the presence of significantly large numbers of coliform organisms, and when other objective findings are satisfactory. Before getting further into this discussion, I shovwld like to give you a few pertinent figures and some background in*ormation about 2h To the origin and size of foreign shellfish shipmerts which have arrived in the United States. Prior to World War iI, few shipments of sheil- fish came into the United States except from Canada and Mexico. Mr. Menzies' paper has already discussed the Canadian situation. in the case of Mexico, most of the shipments were pismo ciams, although an occasional shipment of shucked oysters was offered for entry. In 1952, somewhat over a half million pounds of clam meats were imported from Mexico, principsliy through the San Diego and Los Angeles ports of entry. In 1953, this figure was close to three fourths of a million pounds. It is believed that almost all of these clams are used in production of heat processed clam chowder; apparently, no attempt has been made to distribute the unprocessed clams beyond the State of entry, and thus the question of certification has not arisen. With the expansion of the frozen food industry since the end of World War II, several other foreign countries have developed an interest in the United States as a market for bivalve shellfish, principally frozen clams. Japan, Iceland, Australia, The Netherlands, France, Spain, China, and Panama have ali exported or indicated an interest in exporting frozen clams, mussels, or oysters to us. It appears that the total volume of shellfish shipped to the United States has been relatively small, somewhere in the neighborhood of one or two per cent of our domestic production. It should be borne in mind, how- ever, that all these shellfish, except those produced in Canada, have been faced with the restrictions resulting from lack of certification. There is no easy way to predict what the ultimate volume of shellfish imports might be if that situation did not exist. While it is beyond the scope of this paper, and entirely out of the field of public health, to dwell on such facts as dollar exchange value, tariffs, and the importance to these foreign governments of trade with the United States, it would be a mistake to pass over these factors without mention. Specialists in such matters have analyzed the situation about as follows: International trade in frozen shelifish is now possible on a worldwide basis, and producers of shellfish in far distant countries are eager to help to satisfy what appears to be an increasing demand for shellfish in the United States. The interest of foreign governments stems from the importance of trade to their national economies and the importance that all free world countries attach to closer ties with the United States. It is in our interest to foster such ties and to enable friendly countries to gain strength through trade. Their welfare and ours require that they be aple to earn dollars from their exports to the United States in order to buy the products of ow farms and factories. Japan and Iceland, in particular, must sell us more goods than they now do to pay for all of the American products they need and want. Iceland has virtually nothing except marine products to sell abroad. In the case of Japan, marine products are among the few commodities which can be produced without the use of imported raw materials. In the last few years, the governments of these two countries, -2h6- and of The Netherlanis and Austr es also, have made kmown to the Department of State end to the Public Health Serviee their interest in working out some arrangements Aion would remove at ae restrictions against the marxeving of imported shellfish without endangering the public health. As you may imagine, LHEesEeee, various officials of ovr Denartment of State have developed a2 great interest in this problem, and we are being urged to expand our pre-= sent system of « ac tonnin 610m tO these and ocher foreign countries. The Public Health Service does aot want its fesse system of snellfisa er rige contrel within the United States to act as an artificial.trade barrier against legitimate shellfish shipments which have been produced and packed under conditionms equal to those required of our Own packers, On the other hand, even if the Public Health Service had the authority to do so=-== which it does not--- there would he very great difficulties involved in extending this certification system to other countries. Full knowledge of these difficulties is necessary if those most interested are to face the problem intelligently. The application of our system of Public Health Service endorse- ment of over-all State programs presumes that representatives of the Service keep in fairly close touch with control efforts of the indi- vidual producing States by reasonably frequent consultations with State personnel, cooperative investigations, and check inspections. We believe that we cannot report adequately to the country as a whole our epiniens on the effectiveness of the local procedures unless we maintain this type of contact. This reasoning has been applied even in connection with our agreement with Canada, which specifically in- cludes provision for the exchange of information on methods of pro- duction and handling