Glucose Yeast Extract Agar Research Articles

Vishniacozyma pseudocarnescens sp. nov., a new anamorphic tremellomycetous yeast species.

Three strains belonging to the basidiomycetous yeast genus Vishniacozyma were isolated from marine water samples collected from intertidal zones in Liaoning province, northeast China. Phylogenetic analyses based on the sequences of the small subunit (SSU) ribosomal DNA (rDNA), the D1/D2 domain of the large subunit (LSU) ribosomal DNA (rDNA), the internal transcribed spacer region (ITS), the two subunits of DNA polymerase II (RPB1 and RPB2), the translation elongation factor 1-α (TEF1), and the mitochondrial gene cytochrome b (CYTB) showed that these strains together with 20 strains from various geographic and ecological origins from other regions of the world represent a novel species in the genus Vishniacozyma. We propose the name Vishniacozyma pseudocarnescens sp. nov. (holotype CGMCC 2.6457) for the new species, which differs phenotypically from its close relatives V. carnescens, V. tephrensis, and V. victoriae by its ability to grow at 30 °C and on 50 % (w/v) glucose-yeast extract agar.

Plant Growth-promoting Ability and Pathogen Inhibitory Effect of Actinomycetes Isolated from Fecal Pellets of the Giant Millipede Thyropygus resimus (Diplopoda)

The microbial properties of millipede fecal pellets have been studied mainly in Glomerida (pill millipedes), and much less in the significant majority of other millipede groups. Therefore, the present study examined actinomycetes isolated from the fecal pellets of the non-glomerid giant millipede Thyropygus resimus Attems, 1938 (Spirostreptida) to (1) test their plant growth-promoting ability, and (2) evaluate their potential to control and inhibit plant pathogenic microorganisms. Millipedes were collected from Phu Kum Khao, Kalasin Province, Thailand. A total of 59 actinomycete isolates were obtained and identified as belonging to the genus Streptomyces using 16S rRNA sequencing. The plant growth-promoting properties of the isolates were tested by screening four characteristics: nitrogen fixation, phosphate solubility, siderophore production, and indole-3-acetic acid (IAA) production. A nitrogen-fixation test on nitrogen-free solid malate media (NFM) showed that 54 isolates were capable of fixing nitrogen. Phosphate solubility was tested on double-layered glucose yeast extract agar (GYA) medium containing tricalcium phosphate. This showed that 42 isolates formed a clear zone around the colonies due to phosphate dissolution. Siderophore production was tested on chrome azurol sulfate (CAS) agar. This showed that 55 isolates could grow on this medium and form clear yellow to orange zones around their colonies. IAA production tests revealed that 41 isolates could produce IAA. Based on the combined results of these four tests, eight of the 59 isolates were the most effective in promoting plant growth: KLS-AC04, KLD-AC01, KLD-AC02-1, KLD-AC08, KLD-AC09, KLD-AC16, KLD-AC29-1, and KLD-AC30. Seventeen isolates inhibited the growth of Xanthomonas oryzae pv. oryzae, the causal agent of bacterial leaf blight disease in rice, more effectively than rifampicin (100 ppm), with isolate KLS-AC02-1-1 being the most effective (inhibition zone, 58.25 mm in diameter). Therefore, these isolates can be used for growth promotion and rice disease control in the future.

Toxicogenic fungal profile, Ochratoxin A exposure and cancer risk characterization through maize (Zea mays) consumed by different age populations in the Volta region of Ghana

