The antibiotic effect of culture filtrates of some soil fungi on rhizobial growth in cultures

Summary Four strains ofRhizobium sp. from peanut (Arachis hypogaea L.) root nodules were tested for their sensitivity to metabolites (culture filtrates) of more than ten common soil inhabiting fungi, in yeast extract mannitol agar and broth cultures. Among the rhizobial strains tested strain CB-530, BU-1 and BU-2 were not sensitive to metabolites ofMyrothecium roridum andFusarium moniliforme whereas CB-1024 was sensitive. Culture filtrates ofGliocladium roseum, Thielavia basicola andDrechslera pedicellata inhibited the strains CB-530 and BU-2 but not CB-1024. Most of the soil inhabiting fungi tested were inhibitory to rhizobial growthin vitro and very few were stimulatory, their effect in either case being strain specific.     

Production of plant growth regulators by someFusarium species

About 10 species ofFusarium were screened and tested for their growth regulatory activity on oat coleoptile straight growth test. Culture filtrates of four species contained growthstimulating factors while others showed growth-inhibition responses on oat sections. On the whole,F. moniliforme was found to be produce higher stimulatory effects. Chromatographic analysis revealed the presence of several indole auxin and gibberellin-like plant-growth regulators in their culture filtrates. The quantity and biological activity of each indole spot was also measured.     

Pectolytic enzymes activity and pathogenicity ofGaeumannomyces graminis var.tritici and related Phialophora-like fungi

Gaeumannmyces graminis var.tritici (Ggt), Phialophora sp. (lobed hyphopodia) andPhialophora graminicola vere grown in a liquid medium with pectin and on autoclaved wheat roots (root media) and the activity of pectolytic enzymes in culture filtrates was measured. Most strains of the fungi exhibited polygalacturonate trans-eliminase activity but no pectin methylesterase activity was detected.Ggt polygalacturonase was found in culture filtrates from all the media used whilePhialophora sp. did not exhibit activity of this enzyme in the unbuffered root media. No polygalacturonase activity was demonstrated forP. graminicola. A correlation was found (r=0.548) betweenin vitro polygalacturonase activity and the pathogenicity ofGgt to wheat seedlings.     

Bacterial and fungal metabolites of rhizospheric microflora of cotton antagonistic to Rhizoctonia solani Kühn

Summary Growth response ofRhizoctonia solani Kühn, the damping-off fungus, to metabolites of selected antagonistic rhizospheric bacteria and fungi of some Egyptian cotton varieties, namely, two strains ofBacillus subtilis Cohn,Aspergillus terreus Thom, andAspergillus flavus Link produced in culture media containing nitrate- or ammonium-nitrogen sources, proved the potency ofB. subtilis metabolites in inhibitingR. solani mycelial growth whether from nitrate- or ammonium-nitrogen culture media. Metabolite filtrates ofB. subtilis (II) are more potent than those ofB. subtilis (I). Increasing concentration of bacterial metabolite filtrates resulted in a decreased mycelial dry weight ofR. solani. The bacterial inhibitory factor forR. solani mycelial growth is partially affected by heat. Metabolite filtrates ofA. terreus from nitrate-nitrogen are slightly more potent than from ammonium-nitrogen culture media while an opposite relation is evident withA. flavus metabolites. Growth responses ofR. solani to different experimental dilutions of metabolite filtrates ofA. terreus andA. flavus proved the intervention of the nutritive factor in witholding growth of the damping-off fungus.     

Efficacy ofTrichoderma chitinases againstRhizoctonia solani, the rice sheath blight pathogen

Thirty-five strains ofTrichoderma viride andT. harzianum were screened for their antagonistic ability against the rice sheath blight pathogen,Rhizoctonia solani. The strains that inhibited/overgrew the phytopathogenic fungus were considered effective. Light microscopic studies showed the antagonism of the hyphae of effectiveTrichoderma strains towards their host hyphae. Chitinase activity ofTrichoderma culture filtrates was enhanced, when colloidal chitin was used as the sole carbon source, instead of glucose. Chitinase pattern differed among the four select strains. The chitinase isoforms are induced differentially by carbon sources. The chitin affinity column fraction ofTrichoderma culture filtrate inhibited,in vitro, the growth ofR. solani.     

