Showing papers in "Annals of Microbiology in 2014"

Isolation and characterization of drought-tolerant ACC deaminase and exopolysaccharide-producing fluorescent Pseudomonas sp.

Abstract: The enzyme 1-aminocyclopropane-1-carboxylate deaminase catalyzes the degradation of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of the plant hormone ethylene, into α-ketobutyrate and ammonia. The enzyme has been detected in a limited number of bacteria and plays a significant role in sustaining plant growth and development under biotic and abiotic stress conditions by reducing stress-induced ethylene production in plants. We have screened 32 fluorescent Pseudomonas sp. isolated from rhizosphere and non-rhizosphere soils of different crop production systems for drought tolerance using polyethylene glycol 6000 (PEG 6000). Nine of these isolates were tolerant to a substrate metric potential of −0.30 MPa (15 % PEG 6000) and therefore considered to be drought-tolerant. All of these drought-tolerant isolates were screened for ACC deaminase activity using ACC as the sole nitrogen source, and one (SorgP4) was found to be positive for ACC, producing 3.71 ± 0.025 and 1.42 ± 0.039 μM/mg protein/h of α-ketobutyrate under the non-stress and drought stress condition, respectively. The isolate SorgP4 also showed other plant growth-promoting traits, such as indole acetic acid production, phosphate solubilization, siderophore and hydrogen cyanide production. The ACC deaminase gene (acdS) from the isolate SorgP4 was amplified, and the nucleotide sequence alignment of the acdS gene showed significant homology with acdS genes of NCBI Genbank. The 16S rRNA gene sequencing analysis identified the isolate as Pseudomonas fluorescens. Both sequences have been submitted to the NCBI GenBank under the accession numbers JX885767 and KC192771 respectively.

Mechanisms of phosphate solubilization by fungal isolates when exposed to different P sources

Abstract: The use of phosphate-solubilizing fungi is a promising biotechnological strategy in the management of phosphorus (P) fertilization, as it enables the utilization of rock phosphates (RP) or the recovery of P fixed in soil particles. The objective of our study was to evaluate fungal isolates for mechanisms of solubilization of P-bearing compounds, such as AlPO4, FePO4, Ca3(PO4)2, Araxa RP, and Catalao RP. Four fungal isolates obtained from Brazilian soils were characterized in liquid media: Aspergillus niger FS1, Penicillium canescens FS23, Eupenicillium ludwigii FS27, and Penicillium islandicum FS30. A. niger FS1 was the only isolate able to solubilize all of the P sources, solubilizing 71, 36, 100, and 14 % of the P from AlPO4, FePO4, Ca3(PO4)2, and RPs, respectively. Medium acidification was an effective solubilization mechanism, particularly for Ca3(PO4)2. The other P sources were mainly solubilized through organic acids produced by the fungi. Oxalic acid, produced exclusively by A. niger FS1, and citric acid were decisive factors in the solubilization of AlPO4 and FePO4. Penicillium isolates produced more gluconic acid than A. niger FS1 in all treatments. However, this higher production did not result in higher solubilization for any of the P sources, showing that gluconic acid contributes little to the solubilization of the P sources evaluated. The higher capacity of medium acidification and the production of organic acids with stronger metal-complexation activity are characteristics that confer to A. niger FS1 a wider action on insoluble P sources. Consequently, this isolate qualifies as a promising candidate for application in the management of P fertilization.

Biological detoxification of mycotoxins: a review

Abstract: Mycotoxins are secondary fungal metabolites and are reported to be carcinogenic, genotoxic, teratogenic, dermato-, nephro- and hepatotoxic. Several studies have shown that economic losses due to mycotoxins occur at all levels of food and feed production, including crop and animal production, processing and distribution. Therefore, there is a great demand for a novel approach to prevent both the formation of mycotoxins in food and feed and the impact of existing mycotoxin contamination. Recently, investigators have reported that many microorganisms including bacteria, yeast, moulds, actinomycetes and algae are able to remove or degrade mycotoxins in food and feed. We have reviewed various strategies for the detoxification of mycotoxins using microorganisms such as bacteria, yeast and fungi.

Mechanisms of nisin resistance in Gram-positive bacteria

Abstract: Nisin is the most prominent lantibiotic and is used as a food preservative due to its high potency against certain Gram-positive bacteria. However, the effectiveness of nisin is often affected by environmental factors such as pH, temperature, food composition, structure, as well as food microbiota. The development of nisin resistance has been seen among various Gram-positive bacteria. The mechanisms under the acquisition of nisin resistance are complicated and may differ among strains. This paper presents a brief review of possible mechanisms of the development of resistance to nisin among Gram-positive bacteria.

