STUDIES IN FOREST PATHOLOGY II. THE BIOLOGY OF FOMES PINICOLA (SW.) COOKE By IRENE MOUNCE ASSISTANT PLANT PATHOLOGIST AGRICULTURE CANADA CODE 86/02/21 NO. LIBRARY /BIBLIOTHEQUE OTTAWA KlA PCS DIVISION OF BOTANY H. T. GUSSOW, Dominion Botanist DOMINION EXPERIMENTAL FARMS E. S. ARCHIBALD, Director DOMINION OF CANADA DEPARTMENT OF AGRICULTURE BULLETIN No. Ill NEW SERIES Published by direction of the Hon. W. R. Motherwell, Minister of Agriculture, Ottawa, 1929 DOMINION EXPERIMENTAL FARMS E. S. ARCHIBALD, Director DIVISION OF BOTANY H. T. GUSSOW, Dominion Botanist ECONOMIC BOTANY Botanists J. Adams H. Groh Junior Botanist and Librarian R. A. Inglis PLANT PATHOLOGY Central Laboratory, Ottawa Plant Pathologists. F. L. Drayton J. B. MacCurry A. W. McCallum Assistant Plant Pathologist Irene Mounce Senior Plant Disease Inspector J. Tucker Charlottetown, P.E.I. Assistant Plant Pathologist R. R. Hurst Senior Plant Disease Inspector S. G. Peppin Kentville, N.S. Plant Pathologist • • J. F. Hockey Assistant Plant Pathologist K. A. Harrison Fredericton, NJB. Plant Pathologist J. D. MacLeod Assistant Plant Pathologist J. K. Richardson Ste. Anne de la Pocatiere, Que. Plant Pathologist H. N. Racicot St. Catharines, Ont. Senior Plant Pathologist G. H. Berkeley , Plant Pathologist G. C Chamberlain Assistant Plant Pathologist J. C. Perrault Winnipeg, Man. (Dominion Rust Research Laboratory) Senior Plant Pathologist in charge J. H. Craigie Senior Plant Pathologists Margaret Newton W. F. Hanna Plant Pathologists • • I. L. Conners F. J. Greaney W. L. Gordon Assistant Plant Pathologists A. M. Brown T. Johnson W. Popp B. Peturson Saskatoon. Sask. i&enior Plant Pathologist P. M. Simmonds Plant Pathologists • • G. A. Scott Assistant Plant Pathologists R. C. Russell B. J. Sallons Edmonton, Alta. Plant Pathologists G. B. Sanford W. C. Broadfoot Summerland, B.C. Plant Pathologist H. R. McLarty 4 Assistant Plant Pathologists G. E. Woolliams J. C. Roger Vancouver, B.C. Plant Pathologist Wm. Newton TABLE OF CONTENTS* Pace I. Introduction 3 II. Historical 4 III. Distribution 5 A. Geographical 5 B. Host list 6 IV. Occurrence 8 V. The sporophore 10 VI. Previous cultural studies 11 VII. Cultural methods and conditions of growth 13 A. Original cultures 13 B. Conditions of growth 13 C. Media employed 13 D. Acidity 14 VIII. Spore germination 14 IX. Source of cultures 16 X. Production of sporophores in culture 17 A. On plant decoctions 17 B. On Czapek's synthetic medium 21 C. On wood blocks 23 XI. Effect of variations in temperature and acidity upon mycelial development. ... 26 A. Temperature 26 B. Acidity 28 XII. Variations in cultural characters and in the formation of a line of demarcation in mixed cultures 28 A. Variations in cultural characters 28 B. Mixed cultures and the formation of a line of demarcation 31 XIII. Heterothallism 39 A. Isolation of monosporous mycelia 40 B. Fomes pinicola is a heterothallic bisexual species 41 C. Cultural characters and fruiting of haploid and diploid mycelia 42 D. Sexual strains or geographical races 44 XIV. Destruction of wood 49 XV. Summary 50 XVI. Bibliography 51 XVII. Description of plates 55 'Progress reports have been published in the Report of the Dominion Botanist, Department of Agriculture, Ottawa for 1924, Ym, 1026, and 1927. Digitized by the Internet Archive in 2013 http://archive.org/details/biologyoffomespi111moun STUDIES IN FOREST PATHOLOGY II. THE BIOLOGY OF FOMES PIMCOLA (SW.) COOKE BY IRENE MOUNCE, Assistant Plant Pathologist I. INTRODUCTION It was not until Robert Hartig (25) published the results of his studies of diseased wood and of his experiments with artificial infection that the disease and decay caused by wood-destroying fungi were actually attributed to them. Hartig's discoveries aroused a deep interest in such questions as the distinguishing characters of various rots, the hosts which are most sus- ceptible, the prevalence of certain diseases in certain areas and the conditions which favour their development, prophylactic measures, and, based upon all of these, the formulation of plans for the most economic harvesting of timber. As these studies of wood-destroying fungi progressed, the need of corre- lating the data collected in the field with results of detailed laboratory experi- ments became increasingly evident. Some rots macroscopically indistinguish- able are due to different organisms; some rots vary in appearance on different sub-strata and under varying conditions of growth; and, in the absence of sporophores, the identity of the causal organisms in such cases can be deter- mined accurately only by cultures. Again, no specific method of adequately dealing with a disease can be devised until the mode of dissemination and infection, in fact, until the whole life history of the organism is known. Until recently this phase of forest pathology had received comparatively little attention. However, the work of Long and Harsch (34), of Fritz (21), and of Hubert (28) has supplied criteria for the identification of many wood- destroying fungi from their cultural characters; and the work of Buller (10), Bayliss (3), Rhoads (51), Hiley (27), White (76), Zeller (77), Kauffman (29), and others has supplied us with details of the life histories of such wood- destroying fungi as Polyporus squamosus, Lentinus lepideus, Polystictus versi- color, Polyporus pargamenus, Lenzites saepiaria, Dasyscypha calycina, Fomes annosus, Polyporus dryophilus, Fomes applanatus, and Trametes robiniophila. It is with this type of investigation, in its broader aspects, that this paper i's largely concerned. The subject chosen is Fomes pinicola. Fomes pinicola is one of the very common wood-destroying fungi in both Europe and America. Since it produces large, conspicuously coloured sporophores (plate I) and destroys both sapwood and heartwood of coniferous and deciduous trees, reports of its occurrence and distribution are frequent throughout the literature dealing with the field aspects of forest pathology. In addition, observa- tions on the cultural characters of this fungus have been made by Rumbold (55), Long and Harsch (34), Fritz (21), and Schmitz (57); but beyond this it has received practically no attention. The present paper summarizes the results of those laboratory studies, and then goes on to include such phases as spore germination; the mycelial characters of the fungus when grown on liquid and solid media; temperature as a factor influencing mycelial development; sporo- phore production on artificial media and on wood blocks; a comparative study of monosporous and polysporous mycelia; sexuality; and macroscopic and microscopic features of the decay caused by F. pinicola. * A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy in the University of Toronto. 75474-1 i II. HISTORICAL Because of the frequency of its occurrence on so many different hosts and in so many localities, F. pinicola not only appeared early in mycological litera- ture but also under a variety of names. Murrill (40) gives the following list of synonyms: — Boletus igniarius Scop. Fl. Cam. ed. 2.2:469. 1772. Not B. igniarius L. 1753. Boletus ungulatus Schaeff. Fung. Bavar. 4:88, pi. 137. 138. 1774. BoletmfuLvusSchaeS. Fung. Bavar. 5:89, pi. 262. 1774. Boletus semiovatus Schaeff. Fung. Bavar. 4:92. pi. 270. 1774. Boletus marginatus Pers. Obs. Myc. 2:6. 1799. Boletus pinicola Sw. Sv. Vet. Akad. Handl. 1819:88. 1810. Polyporus marginatus Fries. Syst. Myc. 1:372. 1821. Polyporus pinicola Fries. Syst. Myc. 1:372. 1821. Forties marginatus Gill. Champ. Fr. 1:683. 1878. Fomitopsis pinicola Karst. Rev. Myc. 39:18. 1881. Fomes pinicola Cooke. Grevillea 14:17. 1885. Fomes ponderosus Schrenk. Bull. U.S. Dep. Agr. PL Ind. 36:30. 1903. (Type from Dakota, on Pinus ponder osa.) Fries (20) considered the variations in form, colour, and zonation of the sporo- phores of this fungus on coniferous and deciduous hosts distinct enough to be specific and he has two species, P. pinicola and P. marginatus, based on such differences. Saccardo (56) recognises Fries' species and creates a third F. ungulatus. He gives the following synonyms for each: Fomes pinicola Fr. Elench. p. 105. Hym. Eur. p. 561. Gill. Champ, c. ic. Agaricus sitaneus, subrotundus, fulvus, margine obtuso, pallido, stillante. Boletus fulvus Schaeff. t. 262. B. marginatus Pers. Syn. p. 534. Swartz. Vet. Akad. Handl. 1810. p. 87. B. hornotinus et annosus, pileo unguiato, nigricante, margine cinna- barino. B. semiovatus Schaeff. t. 270. (varietas) . B. igniarius Fl. Dan. t. 953. Pers. Syn. p. 534. B. pinicola Swartz. I.e. p. 88. Wahlenb. Suec. n. 2009. B. marginatus Weinm. Fomes ungulatus (Schaeff.) Sacc. Michelia I. p. 539. Boletus ungulatus Schaeff. Fung. Bat. t. 137 (ab auctoribus non citata), nee Bull. Fomes marginatus Fr. Epicr. p. 468. Hym. Eur. p. 561. Quelet. t. 19, f. 2 (var.) Saccardo states that his F. ungulatus may be only a form of F. pinicola. Later workers, however, including Hedgecock (26), Lloyd (33), Murrill (40), and Overholts (47) consider the three species given by Saccardo as forms of one species which they call either F. pinicola or F. ungulatus, depending on whether or not they accept Fries' Systema mycologicum as the starting point of mycological nomenclature. A study of the sporophores of this fungus, and of the behaviour of monosporus mycelia in paired cultures (section XIII) has led the author to the same conclusion, and throughout this paper Fomes ungulatus (Schaeff.) Sacc. ami F, marginatus have been considered as forms of F. pini- cola (Sw.) Cooke. While collecting on the Queen Charlotte Islands, B.C., where western hemlocks are numerous and Fomes pinicola particularly abundant, groups of sporophores were examined to see whether the majority more closely resembled the applanate form with soft context and pale margin, characteristic of P. mar- ginatus Fr., or the ungulate form with vermilion red margin and hard context characteristic of P. pinicola Fr. Since all of the sporophores were collected from a coniferous host the great majority would be expected to resemble the P. pini- cola form which was supposedly the common one on coniferous hosts, rather than the P. marginatus form which was thought to grow chiefly on deciduous hosts. As a matter of fact both types of sporophores and all gradations between the two were common. Some sporophores had a soft context, some a hard, some had broad, white margins, some vermilion red margins, with many showing intermediate buff-coloured tints. The sporophores were applanate or ungulate in shape, the upper surface rough and zoned, or smooth, completely covered with a shining red lacquer, or entirely dull grey, brown, or black, with all the intermediate forms with vari-coloured margins and vari-coloured zones. Groups of sporophores showing both type forms and many intermediate ones could be collected from one tree. Cultures were made from the inner tissues of a large number of sporophores, but it has not been found possible to associate any specific variations in cultural characters with specific differences in sporophore form. Furthermore, mono- sporous mycelia isolated from cultures 5778, 928, and 586 are all mutually fer- tile when paired with monosporous mycelia isolated from cultures 283, 285A, 285B, and 5770 (See section XIII). In this F. pinicola behaves similarly to other species of Basidiomycetes which have been investigated by Kniep (32),Van- dendries (68-9), Brunswik (9), and Hanna (24). But culture 5778 was made from a sporophore which grew on Prunus serotina, culture 928 was from Betula sp. and collected as typical of the F. marginatus form of Europe, and culture 586 was labelled F. pinicola var. marginatus. The remaining cultures were iso- lated from sporophores which grew on Picea and would formerly, in all prob- ability, have been identified as F. pinicola. The fact that monosporous mycelia from 5778. 