Microbial production and degradation of γ-aminobutyric acid (GABA) in the abalone larval settlement habitat

Abstract Short-term fluctuations in the number of culturable microorganisms and the composition of the bacterial community colonising the phyllosphere of field grown sugar beet were determined at 6 h intervals over a 42 h period. The number of culturable bacteria and Erwinia were found to fluctuate diurnally, increasing in size overnight and declining during daylight hours. In contrast, the number of Arthrobacter and yeasts were constant throughout the study, even after rainfall. Over most of the study the number of filamentous fungi and pseudomonads decreased. The short-term fluctuations in bacterial numbers were similar in magnitude to those detected over most of the previous season. No significant variation was observed in microbial numbers between samples taken from three locations within the field. Fatty acid analysis of isolated bacteria revealed the community to be similar in composition to that detected during the same period the previous year and indicated no significant short-term temporal or spatial variation. To limit the effects of diurnal fluctuations on the number of culturable bacteria detected, it is recommended that in long-term studies, samples be collected at the same time of day on each occasion.     

Dehydrated gelatin drops: a good method for fungi maintenance and preservation

Some techniques have been proposed to maintain fungi culture collection. However, any choice must ensure the cultural stability and its phenotypic characteristics. This work proposes an adaptation of a preservation method considered by few literature reports: the dehydrated gelatin drops method (DGD). A total of 27 strains of fungi of clinical interest, including four dermatophyte fungi isolates, six filamentous non-dermatophyte fungi, five environment isolated filamentous fungi, six dimorphic fungi and six yeasts were maintained by this method for a seven year period at room temperature. After that time, the macro and micro characteristics of each fungus were studied, allowing the evaluation of the DGD method. In our experience, none of the strains maintained by DGD were found to be contaminated by bacteria or other fungi and no apparent changes were observed in morphology or macroscopic features.     

Yeasts and filamentous fungi carried by the gynes of leaf-cutting ants

Insect-associated microbes exhibit a wide range of interactions with their hosts. One example of such interactions is the insect-driven dispersal of microorganisms, which plays an essential role in the ecology of several microbes. To study dispersal of microorganisms by leaf-cutting ants (Formicidae: Attini), we applied culture-dependent methods to identify the filamentous fungi and yeasts found in two different body parts of leaf-cutting ant gynes: the exoskeleton and the infrabuccal pocket. The gynes use the latter structure to store a pellet of the ants’ symbiotic fungus during nest founding. Many filamentous fungi (n = 142) and yeasts (n = 19) were isolated from the gynes’ exoskeleton. In contrast, only seven filamentous fungi and three yeasts isolates were recovered from the infrabuccal pellets, suggesting an efficient mechanism utilized by the gynes to prevent contamination of the symbiotic fungus inoculum. The genus Cladosporium prevailed (78%) among filamentous fungi whereas Aureobasidium, Candida and Cryptococcus prevailed among yeasts associated with gynes. Interestingly, Escovopsis, a specialized fungal pathogen of the leaf-cutting ant-fungus symbiosis, was not isolated from the body parts or from infrabuccal pellets of any gynes sampled. Our results suggest that gynes of the leaf-cutter ants Atta laevigata and A. capiguara do not vertically transmit any particular species of yeasts or filamentous fungi during the foundation of a new nest. Instead, fungi found in association with gynes have a cosmopolitan distribution, suggesting they are probably acquired from the environment and passively dispersed during nest foundation. The possible role of these fungi for the attine ant–microbial symbiosis is discussed.     

In Vitro Interactions Between Yeasts and Bacteria and the Fungal Symbionts of the Mountain Pine Beetle (Dendroctonus ponderosae)

Multi-trophic interactions between prokaryotes, unicellular eukaryotes, and ecologically intertwined metazoans are presumably common in nature, yet rarely described. The mountain pine beetle, Dendroctonus ponderosae, is associated with two filamentous fungi, Grosmannia clavigera and Ophiostoma montium. Other microbes, including yeasts and bacteria, are also present in the phloem, but it is not known whether they interact with the symbiotic fungi or the host beetle. To test whether such interactions occur, we performed a suite of in vitro assays. Overall, relative yield of O. montium grown with microbes isolated from larval galleries was significantly greater than when the fungus was grown alone. Conversely, the yield of G. clavigera grown with these same microbes was less than or equal to when it was grown alone, suggesting that O. montium, and at least some microbes in larval galleries, have a mutualistic or commensal relationship, while G. clavigera and those same microbes have an antagonistic relationship. A bacterium isolated from phloem not colonized by beetles was found to inhibit growth of both G. clavigera and O. montium and appears to be an antagonist to both fungi. Our results suggest that bacteria and yeasts likely influence the distribution of mycangial fungi in the host tree, which, in turn, may affect the fitness of D. ponderosae.     