Impurities of crude glycerol and their effect on metabolite production

Abstract: Glycerol is a valuable raw material for the production of industrially useful metabolites. Among many promising applications for the use of glycerol is its bioconversion to high value-added compounds, such as 1,3-propanediol (1,3-PD), succinate, ethanol, propionate, and hydrogen, through microbial fermentation. Another method of waste material utilization is the application of crude glycerol in blends with other wastes (e.g., tomato waste hydrolysate). However, crude glycerol, a by-product of biodiesel production, has many impurities which can limit the yield of metabolites. In this mini-review we summarize the effects of crude glycerol impurities on various microbial fermentations and give an overview of the metabolites that can be synthesized by a number of prokaryotic and eukaryotic microorganisms when cultivated on glycerol.

The prevalence of antibiotic resistance genes among Aeromonas species in aquatic environments

Abstract: The global rise in antimicrobial resistance (AMR) among bacteria causing infectious diseases is well documented, and the associated risks for human health are well known. There is much less research on AMR with regard to environmental strains, both opportunistic and pathogenic ones. The genus Aeromonas is widely distributed in the environment and causes many variable diseases in fish and humans. Infections in humans are predominantly caused by Aeromonas veronii, A. hydrophila and A. caviae (A. punctata) in a form of bacteremia, gastroenteritis or even septicaemia in immunocompetent and immunocompromised individuals. Different groups of antibiotics are used in the treatment, but studies indicate that fluoroquinolones and cefotaxime are the most efficient. A disturbing consequence of antibiotic overuse is an increasing number of detection of various antibiotic resistance genes (ARG) within this genus. The water environment is one of the major modes of transmission of resistant bacteria from animals to humans, and, thus, the dissemination of antibiotic resistance genes, particularly those located in mobile genetic elements (MGE) occurs in such as plasmids and transposons. This review summarizes recently published information on the type, distribution, and transmission of ARG by MGE, widespread in Aeromonas strains living in various aquatic environments, including wastewater, natural water, aquaculture and urban drinking water. The data available indicate that the opportunistic pathogens like Aeromonas spp. might serve as important vectors of ARG for clinically relevant pathogens present in such bodies of water .

Deciphering the diversity of culturable thermotolerant bacteria from Manikaran hot springs

Abstract: The aim of this study was to analyze and characterize the diversity of culturable thermotolerant bacteria in Manikaran hot springs. A total of 235 isolates were obtained employing different media, and screened for temperature tolerance (40 °C–70 °C). A set of 85 isolates tolerant to 45 °C or above were placed in 42 phylogenetic clusters after amplified ribosomal DNA restriction analysis (16S rRNA-ARDRA). Sequencing of the 16S rRNA gene of 42 representative isolates followed by BLAST search revealed that the majority of isolates belonged to Firmicutes, followed by equal representation of Actinobacteria and Proteobacteria. Screening of representative isolates (42 ARDRA phylotypes) for amylase activity revealed that 26 % of the isolates were positive, while 45 % exhibited protease activity, among which one amylase and six protease producers were tolerant up to 70 °C. BIOLOG-based identification of 13 isolates exhibiting temperature tolerance up to 70 °C, using carbon utilization patterns and sensitivity to chemicals, revealed a high degree of correlation with identification based on 16S rRNA gene sequencing for all isolates, except one (M48). These promising isolates showing a range of useful metabolic attributes demand to be explored further for industrial and agricultural applications.

Bacterial communities in sediments of a drinking water reservoir

Abstract: Sediment microbial communities play crucial roles in biogeochemical cycles of aquatic systems. Knowledge of microbial community structure could aid in our understanding of these roles. In this study, the sediment bacterial community structure of a drinking water reservoir in China was investigated. A large difference in major bacterial groups was observed at different sampling sites. Proteobacteria was the largest bacterial phylum, with a marked shift in the proportions of its major subdivisions. Microorganisms within phylum Proteobacteria might play important roles in various biogeochemical processes.

Characterization of antifungal activity of Paenibacillus ehimensis KWN38 against soilborne phytopathogenic fungi belonging to various taxonomic groups

Abstract: Soilborne fungal phytopathogens cause significant losses in many economically important crops and vegetables. The only way to control these devastating pathogens is by using higher doses of fungicides which not only increase the cost of production but also cause significant damage to the environment. Therefore alternate control measures are always looked for. In the present study, an antagonistic strain was isolated from the soil of the pepper fields around the seashore of Jellanamdo, South Korea and identified as Paenibacillus ehimensis KWN38 based on 16S rRNA sequencing. The strain showed high antifungal activity against six tested fungal pathogens belonging to various taxonomic groups on dual culture plates. Furthermore, the strain produced volatile antimicrobial compounds which had strong fungal growth inhibitory effect. The strain also showed high chitinase, cellulase, glucanase and protease activities. The hyphal morphologies of Rhizoctonia solani AG-1 (IA), Fusarium oxysporum f.sp. lycopersici and Phytophthora capsici were significantly destroyed by the crude enzymes and butanol extract from the culture supernatant and the affected hyphae showed abnormal bending, tip curling, and irregular branching. Hence, Paenibacillus ehimensis KWN38 is considered as a potential biocontrol agent of the soil-borne fungi causing plant diseases which is an important perspective of the present study.

