None of the forty-two one-ascospore cultures produced either chlamydospores or endoconidia and although they have been in culture more than two years they have produced only perithecia and these usually developed sparingly and sometimes not at all. While there may be some difference of growth in the media commonly used for fungi, there has never been the least indication of even the rudiments of chlamydospores or endoconidia. Pea meal agar is the stock medium used and, while the growth on it is usually so delicate as to be readily overlooked, it seems to be a satisfactory one for this fungus, as all the forty-two cultures are still in good condition. The fungus also grows well on corn meal, prune, oat. potato, carrot and bean meal agars. However, there is often a difference in growth of cultures made at the same time and on the same agar. The factor, or factors, bringing about this condition have not been ascertained. With the above media in only a few cultures was there any approach to a cottony mycelium and, while the mycelium was abundant, it still remained delicate, being, in comparison with other fungi, very inconspicuous. MIXED CULTURES OF THIELAVIA BASICOLA ZOPF AND The fact that none of the one-ascospore cultures produced either endoconidia or chlamydospores suggested that the ascospore cultures might be sexually related to the endoconidium-chlamydospore strains without, however, producing any asexual spores With this idea in mind each of three one-ascospore cultures was cultured with eighty-one one-endoconidium cultures, from culture 1351, and with three Thielaviopsis basicola from tobacco, four from violet and one from garden peas. Perithecia were abundantly produced in all. Usually within six days immature perithecia could be detected in the tube placed under the microscope and in eight days almost mature perithecia could be seen with a hand lens. The remaining thirty-nine perithecial cultures were cultured with the eight cultures isolated from tobacco, violet and pea and they were also stimulated to produce an abundance of perithecia. These results seemed to indicate that production of perithecia in this fungus was due to different sexual strains. However, it was early noted that there was a tendency to form perithecia in the one-ascospore cultures even though they were not cultured with the endoconidium-chlamydospore strains. In such cases perithecia were slower in making their appearance and increased in number as the culture grew older. Furthermore the forty-two cultures varied somewhat in this ability of forming perithecia when cultured alone and there was often a difference in cultures of the same strain made at the same time and on the same medium. Ir no case, however, did a single culture alone produce perithecia as quickly or as abundantly as when grown with endoconidium chlamydospore cultures. The factors which cause this difference in production of perithecia have not been ascertained. In autoclaves in which steam is in contact with the tubes a little more moisture at times collects in some tubes than in others, so it was thought that possibly the rigidity of the medium might be a factor. Cultures were made on media containing different concentrations of agar, but these gave no definite results, perithecial formation not being increased in any of them. A few cultures on oat agar, made in the usual way, formed a ring of perithecia along the upper edge of the medium, but these were not general throughout the tube and not always obtained. Possibly a slow drying combined with a favorable medium might be a factor. Occasionally transfers taken from an old dried culture make a more luxuriant growth and form more perithecia than those from newer cultures. For the experiments made on the production of perithecia an ascosporic culture, No. 1603, which alone seldom forms perithecia, was used. MIXED CULTURES OF THIELAVIA BASICOLA ZOPF AND OTHER FUNGI. The writer is indebted to Prof. Thaxter for the suggestion of culturing Thielavia basicola Zopf with other fungi to see if the reaction with them is similar to that with Thielaviopsis basicola (Berk.) Ferraris. First of all, however, Thielavia basicola, culture 1603, the strain which alone rarely produces perithecia, was cultured with the remaining forty-one strains of Thielavia basicola, but there was no additional stimulation in growth or perithecial formation. In this respect the results differ from those obtained by Miss Wineland (32) who found that two one-ascospore cultures of Fusarium moniliforme, grown together, stimulate the production of perithecia. Other fungi were then tried. The same Thielavia basicola culture, No. 1603, was cultured with 120 strains and species, embracing 43 genera, belonging to Phycomycetes, Ascomycetes, Basidiomycetes and Fungi Imperfecti. In all the tests pea meal agar