of shellfish and for inspection visits across the border. From a practical point of view, it has been very easy for the Public Health Service to meet the provisions of our agreement with Canada covering the interchange of information on shellfish sanitation. The capitals of the two countries are only a few hours apart by air, end long distance telephone conversations are relatively inexpensive. Alse, it has cost only a small sum each year far us to keep in clese teuch with operations in the Canadian Maritime Provinces, and in British Columbia on Canada's west coast, by extending routine field trips to these areas while our men are working in Maine and the State of Washing- ton. Health officials in the two countries have many other mutual in- terests, and official contacts are frequently made on matters other than shellfish sanitation. There is, therefore, 2 constant inter- change of information made possibie at relatively low cost. In addition, all of our cooperative efforts with Canada have been built on a long history of parallel development in the two countries, both as to tech- nical procedures and cpeapetsis strative operations. It is easy to see that none of these favorable elements could be duplicated if the concept of - certified dealers and eure orsed" control programs were to be extended to other foreign countries. “29. There are no provisions in the Food, Drug, and Cosmetic Act, under whith the Food and Drug Administration operates, which would make possible any routine international exchange of information about techniques of sanitation control at source, much less provision for setting up any plan of international certification or endorsement of any foreign operating control program. in the view of officials of the Food and Drug Administration, the only justification under the Food, Drug, and Cosmetic Act for that Administration to use its appropriated funds to send a representative to a foreign country would be to gain information considered netessary for the proper enforcement of the Act in comnection with foods or drugs offered for entry into the United States. T understand that such visits have been rare for various reasons. In the first place, a single trip to a foreign country for inspection purposes can only develop information of limited usefulness. In order to carry out the type of inspections performed in this country under authority of the Federal Food, Drug, and Cosmetic Act, it is some- times necessary to visit one or more plants several times during the year. The obvious limitations of funds and personnel make such trips to foreign countries generally unproductive, as compared to the expendi- ture of comparable time and money in inspections in this country. An occasion may arise where a single trip or visit to a foreign country may supply basic information necessary to evaluate fully a particular situation. During January and February of this year, a team was sent, at the invitation and expense of the Government of Japan, to inspect the shellfish industry of that country. The writer was assigned as a sanitary engineer consultant to the Food and Drug Administration for this trip, and, in the company of Mr. L. R. Shelton, a bacteriologist of the Food and Drug Administration, gathered a large amount of in- formation which will be helpful in future considerations of problems involving Japanese shellfish. Aside from the complicated administrative problems outlined previously there are certain other factors which are important in any consideration of this over-all problem: 1. We have already considered the limitations of the objective examination of shellfish at the time of arrival of shipments in this country. If strongly positive bacteriological results are obtained, one may assume that the shellfish were produced or handled under insanitary conditions. However, when bacteriological results are negative, interpretation becomes much more difficult. 2. In spite of a great deal of research, we believe that there has not been established for our own species of shellfish any firm relationship between bacterial content of shucked shellfish in the -250= market and the quality of growing areas and conditions of handling. This is why we have not found it possible to adopt a final bacteriological standard for market quality. As you know, work gen hes been done so far in this field has dealt chiefly with fresh shucked oysters and clams, and has net considered frozen products. Undoubtedly, the freez- ing and prolongedstorage of shelifisk oe ees abroad will have some effect on its apparent bacterial content, to complicate the picture further. a Ca fish, notably clams and missels; are sometimes subject to the accumula- tion of erganic toxins during part of the year. The origin and action of these toxins are fairly well understo od, and a complex administra=~ tive control program is in operacion to oe toxic shellfisn from being used commercially. Adequate test procedures are available and are being improved. However, there is some reason to believe that toxin which sometimes affects certain species of fereign shellfish a not be so well understood, and it is not certain that adequate tests haye been developed. 