Ochrobactrum sp. Pv2Z2 exhibits multiple traits of plant growth promotion, biodegradation and N -acyl-homoserine-lactone quorum sensing

Abstract: Rhizosphere bacteria play a vital role in plant growth, pathogen control, biodegradation and rhizosphere signaling. A motile, rod-shaped bacterium, Pv2Z2, isolated from the nodules of the common bean grown in Tanzanian soil was characterized using a polyphasic approach. The traits assessed included the production of indole-3-acetic acid and N-acyl homoserine lactone (AHL) molecules, solubilization of insoluble phosphate and zinc compounds and biodegradation of a number of toxic compounds. The 16S rRNA sequence of Pv2Z2 (EU399793) showed 99 % homology to Ochrobactrum anthropi isolates (Accession no. AJ867292, AJ867291, AJ867290) from soil samples of wheat root. Phylogenetic analysis showed relatedness to nodulating strain Ochrobactrum cytisi rather than to the clinical/pathogenic type strain of O. anthropi. Moreover, it showed unique fingerprints in the randomly amplified polymorphic DNA (RAPD) and two primers-RAPD assays which were different from those of the pathogenic type strain of O. anthropi. The bacterium produced 6.68 μg/mL-1 indoleacetic acid in the presence of tryptophan, released 25.7 μg/mL-1 phosphorus from inorganic tri-calcium phosphate in the Pikoviskaya’s medium and solubilized zinc sulphate and zinc oxide in the LG1 medium. The production of AHLs (e.g. 3O-C7-HSL, 3OH-C7-HSL) was detected with biosensor strains CV026 using reverse phase thin layer chromatography. The bacterium was able to grow in minimal salt medium supplemented with 100 mg/L each of phenol, 2-bromophenol, 2,4-diamino phenol hydrochloride, 3,4-dimethoxy benzyl alcohol and 4-methoxy benzyl alcohol. Phenol degradation was recorded up to a level of 94 % within 12 days. Inoculation of common bean plants resulted in a significant increase in plant height, fresh/dry weight and nitrogen uptake as compared to non-inoculated plants. The data suggest that the plant growth-promoting and biodegradation potential of this bacterium may be exploited on a large scale. The capacity to produce AHL molecules by members of the Ochrobactrum genus has not been previously reported and needs to be explored in detail.

Effects of photoperiod, salinity and pH on cell growth and lipid content of Pavlova lutheri

Abstract: The aim of the present work was to study the effects of photoperiod, salinity and pH on growth and lipid content of Pavlova lutheri microalgae for biodiesel production in small-scale and large-scale open-pond tanks. In a 250-mL flask, the cultures grew well under 24 h illumination with maximum specific growth rate, μ max , of 0.12 day−1 and lipid content of 35 % as compared to 0.1 day−1 and 15 % lipid content in the dark. The salinity was optimum for the cell growth at 30–35 ppt, but the lipid content of 34–36 % was higher at 35–40 ppt. Algal growth and lipid accumulation was optimum at pH 8–9. Large-scale cultivation in 5-L and 30-L tanks achieved μ max of 0.13–0.14 day−1 as compared to 0.12 day−1 in small-scale and 300L cultures.

Effects of additional Mg2+ on the growth, lipid production, and fatty acid composition of Monoraphidium sp. FXY-10 under different culture conditions

Abstract: The effects of supplementing the culture medium with Mg2+ on the growth, lipid production, and fatty acid composition of Monoraphidium sp. FXY-10 were studied under photoautotrophic, heterotrophic, and mixotrophic conditions. Under the photoautotrophic condition, microalgae supplemented with 100 μM Mg2+ grew significantly better than the control group and exhibited a secondary growth state. The final cell density was 1.25-fold higher than that of the control group (2.98 g L−1), and the peak lipid content reached 59.8 % (control group 52.3 %). Culture under the heterotrophic condition did not significantly increase the growth rate, but the experimental group (100 μM Mg2+ supplementation) achieved a 37.03 % lipid content compared to 28.47 % by the control group. The lipid productivity of the experimental group (100 μM Mg2+ supplementation) was higher, reaching 65.93 mg L−1 day−1 compared with 56.10 mg L−1 day−1 for the group without additional Mg2+. Under the mixotrophic condition, the experimental group achieved a final density of 3.10 g L−1, which was higher than that of the control group (2.98 g L−1). There was also no variation in fatty acid composition between the experimental group and the control group. Under the heterotrophic and mixotrophic conditions, the experimental group produced more than 50% saturated fatty and mono-unsaturated fatty acids, and the degree of unsaturation was <137. This result was relatively lower than that of the control.

