Drechsler, Charles

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Reprinted from SCIENCE, Mny 23, 1952, Vol. 115, New2995, pages 575-5/(i.Widespread Distribution of Delacroixiacoronata and other SaprophyticEntomophthoraceae in Plant DetritusCharles DrechslerDivision of Vegetable Crops and Diseases,Bureau of Plant Industry, Soils, and AgriculturalEngineering, ARA, USDA, Beltsville, MarylandDelacroixia coronata (Cost.) Sacc. & Syd. (1, 2)is one of the few species in the Entomophthoraceaewhich, unlike the many insect parasites belonging inthat family, develop well on various artificial mediacommonly used for cultivating microscopic fungi.When -planted on Petri plates of a fairly transparentmedium like maize meal agar it reveals vigorousmycelial growth, abundant formation of large globoseconidia singly on aerial conidiophores, forcible pro-jection of these conidia some distance from their placeof origin, frequent germination of a conidium byemission of a vegetative germ tube, and equally fre-quent germination by production of a globose sec-ondary conidium on a short, stout aerial outgrowth.It often shows also, though in greatly varying meas-ure, a more distinctive type of reproduction whereina largo globose conidium will simultaneously put forthplural—usually about a dozen—short outgrowths, andon them will give rise terminally to a correspondingnumber of spores, all markedly and in about equaldegree smaller than their parent. Although this multi-plicative reproduction, which led Costantin to erecta separate genus for the fungus, may have lost somevalue as a diagnostic character through its observedoccurrence also in Conidiobolus brefeldianus Couch( 3 ) , it has gained significance with respect to thebasic morphology of the family through the discoveryof two parasitic species, Meristacrum asterospermumDrechsl. (-i) and Ballocephala sphaerospora Drechsl.(5), that reproduce asexually by giving rise only tosmall conidia. In disclosing the large conidium as asporangium with a manner of spore developmentsomewhat like that of Cunninghamella, the multipli-cative reproduction of D. coronata establishes thesmall-spored forms as primitive members of the Ento-mophthoraceae. The resemblances of M. asterosper-mum to Gonimochaete horridula Drechsl. ( 6 ) , whichproduces endogenous immotile spores in aerial hyphaloutgrowths, seem suggestive of remote phylogeneticconnections of the family with lagenidiaceous types,such as Protascus subiliformis Dang. (7) and Hapto-glossa heterospora Drechsl. ( 4 ) , that form endogenousimmotile spores, presumably homologous with zoo-spores, in submerged intramatrical thalli.D. coronata has been encountered at firsthand soinfrequently that it is generally regarded by mycolo-gists as a rare species. During 30 years it has ap-peared adventitiously in my cultures only four times.More recently, however, a procedure designed espe-cially for isolating microscopic fungi that commonlyproduce their spores in a short time and soon shootthem off forcibly has shown the species to be virtuallyubiquitous in leaf mold and in other kinds of plantdetritus resulting from exposure of vegetable mate-rials to slow decay in moist contact with the ground.In this procedure samples of detritus are first freedof their coarser components. Portions of the finerfriable residue are then fastened with soft agar tothe underside of a lid from a sterilized Petri dish. Bygently kneading the mixture of detritus and agar allparticles of decaying material are thoroughly moist-ened and thus made to adhere firmly enough to remainin position when the lid is turned right side up. Thewhole central area of the lid may advantageously beFIG. 1. Petri plate culture with canopy of leaf mold, show-ing bottom view, A, of lid, and horizontal view, B, of culturewith lid in position : a, adhering puddled mixture of leaf moldand soft agar ; b, circular barrier of soft vaseline or heavymineral oil ; c, layer of agar culture medium. Approx x .75.planted with detritus (Fig. 1, a), but an outer zoneabout 15 mm wide should be left clean and as nearlysterile as possible. If small particles have fallen intothe clear zone the whole outer area should be wipedwith a sterile cotton plug slightly moistened in alcohol.Should the detritus appear rather badly infested withmites or other minute crawling animals it may beexpedient to restrict their wanderings by applyingsoft vaseline or heavy mineral oil in a circular