3. in the United States and Canada, certain species of shell- 4. Most of the frozen shellfish which would be shipped to the United States would be cooked before use. In fact, one importer has been investigating the feasibility of introducing clams which would be given some cooking before being frozen for shipment, this product being intended for use as chowder stock. It is unlikely that many frozen shellfish from abroad would be consumed raw. This factor is mentioned, not because we feel that there should be anysignificantly different standards applied to shellfish intended to be heat precessed before sale, but simply because the facts of the matter seem to indicate that any health hazard which might be present in connection with bacter=- ial contamination of frezen shellfish from abroad would be considerably reduced by the cooking process. We do not believe that this expected best treatment should be considered in any way as a cover=up for a filthy item, and, as you know, this attitude is basic to the thinking of the Food and Drug Administration, also. We join with those who are concerned that there are in operation two parallel mechanisms of sanitery control through which imported shellfish on the one hand may be admitted to the country, and on the other hand, their sales may be discouraged. Foreign governments quite naturally find this situation extremely difficult to understand. The reason, of course, is that there are two different sources of legal authority, ore Federal and one State. We hope to develop a workable approach to the problem in time for it to be given full consideration at the forthcoming National Con- ference on Shelifish Sanitation, scheduled to be held in Washington on September 9th and 10th. This Conterence, which is being called by the igeon General of the Public Health Service at the request of the Conference of State and Territorial Health Officers, will review all aspects of shellfish sanitation control in the United States. Since this will be the first over-all reevaluation of our program by the States and the industry in almost 30 years, we look upon it as a significant event. The committee which is developing a detailed agenda and proposed procedures for the Conference includes Mr. Wallace, of your Association. You may be sure that the question of foreign shellfish will be given a prominent place in these forthcoming de- liberations. We urge your interest and participation in this Con- ference, because we aré sure that the decisions made at that time will have a major bearing on future activities of the Public Health Service in the field of shellfish sanitation. ap ape i" rate peer yr to a Bae he ay heal AW ae maton ae abe 744 ; abate sy: 1). ACS (i el eal = nih Pathe oras wi acthan idan kets At letaal cat ' Cary ie Vetta e* they Deki te ales . Bile io, cet ala. bee Lact ped! fe Sete | » ; SI - | a: =) i Graueoeialy, VS an ies L r- Ry bal : orate 43.0, iid bit Me ida cla | la ‘pe ras SS oa i} (iene treticr ko oe & ‘oo @&: > i, afea : ; : roy ¥ v hs ol = a » ie \ 2 a @ —s ; THE DEVELOPMENT OF RECOMMZNDED PRACTICES FOR SANITARY CONTROL OF THE BREADING AND FREEZING OF SHELLFISH (1) Eugene T. Jensen U. S. Department of Health, Edueation, and Welfare, Washington, D.C. Freezing is not a new method of preserving foods. Actually, this process has been used in cold climates since prehistoric times for preservation of perishable foods, particularly fish and meats. Appli- cation of freezing to commercial preservation of foods is, however, a relatively recent development. To a New Englander, Enoch Piper, of Camden, Maine, goes the eredit for the first commercial process for artificial freezing of fish. Almost a hundred years ago, this "down Easter" developed a method in which a salt-ice mixture was used as a refrigerant (2) But a completely feasible, commercial freezing process was not develop- ed until the 1920's, when reliable, mechanical refrigeration equipment became generally available. Since the end of World War II, the frozen food industry, paced by a high degree of public acceptance of its pro- duct, has grown rapidly. Today, frozen foods are sold by almost all grocery stores; many homes have small freezers; and almost all house- hold refrigerators have frozen food storage space. There is no doubt that frozen foods have been well accepted by the consuming public. With some foods, this degree of acceptance has been so complete that sales of canned and fresh products have been adversely affected. Most of you are aware of the tremendous expansion of the frozen, concentrated citrus juice industry (3) Whether freezing will have an equivalent effect on the shellfish in- dustry is problematical; only time will supply the answer. In theory at least, freezing is an almost perfect means of marketing a highly perishable product, such as oysters. Most of you, I believe, have had some experience with the process. These exper- iences, probably, have not been uniformly successful. Frozen oysters, at times, tend to brown, or may pick up a rancid flavor. Consumers have not yet been educated to the fact that oysters are a good summer food; many people still believe the old "R-month" myth. Various types of containers have been tried, freezing processes have been altered, and new packing methods have been adopted. It seems almost inevitable that, eventually, a method will be developed resulting in a frozen pack