Bacterioplankton communities in a high-altitude freshwater wetland

Abstract: Microbial communities play a crucial role in various biogeochemical processes in aquatic ecosystems. However, existing knowledge on microbial communities in the waters of wetlands is still very scant. The objective of the present study was to investigate the bacterioplankton community in the Luoshijiang Wetland, a high-altitude freshwater wetland in the Yunnan-Kweichow Plateau. Water samples were collected from different sites. The bacterioplankton community was characterized using 16S rRNA gene clone library analysis. A spatial variation of the diversity and composition of the bacterioplankton community was observed. Verrucomicrobia and Proteobacteria were the most abundant components. Proteobacteria might play an important role in water self-purification, but the significance of Verrucomicrobia remained unclear. Moreover, Pearson’s correlation analysis showed that Actinobacteria and Gemmatimonadetes were positively correlated with nitrite nitrogen in waters, while Alphaproteobacteria with dissolved phosphorous.

Use of Rhodopseudomonas palustris P1 stimulated growth by fermented pineapple extract to treat latex rubber sheet wastewater to obtain single cell protein

Abstract: Latex rubber sheet wastewater (non sterile wastewater: RAW) was treated efficiently using a stimulated Rhodopseudomonas palustris P1 inoculum with added fermented pineapple extract (FPE) under microaerobic light conditions. Optimization of wastewater treatment conditions using a central composite design (CCD) found that a 3 % stimulated P1 inoculum with 0.9 % added FPE and a 4-day retention time (RT) were the most suitable conditions. Calculations from CCD experiments predicted that a chemical oxygen demand (COD) of 3,005 mg/L could be 98 % removed, together with 79 % of suspended solids (SS) and 72 % of total sulfide (TtS). No H2S was detected, production costs were low and single cell protein (SCP) was a by-product. The results of the verification test had an error of only 4–8 % and confirmed removal of COD (initial COD 2,742 mg/L), SS and TtS at 94 %, 75 % and 66 %, respectively. These values were less than the best set obtained from the CCD experiment (2 % stimulated P1 inoculum, 0.75 % FPE and 4 days RT); upon repeating, this set could reduce 96 % of the COD, 78 % SS and 71 % TtS. The treated wastewater met the standard guidelines for irrigation use and no H2S was detected. The biomass obtaining after wastewater treatment from the best set consisted mostly of R. palustris P1; the biomass of this set had 65 % protein, 3 % fat, 8 % carbohydrate, 14 % ash and 10 % moisture. The results demonstrated that an inoculum of stimulated P1 grew well in RAW supplemented with FPE and could be considered to be an appropriate technology for effectively treating wastewater, with SCP as a by-product.

Bacterial diversity in the central black component of Maotai Daqu and its flavor analysis

Abstract: Maotai Daqu (MTDQ) is a kind of microflora starter for making the distilled liquor Maotai. It mainly contains bacteria, yeasts and moulds. When it comes to its central black component, it is nearly all bacteria. This study aims to clarify the bacterial diversity of the central black component of MTDQ, to analyze the role bacteria plays in Maotai liquor brewing, and to study the flavor components produced during the mixed solid-fermentation of three of the predominant flavor-producing bacteria so as to develop their practical application in other industries. Bacterial diversity was studied using amplified ribosomal DNA restriction analysis; Actinobacteria and Firmicutes were the predominant bacteria. The three flavor-producing strains were identified as Bacillus subtilis, B. licheniformis and B. amyloliquefaciens, which are the predominant bacteria. Their fermentation products include propanoic acid, 1,3-butanediol, acetic acid, methyl ester, etc. Most of which are analogous to the main flavor components of Maotai liquor. The results imply that the sample has diverse bacterial communities. Some bacteria play a very important role in the formation of the unique wine body composition of the Maotai liquor through their biotransformation, and some could be the main origins of the flavor of Maotai-type liquor.