was used and in each case a transfer from one culture of Thielavia basicola was put into a tube with that from one other fungus. Many of the fungi grew so rapidly as to bury the delicate more slowly growing Thielavia basicola and no evidence of its mycelium could be detected, but in others both species survived in the fully mature culture. As a result of these experiments it was found that Thielavia basicola Zopf produces abundant perithecia when it is cultured with Cladosporium fulvum, Aspergillus umbrosus, a strain of Aspergillus glaucus, Eurotium amstelodami and in a less degree with Fusicladium pirinum. In all cases the perithecia produced in these mixed cultures are exactly alike in every respect to those obtained in the original culture isolated from violet and no trace of chlamydospores or endoconidia was found in them. It seems probable that if various other media were used or change of cultural conditions tried that other fungi might react in this same way. It is interesting that results vary with different species of the same genus. While perithecia of Thielavia basicola were abundantly produced when that fungus was cultured with Cladosporium fulvum, isolated from tomato leaf by the writer, three Cladosporium cultures received from Dr. C. L. Shear, two of which were isolated from cranberry and one from blueberry, were unsuccessful. Aspergillus umbrosus, kindly determined by Drs. Charles Thom and Margaret Church, was obtained from peach preserves and Aspergillus glaucus strain and Eurotium amstelodami were received from the same investigators. Eight additional cultures of Aspergillus, received from them and nine others of the same genus, received from Dr. D. H. Linder, as well as one other species from this laboratory, had no effect in stimulating the production of perithecia. Thielaviopsis paradoxa has a striking resemblance to Thielaviopsis basicola, but two cultures received from Dr. J. P. Martin, Honolulu, and one isolated from banana stalk by the writer, were also unsuccessful. Two cultures of Isaria, received from Dr. F. J. Pritchard, had no effect in increasing the number of perithecia. Results, similar to those given above, are recorded by Heald and Pool (13) who found that Melanospora pampeana responded in the production of perithecia when grown with Fusarium moniliforme, Basisporium gallarum and to a smaller extent with Fusarium culmorum. Their description of a one-ascospore culture of Melanospora pampeana as showing a "scanty scarcely distinguishable, white mycelium" might apply equally as well to the one-ascospore cultures of Thielavia basicola Zopf. However, in their one-ascospore culture or in transplants from it they never found perithecia, while on the other hand the one-ascospore cultures of Thielavia basicola Zopf spasmodically produce some perithecia when grown alone on an ordinary culture medium. In opposition to the results of Heald and Pool the writer so far has not been able to get a response of perithecia of Thielavia basicola on cultures of other fungi that have been heated in the autoclave or boiled in the open for twenty minutes. Molliard (16), (17) found that Ascobolus produces perithecia when a bacterium is present in the culture and Sartory announced a similar association necessary for the production of ascospores in a yeast (25) and of perithecia in an Aspergillus (26), (27), (28). The writer's experiments have been sufficiently repeated to demonstrate that Thielavia basicola Zopf, though possessing to some degree the ability to form perithecia when cultured alone, is greatly stimulated in the production of these structures when cultured in association with Thielaviopsis basicola (Berk.) Ferraris, Cladosporium fulvum, Aspergillus umbrosus, Aspergillus of the glaucus group, Eurotium amstelodami and Fusicladimu pirinum. Since Thielavia basicola Zopf does not form either the chlamydospores or endoconidia of Thielaviopsis basicola (Berk.) Ferraris, and since perithecial formation is stimualted equally well by other fungi, there seems to be no real justification for considering Thielavia basicola the perfect stage of Thielaviopsis basicola. Although the frequent association of these two fungi in nature and the fact that their relationship has long been accepted in literature make one reluctant to consider them as two distinct species, yet the cultural evidence is a strong argument in favor of such a view. THE INFLUENCE OF EXTRACTS UPON PRODUCTION OF PERITHECIA. The observations made upon the influence of extracts of some fungi upon production of perithecia are recent, but the experiments have been repeated a sufficient number of times to make them seem worthy of being recorded here. Cultures of several fungi were barely covered with water and allowed to stand about twenty-four hours. The liquid was filtered through paper and