928, and 586 are all mutually fertile with those from cultures 283, 285A, 285B, and 5770 is taken as contributory evidence that all of the sporo- phores, from which these cultures were made, belong to one and the same species. This conclusion is based upon the clamp-connection criterion for the identity of species which Vandendries (68) states as follows, " Si les haplontes de deux carpophores sauvages sont toujours et indefiniment fertiles entre eux ces deux carpophores appartiennent a une meme espece." III. DISTRIBUTION A. GEOGRAPHICAL Fomes pinicola occurs frequently both in Europe and America. Its presence in exsiccati from France, Scandinavia, Saxony, Bavaria, Austria, Hungary, and Italy, indicates its distribution in Europe. It is reported from Uruguay, Cuba, and Asiatic Siberia, and Lloyd (33) has specimens, as well, from Japan, the Philippine Islands, and Mexico. Of its distribution in America Hedgcock (26) says, " it is one of the most widely distributed species in the United States occur- ring, as far as has been observed, in every region where conifers are found," and this is corroborated by the reports of Meinecke (35), Weir (72), Rankin 6 (50,), and others. The same might be said of its distribution in Canada, for the fungus is common in the forests of British Columbia, and is found from Mani- toba to the Atlantic. Weir (74) has collected it in the mountains at altitudes of 6,735, 7,600, and 8,500 feet. B. HOST LIST Although Forties pinicola is most conmmonly found on coniferous trees it occurs on deciduous ones as well. The host list which follows includes 91 species and has been compiled from the reports or collections of Atkinson (1), Boyce (6), Cutler (12), Dudley (13), Farlow and Seymour (16), Graves (22), Faull (17, 18), Hedgcock (26), Long and Harsch (34), Neuman (41), Oudemans (46), Pennington (48), Saccardo (56), von Schrenk (58, 59, 60), Sydow (67), Spaulding (65), Weir (72), and Zeller (75). The list contains all the hosts which have been found recorded for F. pinicola but does not attempt to include every locality from which it has been reported on each host nor the names of every collector who has reported it. Coniferous Hosts Host Locality Collected or reported by 1 Abie, it tt (< « tt (I it tt a Laru tt tt tt it tt Picea ft a tt It (t tt Pinu a Oudemans 2 United States Weir, Hedgcock M 3. ,1 amabilis u « tt 5. 6 tt it u (t Canada, Ontario. United States, Michigan Faull Pennington Hedgcock s tt « q 11) tt tt a 11 u Hedgcock, Weir a 12 tt 13 tt it 1 | « « 1 ") Germany Sydow in United States Hedgcock, Weir 17 18 « u a 19 tt Cl 20. l)l nobilis u tt tt it 22 Farlow and Seymour United States, New Yoik Europe Dudley 9? Saccardo 2d United States Hedgcock " Wisconsin Neuman u Canada, Ontario Faull 25 United States Hedgcock, Weir 26 a n Idaho Canada, Ontario United States Long and Harsch 27. canadensis tt Faull Hedgcock 28 Engelmanni excelsa mariana it << Weir 29 Oudemans •M). United States " Wisconsin Hedgcock, Weir Neuman tt Canada, Ontario United States Faull ril Hedgcock .,., tt 33. sitchensis Canada, British Columbia United States Canada, Ontario United States Cutler Hubert 34. 5 Banksiana M ounce Hedgcock 36. Jcffrcyi » a Coniferous Hosts — Concluded Host 37. Pinus Lambertiana. 38. " Mayriana. 39. " monophylla. 40. " monticola . 41. " palustris. 42. " pinaster. 43. " ponderosa. 44. 45. 46. 47. 48. rigida . Sabiniana. strobiformis . Strobus. sylvestris . 49. 50. 51. " virginiana 52. Pseudotsuga taxifolia. 53. Tsuga canadensis . 54. " heterophylla . M « 55. " Mertensiana. Locality " Wisconsin , Canada, Ontario United I « u Canada, United £ « Canada, United I << Canada, Canada, United ' states Montana New Mexico Washington British Columbia states , Wisconsin New York North Carolina Ontario states Montana British Columbia British Columbia states Collected or reported by Hedge* >ck uu Weir Oudemans Hedgcock, Weir von Schrenk Hedgcock Faull Hedgcock Neuman Faull Saccardo Hedgcock <« Weir Long and Harsch Bartholomew Cutler Neuman Dudley Graves Faull Hedgcock Weir Cutler Cutler Hedgcock Deciduous Hosts OS. 69. 70. 71. 72. 73. 74. 7.-,. 70. 77. Acer saccharinum, " saccharum . . , Alnus glutinosa. u incana " oregona... " tenuifolia. Betula spp " Jontinalis. " lenta lutea occidentalis . u papyrifera . verrucosa.. Cnrya sp Castanea vesca.. . . Fagus atropunicea. " ferruginea . . sylvahca. Magnolia foetida Platanus occidentalis . " orientalis. . . Populus balsamijera. grandidentata . tremula. United States. Adirondacks. Germany, Baden. Canada United States Europe . . United States. Montana. Adirondacks. Montana Canada, Ontario United States, Wisconsin. United States, Wisconsin. United States " Adirondacks. Canada, Quebec Germany, Baden United States. United States. Michigan. N.E. United States. Canada, Ontario Hedgcock Atkinson, Pennington Spaulding Weir Faull Hedgcock Weir Saccardo Hedgcock Spaulding Atkinson, Pennington Hedgcock a Weir Hedgcock Faull Neuman Oudemans Xeuman Oudemans Hedgcock Atkinson, Pennington Mounce Weir Saccardo Hedgcock Oudemans it Hedgcock Pennington Hedgcock Spaulding Faull Saccaido 8 Deciduous Hosts— Concluded Host 80. Popolus Iremuloides . 81. " trichocarpa. 82. Prunus arnum 83. c* Cerasus 84. " persica 85. " serolina. . . . sp. 86. Pyrus communis 87. " Malus 88. Quercus pedunculated. " spp S9. Salix lasiandra frO. Amygdalus persica. . 01. Prunus domestica. . . Locality United States N.E. United States. United States United States. Canada, Ontario. United States, Idaho. Germany, Baden Canada. Selkirk Mts. . United States, Oregon. Collected or reported by Hedgcock, Weir Spaulding Hedgcock, Weir Oudemans Zeller Farlow and Seymour Hedgcock, Pennington Spaulding Faull Oudemans Oudemans Weir << Saccardo Weir Zeller Zeller IV. OCCURRENCE Fomes pinicola usually grows saprophytically, destroying wood in stumps and fallen logs, or in standing trees which have been killed through some other agency, such as fire, bark beetles, or budworm. Although there is evidence to show that it may grow and produce sporophores on living trees, so far, no defi- nite proof of its parasitism, such as White (76) found for F. applanatus by the artificial infection of living trees and by showing, microscopically, that the hyphae attack living tissues, has been obtained. As early as 1894 Nilsson (44, 45) reported F. pinicola attacking the spruce trees of the Norrland and Dalecarlian forests (Kopparberg). He described the gradual browning and cracking of the wood of infected trees, and considered this fungus responsible for their death. In writing of the diseases of conifers in the forests of Sweden the same author states that F. pinicola is parasitic in Southern Norrland and in Dalecarlia, and saprophytic in Southern Sweden. References to the occurrence of F. pinicola on living trees in America are common. In 1901 Atkinson (1) reported, "The fruit-bodies of Polyporus pinicola are sometimes found on the trunks of living hemlocks where it is apparently a wound parasite, entering by means of old knot-holes, branch wounds, fire scars, and similar injuries." Hedgcock (26) finds it attacking " living trees to some extent gaining entrance through heartwood or sapwood as it rots both readily." Graves (22) reports F. pinicola as the cause of a rot of hemlock adding that it is " a dangerous enemy, not only because of its omnivorous habit, but chiefly on account of its ability to attack living as well as dead trees." Meinecke (35, 36) states that it is the commonest timber-destroying pore-fungus in California " exceedingly destructive to fallen timber, and may cause damage to living trees as well;" and, again, " The writer has found it on thrifty sugar pine in Central California, where it was undoubtedly parasitic in the sense of attacking the sound heartwood of living trees through an open fire scar, and extending toward the sapwood." Zeller (79) finds F. pinicola to be the cause of heartrot in the wood of peach {Amygdalus persica) and of prune (Prunus domestica) trees. Neuman (41) found sporophores of this fungus on living and dead trees of hemlock, tamarack, birch, white and red pine, and spruce. The same author gives a 9 detailed description of F. pinicola as a parasite on tamarack. There was a large wound at the base of the tree and, on this, sporophores of the fungus had developed. When the tree was cut down and examined, it was found that near the wound the rot had reduced the entire heartwood and most of the sapwood to a brittle brown condition, and it had spread upward to a height of eight feet from the base of the tree. In spite of this the top of the tree was still green, though it had a sickly appearance, and some of the lower branches were dead or losing their needles. Weir (72) cites a similar instance in which this fungus was found on a wound on a living apple tree. However, as Wreir (75) explains later, " In the writer's opinion the term ' wound parasite ' as it is ordinarly employed is misleading. The parasitism of but few wood-destroying fungi has been investigated. The mere fact that they are found growing from wounds does not imply that they would attack the living cell. Fomes pinicola, one of the most common saprophytes, chiefly on coniferous wood, not unfrequently enters through wounds and destroys the heartwood of living trees, but it would not be considered parasitic." Mr. A. W. McCallum collected, at Gaspe, Quebec, F. pinicola on living Abies balsamea where it caused a typical butt-rot. The fungus had probably entered through a root which was adjacent to infected stumps.* Boyce (6) in writing of decay in Sitka spruce (Picea sitchensis) says, " The red belt Fomes which is most common on snags, windfalls, and other debris, also causes a butt rot in living trees The typical decay is light reddish brown in colour, crumbly and brittle, breaking up into rough cubes with mycelium felts in the cracks." Hubert, in a letter dated January 19, 1926, writes as follows, with reference to a brown rot of spruce, " I know that Fomes pinicola and Polyporus Schweinitzii, both producing brown rots, are common in living Sitka spruce." The writer has seen a sporophore of F. pinicola on a living Douglas fir {Pseudotsuga taxifolia) , and, during the summer of 1925, collected specimens of typical Fomes pinicola rot from two living trees of Sitka spruce at Queen Charlotte City, B.C. Cultures were made from both specimens, and mycelium typical of Fomes pinicola obtained. So far no sporophores have devel- oped in culture, so that identification is not absolute, but there seems little doubt that these results are in accordance with Dr. Hubert's statement. Rankin (50) seems to sum up the opinions cited above when he states that F. pinicola " occurs less frequently in living trees than it does on dead standing trees and logs," but that it may attack trees which have been wounded, or are in gener- ally poor health. In any case F. pinicola is everywhere the cause of serious loss through its rapid destruction of dead standing timber. Von Schrenk (60) describes condi- tions in the Black Hills Forest Reserve, where stands of Western Yellow Pine trees which had been killed by beetles were soon worthless through the activity of this fungus. He found a similar condition in an area which had been burned over four years previously (58). His field observations led him to conclude that, " as a result of the growth of this fungus, where the timber on forest lands has been killed it will be found practically valueless after six or eight years at the most." Von Schrenk's conclusions are substantiated by the observations of Humphrey (53) and others made in Eastern Canada where areas of spruce and balsam had been killed by the spruce bud-worm. Experiments have been made on the utilization of wood infected with F. pinicola, for pulp (54). This fungus belongs to the group which destroys cellulose, and as a re-ult, the yields from both early and advanced stages of rot are very low, and the pulp itself is of little value. *Report of the Dominion Botanist, .Department of Agriculture, Ottawa, for the year 1927, page 40. 