Survey of human pathogenic actinomycetes and fungi in soil from Rome and other Italian areas

As part of a study sponsored by the Ministry of Health of Italy, a research program on pathogenic actinomycetes, keratinophilic and pathogenic fungi in soil was carried out. Two hundred soil samples, collected from different areas of the city of Rome, Calabria, Emilia Romagna, Latium, Apulia, Sardinia, Sicily, Tuscany and Umbria, were examined by several techniques to detect the widest possible variety of pathogenic actinomycetes and fungi. Seven isolates ofNocardia asteroides, four ofActinomadura madurae and one ofNocardiopsis dassonvillei were isolated for the first time from soil in Italy. In addition, numerous isolates ofPetriellidium boydii, Aspergillus fumigatus, A. flavus, A. niger and keratinophilic fungi of the generaMicrosporum, Trichophyton andChrysosporium were also recovered.Presented at the VII Congress of the International Society for Human and Animal Mycology, Jerusalem, Israel, 11–16 March, 1979.     

Insecticidal and nematicidal properties of microbial metabolites

Summary Metabolites from 942 microbial isolates were screened for insecticidal and nematicidal properties. The isolates included 302 streptomycetes, 502 novel actinomycetes including representatives of 18 genera, 28 unidentified aerobic actinomycetes, 70 fungi and 40 bacteria other than actinomycetes. When diluted 10-fold, the metabolites from 55 isolates caused nearly 100% mortality in mosquito larvae (Aedes aegypti) within 24 h. These isolates included 27 isolates ofStreptomyces, four ofActinoplanes, three isolates each ofActinomadura andStreptoverticillium, two isolates each ofMicromonospora, Bacillus andPaecilomyces, and one isolate each ofMicropolyspora, Nocardiopsis, Streptosporangium, Oerskovia, Thermomonospora, Chainia, Pseudomonas, Fusarium, Monilia andSyncephalestrum. Two fungal isolates could not be identified to the generie level. Extracts from the culture broth of 18 isolates caused 100% mortality in mosquito larvae within 15 min to 24 h at a concentration of 1000 ppm. The LC₅₀ of partially purified products from two isolates was 1–2.5 ppm and that of the semipurified preparations from four other isolates was 50 ppm. Valinomycin was identified as an active component in the culture broth from one isolate. The culture broth from 15 isolates of aerobic actinomycetes and 4 of fungi were toxic toPanagrellus redivivus (Nematoda); these included 12 isolates with selective nematicidal properties.Michigan Agriculture Experiment Station, Journal Article No. 12321.     

Plant growth regulatory metabolites from novel actinomycetes

Metabolites from 796 isolates of aerobic actinomycetes were screened for plant growth regulatory properties using an algal bioassay. These included 266 isolates ofStreptomyces, 28 unidentified actinomycetes, and 502 isolates of novel actinomycetes represented by 18 genera. Algal growth inhibition of 30% was observed with 60 isolates, 37 of which belonged to the genusStreptomyces. Among other inhibitors were 8 isolates ofActinomadura, 6 ofActinoplanes, 2 each of the generaThermomonospora, Streptoverticillium, andPromicromonospora, and 3 unidentified. Metabolites from 70 isolates promoted algal growth by 20%. These included 13 isolates ofMicromonospora, 11 ofStreptomyces, 6 ofNocardia, 5 ofActinomadura, and 4 each ofRhodococcus andThermomonospora. Sixteen unidentified isolates; 3 isolates ofPromicromonospora; 2 isolates each ofActinoplanes, Streptosporangium, andOerskovia; and 1 of Thermoactinomyces peptonophilus-like organism andSaccharomonospora viridis also promoted the algal growth by 20%. The plant growth inhibitory properties of 9 actinomycetes and the growth promoting properties of 6 were demonstrable during the secondary screening on higher plants using chemicals extracted from the culture broth. The metabolites fromMicromonospora, Nocardia, Rhodococcus, Streptosporangium, andOerskovia isolates were associated with plant growth promotion only; those fromStreptomyces were most frequently involved with the growth inhibition.This is Michigan Agriculture Experiment Station Journal Article No. 12191.     