Identification and evaluation of a potential biocontrol agent, Bacillus subtilis, against Fusarium sp. in apple seedlings

Abstract: To find a potential biocontrol agent against Fusarium sp. in apple seedlings, an endophytic bacterium strain was isolated from apple tree tissues. The inhibitive efficiency of the isolated strain against the hyphal growth of Fusarium sp. and Rhizoctonia solani was tested. Strain Y-1 showed significant inhibitory effects against Fusarium oxysporum, F. moniliforme, F. proliferatum, F. solani and R. solani. Its antifungal activity against F. oxysporum was the highest, reaching up to 64.90 %. In vivo tests indicated that strain Y-1 effectively protects apple from F. oxysporum infections. The control effect reached 92.26 % when bacterial inoculation was performed 3 days prior to pathogen inoculation. Strain Y-1 could colonize the rhizosphere and tissues within 30 days. It was also able to induce systemic resistance in apple seedlings as shown by the activities of SOD and POD. Strain Y-1 significantly increased the root length, root wet and dry weights, and plant height of the apple seedlings compared with the control group. The homology analysis of the 16S rRNA sequence, together with morphological, physical, and biochemical analyses, revealed that strain Y-1 is Bacillus subtilis.

In vitro antifungal, probiotic and antioxidant properties of novel Lactobacillus plantarum K46 isolated from fermented sesame leaf

Abstract: This study aimed to describe the diversity of antifungal lactic acid bacteria (LAB) in popular traditional Korean fermented food. A total of 22 LAB strains was selected and subjected to a monophasic identical approach using 16S rRNA gene sequence analysis. Antifungal LAB associated with fermented food was identified as Lactobacillus plantarum (9), Lactobacillus graminis (5), Lactobacillus pentosus (4), Lactobacillus sakei (2), Lactobacillus paraplantarum (1), and Leuconostoc mesenteroides subsp. mesenteroides (1). Novel Lactobacillus plantarum strain K46 exhibited comparatively better antifungal activity against several spoilage fungi, and was deposited in the Korean Collection for Type Cultures (KACC91758P). Antifungal substances from the spent medium in which K46 was cultivated were extracted with ethyl acetate. Antifungal activity was assessed using the broth micro dilution technique. Compounds were characterized based on infrared, 13C nuclear magnetic resonance (NMR), and 1H NMR spectral data. The minimum inhibitory concentration (MIC) of the compounds against Aspergillus clavatus, Aspergillus oryzae, Penicillium chrysogenum and Penicillium roqueforti was 2.5 mg/mL and that against Aspergillus fumigatus, Aspergillus niger, Curvularia lunata and Gibberella moniliformis was 5.0 mg/mL. K46 was able to survive gastrointestinal conditions simulating the stomach and the duodenum passage with the highest percentage of hydrophobicity. In addition, its resistance to hydrogen peroxide and highest hydroxyl radical and 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activities, with inhibition rates of 43.53 % and 56.88 %, respectively, were to its advantage. An antimicrobial susceptibility pattern was an intrinsic feature of this strain, thus consumption does not represent a health risk to humans. The results showed the potential of K46 strain as an antifungal, probiotic and antioxidant culture, and hence it was determined to be suitable for application in functional foods.

Wickerhamomyces anomalus AS1: a new strain with potential to improve wine aroma

Abstract: The monoterpenes are the most important contribution to the olfactory profile of wine due to their low odour threshold. These and other aroma-active substances do not generally exist in a free form but are conjugated to mono- or disaccharides, thereby forming water-soluble and odourless complexes. Enzymes that cleave the sugar moieties from the precursors can, therefore, have a major impact on the sensory profile of wine, as they release the volatile aroma compounds. For this reason, we searched for wine yeasts producing glycosidases which are active under oenological conditions. A collection of 100 wine yeasts were screened for glycosidase activities in whole cells and in culture supernatants. Kinetic parameters were determined spectrophotometrically with synthetic model substrates, and hydrolysis of natural glycosides was detected by thin-layer chromatography. A yeast isolate, AS1, was identified as a new Wickerhamomyces anomalus strain which hydrolysed a number of synthetic and natural glycosides under oenological conditions. Citronellol- and nerol-glucosides, among the most frequently occurring aroma precursors in wine, were also cleaved. In contrast to a commercial β-glucosidase, whole cells of W. anomalus AS1 catalysed deglycosylation of arbutin and salicin directly in a white and a red wine. Besides the formation of intra- and extracellular glucoside hydrolases, strain AS1 exhibited arabinosidase and xylosidase activities which are also essential for the release of flavour compounds. Even with limited functionality at oenological conditions, the glycoside hydrolase activities of W. anomalus AS1 may improve aroma development, provided that the reaction occurs over a longer period, as it is the case during wine-making.