10 V. THE SPOROPHORE The rapidity with which Fomes pinicola spreads over large areas of dead standing timber is due, in part at least, to its prolific production of large sporo- phores (plate I), which live for many years and increase their pored surface each year. It is not uncommon to find twenty-five or more of these large brackets on one side of the trunk of a western hemlock, and on Queen Charlotte Islands 121 sporophores were counted on a hemlock trunk (Tsuga heterophylla) , of which only about 40 feet were left standing. The spores are carried in all directions by the wind and sooner or later find any exposed surfaces on the neighbouring trees. Beetle holes afford an easy means of entrance and Spaulding (64) reports that the mycelium of F. pinicola has been found frequently in them. Von Schrenk (58) makes the further suggestion that the holes made by wood-boring insects " may serve to account for the fact that the decay of a large tree takes place with such rapidity." The sporophores of F. pinicola vary a great deal in form and colour. Murrill (40) describes them as ungulate, Overholts (47) as convex or ungulate. They are as often broadly applanate (plate I, fig. 2) as they are ungulate (plate I, fig. 1) even in the same locality and on similar hosts. The upper surface in some specimens is definitely zoned; in others only sulcate with age. There is as wide a variation in colour as there is in form. Young sporophores may have the upper surface entirely covered with a clear, resinous, sticky, varnish-like coating rang- ing in colour from ferruginous and cinnamon-rufous to Hays russet and Kaiser brown (Ridgway, 52) ; or the lower edge of the sporophore may consist of a broad band, even 5 cm. wide, which is identical in colour with the hymenium and in marked contrast to the reddish brown varnished appearance of the upper surface (plate I, fig. 2). In older sporophores the upper surface becomes hard, roughened, and dark in colour, more or less mottled with smoke grey, greyish olive, seal brown, and black, but usually retaining at the margin a band of varying width which exhibits the reddish brown tones found on the young sporophore. These sporophores eventually become very large. Overholts (47) gives the fruit-body measurements as 4-15 by 6-20 cm.; Dr. Faull collected a beautiful specimen at Timagami, Ontario, which measured 20 by 36 cm.; while Schulz (61) found two specimens, on dead stumps near Vigny, which measured 39 by 20 cm. and 45 by 24 cm. respectively. Bataille (2) makes a note of the fact that ammonia causes the hymenium of Polyporus marginatus to become a rose red colour. A similar reaction is obtained when sporophores formed in cul- tures of F. pinicola are exposed to ammonia fumes. In contrast to the variations in the colour of the exterior and in the form of the sporophore the colour and texture of the fruit-body context, the width and colour of the hymenial tubes, and the size of the spores remain fairly constant and offer reliable criteria for the identification of this fungus. The context is corky to woody in texture and more or less concentrically zoned; it ranges in> colour from colonial buff and light ochraceous salmon to shades of pinkish and cinnamon buff; and in younger specimens it may reach a width of 5 cm. or more, while in the later growth of older specimens it gradually decreases until it is only a few millimeters in width. There may be, particularly in younger speci- mens, a layer of context -5-3 mm. wide between the layers of tubes but this is not a constant character. The tubes are concolorous with the context and vary in colour as it does, except that the last formed hymenial layer may be lighter in colour than the others. The hymenium ranges in colour from white, light and pinkish buff, and light pinkish cinnamon, to shades of pinkish cinnamon. Around the pored surface, and concolorous with it, there is a sterile band from 2-5 mm. wide. The pores are small, 4-5 to a millimeter. The spores range in size 11 from 3.4 by 5 /* to 4.5 by 7.3 /i averaging 3.5 by 6.5 /x. They are ovate to subglobose, hyaline, thin-walled, with coarsely granular contents and occasion- ally oil drops (plate II, fig. 1). Complete data regarding the spore discharge period of F. pinicola are not available. From the vicinity of Ouelph, Ontario, Stone (66) reports a visible spore discharge from a sporophore of this fungus in May. The writer has obtained spore casts in the middle of May from sporophores collected by Dr. Fault at Timagami, Ontario, and in September from sporophores on western hemlocks in Stanley Park, Vancouver, B.C. Mr. G. D. Darker of the Depart- ment of Botany, University of Toronto, very kindly set spore-traps on Bear Island, Timagami, and kept records of spore fall from June-August, 1924. He reported the spore discharge continuous throughout June, no evidence of it during July, and a few spores caught in August. This would suggest that F. pinicola, like F. applanatus, has a spore discharge period of several months, but con- firmation is necessary. VI. PREVIOUS CULTURAL STUDIES Cultural studies of Forties pinicola have been made by Brefeld, Rumbold, Long and Harsch, Fritz, and Schmitz. Rumbold was interested particularly in the effects of the acidity of media upon mycelial development; Long and Harsch and Fritz in the cultural characters of the mycelium, and Schmitz in physiological specialisation as exhibited by this fungus. Brefeld (7) tells us no more than that he received his material from Norway and that the spores of the fungus germinated rapidly, developing a large amount of aerial mycelium which in old cultures produced clamp-connections but remained sterile. Rumbold (55) obtained her cultures from sporophore tissue. She describes in a general way the colour and texture of the mycelial mat and notes and figures the fine hypha with their clamp-connections. She grew the fungus on a series of tubes of media whose acidity was so adjusted that when tested with litmus they gave red, red violet, violet, blue violet, or blue colour. Of these the first three were acid to phenolphthalein, the fourth slightly alkaline, and the fifth alkaline. When the mycelium of F. pinicola was grown on this series it was found that the greatest amount of growth occurred on the most acid medium and that the amount of growth decreased with the aridity, until only a slight growth occurred on the neutral medium and none at all on the alkaline one. Long and Harsch (34), too, used mycelium from sporophore tissue' in their study of Fomes pinicola. One strain was obtained from a sporophore which grew on Pseudotsuga taxifolia, the other from a sporophore which grew on Larix occidentalis. No detailed results of their studies are given in their preliminary paper except that both strains of the fungus produced a white mycelium and sporophores. The strain from Pseudotsuga produced a sporophore in malt cultures both in the presence and absence of light, and after a period of 77 days. The strain from Larix produced sporophores only in the light, and on parsnip and prune agars. The sporophore on the prune agar culture required 43 days to develop. Fritz (21) includes Fomes pinicola among the seventeen wood-destroying fungi for which she worked out a system of identification by cultural characters. She studied and has described (1) the macroscopic features of the mycelial growth, such as the texture, colour, rate of growth, and manner of advance on the agar surface; (2) the microscopic features of the aerial and submerged mycelium 12 including the types and colour of the hyphae, the method and frequency of branching, septation, occurrence and type of clamp-connections; and (3) the variations in the character of the mycelium which occur when the fungus is grown on potato dextrose, potato, malt, corn, and Czapek's and dime's synthetic agars. The cultures for this work were obtained from sporophores and wood from dead Populus grandidentata, Betula alba, and Abies balsamea, from sporo- phores from dead Pinus Strobus, Picea mariana, Larix americana, Tsuga carta- densis, and Primus serotina, and from living Abies balsamea. Schmitz (57) studied four strains of Fomes pinicola, which he obtained from sporophores from dead Douglas fir, white fir, western hemlock, and western white pine, with reference to physiological differences in (1) growth characters, (2) intracellular and extracellular enzyme activity, (3) the inhibiting effect of these strains upon one another, (4) growth upon media containing various sub- stances as a source of nitrogen, (5) growth on liquid media, and (6) wood- destroying properties. The four strains of Fomes pinicola were grown in large flasks on carrot. Under these conditions the strain from white fir gave a much heavier and more coriaceous mycelial mat than any of the others, while that from Douglas fir remained fluffy in appearance. The mycelial mats were removed from the flasks, dried and ground, and used in the study of intracellular enzymes, while the liquid in the flasks and juices extracted from the carrots were used in study- ing extracellular enzyme activity of the various strains. Th's study showed that there was a positive and similar action for all strains in curbohydrase and catalase activity; negative results for all strains for urease and amidase; no definite results for rennetase and tannase activity; while slight differences occurred in the relative esterase and glucosidase activity. Similarly a study of intracellular enzyme activity showed positive action and similar results for all strains in carbohydrase and tannase activity, negative in all for protease, and differences in esterase and glucosidase activity. In all mixed cultures the effect of the various strains upon each other was evident. In all plates having two inoculations of the same strain there seemed to be no inhibiting or stimulating effect, the colonies grew into each other and covered the agar surface as if the growth had resulted from a single inoculation. On the other hand whenever plates were inoculated with two different strains the colonies did not intermingle. In these cases a clear line of demarcation remained between the two colonies (cf . plates VI and VII) . In order to study their wood-destroying properties the four strains of F. pinicola were grown for six months on weighed blocks of the following woods: heart and sapwood of Pinus ponder osa, heartwood of P. monticola, Larix occi- dentalis, Pseudotsuga taxijolia, Abies grandis, Tsuga heterophylla, and Picea Engelmanni. At the end of that time the blocks were dried and weighed and the loss of weight calculated. In only one case did the fungus in question cause a more rapid decay on the wood of the host from which it was originally obtained. The white pine strain caused the highest grand average of decay. Schmitz summarises his results by saying, "The writer believes that the data presented are sufficiently conclusive to indicate that there may be considerable physiological variation within the species Fomes pinicola Fr. Whether or not this variation is the result of host influence is not certain." 13 \n. CULTURAL METHODS AND CONDITIONS OF GROWTH A. ORIGINAL CULTURES Cultures of the mycelium of Fomes pinicola were obtained from spores, from sporophore tissue, or from wood which had been decayed by F. pinicola. Small pieces from the interior of a sporophore or from a freshly exposed surface of the diseased wood were removed with sterile forceps and partially embedded in agar slants. Subcultures were made by removing a small piece of mycelium-covered agar with a sterfle loop and placing it on a fresh agar slant. Cultures were made on liquid and solidified media; those on liquid media were set up in 200 cc. Erlenmeyer flasks; those on solidified media were set up in test tubes 15 by 2 2 cm.; except that in one experiment Erlenmeyer flasks were used and, in another, glass culture tubes 2 by 12 inches. B. CONDITIONS OF GROWTH In most cases tube cultures were grown at room temperature and in diffuse light. The tubes were placed on a table in trays which were slanted at an angle of about 15° so that they received light from a window opposite them. For special experiments cultures were grown in incubators at 15°, 22°, 27°, 30°, and 35° C. C. MEDIA 1. Prune Agar. — 120 g. dried prunes were soaked overnight in 400 cc. dis- tilled water and then steamed for one hour. 26 g. agar were dissolved in 750 cc. distilled water by steaming in the autoclave for 30 minutes at 15 pounds pressure. 