which will be as completely successful, from a quality standpoint, as is the fresh, shucked pack. Within the last three or four years, there has been a trend in the industry toward development of a ready-to-cook product, and even an already-cooked product. This trend is simply a phase of the more general =O5 ai drift toward production of prepared or partially prepared foods which has affected the entire food processing industry. Most of you are familiar with the joint State=-Federal-Industry certification system which has been used for many years, and you know that the certification system is applicable to the production and marketing of frozen shellfish (4). Frozen shellfish which have been processed and identified under this sytem, move without difficulty in interstate commerce, as long as the dealer maintains his certificate number . With the advent of the breaded, frozen procuct--- as contrasted with the simply frozen product-=- a new problem arose. Since the breaded product invariably would be cooked before it would be eaten, there was a problem of determining whether it should be included under the certification program or considered exempt from the certification system as a processed food. In 1951, we discussed this question with officials of the Food and Drug Administration, with Mr. Wallace, of the Oyster Institute, with some of the interested members of the industry, and with State officials responsible for shellfish sanitation programs. After care- ful consideration, it was decided that the certification program should be considered applicable to the breaded and prefried products, even though it was certain that all of such products would be cooked before being eaten. I believe it is important that you know the basis for the decision. There were two principal considerations: 1. The basic component of this prepared food is shellfish-- usually oysters, although we understand that some breaded clams have been marketed. Breaded foods are not sterilized during the frying process because of the insulating effect of the breading material, Thus, if the shelifish should come from a contaminated source, some disease-causing bacteria might survive the cooking process. In addition, the sometimes toxic properties of clams are a consideration that we believe must be given considerable weight. 2. The existence oF a market for large quantities of non- certified shellfish would greatly complicate the job of the State enforcement agencies. This is particularly true if you consider that breading plants might be--- and, in fact, are=--- located in the interior States, away from the organized shellfish control activities of the coastai States. In addition, there was a strong feeling that it would be con- fusing to many persons, both in the industry and in the entTorcement -25)- agencies, to apply the certification system to one part of a plant and not to another part of the plant. Similarly, there was a strong belief that receiving States and cities, eventually, would question the sani-~ tary quality of frozen, breaded oysters which did not show evidence of having been processed in a State certified estadlishment. Recent ex- perience has demonstrated that such confusion and marketing difficulties will develop unless the product carries a certification number. The administrative decision to include breaded and/or prefried frozen shellfish in the certification program raised the problem of appropriate sanitary standards. in general, breading is a clean operation; however, the use of the breading material results in some sanitation problems very different from those associated with the shucking-packing operation. In certain respects, the breading plant has the sanitation problems of the ordinary shellfish packing plant and, in other respects, the sanitation problems of a baker, as well. We had no ready knowledge on which to formulate a set of standards, so a process of fitting standards to the industry as it existed was undertaken. Experienced shellfish sanitarians from the Public Health Service, working in cooperation with the State shell- fish control agencies, visited plants which were breading shellfish, and made a careful study of the various steps in the operation. This study of the breading process made it possible to pinpoint those steps which would require the most careful control. Almost a year was spent in making an evaluation of these operations. Our very real appreciation goes to the plant operators who assisted us in making these baseline studies. Using the information which had been obtained by observation and study, a tentative set of "Recommended Practices" was developed. This tentative manual, 32 typewritten pages, was sent to the Public Health Service field offices for review by their food technologists. Many comments and suggested changes were made. The next developmental step consisted of a review of the manual by other official agencies, and by members of the shellfish industry who were interested in the breading process. To accomplish this, over 200 copies of the "Recommended Practices" were sent to all State health departments, the Food and Drug Administration, the Fish and Wildlife Service, the Oyster Institute, the Pacific Coast Oyster Growers Associa- tion, and the National Fisheries Institute. The response