Comparison of bacteriocins production from Enterococcus faecium strains in cheese whey and optimised commercial MRS medium

Abstract: The production of bacteriocins from cheap substrates could be useful for many food industrial applications. This study aimed at determining the conditions needed for optimal production of enterocins SD1, SD2, SD3 and SD4 secreted by Enterococcus faecium strains SD1, SD2, SD3 and SD4, respectively. To our knowledge, this is the first use of cheese whey—a low-cost milk by-product—as a substrate for bacteriocin production by E. faecium; skimmed milk and MRS broths were used as reference media. This cheese manufacturing residue proved to be a promising substrate for the production of bacteriocins. However, the levels of secreted antimicrobial compounds were lower than those achieved by E. faecium strains in MRS broth. Bacteriocin production was affected strongly by physical and chemical factors such as growth temperature, time of incubation, pH, and the chemical composition of the culture medium. The optimal temperature and time of incubation supporting the highest bacteriocin production was determined for each strain. Different types, sources and amounts of organic nitrogen, sugar, and inorganic salts played an essential role in bacteriocin secretion. E. faecium strains SD1 and SD2—producing high bacteriocin levels both in cheese whey and skimmed milk—could be of great interest for potential applications in cheese-making.

Effect of Bisphenol A on human health and its degradation by microorganisms: a review

Abstract: Bisphenol A (BPA), is an industrially important compound and is widely used for the production of polycarbonates and other plastics. Over the past few years, there have been many issues raised all over the world on the use of BPA. BPA is known to possess estrogenic activities; hence, it mimics the role of estrogen once it enters living systems. Thus, it has been placed in the category of compounds called endocrine disruptors. It can cause damage to reproductive organs, thyroid gland, and brain tissues at developmental stages, and most recently it has also been linked to cancer development in humans. Here, in this review, we aim to summarize the various effects of BPA on humans and animals, and at the same time we wish to throw some light on the emerging field of biodegradation of BPA in the natural environment. A few studies conducted recently have tried to isolate BPA-degrading microorganisms from various sites, like water bodies receiving wastes from industries, landfills, etc. In the present scenario, with huge controversies on the use of BPA, we emphasize on bridging the gap between studies, aiming at finding the damage caused by BPA, and the studies which aim at the safe removal of BPA from the environment, with the help of naturally occurring microbes. Once this gap is filled, we will be able to find a way which will allow the use of BPA in manufacturing plastics, without its accumulation in the environment.

Microbial synthesis of spherical nanosilver and nanogold for mosquito control

Abstract: Biosynthesis of metal nanoparticles using microorganisms is an important area of research in nanobiotechnology, which is an emerging eco-friendly science of well-defined sizes, shapes and controlled monodispersity. The present study proposed a green process for the extracellular production of silver (Ag) and gold (Au) nanoparticles (NPs) using the soil fungi Chrysosporium keratinophilum and Verticillium lecanii. The synthesized NPs were formed fairly uniform with spherical shape determined by Transmission Electron Microscope (TEM) and confirmed by Scanning Electron Microscope (SEM). Elemental analysis on single particle was carried by EDX analysis. The results were further supported by UV-vis spectrophotometry. In addition, we have also investigated the effect of synthesized AgNPs and AuNPs against the larvae and pupae of Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti. The efficacy test was performed at different concentrations for periods of different lengths by the probit analysis. The larvae and pupae of Cx. quinquefasciatus, An. stephensi and Ae. aegypti were found highly susceptible to the synthesized AgNPs than the AuNPs. The larvae of Cx. quinquefasciatus and Ae. aegypti were found to be more susceptible to the AgNPs and AuNPs synthesized using the C. keratinophilum and V. lecanii compared with the larvae of An. stephensi. The pupae of Ae. aegypti have shown higher mortality against the synthesized AgNPs than the pupa of Cx. quinquefasciatus, while no adverse effects could be observed in the pupa of An. stephensi. By this approach, it is suggested that this rapid synthesis of nanoparticles would be useful for developing a biological process for mosquito control.

Response surface methodology as a decision-making tool for optimization of culture conditions of green microalgae Chlorella spp. for biodiesel production

Abstract: We have evaluated process optimization and the interactive effects of a number of variables using a Box–Behnken design of response surface methodology (RSM). The process variables nitrate, phosphate, glucose and pH were optimized to enhance the cell growth rate, lipid accumulation and other biochemical parameters of Chlorella spp. The most significant increase in lipid production (dry cell weight basis) occurred at limited concentrations of nitrate and phosphate, 1 % glucose and pH 7.5. The addition of nitrates during the mid-lag and mid-exponential phases produced the maximum inhibitory effect on lipid accumulation and the presence of yeast extract led to a further enhancement of lipid accumulation. Of all the media tested, BG-11 was the best suited medium for algal biomass production and chlorophyll content. A significant increase in algal biomass was observed in BG-11 supplemented with bicarbonate and glucose (1 %). The maximum specific growth rate observed was on 9th day of culturing. Results of optimization of process variables through response surface methodology and optimization of various other conditions reflect cutting edge research directed towards increasing algal biomass and lipid content for biodiesel production using an efficient economical technological approach.