250 cc. of the prune extract were added then to the melted agar, the whole filtered through cotton, tubed, and sterilized in flowing steam for 15 minutes on each of three successive days. 2. Potato Dextrose Agar. — This medium was made according to a formula given to me by Dr. Fritz, viz., 400 g. sliced potatoes and 1 1. distilled water were steamed in an autoclave 30 minutes at 15 pounds pressure and strained. The potato decoction thus obtained was used to replace the unabsorbed water drained from 25 g. shredded agar, which had been soaked overnight in 1 1. dis- tilled water. The agar was then melted by steaming in an autoclave 20 minutes, at 15 pounds pressure, and on removal 25 g. dextrose were added before tubing. To prevent prolonged decomposition of the sugar from the action of heat, the autoclave was heated to steaming before the tubes were placed in it for sterili- zation. They were then sterilized 10 min. at 15 pounds pressure. 3. Carrot, Celery, Bean, Pea, Cornmeal, and Parsnip Agars were made in the same way as (2) with the omission of dextrose and using the following amounts of vegetables per litre: carrot 490 g. ; celery 600 g. ; bean 400 g.; parsnip 490 g. ; pea 400 g.; cornmeal 80 g. 4. Beet Agar was made as for medium (1) using 490 g. of vegetable per litre. 5. Czapck's Synthetic Agar. 0-5 g. magnesium sulphate. 1-0 g. mono-potassium phosphate. 0*5 g. potassium chloride. 0-01 g. ferrous sulphate. 2-0 g. sodium nitrate. 30 0 g. dextrose. 25-0 mm. diam. 1 rudiment, 4x4x2 mm. 1 4x4x3 mm 0. No visible rot ex- cept small aiea on one side. Advanced stage, friable beneath mycelial Layer. Advanced stage of rot. Small amount char- coal-like, friable. Small amount char- coal-like, friable. Block rotted except outside laj er. Centre of block in advanced stage. Advanced stage, centre of block only. Centre of block rot- ted. Small amount in advanced stage. No visible rot. Small amount ad- vanced. Advanced stage of rot interior of block. Block rotted, friable advanced stage. No visible rot. Part of block badly rotted. Interior of block only, advanced rot Most of block friable No visible rot. Small area rotted. Centre of block rotted . Small amount, typi- cal. Very small amount. Small amount near inoculum, friable. Typical rot beneath dense mycelium. Centre of block in advanced stage of rot. No rot. Almost completely rotted. Advanced stage, wood soft, friable, mycelial sheets. Part of block rot- ted. No rot visible. Decay around out- side of block only; just beneath mycel 26 I am very much indebted to Dr. J. H. White of the School of Forestry, University of Toronto, for his kindness in supplying the material used in this experiment. B. LARGE WOOD BLOCKS Blocks of Tsuga heterophylla 2 by 2 by 4 inches were prepared, placed obliquely in 2-quart jars upon layers of water-soaked cotton, plugged, and sterilized. A square of mycelium-covered agar from a culture of Fomes pinicola was then placed upon the upper transverse surface. The cultures were grown in the light and at room temperature, and were moistened with sterile water at intervals of three months. A thin, spreading, downy-silky, white mycelium grew out from the inoculum and gradually covered part of the upper transverse surface and grew slowly over parts of the lower sides of the blocks. This mycelium later became sub-felty to powdery in texture, but otherwise no further change occurred until one year after the inoculations had been made. At that time sporophores began to develop, appearing first as minute hemi- spherical masses of white mycelium which gradually increased in size and some- times coalesced. There were ten of these sporophore rudiments on one block (plate X, fig. 2) six on the upper transverse surface and two on each of two sides; on the other block there were five, all of which were much elongated in shape and occurred on the two under sides of the block. They varied in size from 4-13 mm. in diameter and the largest measured 24 by 15 by 6 mm. When sectioned these mycelial masses exhibited the typical F. pinicola context, corky in texture with tints of massicot yellow. The blocks were not moistened again, and no pores developed. When the blocks were removed nineteen months after they had been inocu- lated they were found to be perfectly dry and, except for shrinkage which was evident from the presence of several large longitudinal cracks, there was no out- ward sign of decay. When they were split open, however, both blocks were found to be rotted from top to bottom (plate X) . The mycelium had spread rapidly in a longitudinal direction and gradually outward so that the central part of the blocks was almost completely destroyed. Except for a very thin layer at the surface the wood was much darker in colour and had contracted in both directions producing longitudinal and transverse cracks which were filled with masses of white hyphse. The wood did not splinter but broke off sharply, and was very friable. In fact all the characters of the typical F. jnnicola rot were present and the red brown wood with charcoal-like fracture held together with sheets of white mycelium did not differ in any way from wood of Tsuga heterophylla rotted by the same fungus under natural conditions. XI. EFFECT OF VARIATIONS IN TEMPERATURE AND ACIDITY UPON MYCELIAL DEVELOPMENT A. TEMPERATURE Prune agar slants were inoculated from a 25-day-old culture of Fomes pinicola (158) and grown, in quadruplicate, at the following temperatures: (a) in a refrigerator (6°-10° C.) ; (b) in incubators at 22°, 29°, 32°, 35° C; and (c) in the light at room temperature. Observations were made and recorded at weekly intervals and these are summarized in table IV. From this it is evident that variations in temperature caused differences in the rapidity of mycelial growth and in the texture and colour of the mycelium developed. The influence of temperature upon rapidity of mycelial growth was very definite. F. pinicola grew very slowly at a temperature of 6°-8° C, rapidity of growth increased with the temperature up to about 29° C; from that point an 27 increase in temperature resulted in a decrease in growth, and a temperature of 35° C. became almost inhibitive (plate IV, figs. 1-4). At first the mycelial mats produced at 29° C. and 32° C. were similar in texture but by the end of the second week those at 29° C. were the most luxuriant, those at 32°C. next, 22° C. next, then 35° C. and finally those at 6°-8° C. and at room temperature. Colour appeared first in the cultures at 32° C, then in those at 35° €., and at room temperature. It was deepest in those cultures grown at 32° C. and 35° C fairly deep in those at room temperature, and very faint in those grown at 29°, 22°', and 8° C. The influence of temperature upon the rate of growth of the mycelium of Forties pinicola is similar to that found by Fritz (21) for F. igniarius, F. roseus, F. foment arms, and P. Schweinitzii; its influence upon colour production in this form, i.e. intensification of colour by increase of heat, is similar to that found for F. igniarius, F. roseus and Lenzites sepnaria. It was also found by Fritz that diffuse light caused a deepening of colour, and the colour present in theinycelia grown at room temperature and in the light is just another instance of that fact. No sporophores were produced, even in the light, by any of these cultures of Fomcs pinicola during the time they were under observation. However, the cultures grown at a temperature of 6° -8° C. in the refrigerator were left there for almost eight months. At the end of that time no apparent change had occurred; the cultures were not dried up, but no sporophores had been1 produced. They were removed then and placed in the light at room temperature and two weeks later two of them had produced pinkish buff, pored areas. Since Long and Harsch (34) have obtained sporophores of F. pinicola in cultures grown in the dark it would seem that the low temperature and not lack of light inhibited sporophore development in this instance. The table which follows summarizes the results of experiments on the influence of temperature upon the texture, rate of growth, and colour production in cultures of F. pinicola. Table IV. — Influence of Temperature on Mycelial Development in Cultures of Forties pinicola Days 6°-8°C. Room 22°C. Xo growth Short downy growth on inoculum . . 8 mm. diameter, downy-silky, white 20 x 22 mm., downy-silky, white. . . 20 x 40 mm., thin, downy, pale pink- ish buff on inoculum. 12 x 18 mm., thin appressed, white. 22 x 45 mm., downy-felty 22 x 62 mm., thin silky-downy to felty, tints of pale pink- ish buff. Medium covered thin, felty, pale pinkish buff. Pale pinkish buff to pinkish buff. 22 x 36 mm., downy, white. 22 x 68 mm. downy-silky to woolly-felty. Medium covered, very dense, downy-felty, drops. Downy growth around sides of tubes, white. Tints of pale pinkish buff. Days 29°C. 32°C. 35°C. 7 U 21 28 35 22 x 50 mm. dense, downy-cottony, white. Medium covered, very dense, woolly-felty. Felted, except base of tube filled with downy-felty growth. Downy growth around sides of tubes, tints of pale pinkish buff. No change 22 x 30 mm., very dense, downy silky, white. 22 x 46 mm. dense, downy- felty. Medium covered close, felty, drops, pale pinkish buff. Close felty, pale pinkish buff. Pale pinkish buff to pinkisli buff. Thin downy growth on inocu- lum only. 22 x 25 mm., fairly thin, cottony. 22 x 40 mm., dense, downy, white. Felted, pale pinkish buff to pinkish buff. Drying out. B. ACIDITY Fomes pinicola grows so well in culture on a number of media, untitrated, that only one experiment was made to test the effect of the initial acidity of the medium upon mycelial development. Czapek's synthetic liquid medium (modi- fied) was brought to a pH value of 4-2, 4-8, 5-2, 5-8, and 6*2, by the addition of monobasic or dibasic potassium phosphate. 75 cc. of each medium were placed in each of six 200 cc. Erlenmeyer flasks. Three of each series were inoculated with F. pinicola mycelium which had been isolated from Picea (219), the remaining three of each series were inoculated with mycelium isolated from Abies balsamea 15776). All of the cultures were grown in diffuse light at room temperature. After two weeks the mycelial growth was still entirely submerged and consisted of a more or less spherical, whitish, translucent mass of hyphse around the inoculum — a mass not more than 3 cm. in diameter, and usually much less. It was quite evident, however, that the greatest amount of growth occurred in the media with a pH value of 4-8 and 5-2, and the least in that with a pH value of 6-2. After four weeks the masses of mycelium had increased in size but were still entirely submerged, except that a small amount of aerial mycelium had developed in the flasks containing media with a pH value of 4-8 and 5-2. The cultures were kept under observation during four months. At the end of that period the media in all the flasks with the exception o'f that with an original pH value of 6-2 had become pale amber in colour. There was slightly more aerial mycelium in the flasks containing media with an original pH value of 4-2, 4-8, and 5-2, but the amount of growth in all cultures was practically equal. Since fungous growth usually causes a change in the acidity of the medium it may be that in those cultures with a high initial acidity a point was reached where further growth was inhibited, while in those which were originally more nearly neutral, growth could continue for a longer period, giving finally a similar result in all flasks. The pH value of the medium at the end of the experiment was not deter- mined, and, since F. pinicola does not grow well in liquid media the experiment was not repeated. However, it does seem definite that F. pinicola prefers an acid medium, as Rumbold (55) found, and preferably one with a pH value of 4-8-5-2. XII. VARIATIONS IN CULTURAL CHARACTERS