to this request for assistance was most gratifying. In fact, several hundred pages of comments were received! Working these comments into a iO page manual was a chore that required several months and some lengthy conferences with interested parties. As a result of this review process, we believe that the recently released "Manual of Recommended Practice for Sanitary Control of the Breading and Freezing of Shellfish" should be a relatively good guide “255+ for maintaining sanitary conditions in shellfish breading establishments. However, we have decided that we will take no action toward final adoption of these “Recommended Practices" until we have had at least one year's operating experience with them in various parts of the country. The Manual, together with its accompanying rating form, has already been used fer the evaluation of oyster breading plants in Virginia. It seems very likely that some changes will be made in the "Recommended Practices" as a result of field use; however, we believe that the changes will not be of a fundamental nature, but will relate instead to the smaller operational details. Those of you who operate and manage shellfish breading plants could be of great assistance to us by becoming thoroughly familiar with the manual as it would apply to your plant. Any errors or dis- ecrepancies should be called to our attention. If you do not have a copy of these “Recommended Practices", and if you are interested in this phase of shellfish packing, you can get a copy from Mr. David Wallace of the Oyster Institute, or you can write directly to the Shellfish Branch of the Milk, Food and Shellfish Sanitation Program, Public Health Service, Washington 25, D. C. The following summary will give you some idea of the principal sanitation requirements of the new manual: Plant Construction and Arrangement: Plants should be con- structed so as to be easily cleaned, and so as to furnish minimum harborage for insects or rodents. In general, this will mean about the same type of construction as has been used in packing rooms. The "Recommended Practices" will require a separate breading room, but will permit its use for other purposes, provided that such operations do not interfere with the breading operation. Adequate screening and lighting are, of course, required. No special heating or ventilating equipment is required, except for special ventilation systems where breading machines are operated. Water Supply: A safe water supply is a fundamental sanitation requirement of any food processing plant. In addition, it will be required that water be piped to all food processing rooms, and that an adequate hot water heating system be provided. Plumbing: The safety of a water supply is intimately related to the quality of the plumbing, and to the skill with which the plumbing is installed. Plumbing which complies with your State plumbing code or with the National Plumbing Code is, therefore, a requirement of the "Recommended Practices." Toilet facilities are an important part of the plumbing in a breading plant. The requirements---somewhat above those of the old shellfish manual-==sre those recommended by the National Plumbing Code. Howevery we believe that modern sanitation, particularly fly control, has removed the need for the intervening vestibule, and this requirement has been deleted. Disease outbreaks have been caused by leakage of sewage from defective overhead sewers. The "Recommended Practices," there- fore require that there be no overhead sewers in food processing areas. Handwashing is perhaps the most important single sanitation item in a food processing plant. ‘To make it easy for workers to wash their hands, an adequate number of handwashing sinks, complete with hot and cold water, soap, and sanitary towels, is a must. Re- search workers have, in the past few years, developed several types of soaps which contain an added bactericide. These special types of soaps are much more effective for killing bacteria than are ordinary soaps, and their use has, thus, been required. Presumabiy, the use of bactericide containing soaps will eventually be required in shucking-= packing plants. Rodent Control: Control of rodents in oyster shucking houses has neyer posed any real problem. It seems likely, however, that rodent control problems will exist in breading plants. It has been required, therefore, that buildings be of ratproof construction, and that rodenticides be properly handled and stored. Equipment Construction: As in previous Manuals there are no specific requirements for equipment construction. Equipment should be constructed of material which will not readily corrode, which is easily cleanable, and which is nontoxic. To make cleaning easy, all joints should be smoothly welded or soldered; corners should be filleted; and there should be no rough or ingecessible areas. The milk industry, in cooperation with the sanitation authorities, has developed specific construction standards fox much of the equipment which they use. Thus, the operstor of a pasteurizing plant can buy equipment which meets the “3-A" Standards with assurance that he is getting equipment which will comply with all sanitation requirements. Perhaps the shellfish industry might profit by this experience by developing its own standards for