The cumulative effect of gamma irradiation on the microbial soil population and its activity

The counts ofAzotobacter, N-fixing clostridia, aerobic cellulose decomposing bacteria, spore formers and actinomycetes were not affected by examined irradiation doses. There were, however, indications of a disturbance in the balance between organisms as evidenced by the rapid increase of actinomycetes and sporeformers during incubation. The effects of various irradiation doses on organic matter decomposition, N-fixing capacity and nitrification power were negligible.     

库木塔格沙漠土壤细菌多样性及具有群体感应抑制活性放线菌筛选

【背景】群体感应抑制剂(quorum sensing inhibitor,QSI)作为抗生素潜在替代品,可有效降低致病菌传染性和毒性。沙漠土壤蕴藏着丰富的放线菌资源,是挖掘群体感应抑制剂的重要来源。【目的】解析库木塔格沙漠土壤细菌群落多样性,筛选并挖掘群体感应抑制活性放线菌资源。【方法】采用Illumina Nova Seq高通量测序技术揭示库木塔格沙漠土壤细菌群落组成,利用可培养方法进行土壤放线菌分离和鉴定;选用紫色杆菌CV026模型筛选群体感应抑制活性放线菌,并对其功能特性进行初步评价。【结果】Illumina Nova Seq高通量测序结果显示,样品土壤细菌涉及23门96目150属,优势菌门为变形菌门(Proteobacteria,61%)、放线菌门(Actinobacteria,28%),其中分枝杆菌属(Mycobacterium)为放线菌门最优势菌属(87.3%),其次为红球菌属(Rhodococcus,6.8%)和丙酸杆菌属(Cutibacterium,0.9%)。可培养方法共分离到108株放线菌,归属9科10属,其中优势菌属为链霉菌属(Streptomyces),占65....     

The influence of germinating seeds and developing roots on re-establishment of micro-organisms in fumigated soil

A laboratory study was made of re-invasion of soils fumigated in closed containers at rates of 3 ml./sq.ft. 10 in. deep with chloropicrin, methyl bromide and a 67:33 mixture of the two materials (MBC–33), using the dilution plate method of determination. Counts after aeration, 3 weeks later, showed that all three fumigants eliminated practically all fungi. Chloropicrin and MBC–33 also killed almost all bacteria and actinomycetes, whereas fumigation with methyl bromide markedly increased numbers of bacteria without materially altering numbers of actinomycetes. Subsequently, in soils fumigated with chloropicrin or MBC–33, fungi recolonized rapidly, but up to 93 % of the colonies isolated were Trichoderma viride. Increase of fungi in soils fumigated with methyl bromide was slower and Penicillium spp. predominated. Bacteria at first declined in numbers, then increased rapidly in soils fumigated with all three materials. In soils fumigated with methyl bromide, actinomycetes, usually considered to include good antagonists, increased but did not exploit their initial advantage over fungi. In soils fumigated with chloropicrin and MBC–33 actinomycetes did not recolonize in appreciable numbers. The greatest variety of fungous genera occurred in isolates from rhizo-spheres of red beet, but the greatest numbers of colonies of both fungi and bacteria were isolated from rhizospheres of pea and pumpkin. None of the crops tested greatly stimulated recolonization by actinomycetes. Compared with the drastic changes brought about by fumigants the influence of seed and root exudates on either the magnitude or the composition of re-invading micro-organisms was slight.     

Characterization of clinical isolates of pathogenic <Emphasis Type="Italic">Nocardia</Emphasis> strains and related actinomycetes in Thailand from 1996 to 2003

In Thailand from 1996 to 2003, 171 strains of pathogenic aerobic actinomycetes from clinical specimens were isolated. Of those strains, 134 were mycolic acid containing actinomycetes, including 96 strains of Nocardia species. Others included 10 strains of Gordonia, 14 strains of Rhodococcus, and 22 strains of Mycobacterium. One strain each of the genera Tsukamurella and Corynebacterium were also isolated. Also identified were 27 strains of non-mycolic acid containing actinomycetes. Our identification studies of 96 strains of Nocardia species showed that significant pathogens in Thailand were N. beijingensis (18 strains), N. cyriacigeorgica (13 strains), and N. farcinica (34 strains); the most prevalent species was N. farcinica (35.4%). We also isolated four strains of N. asiatica, five strains of N. asteroides sensu stricto, four strains of N. nova, seven strains of N. otitidiscaviarum, eight strains of N. transvalensis, and two strains of N. pseudobrasiliensis.     