Toxin profiles of natural populations and cultures ofAlexandrium minutum Halim from Galician (Spain) coastal waters

The toxin profiles of three isolates and natural populations of the PSP agentAlexandrium minutum from several Galician rías (NW Spain) was obtained by HPLC. The toxin content of cultures ofA. minutum is dominated by GTX4 (80–90%) and GTX4 (10–15%) with small amounts of GTX3 and GTX2 (less than 3% of each); similar results were obtained for natural populations ofAlexandrium from three different Galician rías, where a mixture ofA. lusitanicum Balech andA. minutum can occur. Important quantitative differences were found between the three isolates, one being highly and two weakly toxic. The results obtained from these isolates and natural populations ofAlexandrium were very similar to those obtained from HPLC analyses of mussels intoxicated during a PSP outbreak in Ría de Ares (Rías Altas) in 1984, confirming thatA. minutum (previously identified asGonyaulax tamarensis Lebour andAlexandrium lusitanicum) was the PSP agent during the toxic outbreak in May 1984. Toxin profiles obtained from natural populations during different PSP outbreaks in different rías and from cultures are fairly consistent and suggest that at least from the toxin point of view,A. lusitanicum andA. minutum are identical, and that the toxin profile ofA. minutum from Galicia can be used as a biochemical marker.     

Studies on some fungi isolated from the rhizosphere of tomato plants and the consequent prospect for the control of Verticillium wilt

Summary Biological control of Verticillium wilt disease with antagonistic micro-organisms was studied. Antagonism of some fungi, isolated from tomato rhizosphere, toVerticillium albo-atrum R & B. was observedin vitro. A clearly defined zone, in which the growth of the pathogen was inhibited, was observed withPenicillium spp. (includingPenicillium chrysogenum Thom) andFusarium culmorum (S.G. Sm) Sacc., whileTrichoderma viride pers. ex Fries,Gliocladium spp. andPenicillium vermiculatum Dangeard, suppressed the growth ofV. albo-atrum by penetrating, and overgrowing it. OnlyT. viride andP. vermiculatum culture filtrate added to the Dox's agar, reduced the radial growth ofV. alboatrum. Root-dip application of culture filtrates ofT. viride andP. chrysogenum was found to be most effective in controlling the disease, followed by other species ofPenicillium andGliocladium spp. WhileFusarium culmorum provided no control. Improvement of plant height and vigour with a better yield due to culture filtrate treatment occurred. Root-dip application of antagonistic fungal propagules (T. viride, P. chrysogenum) to tomato seedlings was also very effective in controlling wilt in tomato plants grown inV. albo-atrum infested soil. Dedicated to the memory of the late Prof. Ivor Isaac with whom I had the pleasure of working     

Biocontrol potential of Trichoderma species against mango malformation pathogens

Malformation disease of Mango (Mangifera indica L.) caused by Fusarium moniliforme var. subglutinans is one of the most destructive diseases, which is a major production constraint in the mango-growing regions of India. In this study, The bioagents Trichoderma viride (Tr1), Trichoderma virens (Tr2) and Trichoderma harzianum (Tr3) were evaluated in culture with the pathogens to monitor the antagonistic effect and their volatile compound and culture filtrates (non-volatile compound). It was found that all the three isolates of bioagents significantly checked the growth of F. moniliforme var. subglutinans. In dual culture, the best result was obtained with T. harzianum followed by T. virens and T. viride. A similar result was also observed in the case of culture filtrates ofTrichoderma spp. The results clearly showed that inhibition of the growth of the fusaria isolates by T. harzianum was significantly superior to T. viride andT.virens. In case of antifungal activity of volatile compounds released by Trichoderma isolates, it was also observed that T. virens was more superior to T.harzianum and T. viride.     