equipment used in snelifish processing. Certain construction items for breading machines have been specified because of the nature of the breading operation. For example, it has been required that breading machines be easily cleaned, so that they will not harbor insects or rodents. Proper design and construction of the machine can save the plant operator many hours of cleanup work. Refrigeration of the perishable batter has been required, aithough we know that not all machines new on the market provide such refrigeration. aL (s Plant Personnel and Supervision: Most of you are familiar with the requirements for Supervisors in shucking=packing plants. We have found that some person actually in the plant---not the manager in the front offices--omust be responsible for secing that employees wash their hands and wear cleam clothes. If supervision is lax, plant workers will tend to pay less and less attention to these important sanitation measures. Source and Condition of Shelifish: The "Recommended Practices" require that all ghelifish come from a certified or otherwise approved source. Shucked sheilfish must be shipped in approved sealed containers, and must have a tenperature of ho°F. or less om arrival at the breading plant. Breading of frozen oysters has been prohibited. Breading Material: The material used in breading-=-the batter mix and the breading material---should be purchased in containers which will afford a maximm degree of protection to the product. After arrival at the plant, the batter mix and breading materiais should be stored so as to be protected from contamination. Breading Operations: In general, the mechanics of preparing the breaded shellfish have been left to the discretion of the plant operator, although certain limiting guidelines have been established. For example, the perishable batter should be refrigerated; grinding and reuse of lumpy breading materials is prohibited; and reuse of breading materials left over at the end of the day‘’s operation is prohibited. Storage of Containers: There is no convenient way to sterilize paper containers at the breading plant before they are filled. Hence, it is necessary that packaging materials be clean when purchased, and that they be kept clean by storing them where they will not be subject to contamination, Usually, a special storage area will be needed. Cleaning and Sterilizing of Equipment: The requirements for cleaning and sterilizing of equipment are the same as those now in use in the shucking=-packing plants, except that some special problems in sanitizing are posed by the breading process. Adequate cleanup facilities, including a wash sink, detergents, and brushes, are required. Prefried Shellfish: A few operators have been interested in the preparation of breaded prefried shellfish. The sanitation problems involved in the preparation of this product are essentially the same as those in the ordinary breading establishment, except that the presence of grease from the frying process may complicate cleanup work. Adequate ventilation equipment is the key to the problem, and must be provided where deep fat frying is practiced. - Packaging and Labeling: The Food, Drug, and Cosmetic Act re- quires that certain information be shown on each package of food. This information includes: a. The name and address of the packer or distributor. b. The common or usual name of the food. ec. The common or usual name of esch ingredient used in its production, except that spices or flavorings may be declared as such. ad. An accurate declaration of net weight. To insure that the product will be acceptable in all markets, the packer should also indicate his certificate number preceded by the abbreviated name of the State. We have had several complaints about noncertified products which were eventually traced to the type size of the certificate mumber. In some instances, these numbers have been so small that they were virtually impossible to find. Hence, the "Recommended Practices" specifies that these numbers be prominently displayed, and that they be at least 3/16 of an inch high. Also, packages should be conspicuously labeled "PERISHABLE=-- KEEP FROZEN.” The code or date of packing should be placed on each carton, but need not be visible to the ultimate consumer. I should point out that code dating of master cartons does not satisfy this requirement. Freezing of Breaded Shellfish: To avoid excessive growth of bacteria, the preaded shellfish should be frozen as soon as feasible, preferably within a few minutes after packaging. Recording thermom- eters are required for the freeze room. This resume of the shellfish Breading Manual touches on only the most salient points. I must remind you that the requirements contained in the "Recommended Practices" are not yet final, and will doubtless be subject to some readjustment during the coming year. Those of you who are active in this field can be of real help by calling any errors or discrepancies to our attention, so that they may be corrected. If you disagree with any of the provisions of the manual, I suggest that you contact us so that we may discuss these problems with you. With your assistance, we should be able to develop @ manual which will insure a sanitary product of good quality without imposing excessive demands on the industry. -259= (1) (2) (3) (4) (5) Literature Cited Manual of recommended practice for sanitary control of the breading and freezing of shellfish. Div. Sanitary Engineering Serv., Public Health Serv., U. S. Dept. Health, Educ. & Welfare. Tressler, D. K.y and C. F. Evers. 