Maize (Zea mays) is an important staple food crop for the majority of Ghanaians. Maize is mostly contaminated by fungal species and particularly mycotoxins. This work aimed to identify and quantify the incidence of fungal infection and exposure to Ochratoxin A (OTA) as well as the health risk characterization in different age populations due to maize consumption in the Volta region. Maize samples were plated on Dichloran Rose Bengal Chloramphenicol (DRBC) agar, and Oxytetracycline Glucose Yeast Extract (OGYE) agar. All media were prepared in accordance with the manufacturers' instructions. The plates were incubated at 28±2°C for 5-7 days. High-Performance Liquid Chromatography connected to a fluorescence detector (HPLC-FLD) was used to analyze the ochratoxin A (OTA) levels in maize. Cancer risk assessments were also conducted using models prescribed by the Joint FAO/WHO Expert Committee on Additives (JECFA). The maize samples collected from the Volta region contained fungal population between the range of 3.08-4.58 log10CFU/g. Eight (8) genera were recorded belonging to Aspergillus, Trichoderma, Penicillium, Fusarium, Saccharomyces, Mucor, Rhodotorula and Rhizopus. The species diversity includes A. flavus, A. niger, T. harzianum, P. verrucosum, F. oxysporum, Yeast, F. verticillioides, Rhodotorulla sp, A. fumigatus, R. stolonifer, M. racemosus species. Additionally, the ochratoxins level contained in the samples were very noteworthy and ranged from 1.22 to 28.17 μg/kg. Cancer risk assessments of OTA produced outcomes also ranged between 2.15 and 524.54ng/kgbw/day, 0.03-8.31, 0.0323, and 0.07-16.94 for cases/100,000 person/yr for Estimated Daily Intake (EDI), Margin of Exposure (MOE), Average Potency, and Cancer Risks respectively for all age categories investigated. There was very high mycoflora load on the maize sampled from the Volta region, likewise the range of mycotoxins present in the maize grains, suggesting the potential to pose some adverse health effects with the populace of the Volta region.

Excision of endosymbiotic bacteria from yeast under aging and starvation stresses

Although infrequent in our laboratory, growth of bacterial colonies has been observed on top of the purified cultures of yeasts. In this study, the likelihood of bacterial excision from yeast under aging and starvation stresses was assessed using 10 gastric and 10 food-borne yeasts. Yeasts were identified as members of Candida or Saccharomyces genus by amplification and sequencing of D1/D2 region of 26S rDNA. For aging stress, yeasts were cultured on brain heart infusion agar supplemented with sheep blood and incubated at 30 °C for 3–4 weeks. For starvation stress, yeasts were inoculated into distilled water and incubated similarly. After seven days, starved yeasts were cultured on yeast extract glucose agar, incubated similarly and examined daily for appearance of bacterial colonies on top of the yeast's growth. Outgrowth of excised bacteria was observed on top of the cultures of 4 yeasts (Y1, Y3, Y13 and Y18) after 3–7 days. The excised bacteria (B1, B3, B13 and B18) were isolated and identified at the genus level according to their biochemical characteristics as well as amplification and sequencing of 16S rDNA. B1 (Arthrobacter) were excised from Y1 (Candida albicans) upon aging and B3 (Staphylococcus), B13 (Cellulomonas) and B18 (Staphylococcus) were excised from their respective yeasts; Y3 (Candida tropicalis), Y13 (Saccharomyces cerevisiae) and Y18 (Candida glabrata) upon starvation. DNA from yeasts was used for detection of 16S rDNA of their intracellular bacteria and sequencing. Amplified products from yeasts showed sequence similarity to those of excised bacteria. Under normal conditions, yeast exerts tight control on multiplication of its intracellular bacteria. However, upon aging and starvation the control is no longer effective and bacterial outgrowth occurs. Unlimited multiplication of excised bacteria might provide yeast with plenty of food in close vicinity. This could be an evolutionary dialogue between yeast and bacteria that ensures the survival of both partners.

Proteolytic Bacillus spp. associated with tannery industries: Conventional and molecular identification