Williopsis californica, Williopsis saturnus, and Pachysolen tannophilus: novel microorganisms for stereoselective oxidation of secondary alcohols

A screening of 416 microorganisms from different taxonomical groups (bacteria, actinomycetes, yeasts, and filamentous fungi) has been performed looking for active strains in the stereoselective oxidation of secondary alcohols. The working collection was composed of 71 bacterial strains, 45 actinomycetes, 59 yeasts, 60 basidiomycetes, 33 marine fungi, and 148 filamentous fungi. All microorganisms selected were mesophilic. Yeasts were the most active microbial group in the whole-cell-catalyzed oxidation. Williopsis californica, Williopsis saturnus, and Pachysolen tannophilus were the strains of greatest interest, both as growing cells and as resting cells. The oxidation of the alcohols takes place when cells are in the stationary growth phase (after 48 h of culture). These three strains are S-stereoselective for the oxidation of racemic secondary alkanols and show stereospecificity in the oxidation of menthol or neo-menthol, whereas iso-menthol is not oxidized. In the case of the 1-tetrahydronaphtol enantiomers, only the S-enantiomer is oxidized. The three strains were immobilized by entrapment using agarose and agar from algae of the Gracilaria genus. The agarose derivatives displayed significant improvement in the stereospecificity of the reactions.     

Quantitative and qualitative studies of the microflora of barley malt production

Populations of aerobic heterotrophic bacteria, mycelial fungi and yeasts occurring in malting barley were estimated by a plate technique and scanning electron microscopy. There was an increase in the total number of micro-organisms during germination, although populations declined after kilning. Bacteria dominated on all samples, with progressively lower populations of yeasts and filamentous fungi. There was no obvious spatial distribution of micro-organisms on the samples although there appeared to be high populations of bacteria and fungal hyphae on the inner surface of the kernels. The dominant groups of aerobic heterotrophic bacteria were presumptively identified as Alcaligenes sp., Arthrobacter globiformis, Clavibacter iranicum, Erwinia herbicola, Lactobacillus spp. and Pseudomonas fluorescens. The principal filamentous fungi were identified as Aiternaria alternata, Aspergillus glaucus (group), Cladosporium macrocarpum, Epicoccum purpurascens, Fusarium avenaceum, Geotrichum candidum and Penicillium spp. The yeasts isolated most frequently were Candida catenulata, C. vini, Debaryomyces hansenii, Hansenula polymorpha, Kloeckera apiculata, Rhodotorula mucilaginosa, Sporobolomyces roseus and Trichosporon beigelii.     

福建和溪亚热带雨林地表微生物的数量动态

分枯枝落叶层、腐解层和表土层（0—20cm深）三个层次研究福建和溪亚热带雨林的细菌、真菌和放线菌三大微生物类群的数量和月变化。结果表明：三个层次的微生物数量均以细菌最多,次为真菌,后为放线菌。比较三个层次,各微生物类群在各月份都是枯枝落叶层和腐解层数量较多。同一微生物类群在三个层次的数量均有明显而相似的季节变化;不同类群变化规律不尽相同,但基本只有一个高峰。细菌的高峰值在4月份,真菌约滞后一个月在5月份,放线菌则在8月份才出现高峰。各层次的细菌数和真菌数与月凋落物量、月降雨量和月平均气温各有显著的线性正相关（p＜0．01或0．05）,而放线菌则仅与月平均气温显著相关。     

Nine-year microflora study of an isolator-maintained immunodeficient child

A male child, maintained in a controlled environment, was monitored each month for bacteria, yeasts, and filamentous fungi recovered from the mouth, nasal passages, feces, and nine body surface sites. Three natural microbial categories became apparent. Incident microorganisms were recovered from within the isolator but did not establish permanent residence. Of the 53 incident types isolated, 20 were filamentous fungi and 4 were yeasts. Some genera, such as Fusobacterium, Lactobacillus, Neisseria, and Rothia, which were commonly found in the reference group, did not become permanent inhabitants. Transient microorganisms were repeatedly recovered but could not be demonstrated within the isolated environment at the end of the study. The loss of only a few of the 19 transient species could be associated with antimicrobial therapy. Permanent microorganisms consisted of Pencillium citrinum and 17 bacterial types, of which alpha-hemolytic streptococci, Staphylococcus edpidermidis subgroups II and V, Micrococcus groups 1 and 2, Clostridium bifermentans, and Propionibacterium acnes were recovered throughout the entire 9 years of the study. The number of CFUs recovered from each sample type was generally not unlike that from the reference group of healthy male adults. Also, the number of different aerobic species recovered from the feces was within the normal range of that of the reference group. In contrast, the number of different species recovered from all other samples was less than that commonly found in the reference group.     