1947. The freezing preserva- tion of foods. Avi Publ. Co., Inc., N. Y. Frozen foods, a $700 million business last year. Chem. & Engineer- ing News 29(11), March 12, 1951. Manual of recommended practice for sanitary control of the shell- fish industry. Public Health Serv. Publ. No. 33, Public Health Serv., U.- S. Dept. Health, Educ., & Welfare. Report of the Coordinating Committee for the National Plumbing Code, U. S. Dept. Commerce, 1951. -260- Papers Presented at the Convention but Published Elsewhere: Moulton, J. M., and G. W. Coffin. 1954. The distribution of Venus larvae in Orr's Cove plankton over the tide cycle and during the summer and early fall of 1953. Res. Bull. 17, Dept. Sea & Shore Fish., Me. =261< ne a vf a - vena oi =, on _ : ul ay a ’ boas en Ays ie pe gt ‘near ea itis am +s : a am Goins: Pay) es eee ; oy meet alin, | Riera See Caeeaicald Pa , v. foam ber DIRECTORY OF MEMBERS OF THE NATIONAL SHELLFISHERIES ASSOCIATION (To April, 1955) Aldrich, Dr. Frederick A., Assistant Curator of Limnology, Academy of Natural Sciences, 19th. and the Parkway, Philadelphia 2, Pa. Allen, Dr. J. Francis, Department of Zoology, University of Maryland, College Park, Md. Andrews, Dr. Jay D., Oyster Biologist, Virginia Fisheries Laboratory, Gloucester Point, Va. Atlantic Biological Station, Fisheries Research Board of Canada, St. Andrews, N. B., Canada. Baker, Byron B., Jr., 7222 Marywood Street, Landover Hills, Md. Ball, Eric T., 212 Summit Street, New Haven 13, Conn. Baptist, John P., U. S. Fish and Wildlife Service Shellfish Laboratory, Beaufort, N. C. Beaven, G. Francis, Maryland Department of Research and Education, Solomons, Md. Berry, W. R., Department of Health, 301 Essex Building, Bank and Plume Streets, Norfolk 19, Va. Blount, F. Nelson, Blount Seafood Corporation, 383-393 Water Street, Warren, R. I. Butler, Dr. Philip A., Chief, Gulf Oyster Investigations, U. S. Fish and Wildlife Service Shellfish Laboratory, P. 0. Box 1826, Pensacola, Fla. Carriker, Dr. Melbourne R., Department of Zoology, University of North Carolina, Chapel Hill, N. C. Chanley, Paul E., U. S. Fish and Wildlife Service Biological Laboratory, Milford, Conn. Chestnut, Dr. A. F., Institute of Fisheries Research of the University of North Carolina, Morehead City, N.- C. Chipman, Dr. Walter, Director, U. S. Fish and Wildlife Service Shellfish Laboratory, Beaufort, N. C. =262= Collier, Dr. Albert, Chief, Gulf Fishery Investigations, U. S. Fish and Wildlife Service, Fort Crétkett, Galveston, Tex. Cronin, Dr. Eugene, Director, Maryland Department of Research and Education, Solomons, Md. Currier, Wendell, Assistant to the Vice-President, Research and Develop- ment, Campbell Soup Co., Camden, N. Jd. Darling, J. S. & Son, P. 0. Box 412, Hampton, Va. Davis, Harry C., U. S. Fish and Wiidlife Service Biological Laboratory, Milford, Conn. Dawson, C. k., institute of Marine Research, Port Aransas, Tex. Deiler, Frederick G., Biologist, Freeport Sulphur Co., Port Sulphur, La. Dow, Robert L., Director of Marine Research, Department of Sea and Shore Fisheries, Vickery-Hill Building, Augusta, Maine. Dumont, William H., U. S. Fish and Wildlife Service, Washington 25, D.C. Dunnington, Elgin W., Department of Research and Education, Solomons, Md. Ellison, William A., Director, Institute of Fisheries Research of the University of North Carolina, Morehead City, N. C. Engle, James B., Chief, Chesapeake Shellfish investigations, U. 5. Fish and Wildlife Service, P. 0. Box 151, Annapolis, Md. Fahy, Dr. William, Institute of Fisheries Research of the University of North Carolina, Morehead City, N. C. Flower, Frank M. & Sons, Growers of Pine Island Oysters, Bayville, Long Island, N. Y. Fox, Leo, Department of Public Health, 511 A State House, Boston 33, Mass. Galtsoff, Dr. Paul S., Director, U. S. Fish and Wildlife Service Shelifish Laboratory, Wocds Hole, Mass. Ganaros, Anthony E., U. S. Fish and Wildlife Service Biological Laboratory, Milford, Conn. Gibbs, Harold N., A-71 Sowams Road, Barrington, R. I. Glancy, Joseph B., Shellfish, Inc., Box 212, West Sayville, Long Island, Ns Ye =263= Glude, John B., Chief, Clam Investigations, U. S. Fish and Wildlife Service, Boothbay Harbor, Me. Green, Richard S., Chief, Shellfish Sanitation Branch, Public Health Service, Room 4113, DHEW Building, South 3rd. and C Streets, S. W., Washington 25, D. C. Greenwich Oyster Company, Greenwich, N. J. Grice, Dr. George D., Oceanographic Institute, Florida State Univ- ersity, Tallahassee, Fla. Gunter, Dr. Gordon, Acting Director, Institute of Marine Science, Port Aransas, Tex. Gustafson, Dr. Al, Chairman, Department of Biology, Bowdoin College, Brunswick, Me. Hammer, Ralph, Maryland Department of Tidewater Fisheries, State Office Building, Amapolis, Md. Hanks, James E., U. S. Fish and Wildlife Service Biological Lab- oratory, Milford, Conn. Harrison, George T., President, The Tilghman Packing Co., Tilghman, Md. Haskin, Dr. Harold H., Director, Oyster Research Laboratory, Bivalve, N. J.y and Department of Zoology, Rutgers University, New