AND IN THE FORMA- TION OF A LINE OF DEMARCATION IN MIXED CULTURES A. VARIATIONS IN CULTURAL CHARACTERS OF MYCELIUM FROM DIFFERENT SOURCES Throughout the description of cultures of the mycelium of F. pinicolai frequent references have been made to the variations which occur in rate of growth, in amount and texture of mycelium, in colour, in zonation, and in sporophore production when comparative cultures are made of mycelia from various sources. These differences in cultural characters are, apparently, due to individual variations rather than to host influence. At least it has not been found that one type of mycelium is isolated, constantly, from one type of fruit-body, from one host, or from different hosts in one locality. To study variations in cultural characters fifty-seven cultures of F. pinicola including 158-306, and 5G55- 5778, together with a number of monosporous and polysporous origin, were sown in quadruplicate on prune agar, and grown in an incubator at 22-5° C. At the end of seven days the mycelium was uniformly downy with practically no appressed advancing zone; the average amount of 29 growth in all tubes was 31-7 mm. Culture 5772 (from Larix) and 280 (Picea) grew most rapidly; 5655 (Pojmlus) was much denser and silkier in texture than the average; 282 {Picea mariana) produced by far the densest mat; while 284 C. (P. mariana) gave the least growth, producing a thin, appressed to powdery mycelium. F. pinicola does not produce much colour on prune agar at any time, and still less in the absence of light. However, there were traces of colour in 302 {Pinus) and in six cultures from Picea. At the end of ten days the medium in most tubes was three quarters or completely covered with mycelium. Growth was much denser than the average in cultures 268 and 282 (P. mariana). Zonation was present in cultures 267, 274. 280, 281, 283, 283A. 284, 284B, 284B + +, 284D, and 285B (all from P. mariana) and in 305 {Betula alba). Colour was entirely absent from cultures 269, 270, 282, 283, 283A, 284A ++, 284D, 285C, 285E + +, and 287 (all from P. mariana), and from 160 and 5772 {Larix), and from 302 {Pinus). Varying amounts of pale pinkish buff, pale ochraceous salmon, pinkish buff, and light pinkish cinnamon were present in the remaining cultures. Colour was deeper than the average in 219, 268, and 271 {Picea), and especially in 300 {Picea), which was uniformly pinkish buff throughout the culture. Culture 284 {Picea) still gave the least growth, while 5655 {Populus) was unique with its pure white mycelium, uniformly dense and velvety. 282 {Picea mariana) resembled it somewhat, but the advancing zone was more downy-silky, and the remainder somewhat felted. When the cultures were 18 days old they were divided into the following groups and examined for similarities and differences: (1) cultures from sporo- phores from the same host species; (2) cultures from sporophores from a single tree; (3) cultures from sporophores and from the spores they produced; (4) cultures alike in macroscopic characters. 1. Cultures of F. pinicola from sporophores ivhich grew on the same host species. {a) on Larix laricina. (160 and 5772). — The cultures from both sources were so nearly alike that it was impossible to separate them by mycelial differ- ences (plate IV, figs. 4-8). The mycelium was pure white, dense, downy, and later irregularly felted. (6) on Picea mariana. (267-286). — In this series of 148 cultures from 37 sources there was no culture quite as unique as 5655 {Populus), but there were all gradations from the slow-growing, thin, powdery, mycelium such as 284C, to the rapidly spreading, dense, downy-felty mycelium of 280, which gave next to the best growth of anv culture. There were similar gradations in colour; cultures 282, 284D, and 285D were pure white, while 286, 286A, and 268 were deeply coloured. It is obvious, then, that in this series from Picea mariana there is no definite type of culture associated with this host. (c) on Picea canadensis. (287, 300, 303). — 287 produced a very dense, downy growth, white, with the merest trace of colour. 300 produced a much thinner growth, felted, and exhibiting the deepest colour of any culture. 303 produced a mycelium with the texture of 300 but with the colour of 287. {d) on Populus grandidentata. (304, 56551. — These two cultures were absolutely different in every respect, the mycelium of 5655 was pure white in colour, dense, uniform, velvety in texture, and showed definite zonation; 304 was pale pinkish buff, with points of pinkish buff and cinnamon buff at the base, of moderate thickness, downy to tufted, felted in some cases, and without any sign of zonation. (e) on Pinus Strobus. (302, 306).— These two cultures were alike in tex- ture, both were white with tints of pale pinkish buff, but 306 was slightly denser and had developed more pale pinkish buff at the tip of the culture. 30 (/) on Betula papyrifera. (305, 5657) . — These two cultures were similar in texture, downy and irregularly felted, but the colour was much deeper in 305. (g) on Abies balsamea. (5775, 5776). — Culture 5775 was thin, downy to subfelty, and white in colour; 5776 was denser, felted, and white tinted with pale pinkish buff. The evidence presented shows that cultures from sporophores growing on one species of host do not, necessarily, or even usually, resemble one another very closely; i.e. although there are many different types of cultures it has not been possible to associate one type with cultures from any one host. 2. Cultures isolated from several different sporophores which grew on one tree and from the infected wood of the tree itself. All of the material used in this series came from Picea mariana and was collected at Timagami, Ont., by Dr. Faull. (a) Cultures 283 and 283A, from infected wood and from the context of a sporophore which grew upon it. The cultures in these two series were identical in every way, markedly so in the presence in both series of two denser zones of mycelium, corresponding, probably, to two exposures to sunlight. (b) 284 (from infected wood), 284A, B, C, D, E, (sporophores which grew on 284). Culture 284, 284B, and 284D were identical in every way, downy in texture, and showing two zones of mycelium which was denser and somewhat more deeply coloured than the rest, 284A resembled this group in texture, but grew more slowly. 284C was unique. It grew very slowly, producing a thin, subfelty mycelium. 284E produced a mycelium which was denser than the others and deeper in colour, while zonation was almost entirely absent. (c) 285A, B, C, D. All of these cultures were somewhat similar, white in colour, or only faintly shadowed with pale, pinkish buff, downy in texture, and of moderate thickness; yet they were distinct enough that the twenty tubes could be sorted into their respective groups without much difficulty. Cultures of 285B showed zonation, those of 285C were more evenly downy and grew around the sides of the tubes, those of 285D were denser and more felted, those of 285 were felted, but not as dense as 285D. (d) 286 (from infected wood), 286A and 286B (from sporophores which grew on 286) . 286 and 286A were indistinguishable, producing a heavy, downy to woolly growth, and a very deep colour. 286 B was similar in texture but denser, and with no hint of colour. Again, the evidence from comparative cultures shows that even mycelia from a number of sporophores from one tree, and from the infected wood of that tree do not, necessarily, resemble one another very closely. 3. Cultures isolated from several different sporophores and from the spores which they produced. (a) 268 and 268++ (spores from 268). These two mycelia were prac- tically identical; they could be distinguished only by the fainter colour at the bases of the tubes containing 268 ++. (b) 269 and 269++. These two mycelia were identical, uniformly downy- felty, and white. (c) 282 and 282 ++. These two mycelia were entirely different. 282 was pure white, woolly-felted in texture, and very dense; 282++ was tinted with pale pinkish buff, downy, and not so dense. (d) 283A and 283A ++. These two mycelia were identical in texture, fairly thin, and subfelty, but 283A was pure white, 283A ++ was uniformly pale pinkish buff. 31 (e) 284B, and 284B -f +. These two mycelia differed mostly in texture, 284B was less dense, and more uniform in texture than 284B -\ — |-, which was almost woolly and1 definitely zoned. (/) 285A, and 285A-| — f-. These two mycelia were practically identical in texture and colour. Hence, the mycelium obtained from sporophore context may differ from that obtained from spores from that sporophore in colour, in density, or in texture, or in all three; though in many cases the mycelia from two such sources resemble one another closely. It has been shown that mycelia from sporophores from the same host species, or from sporophores from the same tree, or from sporophores and the spores which they produced, need not resemble one another closely in culture. Some idea of the amount of variation in cultures of F. pinicola may be gleaned from plate VIII, figs. 5-8. Fig. 5 illustrates a mycelium isolated from a sporophore which grew on Populus grandidentata. The growth is dense, and uniformly felted. Fig. 6 illustrates a mycelium isolated from Pinus Strobus and shows a dense, uniformly cottony-woolly mycelium which has begun to grow around the sides of the tube. Fig. 7 represents a mycelium isolated from Pinus sp. (New Brunswick), and shows a thin, even, downy mycelium; while fig. 8, isolated from Tsuga heterophylla (Vancouver, B.C.), shows a downy mycelium with irregular, thickened areas of woolly mycelium, which became much denser at the tip of the culture. On the other hand, figs. 1-4 show mycelia with a fairly uniform, downy growth of moderate density, yet (1) was isolated from Tsuga heterophylla, Vancouver, B.C., (2) from Pinus collected in New Bruns- wick, (3) from Tsuga heterophylla collected on Queen Charlotte Islands, B.C., and (4) from Picea canadensis, Timagami, Ontario. In this group, then, there are mycelia from localities as far removed as Vancouver, B.C., Timagami, Ont,, and New Brunswick, yet the cultures are quite similar. Since, then, it has not been found that one type of mycelium is isolated, constantly, from one type of fruit-body, from one host, or from different hosts in one locality, the differences in cultural characters have been considered to be^due to individual variation, rather than to the influence of any specific factor. Yet, as Dr. Fritz (21, p. 225) pointed out, these differences are not great enough to render it impossible to identify F. pinicola in culture, with a fair degree of accuracy. Observations on the rate and mode of growth, colour production, sporophore formation, and the size and types of hyphae produced in culture, combine to give a very fair means of determining this species despite the variations. B. MIXED CULTURES AND THE FORMATION OF A LINE OF DEMARCATION A great deal of work has been done by Zeller and Schmitz (78), Brown (8), Porter (49), and others on mixed cultures of fungi, but their interest lay, primarily, in the behaviour of fungi when different species, genera, or families, were mixed. Thus Schmitz and Zeller worked with such forms as Lenzites, Mendius, Daedalea, Trametes, Pleurotus, and Polyporus: Brown used organ- isms representative of all the great fungous groups. Brown (p. 126) states that, "The question of the intermingling or non- intermingling of two fungal colonies (of the same or different species) is intimately connected with the question of staling. . . . The phenomenon is not one of absolute incompatibility, and investigation would probably show that any fungus could be made to intermingle with any other under appropriate conditions." Porter describes in detail five types of reaction which occur when fungi are grown in the presence of other fungi: type A mutually intermingling, 32 not common; type B, growth superficial over the contending organism; type C, slight inhibition, ("This is the prevailing type when any organism is