Quelques champignons pathogènes,isolés du sol roumain,par la méthode de l'inoculation intrapéritonéale à la souris

Resumé En injectant, selon le procédé d'Ajello et al., dans la cavité péritonéale de la souris blanche, une suspension de terre prélevée de divers endroits et en ensemençant après un délai de 6 à 8 semaines des fragments du foie, de la rate, des poumons et des reins de l'animal inoculé, les auteurs ont décelé dans le sol roumain la présence des champignons suivants:Geotrichum candidum, Trichophyton mentagrophytes, Keratinomyces ajelloi (une souche pathogène) etCoccidioides immitis. Selon les auteurs, cette méthode d'isolement est capable de compléter la méthode deVanbreuseghem pour l'isolement des dermatophytes du sol. La présence duCoccidioides immitis dans le sol roumain mérite une mention spéciale, la coccidioidomicose étant, jusqu'en 1961, inconnue en Roumanie.

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Summary By injecting the supernatant from aqueous suspensions of different rumanian soil specimens intraperitoneally into white mice and subsequent culture of their livers and spleens, according to the procedure ofAjello et al., the authors were able to recover the following fungi:Geotrichum candidum, Trichophyton mentagrophytes, Keratinomyces ajelloi (a pathogenic strain) andCoccidioides immitis (two strains). In the author's opinion, this indirect isolation procedure may complete the method developed byVanbreuseghem for isolating dermatophytes from soil. The presence ofCoccidioides immitis in the rumanian soils needs special mention owing to the fact that coccidioidomycosis was unknown in Rumania until 1961.

     

Populations of bacteria and actinomycetes associated with sclerotia ofPhymatotrichum omnivorum buried in Houston black clay

Sclerotia ofPhymatotrichum omnivorum (Shear) Duggar (the causal agent of root rot of cotton) were produced in the laboratory and then buried at a depth of 45 cm at three sites in Texas situated on Houston black clay soils with various cropping histories. The sites included a native grassland prairie, a field in continuous cotton production, and a field in which cotton, corn, and sorghum were grown in rotation. Samples of sclerotia were retrieved monthly over a 12 month period. Populations of bacteria and actinomycetes were enumerated using dilution-plate techniques and isolates were screened (in vitro) for their ability to produce substances inhibitory toP. omnivorum. The sclerotia supported large numbers of bacteria (including fluorescent pseudomonads) and actinomycetes. Numbers associated with sclerotia ranged from 10⁶–10⁹ cells per gram of sclerotia plus adherent soil and were 2–3 orders of magnitude greater than numbers from soil at the same depth but free of sclerotia. Bacteria and actinomycetes antagonistic toP. omnivorum were isolated from sclerotia buried at each of the three sites. Up to 26% of the isolates inhibited growth ofP. omnivorum.     

Selection of antagonistic soil microbes against a few fungal pathogens of piper betle L

Summary Against the fungal pathogens,Phytophthora parasitica, Sclerotium rolfsii, Colletotrichum capsici andGlomerella cingulata responsible for leaf and foot rot of betel vines, 3 fungus, 9 actinomycetes and 4 bacterial antagonists were screened out from 61 fungus, 28 actinomycetes and 4 bacterial organisms isolated from 3 sources of soils. One each from the 3 groups of antagonists when further tested, was found quite effective against the pathogens towards neutral side of H-ion concentration. The antagonists (P. citrinum, Streptomyces sp. and bacterial organism B-7) were also found effective against 18 among 32 isolated fungus organisms from potted soil (collected from betel vine soil) and except one which happened to beP. citrinum, the rest had no adverse effect upon their growth.This work is a part of a scheme supported by Food and Agriculture Council of Pakistan and conducted in Jute Research Institute.     

The differential mortality ofTrifolium repens andPhleum pratense seedlings in relation to temperature

Summary 1. The pathogenic potential of test soils towards germinating seeds ofTrifolium repens andPhleum pratense showed marked variation with time, but no apparent relation to seasonal changes or to the condition of the soil.2. The two species differed in their response to pathogenic factors and the ratioT. repens :P. pratense in the seedlings which emerged showed marked variation.3. Both pre- and post-emergence mortality ofT. repens occurred, whileP. pratense responded to low soil temperature mainly by failure to germinate, through death, and more often enforced dormancy of the seed.4. Chilling the seed in wet soil before germination increased mortality considerably.P. pratense stands were reduced at temperatures below 10°C. Losses ofT. repens were small above 5°C, but at 0°C and below losses were heavier than ofP. pratense.5. In sterilised soil, establishment was consistently better than in corresponding unsterilised soils. Fungicide treatment of the seed in unsterilised soil resulted in improvement of stands in most cases, but less than soil sterilisation.6. The improvement in emergence due to soil sterilisation or fungicide treatment was much the same after all pre-chilling treatments.7. The relationship of germination behaviour and liability to pathogenic attack under different temperature conditions is discussed in the light of experience in the field.     