Bioactivity of endophytic Trichoderma fungal species from the plant family Cupressaceae

Abstract: Trichoderma fungal species are universal soil residents that are also isolated from decaying wood, vegetables, infected mushroom and immunocompromised patients. Trichoderma species usually biosynthesize a plethora of secondary metabolites. In an attempt to explore endophytic fungi from healthy foliar tissues of the plant family Cuppressaceae, we explored Cupressus arizonica, C. sempervirens var. cereiformis, C. sempervirens var. fastigiata, C. sempervirens var. horizontalis, Juniperus excelsa, Juniperus sp. and Thuja orientalis plants and recovered several endophytic Trichoderma fungal strains from Trichoderma atroviride and Trichoderma koningii species. We found that the host plant species and biogeographical location of sampling affected the biodiversity and bioactivity of endophytic Trichoderma species. Furthermore, the bioactivity of Trichoderma isolates and the methanol extracts of their intra- and extra-cellular metabolites were assessed against a panel of pathogenic fungi and bacteria. Fungal growth inhibition, conidial cytotoxicity, minimum inhibitory concentration and minimum bactericidal concentration were evaluated and analyzed by statistical methods. Our data showed that both intra- and extracellular secondary metabolites from all endophytic isolates had significant cytotoxic and antifungal effects against the model target fungus Pyricularia oryzae and the cypress fungal phytopathogens Diplodia seriata, Phaeobotryon cupressi and Spencermartinsia viticola. Further research indicated their significant antimicrobial bioactivity against the model phytopathogenic bacteria Pseudomonas syringae, Erwinia amylovora and Bacillus sp., as well. Altogether, the above findings show for the first time the presence of T. atroviride and T. koningii as endophytic fungi in Cupressaceae plants and more importantly, the Trichoderma isolates demonstrate significant bioactivity that could be used in future for agrochemical/drug discovery and pathogen biocontrol.

Impact of environmental variables on the isolation, diversity and antibacterial activity of endophytic fungal communities from Madhuca indica Gmel. at different locations in India

Abstract: A total of 1,897 isolates of endophytic fungi from Madhuca indica Gmel., representing 40 morphologically distinct fungal taxa were obtained from 2,700 segments of stem, bark and leaf from three different locations (Loc 1, Loc 2 and Loc 3) in Uttar Pradesh, India. Out of 40 taxa, 28 were identified microscopically and the remaining 12 by molecular methods. Coelomycetes (62.41 %) were the most prevalent fungal group followed by hyphomycetes (28.89 %) and ascomycetes (8.70 %). Colonisation frequency (CF) was greater in stem (82.55 %) than in leaf (65.00 %) and bark (63.22 %). Due to the dominance of a few taxa, species richness and Shannon and Simpson diversity indices were lower in stem than in leaf and bark at each location. Interestingly, less rainfall and lower temperatures disfavoured the overall colonisation of fungal endophytes at Loc 2. The stem samples from all locations were very similar in their endophytic composition, whereas bark and leaf samples showed differences. The dominant endophytic fungi isolated were Phomopsis sp. 1 (9.185 %), and Colletotrichum gloeosporioides (7.00 %). Principal component analysis showed 55 % tissue specificity with 51.08 % maximum variance. Antibacterial activity revealed that 58.33 % endophytic fungi were active against at least one or more bacterial pathogens, whereas the crude extract of five endophytic fungi inhibited the growth of five or more than five (50 %) of the pathogens tested. This report illustrates the value of having an adequate sample size from different tissues and different locations for species and chemical diversity in search of novel natural products.

Zeta potential as a measure of the surface charge of mycobacterial cells

Abstract: The surface charge of bacteria is closely related to their envelope structure and interactions with surfaces in natural environments. The aim of this study was to estimate the effect of experimental conditions on the zeta (ζ) potential of mycobacterial cells as a measure of their cell-surface charge. We observed that Mycobacterium smegmatis mc2155 cells at physiological conditions displayed a high and stable ζ potential (−42.9 ± 5.9 mV) which increased from the late-exponential phase of growth and at pH levels of >8.0. The optimal conditions for estimating the surface charge of mycobacteria using the ζ potential occurred when cells were harvested during the exponential growth phase (OD595 0.3–0.5) and then dispersed in solutions with pH levels of 7.0–10.0. These optimal conditions of ζ potential measurements were useful for differentiating between the virulent M. tuberculosis H37Rv strain and various non-virulent mycobacterial strains at pH 9.8. This study is the first to use zetametry to estimate the cell-surface charge of M. tuberculosis cells. We expect that the experimental conditions presented in this work will have further applications to estimate the cell-surface charge of other wild-type or genetically modified mycobacterial species and thereby further our understanding of the physicochemical interactions of mycobacteria with external surfaces in natural environments.