band(Fig. 1, 6) around the central area. The lid is thengently placed on the lower half of a sterilized Petridish containing a firm plate of moderately transpar-ent, sterile.agar culture medium (Fig. 1, c ) . WhereverD. coronata is present in the canopy of moist vege-table detritus it produces conidiophores from whichconidia are forcibly propelled. After falling on theagar below, many of the conidia germinate vegeta-lively, thereby giving rise to scattered mycelia. Asthese inycelia are usually free of alien organisms theyreadily yield pure cultures when transferred to sterileagar slants. More than a dozen pure cultures, alloriginating from separate conidia, may often he ob-tained from a single Petri plate within 2 days afterit has been covered with a detritus canopy.In addition to D. coronata other members of theEntomophthoraceae capable of "saprophytic" growthdevelop in canopied plate cultures, though in lesserquantity. The genus Conidiobolus would seem onlymeagerly represented in the collection of cultures iso-lated from leaf mold taken up in different localitiesin Maryland and Virginia late in the autumn of 1951.A species of Basidiobohis has been obtained ratherfrequently from dark, soggy leaf mold taken fromwet, wooded areas near Beltsville, Md. When it isgrown on maize meal agar the species produces nu-merous zygospores with the curious paired beaklikeprotuberances long familiar in textbook illustrationsof B. ranafum Eidam (8), though the thick wall sur-rounding the individual zygospore seems always com-pletely colorless. Its asexual reproduction has so farnot been observed directly, yet some production andforcible discharge of conidia must in each instanceprecede its appearance in a canopied agar plate cul-ture. Moreover, when a sizable slab of maize mealagar occupied by its vegetative mycelium is affixedadhesively to the ceiling of a Petri dish containingsterile agar culture medium, the fungus will begingrowing in the agar below quite as certainly, thoughnot as promptly, as will D. coronata under similarconditions. By such affixture above a sterile agar plateentomophthoraceous mycelia that in a canopied cul-ture have become contaminated with bacteria or beenovergrown by alien fungi may often be easily purified.At temperatures near 20° C myeelial strands ex-tended downward here and there from a canopy ofdetritus, mainly b}' members of the Mucorales, willusually reach the agar floor in 2% or 3 days after theculture was prepared. The culture thereupon becomesrather quickly overgrown by miscellaneous microor-ganisms and soon is greatly impaired in usefulness,if not utterly ruined. Success in isolating entomoph-thoraceous forms by the procedure here recommendedmust be achieved early. Newly collected moist detritusshould, as a rule, provide most rapid development.However, material collected in an air-dry or nearlyair-dry condition has given very satisfactory resultsfor several months—far better results than moist orwet material that after being collected was kept in atight container in a warm laboratory for 2 or 3 weeksbefore it was used. The pronounced deterioration ofthe material in the tight container evidently resultedfrom excessive development of species of Trichoderma,Aspergillus, and Penicillmm, as well as of members of 'the Mucorales, during the brief period of storage.Where moist plant detritus cannot be used immedi-ately the saprophytic entomophthoraceous forms andother components of its natural microflora, includingmany predaceous and parasitic fungi destructive torhizopods and eelworms, can be preserved fairly wellby allowing it to dry out slowly at relatively lowtemperatures.ReferencesI.^-COSTANTIN, J. Bull. xnc. muci)l. France, 13. 38 (1897).2. SACCAEDO, P. A., and STDOW, P. In P. A. Saccnrdo (Ed. ) .Sylloge fungorum, 14. 457 (1899).3. COUCH, J. N. Am. J. Botany, 26, 119 (1939).4. DRECHSLER, C. J. Wash. Acad. 8ci., SO, 240 (1940).5. . Bull. Torrey Botun. Club, 78, 183 (1951).G. Ibid., 73, 1 (1946).7. DANOEARD, P. A. Botanlste, Le_, 9, 23 O903).8. EIDAM, E. Beitr. Biol. Pflanz., 4, 181 (1886).Manuscript received December 31, 1951.

Widespread Distribution of Delacroixia Coronata and Other Saprophytic Entomophthoraceae in Plant Detritus

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Widespread Distribution of Delacroixia Coronata and Other Saprophytic Entomophthoraceae in Plant Detritus

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