A comparative analysis of microflora during biofilm development on grape seeds exposed to methanol in a biofilter

The attachment of microorganisms onto biotic surfaces to form biofilm structures on the support media of a biofilter has great impact on biodegradation systems. This study examined the composition of the microbial community that developed on grape seeds (GS) used as support media in methanol degradation biofilters. They were analyzed using conventional microbiology techniques and API galleries. Analysis of microbial counts showed that, in GS before methanol exposure, bacteria and filamentous fungi were predominant over yeasts. In contrast, GS exposed to methanol exhibited more bacteria and yeasts than fungi. Most of the Gram-negative bacteria were the Pseudomonas genus, Bacillus staerothermophilus, Bacillus amyloliquefaciens, and Bacillus pumilus. Rhodotorula mucilaginosa was the primary yeast found. The filamentous fungi Aspergillus sp. Cladosporium cladosporioides, Fusarium sp., and Alternaria sp. were also detected. No Gram-positive bacteria growth was found on GS exposed to methanol. Using scanning electron microscopy, biofilm formation on the GS was examined to reveal the presence of both prokaryotic and eukaryotic microorganisms as biomass accumulation was visible on the seeds. Seeds exposed to methanol for 90 days showed a mature biofilm with cuticle and epidermal layer decline, as well as biofilm dissolution into grape seed integuments.     

Nine-year microflora study of an isolator-maintained immunodeficient child.

A male child, maintained in a controlled environment, was monitored each month for bacteria, yeasts, and filamentous fungi recovered from the mouth, nasal passages, feces, and nine body surface sites. Three natural microbial categories became apparent. Incident microorganisms were recovered from within the isolator but did not establish permanent residence. Of the 53 incident types isolated, 20 were filamentous fungi and 4 were yeasts. Some genera, such as Fusobacterium, Lactobacillus, Neisseria, and Rothia, which were commonly found in the reference group, did not become permanent inhabitants. Transient microorganisms were repeatedly recovered but could not be demonstrated within the isolated environment at the end of the study. The loss of only a few of the 19 transient species could be associated with antimicrobial therapy. Permanent microorganisms consisted of Pencillium citrinum and 17 bacterial types, of which alpha-hemolytic streptococci, Staphylococcus edpidermidis subgroups II and V, Micrococcus groups 1 and 2, Clostridium bifermentans, and Propionibacterium acnes were recovered throughout the entire 9 years of the study. The number of CFUs recovered from each sample type was generally not unlike that from the reference group of healthy male adults. Also, the number of different aerobic species recovered from the feces was within the normal range of that of the reference group. In contrast, the number of different species recovered from all other samples was less than that commonly found in the reference group.     

Yeasts and fungi in excrements of the sand hill snail,Theba pisana

Eight microbial species were isolated and identified from excrements of the sand hill snail,Theba pisana, viz. 3 filamentous fungi, 3 basidiomycetous yeasts and 2 yeast-like ascomycetous fungi.     

The challenge for identifying the fungi living inside mushrooms: the case of truffle inhabiting mycelia

The Tuber ascomata (truffles) are a microhabitat for bacteria, viruses, and fungi (yeasts and filamentous fungi). In this survey, we tried to develop a method that would make it possible to define the mycobiome of the truffle-inhabiting filamentous fungi using culture independent molecular methods. The nested quantitative Real-Time PCR allowed us to demonstrate that each truffle is home to multiple species of filamentous fungi and that their DNA is present within the healthy ascoma at the ratio of 10^?6 compared to that of the truffle. Probably due to their insignificant presence, Denaturing Gradient Gel Electrophoresis of the amplification of ITS amplicons showed only those of the host. Based on the results, the possibilities of being able to detect the fungicolous fungi present in very small amounts within a fungal host are discussed.     

The influence of bacterial inoculants on the microbial ecology of aerobic spoilage of barley silage.