grown with another individual of the same species ") ; type D, growth around the contending organism; type E, mutual inhibition at a considerable distance. He suggests (P. 174) three possible explanations for the morphological changes which occur in mixed cultures: " (a) The nutrients may be exhausted, (b) The distortions may be due to change in the osmotic equilibrium of the medium induced by the metabolic activities of the growth process, (c) Certain poison- ous products may be created by fungous growth capable of producing mal- formations and creating a zone through which fungous filaments can not pass." He demonstrates, among other things, that inhibitions are not so marked in media rich in nutriments, that they vary but slightly with changes in the amount of inoculum, in time of inoculation, or in depth of medium, and that a "common cause of the inhibitory action in the cases studied was determined to be the presence of some product formed during growth." Schmitz (57), in working with strains of Fomes pinicola, found that there was no intermingling when four cultures of this fungus from four different hosts, Douglas fir, white fir, western hemlock, and western white pine, were paired with one another. Cayley (11), working with Diaporthe perniciosa, goes still further and finds mutual aversion between monospore mycelia. She finds that " the phenomenon of aversion between two antagonistic strains has occurred on all media so far used irrespective of the depth of the medium, the distance at which inocula are placed or the age of the cultures when tested." Aversion was found usually to occur "between mono-mycelia from different hosts whether of the same variety, different varieties, or different species, but mono-mycelia isolated from such widely different hosts as apricot and plum have been found to meet; on the other hand averting strains have been found in different perithecia on the same host, thus showing that the type of strain is irrespective of the variety of host. It is quite possible that the occurrence of averting strains on the same host may be due to multiple infection from two or more different sources, and not to the splitting up into physiological strains in the host plant." Smith (62) notes an interesting fact, namely, the presence of black lines in some crustaceous lichens. "A patch of crustaceous lichen on tree or rock may belong to one species and yet be composed of many individuals which have started from different centres, each growing centrifugally. The dark lines chiefly occur when the different individuals encounter each other. A striking instance of such intersecting lines occurs in the thallus of the well-known Rhizocarpon geographicum. Strong boundary lines also frequently divide different species inhabiting the same substratum." In view of these interesting results it was thought worth while to extend Schmitz' studies of mixed cultures of Fomes pinicola. With the material avail- able it was possible to test the effect on one another of organisms from many different hosts, or from the same host growing in the same or different localities, of organisms from the same individual tree, and of organisms from single spores from the same sporophore. Some experiments were made as well to test the effect of variations in temperature, amount of light, and concentration of nutrients in the medium, on the behaviour of paired cultures. Unless otherwise stated all paired cultures were made on Petri plates 10-12 by 1-12 cm. containing a layer of malt agar about 0-5 cm. thick. The small pieces of mycelium used as inocula were placed about 1-5 cm. apart and near the centre of the plate. Cultures were incubated at 25° C. for from 15 to 20 days. 33 (1) Various Hosts and Various Localities. — In this series all possible pair- ings of seventeen different cultures of F. pinicola were made, using cultures from the following sources: — Cultuie Number Locality 305 635 5778 304 5775 160 5772 300 303 286a 6S4 302 5769 306 558 509 586 Bctula alba Facjus grandijolia Primus scrotina Popitlus grandidentata Abies balsamca Larix laricina LartX laricina Picea canadensis Picea canadensis Picea mariana Pinus sp Pinus Strobus Pinus Strobus Pinus Strobus Tsuga heterophylla Tsuga heterophylla ? Timagami, Ontario. Chelsea, Quebec. York Mills, Ontario. Timagami, Ontario. Guelph, Ontai io. Guelph, Ontario. Guelph, Ontaiic. Timagami, Ontario. Timagami, Ontario. Timagami, Ontario. St. John, New Brunswick. Timagami, Ontario. Guelph, Ontario. Guelph, Ontario. Vancouver, B.C. Queen Charlotte Island, B.C. France. The results are given in table V where the sign + indicates the presence of -a definite line between the two mycelia and sign 0 the complete inter- mingling of the two mycelia. Table V -All Possible Pairings of Seventeen Cultures (Diploid) of Fomes pinicola from various Hosts and various Localities 305.. 635.. 5778.. 304.. 5775.. 160.. 5772.. 300.. 303.. 286A 694.. 302.. 5769.. 306.. 558.. 509.. 586.. 00 iO . -* t>. o t^ o CO co •* CNJ CO CO 00 cs O eo t^ o t^. co r-~ o o 00 05 o t~- o «o o CO c© id CO »o "H »o co CO c >o to ~ --c i^ to o t^ t- 00 CO X X X X x X X s. x X X X X co X X X CO ■-Z X X 4 \ 4 4 + 4 5 4 + 4 4 + 4 • 1 I + r + 0 | + + 4 Table X. — Fomes pinicola. All possible pairings of ten monosporous myeclia from culture No. 285B with ten monosporous mycelia from culture No. 285 A. 285A 28 5B " 2 ~3 4 1 "I 7 1 i + T 4 4 + 4- 4- f 2 + + 4 + + + + h .; -i 1 + + -f I + ± i 1 ': + + + f -f ■+ 1 + + + 1 + + + + _!_ T + + + + H + ! ! ":l -1 4 4 •• i 4 f + 4 ! 4 + 4 9 1 I 4 4 -1 4 H; 4 10 ! h + 4 i ! + 4- In every pairing but one (285A-1 x 586-2) clamp-connections were pro- duced; that is each source belonged to a different sexual strain or geographical race, and these strains or races were all mutually fertile. (c) Material from Sweden. — A year and a half later a culture of Fomes pinicola (F. marginatus) from Betula sp. was received from Stockholm, Sweden, through the kindness of Mr. E. J. Eliason. Monosporous mycelia were isolated and all possible pairings were made between five monosporous mycelia from this source and five from each of the sources in (a) and (b). Clamp-connections were formed in all but six of the three hundred pairings made; that is, in 98 per cent of the pairings, strains from sources (a) and (b) were completely cross fertile with those from (c), and table XI is typical of the results obtained. Table XI. — Fomes pinicola. All possible pairings of five mono- sporous mycelia from culture No. 285B (Timagami, Ont.) with five monosporous mycelia from culture No. 928 (Sweden). 285B 928 8 1 4 4- + 5 + + 0 + + 7 + + il + + 2 3 4- + 4- + + + + + + 4- 4 4- 4- + + + Table XII. — Fomes pinicola. All possible pairings of five mono- sporous mycelia from culture No. 283 (Timagami, Ont.) with five monosporous mycelia from cul- ture No. 928 (Sweden) 283 928 6 7 1 + 9 + + 4- + + il 4- 15 4- + + + 16 4- 4- + 4- 4- + 8 1 + + + ~ 2 Four of the six exceptions included mycelium 283-11 (see table XII) and two, mycelium 5770-1. Since these two monosporous mycelia behaved as all the others used in the series of pairings (a) and (b), and refused to behave nor- mally in all subsequent series, it seems probable that their behaviour was due to loss of vigour, and that the six exceptions may be disregarded on that account, and the strain from Stockholm considered completely cross fertile with those from Ontario and from France. 46 {d) Material from British Columbia. — In the autumn of 1925 cultures were made from a large number of sporophores of Fomes pinicola collected in British Columbia. All of these cultures, however, refused to produce sporo- phores and spores, despite every effort that was made. Vandendries (71a) found, in working with cultures of Coprinus micaceus, a similar variation in readiness to produce sporophores. It was not until two years later that one culture, 562C, eventually shed spores. Nineteen monosporous mycelia were isolated and all possible pairings made of ten of them. The results did not differ from those shown in table VIII, and F. pinicola from this source too was shown to be heterothallic and bisexual. Then five monosporous mycelia from 562C were paired with five monosporous mycelia from every other source in (a) , (b), and (c). The results were entirely unexpected for, with seven exceptions, clamp-connections were not produced in any of the 175 pairings made, and table XIII is typical of the series. Table XIII. — Fomes pinicola. All possible pairings of five monosporous mycelia from culture No. 5778 (York Mills, Ont.) with five monosporous mycelia from culture No. 562C (Vancouver, B.C.) 5778 Table XIV. — Fomes pinicola. All possible pairings of five monosporous mycelia from culture No. 586 (France) with five monosporous my- celia from culture No. 562C (Vancouver, B.C.) 586 562C 7 8 9 £ 4 _5_ _6_ 7_ 10 11 562C T7 - 16 17 18 19 1 £ j>_ 7 ± ± _8_ 9 + + + + + The seven exceptions occurred in the 562C x 586 pairings and are shown in table XIV. Even in this case there were only two of the five mycelia of 586, i.e. 586-7 and 586-9, involved. Hence, in this series, where monosporous mycelia Irom British Columbia were paired with monosporous mycelia from Ontario, France, and Sweden, only four per cent of the pairings were fertile, and com- plete sterility is the rule, though there are a few exceptions. (e) Summary. — All the pairings made with monosporous mycelia of Fomes pinicola from different sources are summarized in table XV and several interest- ing facts are revealed. 47 o> ec J oooooc OOOOOOO^OOCNOC CO »n IC >C »C >C CX5 'O 00 *j — c£ B i.~> »r> in to O iO »0 to to to © to to to to i—" to to CO to tr; OS OOOOOt^O t^ r^3 •— » CMCMOI O -— ' • a h H 3 < «- 0] v .8 | iO >C o >o o >o iC >o "5 >c o >o W "5 «5 U5 lO i(5 »o >o ><; CMCNCMCM©CMCMCMCMCM©cMCMCMCMCMeMCMCMCMCN lO lO lO lO >o lO »o >o >o o t^ CMCMCNCMCMCN1CM t^ h5 i I T-C 1— 1 1-1 p o « 3 •< a > "C . 0 z < >> 5 B O 1 * c c 3 "g "3 Co :o ot.^h* 3 tn C S cOSC co ._: 3 co.^ 3 co O H -1^ 3 o ^ e g3 3 C 3 o CO ^ : O o 33 ,c"S gj.gS.gS §, ,dS §, .dS cl ^ ^ «'.§ W 3 O.S 3 O 3 O .5 3 O.S 3 O £ .1 1s|| OP ti P o 5 < h H O^fecx « • 2 e a .§ "S > 5 e '.5 •^8 :■« •S 8 ^3 e e C3 S S o « O o e . o Q e 03 o w I3 c V i.09 I ft00 e «0 S CO =0 X co ~ ?: - x u03 2- 3 e > iOK3iOiflO>Oir:>rjiflicO»OinK5iOiOiOiOW5iOiO lO to to to to to to § £ sa (T o ^ H X X KK««XXXXXXXXX«X«HX« X X X X X X X fc O J5 — ■+-» « fis -< &H g 4> s 3 S 6 2 o CC PQ 2 P 3 o O a 5 3 B a | 3 . - o g2 3 3 3 3 03 3 3 3 3 3 8 H 3§ ' "% 3 o o 3 e3 H > B 05 Si i e H ;;;;;; -5 o a : o • ^333333 1 CO O w : c : ••* p. S 3 5 3 5 S S a 33 332 « M 3 e 3+3 iCS3 3 - 3 3 > a H o a £ ^K cq ^ < cu H > 2 £ £ lOiOiOiflO^iOiCiflKJOiOifliCiOifliOiCiCiCiO to to to to to tO to "- ' •^ 48 1. Clamp-connections were produced in each of the 325 pairings using material from deciduous and coniferous trees from various localities in Ontario. That is material from each source belonged to a different so-called sexual strain or geographical race and these strains or races were all mutually fertile. The particular case of pairings between 285A and 285B is rather interest- ing. Any monosporous mycelium from sporophore 285A forms clamp-connec- tions when paired with any monosporous mycelium of 285B, table X, showing that they belong to two different sexual strains, Yet sporophores 285A and 285B were growing at the same time on a black spruce. In working with Coprimes lagopus Hanna found (24, p. 442) that in pairings made of monosporous mycelia from two fruit-bodies arising from the same compound mycelium the mycelia " react together in every respect as though they had been isolated from a single fruit-body ". Apparently then there was more than one source of infection in this tree and the mycelium which produced 285A was of different origin from that which produced 285B. This theory is supported by the fact that a black line is produced when cultures 285A and 285B are grown in the same Petri plate culture. No such line is formed when the two inocula come from the same source, and frequently none develops when the two inocula come from spores and the tissue of the sporophore, or from the decayed wood on which the sporo- phore developed (see section XIII B). This result emphasizes the care which is necessary in studying, for instance, the progress of decay in individual trees. 