Samples collected from different stages of the tannery industry were found to be alkaline (pH 7.52 to 12.11). A good number of bacteria were found to be associated with the samples. The bacterial count ranged in between 1.34×105 to 3.44×105 cfu/ml and 1.04×105 to 6×105 cfu/ml on nutrient agar (NA) and peptone yeast extract glucose agar (PYG) medium, respectively. The maximum bacterial count was observed in bating stage while the minimum count was in the deliming stage. Primarily, 70 isolates were selected based on their different colonial morphology. After heat shock test 27 isolates were finally selected for identification and proteolytic potential. All the selected isolates were the members of different species of the genus Bacillus. The conventionally identified species were B. stearothermophilus (9), B. subtilis (4), B. brevis (3), B. pumilus (3), B. alcalophilus (2), B. badius (2), B. firmus (2) and B. lentus (2). Four important proteolytic isolates of Bacillus were selected for molecular identification. The isolates were confirmed as Bacillus subtilis strain B20 (L/P/2/1), B. subtilis strain PB18 (D/P/3/1), Bacillus sp. strain BVC38 (D/P/3′/2) and B. amyloliqefaciens strain Egy25 (B/N/3′/1). Except B/N/3′/1 all the conventional identification was in accordance with the molecular identification as the isolate B/N/3′/1 was conventionally identified as B. pumilus (B/N/3′/1). All the isolates showed positive proteolytic activities on skim milk agar and the zone ratio was in between 2.61 ± 0.44 and 6.42 ± 0.95. These isolates could be commercially utilized in the tannery and detergent industries for their proteolytic activity.

Selective Isolation and Phenotypic Characterization of Bacteria and Actinomycetes from Oil-contaminated Soils

Large numbers of bacteria and actinomycete strains were isolated from two contaminated soils with high total petroleum hydrocarbons (TPH) content. From five selective media formulations used, nutrient agar and glucose yeast extract agar showed the best results. Bacterial colony forming units per dry gram soil (ufc/g) ranged from 5.14x103 to 4.20x1011, whereas for actinomycetes was from 8.99x104 to 9.2x107 ufc/g. Seventy-three bacteria and ninety-three actinomycete isolates were recovered from the two soils. Numerical taxonomy analysis of phenotypic data shows that at 90% similarity, the bacteria could be divided into sixteen multimembered and fifteen single-membered phenetic-groups, while the actinomycetes were separated in twenty multimembered and four single-membered phenetic-groups. Out of the one hundred and sixty-six strains, only ten were unable to use gasoline or oil as sole carbon and nitrogen sources. Most of the isolates could degrade both hydrocarbons. The results of this study show that bacteria and actinomycetes are present in large numbers in high TPH contaminated soils, with an extraordinary degree of diversity and that they can use oil or its derivatives as nutrients.

First Report of Achlya oblongata Infection in Freshwater-Reared Asian Seabass Lates calcarifer.

In September 2014, a freshwater oomycete was first isolated from Asian Seabass Lates calcarifer fry that were reared in freshwater at a fish hatchery in Sabah, Malaysia. A fungal strain was isolated from infected fry by using glucose yeast extract (GY) agar. From morphological identification, the strain belonged to the genus Achlya based on the mode of zoospore release. Molecular phylogenetic analysis of the internal transcribed spacer region sequences from the strain showed high similarity (99-100%) to Achlya oblongata. The isolate was able to grow on GY agar incubated at 15-35°C, in GY broth adjusted to pH 3.0-11.0, and in up to 1.0% NaCl. This is the first report of Achlya infection in freshwater-reared Asian Seabass in Malaysia.

Use of two fungal species to induce agarwood resin formation in Gyrinops walla

Gyrinops walla Gaertn. is the only species growing in Sri Lanka that belongs to the agarwood family, Thymelaeaceae. Although agarwood resin induction and extraction from Aquilaria species of the same family have been practised for many decades in Southeast Asian region, the ability of producing agarwood resins in G. walla was discovered recently. Since previous studies were on agarwood resins formed due to natural causes, the present study was conducted to identify the potential fungal species that are capable of artificially inducing agarwood resin formation in G. walla. Since this is the first ever study conducted on artificial inducement of agarwood resin formation in G. walla, Aspergillus niger and Fusarium solani were selected owing to their high abundance in the naturally formed agarwood resinous tissues collected from 25 G. walla trees. Both fungal species were separately grown in yeast extract glucose agar and used to inoculate healthy G. walla trees under aseptic conditions. Three holes were made for each tree and 2 g of fungal culture including the medium were placed in each hole. Tissue discoloration, characteristic aroma, resin content and resin constituents were checked at 10 cm intervals above and below the inoculation points for a period of 1 year. Results revealed that tissue discoloration and resin content were higher in the trees inoculated with A. niger. Other than at 10 cm above and below the inoculation points, samples collected at all locations had significantly higher resin contents when inoculated with A. niger compared to F. solani. Sixteen agarwood resin constituents, which were also recorded in Aquilaria species, were identified from the discolored tissues using GC–MS analysis. Jinkohol, agarospirol and 2(2-phenyl) chromone derivatives were found in all discolored tissues collected at 10-cm intervals of the trees inoculated with both fungi. β-Seline, γ-eudesmol and valerenal were found in nine of 10 sample points on the stem. γ-Elemene was recorded only in one sample. The characteristic aroma during burning was stronger for dark-colored tissues than the light-colored ones. The present study confirmed the potential use of certain fungal species to induce agarwood resin in G. walla and that A. niger is more effective than F. solani.