then through a Berkefeld filter, thus avoiding sterilization by heat; for, as already stated, cultures of the various fungi, which in living condition stimulate the production of perithecia, after heating, no longer have this stimulating effect. In some cases the liquid only was used and in others the mat of mycelium was crushed in a mortar, squeezed through cheese cloth and the juice thus obtained also filtered with the extract. There is some indication that the latter method is the better one, but this point has not been fully determined. All the extracts from fresh fungi, so far tried, have been made with water only and sterilized by passing the liquid through a Berkefeld filter. Cultures of Thielavia basicola Zopf, No. 1603, were made in the usual way upon pea meal agar and some of the extract, prepared as above described, poured into the tube and allowed to flow over the transplant. In every case the fungus from which the extract was made was also grown upon pea meal agar to avoid conflicting results which might arise from the introduction of some other medium. Check cultures on pea meal agar alone were also made in every series. Nine different extracts of Thielaviopsis basicola (Berk.) Ferraris were made and each one caused a marked stimulation both in growth and perithecial production, in the majority of cases fully equaling in numbers the perithecia formed in mixed cultures of the two living fungi. Following the use of extracts a stimulation in growth is noticeable within a few days, but the formation of perithecia is slower than when two living fungi are cultured together. However, in every series, the checks have no perithecia or only a few scattered ones, while the cultures treated with the extracts have produced them abundantly. The age of the fungus from which the extract is made, amount of water necessary for extracting and the optimum length of time required for extraction are probable factors which affect the number and rapidity of perithecial development. Three different extracts were made from Cladosporium fulvum and one from Aspergillus umbrosus. In all cases these extracts also greatly stimulated growth and perithecial production. As when grown with the living fungi, Thielavia basicola produces no asexual spores when treated with any of the extracts so far used. The effect of heat upon the extracts was tried both in the autoclave at twenty pounds pressure for twenty minutes and by boiling in the open for twenty minutes. Without exception both methods of heating have so far completely destroyed the stimulating power of the extract. Extracts from Aspergillus of the glaucus group, Eurotium amstelodami and Fusicladium pirinum have not yet been tried. As stated above, Thielaviopsis paradoxa, in living condition, failed to stimulate the production of perithecia and it was also suggested that the failure with many fungi in living condition might be due to the smothering of the more delicate Thielavia basicola. If this is the situation, then, extracts from such luxuriant fungi might act as stimulants even though the living fungus gave no result. While one experiment is not sufficient to be conclusive it may be stated here that an extract made from Thielaviopsis paradoxa also greatly stimulated growth and perithecial formation. The case is the same with Saccharomyces cerevisiae. Cultures of yeast were made on pea meal agar and mixed cultures were made from them and Thielavia basicola. In each case the yeast completely covered the Thielavia basicola transplant before it had time to grow; but two different extracts made from fresh yeast cultures, stimulated growth and perithecial production. The stimulating factor in the yeast extract was also completely destroyed by heating. To determine whether commercial extracts of fungi contained the stimulating factor, 1%, 2%, 4% and 6% water solutions of Taka-diastase were made and filtered through a Berkefeld filter. Cultures of Thielavia basicola were treated with these by the same method as with the preceding extracts. The resulting growth far surpassed that following the use of any other extract so far used. The mycelium became very abundant and made almost a felt-like growth. Later, perithecia developed. The type of growth following the addition of Taka-diastase differs so markedly from that in any medium or extract which has been tried that it suggests the presence of two factors, one an unusually potent stimulant for growth and another stimulating perithecial formation. XXXIX. The nature of the factors in any of the extracts above considered, causing such a striking increase in growth and production of perithecia of Thielavia basicola, is beyond the scope of the problem at hand. The results with the mixed cultures of Thielavia basicola Zopf and of other fungi are an indication that this fungus should |