2. In 350 pairings of European and Canadian strains of Fomes pinicola, omitting strain 562C, only six of the number failed to produce clamp-connec- tions; i.e. the European and Canadian strains used were mutually fertile and, by the clamp-connection criterion, have been shown to be identical.* 3. Culture 586 from France was labelled Fomes pinicola, var. marginatus, and culture 928 from Betula sp. was submitted by E. J. Eliason, Stockholm, as typical of the F. marginatus form. With the exception of culture 562C, all pair- ings of the Fomes pinicola and F. marginatus forms were fertile so that, again, by the clamp-connection criterion, these two forms have been shown to be identical. 4. In pairings of monosporous mycelia of culture 562C from British Columbia with those from all other sources in Canada, Sweden, and France, 94 per cent of them remained sterile. This strain of F. pinicola duplicates in part results obtained by Vandendries with Coprinus micaceus. He found that sterility was the rule when monosporous mycelia from material from different geographical areas were paired; "... toute souche est fertile pour tous les individus de sa region, de meme qu'elle est sterile pour tous les individus des regions eloignees ". However, experiments made so far with strains of F. pini- cola from various regions indicate that, with this fungus, fertility is the rule, and sterility the exception. ^Recently monosporous mycelia have been obtained from culture 694, isolated from a sporophore of Fomes pinicola which grew on Pinus sp., York county, New Brunswick. These monosporous mycelia were paired with monosporous mycelia from each of the other sources in both Canada and Europe. Clamp-connections were formed in every pairing except those with culture 562C from British Columbia. 49 XIV. DESTRUCTION OF WOOD Fomes pinicola causes a rot of sapwood and heartwood of coniferous and deciduous trees, but is particularly destructive to the former. It probably does most damage in stands of trees which have been killed by other agencies. Under such conditions it works with remarkable rapidity and soon renders the timber worthless. The rot which it causes is quite characteristic, and readily recognizable, especially in the more advanced stages (plate I). At first it is characterized by a darkening of the wood, due to the removal of cellulose by the fungus. Even at this stage it is impossible to get a smooth surface with a plane, for the wood becomes brittle and breaks away irregularly. The removal of cellulose results in shrinkage of the wood causing the formation of longitudinal and horizontal cracks which become more and more numerous. Gradually these spaces are filled with wefts of white mycelium and in the final stages the wood is reduced to a mass of wood with charcoal-like fracture which absorbs water readily, but it is frequently dry and brittle, and always very friable. This mass is held together by the mycelial felts, which consist of closely intertwined fungous hyphae and are similar to the mats formed on agar cultures. Badly decayed wood of this kind gives the characteristic red colour with phloroglucin and hydrochloric acid, but remains unchanged in chlor-zinc iodine, indicating the absence of cellulose in decayed wood. Sections of the wood of Abies and Picea, which had been rotted by Fomes pinicola, were cut and examined. Evidence of the shrinkage which had occurred was common in most sections, particularly in the" form of radial cracks in transverse sections. Vertical shrinkage seemed to result frequently in the separation of the cells of medullary rays causing lengthwise cracks in the rays as seen in longitudinal radial sections. Hyphse were frequent both in the tracheids and medullary ray cells. They were all hyaline and thin-walled, but of two distinct types (1) the usual branched hyphee, with clamp-connections, varying from 6-7 ju, to 2-3 /* in width, but com- monly 2-3 n; and (2) hyphse which were exceedingly fine, sparsely septate, and very much branched. They developed so profusely that they completely filled the lumen of the tracheids. The work of the fungus in medullary ray cells is evident not only from the presence of hyphse which grow in every direction through the rays but also from the presence of bore holes and a great number of enlarged pits. Hyphae were found frequently passing through these enlarged pits and entering the adjoining tracheid. Not uncommonly the cells of the medullary rays were filled with a deeply staining granular substance. Similarly, in the tracheids, bore-holes made by the fungus and enlarged pits were very common. Frequently longitudinal slits occurred in the cell wall on both sides of bordered pits and bore-holes. Hyphae passed from one tracheid to another in radial or tangential direction. They made use of the bordered pits, even of the ones in the tangential walls of the summer wood, but at the same time bore-holes were frequently found passing through the border of a bordered pit. The most characteristic feature, microscopically, both in longi- tudinal and transverse sections was the presence of rows and groups of cells in which the lumen was completely rilled with a very densely woven mass of the fine, much-branched hyphse, so closely packed that, in cases where the cell wall had broken away, the mycelium still retained the shape of the cell. Sections of the wood o'f Tsuga which had been inoculated with Fomes pini- cola and grown in culture showed similar evidences of the work of this fungus in the presence of bore-holes, hyphae with clamp-connections, splitting of the cell wall, and the presence of tracheids completely filled with mycelium. 75474-4 50 XV. SUMMARY 1. A list of 91 hosts is given for Fomes pinicola (Sw.) Cooke. [Fomes marginatus (Fr.), Fomes ungulatus (Schaeff.) Sacc] 2. Spores of F. pinicola were sown in twenty-five different substrata and found to germinate in many of them. Germination took place at temperatures ranging from 8° to 35 °C. Light retarded but did not inhibit germination. 3. Sporophores of F. pinicola have been obtained in cultures made on malt and on prune agar, and on Czapek's synthetic liquid medium containing various substitutes for the dextrose of the formula ; and on eleven varieties of coniferous and deciduous wood, with rudimentary sporophores on fourteen others. The context of the sporophores of F. pinicola produced in culture was typical both in colour and texture, and the spores produced were viable. 4. A Foraes-type of fruit-body with three definite pore-layers, each of which shed spores, developed in a culture on wood of Pinus divaricata. 5. The mycelium of F. pinicola grew at temperatures ranging from 8° to 35 °C. with an optimum temperature of about 27° to 29 °C. Growth was retarded at both the maximum and minimum temperatures. Sporophore production was inhibited at 8°C. The mycelium grew best in an acid medium, preferably one with a pH value of 4-8 to 5-2. 6. Young cultures of mycelia from various sources differ from one another in rate of growth, colour production, and texture of the mycelial mat; older cultures tend to become more uniformly white and felted. These differences have been ascribed to individual variation rather than to host influence. Despite these variations a study of the macroscopic and microscopic characters of the mycelium made it possible to identify this fungus with a fair degree of accuracy. 7. A study of mixed cultures of F. pinicola gave no definite clue as to the cause of the frequent formation of a line of demarcation or a space of aversion between two mycelia from different sources. 8. A method of isolating monosporous mycelia of F. pinicola or of similar forms is described. 9. F. pinicola is a heterothallic bisexual species. 10. Many so-called sexual strains, or geographical races, which were com- pletely cross fertile, were found. 11. Monosporous mycelia of F. pinicola which have been kept in culture for five years, have remained in the haploid condition. In this they are unlike monosporous mycelia of two heterothallic bisexual species of Coprinus, C. Rostrupianus investigated by Miss Newton and C. radians investigated by Vandendries, many of which were found to change spontaneously from the hap- loid to the diploid condition. 12. Monosporous mycelia of F. pinicola isolated from sporophores from deciduous hosts were mutually fertile' with those from sporophores from coni- ferous hosts; monosporous mycelia from cultures from France and Sweden were mutually fertile with monosporous cultures from all Canadian strains except that from British Columbia. These results, interpreted in the light of the clamp- connection criterion for the identity of species, furnish experimental evidence in support of the generally accepted conclusions (1) that F. marginatus and F. pinicola are one and the same species, and (2) that the European and American forms of this fungus are identical. 51 13. Monosporous mycelia of a strain of F. pinicola from British Columbia remained completely sterile when paired with monosporous mycelia from any other Canadian strain or from the strain from Sweden, and were only partially fertile with those from a strain from Franco. These results duplicate, in part, those obtained by Vandendries with Coprinus micaceus. 14. The macroscopic and microscopic characters of rot caused by F. pinicola are described. Typical rot was obtained in artificial culture. This investigation was begun at the Department of Botany, University of Toronto, during the tenure of a studentship and fellowship granted by the National Research Council, and completed at the Division of Botany, Experi- mental Farm, Ottawa. I wish to express my gratitude to Professor J. H. Faull for his kindness in supplying a great part of the material used, and especially for his valuable advice and criticism throughout the progress of the work; to Prof. A. H. R. Buller, who suggested the problem; and to Mr. H. T. Gussow, Dominion Botanist, for his unfailing interest in the investigation. I am also indebted to Dr. C. W. Fritz, Dr. A. H. Hutchinson, Dr. J. H. White, Mr. E. J. Eliason, and the Forest Products Laboratory, Vancouver, B.C., for material, and to Mr. C. W. Lowe for the photographs used in plate I. XVI. BIBLIOGRAPHY i. Atkinson, G. F. Studies of some shade tree and timber destroying fungi. Cornell Univ. Agr. Expt. Sta. Bui. 193: 199-235. 1901. . 2. Bataille, F. Miscellanees mycologiques. Bull. Soc. Mycol. Fr. 25: 79. 1902. Bot. Centralb. 3: 542. 3. Bayliss, Jessie S. The biology of Polystictus versicolor Fr. Jour. Econ. Biol. 3 (1) : 1-24. 1908. 4. Bensaude, Mathilde. Recherohes sur le cycle evolutif et la sexualite chez les Basidio- mycetes. 156 pp. Nemours. 1918. 5. Boyce, J. S. The dry-rot of incense cedar. U.S. Dept. of Agr. Bull. 871 : 1-58. 1920. 6. Boyce, J. S. Diseases of commercially important conifers in the Pacific Northwest. U.S. Forest Service, Study Course 1923, District 6, Course V, Enemies of the Forest, Part I— Disease. February 26, 1923. Reissued September 17, 1926. 7. Brefeld, 0. Untersuchungen aus dem Gesammtgebiete der Mycol ogie. 8: 111-112. 1889. 8. Brown, William. Experiments on the growth of fungi in culture media. Ann. Bot. 37 (145): 105-129. 1923. 9. Brunswik, H. Untersuchungen iibetr die Geschlechts- und Kernverhiiltnisse beJ der Hymenomyzetengattung Coprinus. Botandsche Abhandlungen. 5: 1-152. 1924. 10. Buller, A. H. R. The biology of Polyporus squamosus Huds. Jour. Econ. Biol. 1 (3) : 101-138. 1906. 11. Cayley, Dorothy M. The phenomenon of mutual aversion between mono-spore mycelia of the same fungus (Diaporthe perniciosa Marohal) with a discussion of sex- heterothallism in fungi. Jour. Genetics. 13 (3) : 353-370. 1923. 12. Cutler, N. L. A contribution to the knowledge of the tree-destroying fungi of the Van- couver forestry district. Abs. in Phytopath. 