The occurrence of antagonistic actinomycetes in bulgarian soils and their antibiotic properties

Of 1448 actinomycete strains isolated from different types of soils 46% exhibited an antibiotic activity. Strains exhibiting a narrow antibiotic spectrum were more usual than those exhibiting a broad spectrum. An antifungal effect was the most common property. Strains exhibiting a considerable activity against pathogenic, phytopathogenic and dermatophytic microorganisms were found among isolated cultures. Fifty-three antagonistic actinomycetes were classified in 29 species.Actinomyces candidus, Actinomyces flaveolus, Actinomyces flavoviridis andActinomyces griseovariabilis were the most common. The antibiotic spectrum of individual strains belonging to the same species was qualitatively different in most cases.     

Studies on strains of Microsporon gypseum isolated from soil

Summary 180 samples collected from different types of soils were examined for presence of dermatophytes.17 strains ofM. gypseum isolated from these samples were studied for their morphology, perfect stage formation and animal pathogenicity. Two isolates could be induced to produce perfect stage,Nannizia incurvata and only one isolate was pathogenic to guinea pig.     

Distribution of pathogenic vibrios and other bacteria in imported frozen shrimps and their decontamination by gamma-irradiation

The distribution of pathogenic vibrios and other bacteria in eight samples of imported frozen shrimps and the effect of irradiation on these bacteria were investigated. Total aerobic bacteria were at 2×10⁴ to 4×10⁶/g. Coliforms consisted mainly ofEnterobacter. No salmonella were detected. A total of 66 isolates, includingVibrio parahaemolyticus, V. mimicus, V. alginolyticus, V. vulnificus, V. fluvialis and a few ofListeria monocytogenes, were obtained. The gamma-radiation dose needed to reduce by 10^–4 the number of vibrio isolates andAeromonas hydrophila was about 3 kGy in frozen shrimps, whereas about 3.5 kGy was required forL. monocytogenes.     

西沙石珊瑚共附生放线菌的分离培养及抗菌活性评价

【背景】珊瑚礁生态系统是海洋中一类极其重要的生态系统,健康珊瑚礁中丰富的共附生放线菌群体是珊瑚抵御各种致病菌的重要防线,因此,这类放线菌是寻找抗菌活性分子的重要资源,其药用潜力巨大。【目的】从西沙石珊瑚样品中分离共附生放线菌,并从中筛选具有良好抗菌活性的菌株。【方法】通过稀释涂布法分离珊瑚共附生放线菌,并根据16S rRNA基因序列构建系统发育树进行菌种鉴定;通过平板对峙法对放线菌进行抗菌活性筛选并确定目标菌株;将目标菌株涂布于不同氯化钠浓度的ISP2固体培养基上培养,测试其盐度耐受能力;通过平板对峙法对该菌株发酵产物的热稳定性和光稳定性进行测试;采用NanoPore和Illumina方法完成目标活性放线菌全基因组测序,并通过antiSMASH在线分析预测其次级代谢产物生物合成基因簇及其结构类型。【结果】从6份西沙石珊瑚样品中分离得到104株可培养放线菌,根据菌落形态和分离来源去重后对其中27株放线菌进行16S rRNA基因序列测序,通过序列比对和系统发育树分析将菌株初步鉴定为盐孢菌属(Salinispora)(25株)、链霉菌属(Streptomyces)(1株)和戈登菌属(Gord...     