The aerobic decomposition of barley silage treated with two inoculants (LacA and LacB) containing mixtures of Lactobacillus plantarum and Enterococcus faecium was investigated over a 28-day period. Initially, yeast and bacterial populations were larger in silage inoculated with LacA than in silage treated with LacB or water alone (control). Differences in the succession of yeasts in silage treated with LacA were observed relative to the other two treatments. From silage treatment with LacA, Issatchenkia orientalis was the most prevalent yeast taxon over all of the sample times, and the filamentous fungus Microascus brevicaulis was also frequently isolated at later sample dates (> or = 14 days). In contrast, Saccharomyces exiguus was the most prominent yeast recovered from silage treated with LacB and water alone on days 2 and 4, although it was supplanted by I. orientalis at later sample times. Successional trends of bacteria were similar for all three treatments. Lactobacillus spp. were initially the most prevalent bacteria isolated, followed by Bacillus spp. (primarily Bacillus pumilus). However, the onset of Bacillus spp. prominence was faster in LacA silage, and Klebsiella planticola was frequently recovered at later sample times (> or = 14 days). More filamentous fungi were recovered from LacA silage on media containing carboxylmethylcellulose, pectin, or xylan. The most commonly isolated taxa were Absidia sp., Aspergillus flavus, Aspergillus fumigatus, Byssochlamys nivea, Monascus ruber, Penicillium brevicompactum, Pseudoallescheria boydii, and M. brevicaulis. The results of this study indicated that the two bacterial inoculants incorporated into barley at the time of ensilage affected the microbial ecology of silage decomposition following exposure to air. However, neither of the microbial inoculants effectively delayed aerobic spoilage of barley silage, and the rate of decomposition of silage treated with one of the inoculants (LacA) was actually enhanced.     

Characterization and analysis of antibiotic activity of some aquatic actinomycetes

Nine different isolates of aquatic actinomycetes identified as Streptomyces spp. were studied for their morphological and cultural characteristics. One of these isolates (C4-S, Streptomyces violaceusniger) was extensively studied for its inhibitory effect against a wide range of Gram-positive, Gram-negative bacteria, Mycobacterium vaccae ATCC 29678, Candida albicans and several food associated filamentous fungi and yeasts. Most of these were characterized by flexous sporophore morphology and their inability to produce cultural pigments. Bioassay results indicated that S. violaceusniger of 10 days culture age was highly active against Gram-positive cocci and bacilli with an inhibition zone of 16-25 mm, and slightly active against M. vaccae ATCC 29678 with an inhibition zone of 5-10 mm. The inhibitory effect was slight against Escherichia coli, Aspergillus niger 1 and C. albicans with an inhibition zone of 8-10 mm for each of them. There was no inhibitory effect of S. violaceusniger against other Gram-negative bacteria, filamentous fungi and yeast. The nature of the active molecule produced by S. violaceusniger showed a maximum absorption in the UV region at 210-260 nm.     

天山云杉天然林不同林层的空间格局和空间关联性

天山云杉(Picea Schrenkiana)林是天山林区的重要针叶林。研究调查了分别代表天山云杉增长型种群和成熟型种群的两块标准地,应用Ripley's K系数法研究了天山云杉更新层、低林层、中林层和高林层的空间格局和不同林层的空间关联性,并应用Moran's I系数法研究了不同林层的林木高度的空间自相关性。结果表明:所有林层的天山云杉活立木都是显著聚集的,只是发生聚集格局的强度和聚集尺度不同。更新层天山云杉具有最大的聚集强度,从更新层到低林层,聚集强度急速降低。两块标准地中天山桦(Betula tianschanica Rupr.)、天山柳(Salix tianschanica Rgl.)和天山云杉活立木的空间关联性的表现不同。研究结果表明:增长型种群中天山桦、天山柳在所有尺度上都抑制天山云杉的生长;而成熟型种群中天山桦、天山柳在1-10 m尺度内对天山云杉种群的生长没有影响,10-40 m表现出正相关,40 m以上没有表现出明显的关联性;同时天山桦、天山柳基本上在所有尺度都抑制天山云杉的更新。在更新层和低林层间表现出正相关;而更新层和枯死木之间的空间关联性为正相关。     