Expression of elastinolytic activity among isolates inAspergillus SectionFlavi

A survey of the distribution of elastinolytic potential among 32 culture collection isolates ofAspergillus flavus, A. oryzae, A. parasiticus, A. sojae, A. nomius, andA. tamarii revealed this character to be highly conserved withinAspergillus SectionFlavi. Furthermore, 144 isolates ofA. flavus from environmental samples from six separate regions of the United States produced elastase on solid medium. Most previously described polymorphisms in elastinolytic potential were attributed to the toxicity of borate buffers. Replacement of borate with HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) resulted in detection of elastase production on solid medium by all tested fungal isolates except two that had been in culture over 50 years. In liquid culture, only isolates ofA. flavus, A. tamarii, andA. oryzae accumulated elastase activity. Although isoelectric focusing revealed only one isoform (pI 9.0) of elastase in these culture filtrates, elastinolytic activity in filtrates was partially inhibited by both 1,10-phenanthrolene (2 mM) and phenylmethylsulfonylfluoride (2 mM), suggesting the presence of both metallo and serine elastinolytic proteinases.Abbreviation IEF isoelectric focusing The US Government's right to retain a non-exclusive, royalty-free licence to any copyright is acknowledged.     

Effect of rhizosphere fungi on seed germination

Summary The effect of culture filtrates of some common rhizosphere fungi on the seed germination ofDichanthium annulatum, Bothriochloa pertusa andSetaria glauca was studied. Surface sterilized seeds were soaked in the fungal culture filtrates for 24 hours and after several washings in distilled water transferred to moist filter paper in Petri dishes and the percentage germination recorded. No significant inhibition or stimulation of germination was noticed in any of the treatments. The only exception wasTrichoderma viride which inhibited the germination ofD. annulatum seeds to some extent.     

Two new species ofTrichogramma [Hym.: Trichogrammatidae] from the U.S.A.

Two new species ofTrichogramma from the U.S.A. are described.T. parkeri sp. n., a morphologically distinct species, was reared from an egg ofHeliothis zea (Boddie), whileT. platneri sp. n. which is morphologically indistinguishable fromT. minutum Riley, was reared from eggs ofCydia pomonella (L.). Reproductive incompatibility ofT. platneri andT. minutum is discussed.     

Binding of Host-Associated Treponeme Proteins to Collagens and Laminin: A Possible Mechanism of Spirochetal Adherence to Host Tissues

The polypeptides of seven strains of human treponemes were investigated by immunoblot analysis for their binding to the human placental collagens and laminin. Of the treponemal polypeptides, eleven polypeptides, 45-kDa, 49-kDa, and 62-kDa polypeptides from T. pallidum ATCC 27087, a 48-kDa polypeptide from T. phagedenis biotype Reiter, 51-kDa and 53-kDa polypeptides from T. vincentii ATCC 35580, 30-kDa, 53-kDa and 63-kDa polypeptides from T. socranskii subsp. buccale ATCC 35534, a 52-kDa polypeptide from T. denticola ATCC 35405, and a 53-kDa polypeptide from T. denticola ATCC 33520 possessed an ability to bind to the laminin, type I, III, IV, or V collagen. An intermediate-sized human oral isolate strain G7201 did not possess any laminin- or collagen-binding polypeptides. Immunoelectron microscopy using intact treponemal cells with a single collagen-binding polypeptide and the corresponding antisera demonstrated that the 51-kDa and 53-kDa polypeptides from T. vincentii, the 53-kDa polypeptide from T. socranskii subsp. buccale ATCC 35534 and the 52-kDa polypeptide from T. denticola ATCC 35405, were outer envelope proteins.     