Mycelial Growth and Fruit Body Nutritional Composition of Pleurotus Species Grown on Different Lignocellulosic Waste-based Media

Pleurotus species are edible mushrooms that possess the ability to degrade lignocellulose and are cultivated on lignocellulosic substrates to produce fruiting bodies that are appreciated for their nutritional and culinary values. Radial growth rate (ur), mycelial biomass, and intracellular glycogen and protein contents were evaluated in the colonies of two strains of P. ostreatus (strains 26 and 50) and one strain of P. pulmonarius (strain 35) grown on barley straw extract agar, corn stover extract agar (CSEA), wheat straw extract agar, and glucose-yeast extract agar (GYEA). The nutritional composition of the Pleurotus fruit bodies cultivated on barley straw, corn stover, and wheat straw were also determined. Of all of the strains, P. ostreatus 50 showed the highest ur on the CSEA and GYEA, and P. ostreatus 26 showed the greatest mycelial biomass production on the lignocellulosic waste-based agar media (LWAM). All of the Pleurotus strains showed the greatest and lowest glycogen contents on the CSEA and GYEA, respectively. These results showed that the LWAM are more suitable for mycelial growth than a glucose-based agar medium, and that Pleurotus fruit bodies cultivated on corn stover had the highest nutritional value compared with fruit bodies grown on the wheat and barley substrates.

Isolation and molecular characterization of bacteria from the gut of Eisenia fetida for biodegradation of 4,4 DDT

There is a growing interest in the bio-application of microorganisms from the gut of earthworms on the bio-transformation of persistent and toxic pollutants during vermifiltration. Earthworms harbor beneficial microbes in their gut which symbiotically aid in biodegradation of pollutants in their drilosphere. Dichlorodiphenyltrichloroethane is an example of polychlorinated hydrocarbons and metabolites thereof that are persistent in the environment and are toxic to humans and animals. This study aimed at isolating and identifying 4,4 dichlorodiphenyltrichloroethane degrading microorganisms in the gut of Eisenia fetida acclimatized to sewage. Six pure isolates obtained from gut contents were cultured in minimum salt media supplemented with 15 mgL-1 dichlorodiphenyltrichloroethane followed by glucose yeast extract agar sprayed with 1% 4,4 DDT in ether (v/v). Two pure isolates positive for 4,4 dichlorodiphenyltrichloroethane biodegradation were inoculated on minimum salt yeast extract media containing 15 mgL-1 4,4 dichlorodiphenyltrichloroethane. The resulting metabolites were identified using Gas Chromatography. The positive isolates were identified using 16S rRNA gene analysis as Rhodococcus genus and the Bacillus genus, exhibiting 88.36% and 85.22% 4.4 DDT degradation respectively. The study demonstrated DDT-degradation by bacteria from the gut of Eisenia fetida. These findings can be useful in optimization of vermifilters for biodegradation of dichlorodiphenyltrichloroethane and other xenobiotics.

Study of fungicidal properties of colloidal silver nanoparticles (AgNPs) on trout egg pathogen, Saprolegnia sp.