13 (6) : 294. 1923. 13. Dudley, P. H. Fungi destructive to wood. 41st Ann. Rep. N.Y. State Mus. Nat. Hist. 1887. 14. Duggar, B. M., J. W. Severy, and H. Schmitz. Studies in the physiology of the fungi. IV. The growth of certain fungi in plant decoctions. Ann. Miss. Bot. Gar. 4 (2) : 165-173. 1917. 52 15. East, E. M., and H. J. Mangelsdorf. A new interpretation of the hereditary behaviour of self-sterile plants. Proc. Nat. Acad. Sci. 11: 166. 1925. 16. Farlow, William Gibson, and A. B. Seymour. A provisional host-index of the fungi of the United States. Cambridge. 1888-91. 17. Faull, J. H. Forest Pathology. Report of Minister of Lands, Forests, and Mines, Prov of Ontario, 119-125. 1919. 18. Faull, J. H. Forest Pathology. Report of Minister of Lands, and Forests, Prov. of Ontario. 197-207. 1923. 19. Ferguson, M. C. A preliminary study of the germination of the spores of Agaricus campestris and other basidiomycetous fungi. U.S. Dept. Agr. Bur. PL Ind. Bull. 16. 1902. 20. Fries, Eldas, M. Systema mycologieum I: 372. 1821. 21. Fritz, C. W. Cultural criteria for the distinction of wood-destroying fungi. Trans. Roy. Soc. Can. Sec. V, 191-288. 1923. 22. Graves, A. H. Notes on diseases of trees in the Southern Appalachians. III. Phytopatih. 4 (2): 63-72. 1914. 23. Hanna, W. F. The dry-needle method of making monosporous cultures of Hymeno- mycetes and other fungi. Ann. Bot. 38: 791-795. 1924. 24. Hanna, W. F. The problem of sex in Coprinus lagopus. Ann. Bot. 39 (154) : 431-457. 1925. 24a. Hanna, W. F. Sexual stability in monosporous myoelia of Coprinus lagopus. Ann. Bot. 52 (166) : 379-388. 1928. 25. Hartig, R. Zersetzungserscheinungen des Holzes der Nadelholzbaume und der Eiche. 1878. 26. Hedgcock, G. G. Notes on some diseases of trees in our National forests IV. Phytopath. 4: 181-188. 1914. 27. Hiley, W. E. The fungal diseases of the common larch. Clarendon Press, Oxford. 1919. 28. Hubert, E. E. The diagnosis of decay in wood. Jour. Agr. Res. 29 (11) : 523-567. 1924. 29. Kauffman, C. H., and H. M. Kerber. A study of the white heart rot of locust, caused by Trametes robiniophila. Am. Jour. Bot. 9 (9) : 493-508. 1922. 30. Kniep, Hans. Beitrage zur Kenntnis der Hymenomyceten, III, IV, and V. Zeitschr. f. Botanik. 7: 369-398, 1915; 8: 353-360, 1916; 9: 101-119, 1917. 31. Kniep, Hans. Ueber morpholbgische und physiologische Geschlechtsdifferenzilemng. Ver- handl. der Physikal-Med. Gesellschaft zu Wiirzburg. 46: 1-18. 1919. 32. Kniep, Hans. Ueber Geschlechtsbestimmung und Reduktionsteilung. Ibid. 47. 1922. 33. Lloyd, C. G. Synopsis of the genus Fomes. Cincinnati, Ohio. January, 1915. 34. Long, W. H., and R. M. Harsch. Pure cultures of wood-rotting fungi on artificial media. Jour. Agr. Res. 12 (2) : 33-82. 1918. 35. Meinecke, E. P. Forest tree diseases common in California and Nevada. U.S. Dept. Agr. Forest Serv. Field Manual. 1914. 36. Meinecke, E. P. Forest pathology in forest regulation. U.S. Dept. of Agr. Bui. 275. 62 pp. 1916. 37. Morgan, T. H. Recent results relating to chromosomes and genetics. Quart. Rev. Biol. 1 (2) : 186-211. 1926. 38. Mounce, Irene. Homothallism and heterothallism in the genus Coprinus. Trans. Brit. Myc. Soc. 8: 256-269. 1922. 39. Mounce, Irene. A preliminary note on Fomes pinicola (Sw.) Cke. and Pholiota adiposa Fr. — two heterothallic species of wood-destroying fungi. Abstract. Phytopath. 16 (10): 757-8. 1926. 40. Murrill, W. A. North American Flora. 9 (2) : 72-131. 1908. 41. Neuman, J. J. The Polyporacese of Wisconsin. Wis. Geol. Nat. Hist. Survey. Bull. 33: 1-206. 1914. 42. Newton, Dorothy E. The bisexuality of individual strains of Coprinus Rostrupianus. Ann. Bot. 40: 1-24. 1926. 53 43. Newton, Dorothy E. The distribution of spores of diverse sex on the hymenium of Coprinus lagopus. Ann. Bot. 40 (160) : 891-917. 1926. 11. Nilsson, Alb. Om barrtradsrotor och deras upptradande i vara skogar. (Ueber Nadcl- holzfaule und ihr Auftreten in den Schwedischen Waldern.) Tidskrift for Skogshu- sfiHning. Heft 1. 15 pp. Stockholm. 1896. From Botanisches Central bkvtt, Bd. 66: 328-329. 1896. 46. Nilsson, Alb., und K. G. G. Norling. Skogsundersokningar i Norrland oeh Dalarne, somniaren 1894 utforda pa uppdrag af k. Domanstyrelsen. (Untensuehungen del Walder Norrlands und Dalekarliens, im Auftrage der k. Direction der Dorruinen im Sommer 1894 ausgerfiihrt.) Bihang till Domanstyre'Isens underd. Berattelse. 38 pp. Stockholm. 1895. Bot. Cejntralblatt. Beihefte VI: 41-45. 1896. 46. Oudemans, C. A. J. A. Enumeratio systematica fungorum. 1919. 47. Overholts, L. 0. Polyporacece of the middle western United States. Wash. Univ. Stud. 3 (1): 1-98. 1915. 4S. Pennington, L. H. Fomes pinicola Fr. and its hosts. Midh. Acad. Sci. Rept. 9: 80-82. 1907. 49. Porter, Charles Lyman. Concerning the characters of certain fungi) as exhibited by their growth in the presence of other fungi. Amer. Jour. Bot. 11 (3) : 168-188. 1924. 50. Rankin, W. Howard. Manual of tree diseases. Macmillan Co., New York. 1918. 51. Rhoads, Arthur S. The biology of Polyporus pargamenus Fries. N.Y. State Coll. of Forestry, Tech. Publ. 11: 1-197. 1918. 52. Ridgway, Robert. Colour standards and colour nomenclature. Washington. 1912. 53. Rue, John D., R. N. Miller, and C. J. Humphrey. Decayed wood for sulphite pulp. Paper presented to Can. Pulp and Paper Assoc, Montreal, Jan. 24, 1924. 54. Rue, J. D., R. N. Miller, and C. J. Humphrey. The utilization of decayed wood in the chemical processes. (Paper Trade Journ., May 15, 5 pp. 1924.) Rev. Appl. Myc. 4(1): 5-6. 1925. 55. Rumbold, C. Beitrage zur Kenntniss der Biologie holzzerstorender Pilze. Naturw. Ztschr. Forst. u. Landw. 6 (2) : 81-140. 1908. 56. Saccardo, P. A. Sylloge fungorum omnium hucusque cognitorum. 6: 167. 1888. 57. Schmitz, Henry. Studies in wood decay V. Physiological specialization in Fomes pinicola Fr. Amer. Jour. Bot. 12 (3) : 163-177. 1925. 58. Von Schrenk, Hermann. Fungous diseases of forest trees. Year Book of U.S. Dept. of Agr. Pp. 199-210. 1900. 59. Von Schrenk, Hermann. Some diseases of New England conifers. U.S. Dept. Agr. Div. Veg. Physiol, and Path. Bui. 25: 1-56. 1900. 60. Von Schrenk, Hermann. The " bluing " and " red rot " of the western yellow pine, with special reference to the Black Hills Forest Reserve. U.S. Dept. Agr. Bur. PI. Ind. Bui. 36: 1-40. 1903. 61. Schulz, R. Mitteilung iiber einige ungewohnlich grosse Polyporaceen. (Verh. bot. Ver. Pr. Brandenburg, pp. 73-75. 1916) Bot. Centr. 137 p. 314. 1917. 62. Smith, A. L. A handbook of tihe British lichens. P. 5. London. 1921. 63. Spaulding, Perley. A disease of black oaks caused by Polyporus obtusus Berk. Rept. Miss. Bot. Gar. 16: 109-116. 1905. 54. Spaulding, Perley. Two fungi growing in holes made by wood-boring insects. Rept. Miss Bot. Gar. 15: 73-77. 1904. 65. Spaulding, Perley. Diseases of eastern hemlock. Proc. Soc. Amer. Foresters. 9: 251. 1914. 66. Stone, R. E. Visibility of spore dissemination in Fomes pinicola. Trans. Brit. Myc. Soc. 6 (3) : 293. 1919. 07. Sydow, Hans u. Paul Sydow. Mycotheca germanica. Berlin. 68. Vandendries, Rene. Nouvelles reoherches sur la sexualite des Basidiomycetes. Bull. Soc. Roy. Bot. Belg. 56 (1) : 1-25. 1923. 09. Vandendries, Rene. Contribution nouvelle a Petudc de la sexualite des Basidiomycetes. La Cellule. 35 (1) : 129-157. 1924. 54 69a. Vandendries, Rene. L'tetercHhomothallisTiie dans le genre Coprinus. Bull. Soc. Roy. Bot. Belg. 57 (2) : 1-8. 1925. 70. Vandendries, Rene. La tetrapolarite sexuelle des Coprins. Bull. Soc. Roy. Bot. Belg. 58 (2) : 1-7. 1926. 71. Vandendries, Rene. Les mutations sexuelles, I'hetero-homothallisme et la sterilite entre races geographiques de Coprinus micaceus. Bull, de la Classe des Sciences dq l'Acad. Roy. de Belgique, t. IX, 1927. 71a. Vandendries, Rene. Nouvelles recherches experimental es sur le comportement sexuel de Coprinus micaceus. Bull, de la Classe des Sciences de l'Acad. Roy. de Belgique. t. IX. 1927. 72. Weir, James R. Notes on some wood-destroying fungi wlhio'h grow on both coniferous and deciduous trees. Phytopath. 4 (4) : 271-276. 1914. 73. Weir, James R. Montana forest tree fungi I. Polyporacece. Mycologia 9 (3) : 129-137, 1917. 74. Wear, James R. Altitudinal range of forest fungi. Mycologia 10 (1): 4-14. 1918. 75. Weir, James R. Concerning the introduction into the United States of extra4imital wood-destroying fungi. Mycologia 11 (2) : 58-65. 1919. 76. White, J. H. On the biology of Forties applanatus (Pers.) Wallr. Trans. Roy. Can. Inst. 12: 133-174. 1919. 77. Zeller, S. M. Studies in the physiology of the fungi II. Lenzites scepiaria Fries, with special reference to enzyme activity. Ann. Mo. Bot. Gar. 3: 439-512. 1916. 78. Zeller, S. M., and Henry Schmitz. Studies in the physiology of the fungi VIII. Mixed Cultures. Ann. Mo. Bot. Gar. 6 (3) : 183-192. 1919. 79. Zeller, S. M. Wood decay in orchard trees in Oregon. Oreg Agric Expt Sta. Crop Pest and Hort. Rept. 3: 132-138. 1921. XVII. DESCRIPTION OF PLATES 5G Plate I Fig. 1 (upper). — Transverse section of block of Tsuga heteropltyUa sfhowiing ungulate sporo- phore of Fomes pinicola and advanced stage of rot. f nat. size. Fig. 2 (lower). — Longitudinal section of block of Tsuga heterophylla showing applanate sporophore and stages of rot. £ nat. size. Plate I 58 Plate II Fig. 1. — Spores of Fomes pinicola. x 800. Figs. 2-5. — Germinating spores of F. pinicola. x 490. In 1% lactose. Figs. 6, 7. — Germinating spores of F. pinicola. x 490. In prune agar 3 days. Fig. 8. — Germinating spore of F. pinicola. x 490. Same spore as fig. 6, 24 hours lateir. Fig. 9. — Germinating spore of F. pinicola. x 490. In beet agar 2 days. Fig. 10. — Germinating spore of F. pinicola. x 490. In prune agar 3 days. Fig. 11. — Germinating spore of F. pinicola. x 490. In parsnip agar 2 days. Figs. 12-14. — Types of thin-walled hyphse with clamp-connections. From aerial mycelium of tissue culture on prune agar, x 800. Fig. 15.— As figs. 12-14. x 490. Fig 16. — Thick-walled fibre-like hypha from aerial mycelium of same culture as fig. 12. x 800. Figs. 17-18. — Types of thin-walled, irregularly branching hyphse from submerged mycelium of a tissue culture on prune agar, x 490. Figs. 19-21. — Chlamydospores from submerged mycelium of a month-old tissue culture on malt agar, x 800. Figs. 22-23. — Types of thin-walled hyphse from submerged growth of a monosporous myce- lium on malt agar, x 800. Fig. 24. — Fibre-like hypha from submerged growth of a monosporous mycelium on prune agar, x 800. Fig. 25. — Thdn-walled hypha with clamp-connections obtained by pairing two monosporous mycelia, 158-2 x 5770-2, on Czapek's synthetic agar, x 800. Fig. 26. — Hypha with yellow-brown coarsely granular contents from black line area in a paired culture of 283 x 286A. x 800. Plate II 60 Plate III Cultures of Fomes pinicola grown in diffuse lilght at room temperature. Nat. size. 1^^^&!^a^3)^ fdtGd myCeliura' ^ophores, and spore-deposits (mask- F^^^dHf 2tube.SP°rOPh0re Wkh C°arSe POreS- Indicati'on of ^ore-deposit on right- Fig. 5.— Culture 158. Sporophore and typical growth on Czapek's synthetic agar. Fig. 6.— Culture 5770 from Picea mariana showing pores and spore-deposit. Fig. 7.— Culture 5775 from Abies balsamea showing pores and spore-deposit. Fig. 8.— Culture 5657 from Betula alba showing pores and spore-deposit. Fig. 9.-Culture 5778 from Prunus serotina showing pores and spore-deposit. Fig. lO.-Culture, 5769 from Pinus Strobus showing pores and spore-deposit. Figs. 6-10.— The similarity of texture of mycelial mat in old cultures is shown. Plate III 5 8 10 62 Plate IV Figs. 1-4. — Fomes pinicola. Ten-day old cultures of 158 on prune agar. Nat. size. Fig. 1.— Incubated at 35°C. Fig. 2.— Incubated at 31 °C. Fig. 3.— Incubated at 29°C. Fig. 4.— Incubated at 22°C. Figs. 5-8. — Cultures of F. pinicola from Larix. On prune agar. Fig. 5.— Culture 5772. Figs. 6-8.— Culture 160. Plate IV 4 1 IK j,^> 1 CAL/BCA OTTAWA K1A . 0C5 39073 00215603 4 STUDIES IN FOREST PATHOLOGY I. Decay in Balsam Fir (Abies balsamea, Mill.) : by A. W. McCallum, Forest Pathologist. OTTAWA F. A. ACLAND PRINTER TO THE KINGS MOST EXCELLENT MAJESTY 1929