Carbon metabolism and morphogenesis of aspergillus fonsecaeus,II. nutritional status of culture filtrate and dissimilation by mycelial mat

Summary Culture filtrates ofA. fonsecaeus after its growth on glucose nitrate medium were studied for their nutritional status. It was found that culture filtrates support only the vegetative growth. This led to the conclusion that conditions for vegetative growth are different from those of asexual reproduction in this fungus.Efficiency of the fungus in utilization of glucose has been expressed in a ratio and it was found that economic coefficient in the case of this fungus ranges from 30 to 75.As regards the effect of culture filtrate on root elongation of different germinated seeds, it was found that in some cases it is stimulatory and in others it is inhibitory.As regards dissimilation of glucose, it brings about more rapid decomposition of glucose in the replacement culture than in the complete medium and in both cases one of the major metabolic products is oxalic acid.     

Production of pectinases and cellulase by six Pythium species isolated from necrotic strawberry roots

Polygalacturonase and polymethylgalacturonase activity was detected in filtrates ofPythium irregulare, P. perniciosum, P. sylvaticum, andP. dissotocum but not in filtrates ofP. rostratum andP. acanthicum. OnlyP. irregulare, P. perniciosum andP. dissotocum elaborated pectin methyl esterase and in trace amounts. None of the species produced detectable pectintrans-eliminase and macerating enzyme activity.P. sylvaticum was the only species of four tested which produced cellulase.Cooperative Investigations, Agricultural Research Service, United States Department of agriculture and Plant Industries department, School of Agriculture, Southern Illinois University, Carbondale, Ill.     

Absence of a role for lytic microorganisms in the decline of bacteria andSaccharomyces introduced into soil

The populations ofKlebsieila pneumoniae, Escherichia coli, Enterobacter aerogenes, andPseudomonas sp. fell following their addition to soil, but species lysing these gram-negative bacteria were not detected. The numbers ofStaphylococcus aureus andMicrococcus flavus fell by more than four orders of magnitude and ofSaccharomyces cerevisiae by more than two orders after their addition to soil. Organisms lysing these gram-positive bacteria were present in soil, but their numbers did not increase as a result of the additions. Lytic activity againstS. aureus was detected in soil filtrates, but this activity was not enhanced by inoculation of soil with the bacterium. Addition of cycloheximide to soil suspensions delayed the fall in abundance ofM. flavus but did not suppress the lytic populations. We conclude that lysis is not responsible for the decline of bacteria orS. cerevisiae added to soil.     

Test for allelopathic interactions between two marine microalgal species grown in intensive cultures

Both high pH and cell senescence are believed to lead to the production of toxic extracellular metabolites in freshwater microalgae. However, there was no evidence for allelopathic suppression of photosynthesis when filtrates of either of two marine microalgae,Phaeodactylum tricornutum andDunaliella tertiolecta, were mixed with whole cultures of the other species. This was true even when filtrate ofP. tricornutum sample was derived from a culture at high pH or from one in various stages of senescence. It is believed that the major factor leading to the dominance ofP. tricornutum in intensive outdoor cultures is the unique ability of this alga to tolerate pH levels above 9.5, not the allelopathic inhibition of competing species.     

Enhancement of anthocyanin production in callus cultures ofDaucus canota L. under the influence of fungal elicitors

Aqueous extracts of fungal biomass ofAspergillus niger, A. flavus, Penicillium notatum, Fusarium oxysporum and the filtrates of their culture media were analysed for elicitation capability to enhance anthocyanin production in callus cultures ofDaucus carota. The mycelial extract ofA. flavus at the 2.5% level gave maximum elicitation, which resulted in a two-fold increase in anthocyanin with maximal productivity of 23.7% on a dry weight basis. Whereas the media filtrates ofA. flavus induced a 1.25-fold increase in anthocyanin production with a yield of 20.6% on a dry weight basis,P. notatum andF. oxysporum were not as effective asA. flavus orA. niger. The contact time and the concentration required for maximal elicitation of anthocyanin differed with the type of elicitor used. Qualitative analysis of anthocyanins revealed the presence of cyanidin glycosides in control and elicited cultures.     