Silver nanoparticles (AgNPs) are known to have bactericidal and fungicidal effects. Since, there is few information available on the interaction of colloidal nanosilver with fish pathogens. Hence, the current study investigated the effects of colloidal AgNPs on the in vitro growth of the fish pathogen Saprolegnia sp.. Before the experiments, various important properties of AgNPs were well-characterized. The antifungal activity of AgNPs was then evaluated by determining the minimum inhibitory concentrations (MICs) using two-fold serial dilutions of colloidal nanosilver in a glucose yeast extract agar at 22oC. The growth of Saprolegnia sp. on the AgNPs agar treatments was compared to that of nanosilver-free agar as controls. The results showed that AgNPs have an inhibitory effect on the in vitro growth of the tested fungi. The MIC of AgNPs for Saprolegnia sp. was calculated at 1800 mg/L, which is equal to 0.18 percent. It seems that AgNPs could be a proper replacement for teratogenic and toxic agents, such as malachite green. In addition, the indirect use of AgNPs could be a useful method for providing new antifungal activity in aquaculture systems.

Antimicrobial and Cytotoxic Activities of Secondary Metabolites Obtained from a Novel Species of Streptomyces

With an aim to isolate actinomycetes having antimicrobial and cytotoxic activities, five colonies of actinomycetes were collected from soils of a vegetable garden of Rajshahi District, Bangladesh and designated as ANTS-1 to ANTS-5. Initial screening for antibacterial activity showed that the isolate ANTS-1 produced potent antimicrobial metabolites in the culture media. The crude ethyl acetate extract obtained from the yeast-extract glucose agar (YEGA) medium showed strong activity against pathogenic bacteria and moderate activity against fungi. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of the extract against the test bacteria were in the range of 1-2 ?g/ml and 4-8 ?g/ml, respectively. The LC50 value of the crude extract against brine-shrimp nauplii was only 0.4?g/ml indicating its potent cytotoxic activity. One component of the extract was identified as actinomycin D by interpretation of the spectral data and by comparison with published values. DOI: http://dx.doi.org/10.3329/bpj.v16i1.14485 Bangladesh Pharmaceutical Journal 16(1): 15-19, 2013

Hexavalent chromium reduction by aerobic heterotrophic bacteria indigenous to chromite mine overburden

Microbiological analysis of overburden samples collected from chromite mining areas of Orissa, India revealed that they are rich in microbial density as well as diversity and dominated by Gram-negative (58%) bacteria. The phenotypically distinguishable bacterial isolates (130) showed wide degree of tolerance to chromium (2–8 mM) when tested in peptone yeast extract glucose agar medium. Isolates (92) tolerating 2 mM chromium exhibited different degrees of Cr+6 reducing activity in chemically defined Vogel Bonner (VB) broth and complex KSC medium. Three potent isolates, two belonging to Arthrobacter spp. and one to Pseudomonas sp. were able to reduce more than 50 and 80% of 2 mM chromium in defined and complex media respectively. Along with Cr+6 (MIC 8.6–17.8 mM), the isolates showed tolerance to Ni+2, Fe+3, Cu+2 and Co+2 but were extremely sensitive to Hg+2 followed by Cd+2, Mn+2 and Zn+2. In addition, they were resistant to antibiotics like penicillin, methicillin, ampicillin, neomycin and polymyxin B. During growth under shake-flask conditions, Arthrobacter SUK 1201 and SUK 1205 showed 100% reduction of 2 mM Cr+6 in KSC medium with simultaneous formation of insoluble precipitates of chromium salts. Both the isolates were also equally capable of completely reducing the Cr+6 present in mine seepage when grown in mine seepage supplemented with VB concentrate.

Isolation and molecular identification of actinomycetes from mycetoma patients in Sudan