Induction of competence in nonencapsulated and encapsulated strains ofHaemophilus influenzae

A simple procedure for induction of competence in nonencapsulated and encapsulated strains ofHaemophilus influenzae is described, which consists of growing cells without shaking in brain-heart infusion broth under aerobic conditions. Competence emerged at the end of the exponential phase and reached a peak at the stationary phase. InH. influenzae Rd competence was maintained for at least 6 h at 37°C, whereas in two encapsulated clinical isolates ofH. influenzae type b a decrease in competence was observed after 4 h. Competence was maintained for 24 h at 22°C and 4°C as well as by freezing the cells in 15% glycerol and storing them at –70°C. Transformation frequencies of three chromosomal markers—streptomycin, nalidixic acid, and erythromycin resistance—were 0.5% to 1% inH. influenzae Rd and about tenfold lower in the two encapsulated clinical isolates ofH. influenzae type b. The advantage of this procedure is that it is simpler than the previously described procedures and yields stable, highly transformable cells. Unlike the standard M IV method, the static aerobic procedure does not interfere with the capsule synthesis and can be used for testing transforming activity of encapsulated virulent isolates ofH. influenzae.     

A modified sucrose fractionation procedure for the isolation of frankiae from actinorhizal root nodules and soil samples

Summary The isolation and pure culture of the symbiotic nitrogen-fixing frankiae has always been difficult. In the past the isolation of these actinomycetes directly from soil samples has proven impossible and isolations from root nodules of many genera has been only poorly successful. We report here a modified sucrose fractionation procedure which increased the success of isolations from root nodules and which permitted the isolation ofFrankia directly from soil samples. Crushed nodule suspensions or soil suspensions were incubated briefly in 0.7% phenol (carbolic acid) just before application to a sucrose density gradient. This phenol incubation decreased the number of contaminating eubacteria and fungi but more importantly increased the number ofFrankia developing on the isolation plates. If the phenol incubation was used solely without sucrose fractionation noFrankia were isolated, suggesting the death of the organisms due to phenol toxicity. The use of selective nitrogen-deficient media proved important for the isolation of frankiae from soils.     

Selective isolation,evaluation and characterization of antagonistic actinomycetes against <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

The baiting bag method was found to be useful for isolating antagonistic actinomycetes from terrestrial habitat. Out of total 110 actinomycetes isolated from rhizospheric and non-rhizospheric soil of Indo Gangetic Plains (IGP) of India, 9 isolates exhibited aggressive antagonism against Rhizoctonia solani, screened through dual culture, well diffusion and sealed plate technique. Maximum growth inhibition was recorded up to 50% under well diffusion (S. toxytricini vh22) and 52.6% in a direct confrontation (Actinomycetales bacterium vh41). Whereas maximum disease suppression (53.33%) under green house condition was achieved on seedling treated with S. tricolor vh85. Scanning electron microscopy of antagonists and pathogen interaction exhibited pore formation and hyphal degradation of test pathogen. Physiological and molecular characterization of selected isolates showed wide diversity and uncommon species has been encountered through the selective isolation technique.     

Efficiency of various selective media in determining Pythium population in soil

Of 15 selective media recommended for isolation and enumeration ofPythium spp. directly from soil, corn meal agar (CMA) supplemented with agar, sucrose, minor elements, thiamine, rose bengal, pimaricin, pentachloronitrobenzene and vancomycin (MPVM) was the most efficient. Streptomycin (30–50 ppm) and rose bengal (33–60 ppm) as used in certain tested media effectively suppressed development of bacteria and actinomycetes. However, these chemicals adversely affected germination of spores and mycelial growth and thereby the recovery ofPythium spp. from soil. Media containing pimaricin (5 to 100 ppm) were more effective than those with nystatin (40 ppm) in suppressing development of nonphycomycetous fungi on isolation plate. MPVM with pimaricin at 5 ppm was more efficient than that with 10 ppm of the antibiotic in recoveryingPythium from soil. However, there was no difference in recovery ofPythium by this medium containing rose bengal at 5 ppm or at 10 ppm, butPythium colonies were more dense and better delineated when the medium contained 10 ppm of rose bengal. CMA containing pimaricin (5–100 ppm) and vancomycin (200 ppm) permitted occasionally development of a large number ofMortierella and bacterial colonies from certain soils, that interfered with accurate determination of colonies of certainPythium spp. on the plates. Vancomycin at 300 ppm, as used in MPVM, substantially reduced development of bacterial colonies compared to 200 ppm of the antibiotic. Surface-soil dilution-plate was more effective than the soil-dilution-plate method in reducing bacteria andMortierella colonies on isolation plates without affecting recovery ofPythium. The importance of basal medium, complement of antimicrobial agents, and isolation methods for efficiency of selective medium in recovery ofPythium spp. directly from soil is discussed.     

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.