Antagonistic interactions between fungal pathogens and phylloplane fungi of guava

Dominant phylloplane fungi of guava (Psidium guajava L.) were screened for their antagonistic activities against the two pathogens,Colletotrichum gloeosporioides andPestalotia psidii, bothin vitro andin vivo. Culture filtrates ofAspergillus niger, Fusarium oxysporum andPenicillium citrinum caused more than 50% growth inhibition ofC. gloeosporioides. Filtrates ofCephalosporium roseo-griseum andF. oxysporum were most effective in reducing the growth ofP. psidii. Volatiles produced from the cultures ofA. niger, F. oxysporum, P. citrinum andP. oxalicum inhibited the growth ofC. gloeosporioides, whereas volatiles fromC. roseo-griseum, F. oxysporum andTrichoderma harzianum inhibited the growth ofP. psidii. The inhibitory effect of volatiles decreased with increase in incubation time. In general, the maximum effect of volatiles was noticed after 48 h incubation. Different grades of colony interactions in dual cultures were recognised between the two pathogens and the phylloplane fungi examined. Maximum inhibition ofC. gloeosporioides was caused byAureobasidium pullulans, Cladosporium cladosporioides, epicoccum purpurascens, F. oxysporum andMyrothecium roridum, whereasAspergillus terreus, C. roseo-griseum andP. oxalicum significantly reduced the growth ofP. psidii. Application of a spore suspension of each test fungus inhibited lesion development of guava leaves caused by the test pathogensin vitro. Inhibition was more pronounced when the spore concentration was increased.A. pullulans, C. cladosporioides, E. purpurascens, F. oxysporum, andT. harzianum were found to be strongly antagonistic toC. gloeosporioides. A. niger, A. terreus, C. roseo-grisem andT. harzianum were strongly antagonistic toP. psidii.     

Role of serum in survival ofTreponema pallidum in tissue culture

Summary During attempts to cultivateTreponema pallidum, it was determined that length of time for survival of virulent treponemes was highly dependent on the quality of the fetal bovine serum (FBS) used as a protein supplement in the culture medium. Eighteen lots of commercial FBS were tested for their ability to maintain survival ofT. pallidum in cultures of cottontail rabbit epithelial (SflEp) cells. All were capable of supporting growth of these cells. However, in tests on five of the lots, attachment of treponemes to the SflEp cells was either extremely poor or the 50% survival time (ST₅₀) was less than 5 days. With two of these lots, no treponemes survived for 5 days. By contrast, in tests with 11 of the FBS lots, the ST₅₀ of the treponemes was 12 days or greater; however, there was a great variation in the number of treponemes that attached. Selection of lots of FBS for ultimate experimental use was based on their influence both to extend length of time for survival of treponemes and to increase the number of treponemes that attached to the SflEp cells during that period.     

Inhibition of fungi,actinomycetes and bacteria by Stachybotrys atra

The effect ofStachybotrys atra on fungi, actinomycetes and bacteria was studied in agar culture. Of the 27 genera and 52 species of fungi, 6 species of actinomycetes and 5 genera and 10 species of bacteria tested,S. atra was found to inhibit 95.7 % fungi, all the actinomycetes and 83.3 % of bacteria.Aspergillus ustus, 2 unidentified species ofFusarium, Rhizobium trifolii andPseudomonas spp. were however not inhibited. In experiments in soil,S. atra inhibited the growth ofTrichoderma viride. S atra was ineffective whereT. viride andS. atra were inoculated simultaneously or whereT. viride was added 7 days earlier. Besides,S. atra was not found to controlMacrophomina phaseoli infection on cotton. Soil culture ofS. atra showed phytotoxic effect on cotton seedlings.S. atra produced a dark brown to black thermostable toxic metabolite in the medium. The culture filtrate retarded the growth ofM. phaseoli in agar culture. Despite the importance ofS. atra as a fungistatic agent with wide antimicrobial spectrum, the use ofS. atra as a protectant against plant pathogens seems to be limited because of its phytotoxicity on cotton.