Introduction: In order to minimize chance of amputation due to actinomycetoma, it is important to correctly identify the causative agents. Microscopic examination of grains is not definite and further confirmatory diagnostic tests are needed. This study aims to investigate the prevalence of actinomycetoma and to explore the usefulness of strb1 gene in the diagnosis of the disease. Materials & Methods: The present study is a prospective cross-section laboratory-based study in which clinical samples (n = 100) from patients with mycetoma lesions were collected. The samples were cultured on Lowenstein Jensen and glucose yeast extract agar media. Grown colonies were initially identified using Gram's stain, Ziehl Neelsen stain, and selected biochemical reactions. Confirmation was done by the analysis of polymerase chain reaction amplified strb1 gene. Results: Actinomycetoma was represented by a high ratio (12%) among the study population. Nine out of the 12 isolates (75%) were found to belong to the genus Streptomyces; whereas three isolates (25%) were identified as Nocardia spp. on the basis of phenotypic and mycolic acid contents. Conclusion: It could be concluded that actinomycetoma exists with significant prevalence (12%) among patients investigated in the present study. Streptomyces is the most important etiological agent of actinomycetoma compared to Nocardia.

Prevalence of Nocardia species among HIV-positive patients with suspected tuberculosis

The objective of the study was to determine the frequency of nocardiosis in HIV-positive and HIV-negative individuals clinically suspected of having tuberculosis (TB). The study population (n = 171) were those who attended chest hospitals in Khartoum State, Sudan, between January and March 2010. The patients suffered from pulmonary infections with positive acid-fast bacilli. Blood (n = 171) and sputum (n = 171) samples were collected simultaneously. Blood samples were tested serologically for the presence of antibodies using HIV/Intensified Combination Prevention (ICP) test and sputum were cultured onto Lowenstein Jensen slants according to standard methods. Isolates showing rapid growth characteristic of Nocardiae were subcultured and subsequently identified using glucose yeast extract agar medium. All candidates in the study population (n = 171) suffered from pulmonary infections, nocardiosis was diagnosed in 4% (n = 7), HIV-positive cases were 17 (9.9%). Five Nocardia species were isolated from HIV-negative patients whereas two were from HIV-positive patients. Nocardia spp. cause pulmonary infections (4.09%) in both immunocompetent (2.92%) as well as immunocompromised (1.17%) patients who attend chest clinics in Sudan.

Differentiation in Fatty Acid Profiles of Pigmented and Nonpigmented Aurantiochytrium Isolated from Hong Kong Mangroves

Twenty-five thraustochytrids that belong to the genus Aurantiochytrium were isolated from subtropical mangroves in Hong Kong. Although they have similar morphological and physiological characteristics, they have different colors on a yeast extract-glucose agar plate, which were largely ignored before. On the basis of the differences in their colony color, 25 Aurantiochytrium strains were further classified into pigmented and nonpigmented subgroups and their fatty acid profiles were analyzed and compared. In general, nonpigmented Aurantiochytrium strains were found to contain biomass concentrations and growth yield coefficients statistically higher than pigmented Aurantiochytrium strains (p < 0.01). Among all isolates, a significantly higher content of polyunsaturated fatty acid (PUFA, 123.41-179.64 mg/g) was found in the nonpigmented Aurantiochytrium (p < 0.01), whereas the pigmented strains contained a higher amount of saturated fatty acids. Docosahexaenoic acid (DHA) was identified as the most abundant PUFA in both nonpigmented and pigmented Aurantiochytrium. According to the result of principal component analysis, the contents and composition of saturated fatty acids and PUFAs are the major varieties to distinguish these two Aurantiochytrium groups, especially the contents of C15:0, C13:0, C16:0, C17:0, and DHA. With a rapid growth rate and high DHA yield, the strain from the nonpigmented Aurantiochytrium group was regarded as the ideal candidate for PUFA production.

A new preferential medium for enumeration and isolation of desert actinomycetes

In order to facilitate the discovery of novel actinomycetes from the Egyptian deserts, which can be useful as new sources for bioactive metabolites, different media for enumeration and isolation of desert actinomycetes have been tested. For this purpose, 30 soil samples from different six sites representing the Western and Eastern deserts of Egypt were collected. The two deserts are considered hyper-arid and the soil characteristics were determined. The media used were glucose–yeast extract agar, soil extract agar and a new minimal medium (MM) containing glucose, yeast extract and mineral salts. The effects of the soil characteristics on the total viable actinomycete counts on the three media were evaluated. The results showed that the highest actinomycete count in samples from five out of six sites was obtained on MM. Also MM was more selective for actinomycetes and significantly decreased the number of fungal colonies and to a lower extent the number of bacterial colonies. Moreover, it supported the development of different and diverse groups of actinomycetes. From the results obtained in this study, MM is a new useful medium for enumeration and selective isolation of actinomycetes from the desert soils.