Microbial characterization during composting of municipal solid waste

This study investigates the prevailing physico-chemical conditions and microbial community; mesophilic bacteria, yeasts and filamentous fungi, bacterial spores, Salmonella and Shigella as well as faecal indicator bacteria: total coliforms, faecal coliforms and faecal Streptococci, present in a compost of municipal solid waste. Investigations were conducted in a semi-industrial pilot plant using a moderate aeration during the composting process. Our results showed that: (i) auto-sterilization induced by relatively high temperatures (60–55°C) caused a significant change in bacterial communities. For instance, Escherichia coli and faecal Streptococci populations decreased, respectively, from 2×10⁷ to 3.1×10³ and 10⁷ to 1.5×10³ cells/g waste dry weight (WDW); yeasts and filamentous fungi decreased from 4.5×10⁶ to 2.6×10³ cells/g WDW and mesophilic bacteria were reduced from 5.8×10⁹ to 1.8×10⁷ bacteria/g WDW. On the other hand, the number of bacterial spores increased at the beginning of the composting process, but after the third week their number decreased notably; (ii) Salmonella disappeared completely from compost by the 25th day as soon as the temperature reached 60°C; and (iii) the bacterial population increased gradually during the cooling phase. While Staphylococci seemed to be the dominant bacteria during the mesophilic phase and at the beginning of the thermophilic phase, bacilli predominated during the remainder of the composting cycle. The appearance of gram-negative rods (opportunistic pathogens) during the cooling phase may represent a serious risk for the sanitary quality of the finished product intended for agronomic reuse. Compost sonication for about 3 min induced the inactivation of delicate bacteria, in particular gram-negatives. By contrast, gram-positive bacteria, especially micrococcus, spores of bacilli, and fungal propagules survived, and reached high concentrations in the compost.     

Cultivable Hydrocarbonoclastic Microbial Community from Historically Polluted Soil: Tests for Consortium Development

Contamination of soil by petroleum products is a global concern. Microbial-based bioremediation strategies could be cost-effective alternatives to other remediation methods; however, they should be site-tailored to guarantee effectiveness and success. The present paper illustrates data on autochthonous microbial communities in a hydrocarbon pollute soil, considering not only bacteria, but also yeasts and filamentous fungi as strains with pollutant degradation capability. The interesting strains were identified as members of the genus Pseudomonas, Rhodotorula, Yarrowia, Penicillium and Aspergillus. Preliminary tests for consortium assemblage were carried out to evaluate the selected strains as possible inoculum in biotreatment via autochthonous bioaugmentation (ABA).     

Effects of Heavy Metal Pollution on Oak Leaf Microorganisms

During the growing season, comparisons were made of the leaf surface microflora of (i) two groups of mature oak trees, one in the vicinity of a smelting complex contaminated by heavy metals and the other at a relatively uncontaminated site, and (ii) two groups of oak saplings at the uncontaminated site, one of which was sprayed with zinc, lead, and cadmium to simulate the heavy metal pollution from the smelter without the complicating effects of other pollutants. Total viable counts of bacteria, yeasts, and filamentous fungi (isolated by leaf washing) were generally little affected by the spraying treatment, whereas polluted leaves of mature trees supported fewer bacteria compared with leaves of mature trees at the uncontaminated site. Numbers of pigmented yeasts were lower on polluted oaks and on metal-dosed saplings compared with their respective controls. Polluted leaves of mature trees supported both greater numbers of Aureobasidium pullulans and Cladosporium spp. and a greater percentage of metal-tolerant fungi compared with oak leaves at the uncontaminated site. There were no significant overall differences in the degree of mycelial growth between the two groups of saplings or the mature trees.     

Assay for Lipolytic and Proteolytic Activity Using Marine Substrates

Nondestructive assay procedures for determining microbial lipolytic and proteolytic activity on marine substrates were developed and tested with 287 isolates of bacteria, filamentous fungi, and yeasts. A definite substrate specificity was noted when the enzymatic activities on marine and nonmarine substrates was compared. Of 170 lipolytic isolates, 14 were only active on menhaden oil, 11 could hydrolyze menhaden oil and Tween 80 and/or tributyrin, and 145 isolates could only hydrolyze one or both of the nonmarine lipids. Of the 198 proteolytic isolates, 10 were specific for codfish extract, 152 were active against the marine substrate plus casein and/or gelatin, and 36 were specific for nonmarine substrates.     

The occurrence of fungi in hospital and community potable waters

The prevalence of fungi was investigated in 126 potable water samples (84 hospital and 42 community samples), in parallel with the standard pollution indicator micro-organisms. Filamentous fungi were isolated from 104 of 126 (82.5%) samples and yeasts from 14 (11.1%), whereas their mean counts were 36.6 and 4.4, respectively. Fungi were isolated from 95.2% of community and 76.2% of hospital water samples, with the difference being statistically significant (P < 0.05), while yeasts were isolated from 9.5 and 11.9%, respectively. Prevailing genera were Penicillium spp., isolated from 64, Aspergillus spp., from 53, and Candida, from nine of the examined samples. Colony-forming units of yeasts were significantly correlated with those of total and faecal coliforms, whereas the counts of filamentous fungi were significantly correlated with total heterotrophic bacteria counts. These results suggest that tap water is a potential transmission route for fungi both in hospitals and the community in the examined region and may pose a health hazard mainly for the immunocompromised host.