Brunswick, N. J. Haven, Dexter, Virginia Fisheries Laboratory, Gloucester Point, Va. Hayes, E. C., Jr.y Assistant Director, Department of Agriculture and Conservation, Veterans Memorial Building, 83 Park Street, Providence 2, R. I. Hedrick Brothers Oyster Company, 730 Auster City Street, New Orleans, La. Hewatty, Dr. Willis G., Biology-Geology Department, Texas Christian University, Fort Worth, Tex. Heydecker, Wayne D., Atlantic States Marine Fisheries Commission, 22 West First Street, Mount Vernon, N. Y. Hofstetter, Robert P., Bay Oaks Addition, La Porte, Tex. Hopkins, Dr. Sewell H., Biology Research Laboratory, Texas A. & M. Research Foundation, College Station, Tex. ~26 = Huber, L. Albertson, Hydrographic Engineer, 297 H. Commerce Street, Bridgeton, N. J. Jensen, Eugene T., Shellfish Branch, U. S. Public Health Service, Washington 25, D. C. Kahan, Archie M., Executive Director, Texas A. & M. Research Foundation, College Station, Tex. Lamson, P. G., Publisher of “Atlantic Fisherman", Goffstown, N. H. Lednum, J. M., Town Engineer, Town of Islip, N. Y. Lester & Toner, Inc., % Royal Toner, Fulton Market, New York 38, N. Y. Lindsay, Cedric, Shellfish Laboratory, Fisheries Department, Washington State, Quilcene, Wash. Littleford, Dr. Robert A., Department of Zoology, University of Mary- land, College Park, Md. Logie, R. R., Department of Zoology, Rutgers University, New Brunswick, N. Jq Loosanoff, Dr. Victor L., Director, U. S. Fish and Wildlife Service Biological Laboratory, Milford, Conn. Lunz, G. Robert, Director, Bears Bluff Laboratories, Wadmalaw Island, S.C. Mackin, Dr. J. G., Director, Marine Laboratory, University of Texas Medical Branch, Galveston, Tex. ; Macomber, Ronald, U. S. Public Health Service, 11 Prescott Avenue, Montclair, N. J. Manning, Joseph H., Chesapeake Biological Laboratory, Solomons, Md. Mansueti, Romeo, Maryland Department of Research and Education, Solomons, Md. Marshall, Dr. Nelson, Saunders Point, Niantic, Comn. McConnell, James L., Department of Wildlife and Fisheries, New Orleans, La. McConnell, James N., Director, Division of Oysters and Water Bottoms, Department of Wildlife and Fisheries, New Orleans, La. McHugh, Dr. L. H., Director, Virginia Fisheries Laboratory, Gloucester Point, Va. ~265- Menzel, Dr. R. Winston, Oceanographic Institute, Ficrida State Uni- versity, Tallahassee, Fla. Messer, Richard, Director, Division of Engineering, Department of Health, 713 State Office Building, Richmond 19, Va. Miles, J- H. & Co., Inc., Norfolk 1, Va. Nelson, J. Richards, President, The F. Mansfield & Sons Co., 610 Quinnipiac Avenue, New Haven, Conn. Nelson, Dr. Thurlow C., Department of Zoology, Rutgers University, New Brunswick, N. J. New Jersey Department of Conservation, Trenton, N. J. Perlmutter, Dr. Alfred, Bureau of Marine Fisheries, Conservation Department, State of New York, 65 West Sunrise Highway, Freeport, N. Y. Pomeroy, Dr. Lawrence, Marine Biology Laboratory of the University of Georgia, Sapelo Island, Ga. Pritchard, Dr. Donald W., Director, Chespeake Bay Institute of the Johns Hopkins University, Box 426A, R.F.D. #2, Annapolis, Md. Ray, Dr. Sammy M., Biology Department, Rice Institute, Houston, Tex. Rego, John L., Director, Department of Agriculture and Conservation, Veterans Memorial Building, 83 Park Street, Providence 2, Rs Le Rice, Dr. Theodore R., U. S. Fish and Wildlife Service Shellfish Laboratory, Beaufort, N. C. Ropes, John W., U. S. Fish and Wildlife Service, 29 Linden Street, Salem, Mass. Russell, Henry D., Springdale Avenue, Dover, Mass. Sangree, Dr. John B., Glassboro State Teachers College, Glassboro, N. J. Sieling, Fred W., Department of Research and Education, Snow Hill, Md. Smith, Dr. F. G. Walton, Director, The University of Miami Marine Laboratory, Coral Gables 46, Fla. Smith, Osgood R., U. S. Fish and Wildlife Service, 13 State Street, Newburyport, Mass. ~266~ Sollers, Allon A., 1305 Park Avenue, Baltimore 17, Md. Sprague, Victor, Hiawassee, Ga. Truitt, Dr. Reginald V., Maryland Department of Research and Education, Solomons, Mc. Udell, Harold, Bureau of Marine Fisheries, New York Conservation Department, Freeport, Long Island, N. Y. Virginia Commission of Fisheries, Newport News, Va. Wallace, Dana H., Shelifish Specialist, Department of Sea and Shore Fisheries, Vickery-Hill Building, Augusta, Me. Wallace, David H., Director, Oyster Institute of North America, and Executive Secretary of the Oyster Growers and Dealers Associa- tion, 6 Mayo Avenue, Bay Ridge, Annapolis, Md. Webster, John R., U. S. Fish and Wildlife Service, P. O. Box 151, Annapolis, Md. Weiss, Dr. Charles M., Sanitary Chemistry Branch, Medical Laboratories, Army Chemical Center, Md. Welch, Walter R., Clam Investigations, U. S. Fish and Wildlife Service, Boothbay Harbor, Me. Westley, Ronald E., Shellfish Laboratory, Fisheries Department, Washington State, Quilcene, Wash. Whaley, Horace A., Chesapeake Bay Institute of the Johns Hopkins University, Van Buren Street, Annapolis, Md. Wolman, Abel, Johns Hopkins University, Whitehead Hall, Baltimore 18, Md. Wright, Thomas J., Chief, Division of Fish and Game, Veterans Memorial Building, 83 Park Street, Providence 2, R. I. Wurtz, Charlies B., 3247 Disston Street, Philadelphia OQ, Pa. ne MBL/WHOL LIBRARY AION TI WH LACL ? racqvasans neste yeNe Ns . ON coun’