Pangola grass colonized with Scytalidium thermophilum for production of Agaricus bisporus

This work had the dual objective of selecting a substrate for rapid mycelial growth of Scytalidium thermophilum and then comparing the growth and production of a brown variety of Agaricus bisporus on substrate non-colonized and colonized with S. thermophilum. Mycelial growth of S. thermophilum at 45 °C was significantly greater on potato dextrose yeast extract agar (0.58 mm/h) as compared to malt extract glucose agar (0.24 mm/h) and yeast extract glucose agar (0.44 mm/h). On cereal grain, S. thermophilum grew significantly faster on rice (0.31 mm/h) compared to sorghum (0.22 mm/h) and millet (0.18 mm/h). It also grew faster on Pangola grass (0.49 mm/h) compared to corncobs (0.30 mm/h) and sawdust (0.18 mm/h). Colonization of Pangola grass with S. thermophilum was influenced by the addition of calcium salts in the form of gypsum, hydrated lime and ground limestone. For production of A. bisporus, biological efficiency (BE) on pasteurized Pangola grass pre-colonized by S. thermophilum for 4 days at 45 °C was more than twice (26.4%) that on grass non-colonized by S. thermophilum (11.0%). The addition of 2% hydrated lime to Pangola grass prior to colonization by S. thermophilum resulted in an additional doubling of BE of mushroom production (48.1%). These results show the possibility of developing a non-composted substrate method for producing A. bisporus without autoclaving the substrate.

Evaluation of direct plating methods to enumerate Alicyclobacillus in beverages

Ten agar media were evaluated for their suitability to support spore germination and colony development by six strains of Alicyclobacillus acidoterrestris, three strains of Alicyclobacillus acidocaldarius, and one strain of Alicyclobacillus cycloheptanicus. The influence of plating method (pour versus spread), incubation temperature (43 °C and 50 °C), and incubation time (up to 10 days) on colony development were determined. K agar, Alicyclobacillus medium (ALI agar), and Bacillus acidoterrestris thermophilic (BAT) agar recovered the highest numbers of spores. Orange serum agar and Hiraishi glucose yeast extract agar were the least suitable. Overall, surface plating was superior to pour plating and, with the exception of one strain of A. acidocaldarius which grew better at 50 °C, incubation of K agar, ALI agar, and BAT agar plates at 43 °C or 50 °C resulted in recovery of equivalent numbers of spores. Essentially all viable spores were detected on media incubated for 3 days at 43 °C. The ability of one strain of each Alicyclobacillus species to grow in ten non-carbonated commercially manufactured beverages at 30 °C and 43 °C was markedly affected by the composition of the beverages. Results show that surface plating samples on BAT agar, followed by incubating plates at 43 °C for 3 days provide the most suitable conditions to enumerate ten strains of three species of Alicyclobacillus most commonly responsible for spoilage of beverages.

Deinococcus mumbaiensissp. nov., a radiation-resistant pleomorphic bacterium isolated from Mumbai, India

A radiation-resistant, Gram-negative and pleomorphic bacterium (CON-1) was isolated from a contaminated tryptone glucose yeast extract agar plate in the laboratory. It was red pigmented, nonmotile, nonsporulating, and aerobic, and contained MK-8 as respiratory quinone. The cell wall of this bacterium contained ornithine. The major fatty acids were C16:0, C16:1, C17:0, C18:1 and iso C18:0. The DNA of CON-1 had a G+C content of 70 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that CON-1 exhibited a maximum similarity (94.72%) with Deinococcus grandis. Based on the genotypic, phenotypic and chemotaxonomic characteristics, the bacterium CON-1 was identified as a new species of the genus Deinococcus, for which the name Deinococcus mumbaiensis sp. nov. is proposed. The type strain of D. mumbaiensis is CON-1 (MTCC 7297(T)=DSM 17424(T)).