Lactobacillus salivarius and L. gasseri down-regulate Aggregatibacter actinomycetemcomitans exotoxins expression

Abstract: Beneficial microbes, such as lactobacilli establish a symbiosis with the host and confer health-associated effects, by limiting the growth of indigenous pathogens and challenging microbes introduced by altered foods. Nevertheless, there is scarce information on the effects of beneficial microbes on the virulence properties of bacterial species associated with oral diseases, such as periodontitis. Aggregatibacter actinomycetemcomitans is a Gram-negative species highly implicated in the etiology of localized aggressive periodontitis. The objective of this study was to investigate the effect of lactobacilli on the expression of the two major virulence factors of A. actinomycetemcomitans. Lactobacillus salivarius and L. gasseri were selected as beneficial species. The gene expressions of leukotoxin (LtxA) and cytolethal distending toxin (CdtB) by A. actinomycetemcomitans were analyzed in response to challenge by lactobacilli cell-free supernatants. Neither lactobacilli affected the growth, but strongly attenuated the expressions of both CdtB and LtxA in the two A. actinomycetemcomitans strains tested. This reduction of the expression of these two exotoxins was time-dependent. These fundamental findings may indicate that lactobacilli can reduce the virulence of putative opportunistic oral pathogens, and may provide insights to future therapeutic approaches for the respective diseases.

Isolation and identification of antimicrobial secondary metabolites from Bacillus cereus associated with a rhabditid entomopathogenic nematode

Abstract: The cell-free culture filtrate of Bacillus cereus associated with an entomopathogenic nematode, Rhabditis (Oscheius) sp., exhibited strong antimicrobial activity. The ethyl acetate extract of the bacterial culture filtrate was purified by silica gel column chromatography to obtain six bioactive compounds. The structure and absolute stereochemistry of these compounds were determined based on extensive spectroscopic analyses (LCMS, FABMS, 1H NMR, 13C NMR, 1H −1H COSY, 1H −13C HMBC) and Marfey’s method. The compounds were identified as cyclo(D-Pro-D-Leu), cyclo(L-Pro-D-Met), cyclo (L-Pro-D-Phe), cyclo (L-Pro-L-Val), 3,5-dihydroxy-4-ethyl-trans-stilbene, and 3,5-dihydroxy-4-isopropylstilbene, respectively. Compounds recorded antibacterial activity against all four tested bacteria strains of Bacillus subtilis, Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. 3,5-dihydroxy-4-isopropylstilbene recorded activity only against Gram-positive bacteria while cyclo(L-Pro-L-Val) recorded no antibacterial activity. Best antibacterial activity was recorded by 3,5-dihydroxy-4-ethyl-trans-stilbene (4 μg/ml) against Escherichia coli. The six compounds recorded significant antifungal activities against five fungal strains tested (Aspergillus flavus, Candida albicans, Fusarium oxysporum, Rhizoctonia solani and Penicillium expansum) and they were more effective than bavistin, the standard fungicide. The activity of cyclo(D-Pro-D-Leu), cyclo(L-Pro-D-Met), 3,5-dihydroxy-4-ethyl-trans-stilbene, and 3,5-dihydroxy-4-isopropylstilbene against Candida albicans was better than amphotericin B. To the best of our knowledge, this is the first report of antifungal activity of the bioactive compounds against the plant pathogenic fungi Fusarium oxysporum, Rhizoctonia solani, and Penicillium expansum. We conclude that the Bacillus cereus strain associated with entomopathogenic nematode is a promising source of natural bioactive secondary metabolites which may receive great benefit as potential sources of new drugs in the agricultural and pharmacological industry.

Biosynthesis of poly-β-hydroxybutyrate (PHB) with a high molecular mass by a mutant strain of Azotobacter vinelandii (OPN)

Abstract: The aim of this study was to characterize the influence of the aeration conditions on the production of PHB and its molecular mass in a mutant strain of Azotobacter vinelandii (OPN), which carries a mutation on ptsN, the gene encoding enzyme IIANtr, previously shown to increase the accumulation of PHB. Cultures of A. vinelandii wild-type strain OP and its mutant derivative strain OPN were grown in 500-mL flasks, containing 100 and 200 mL of PY sucrose medium. PHB production and its molecular mass were analyzed at the end of the culture. The molecular mass (MM) was significantly influenced by the aeration conditions and strain used. A polymer with a higher molecular weight was produced under low aeration conditions for both strains. A maximal molecular mass of 2,026 kDa (equivalent to 3,670 kDa measured by GPC) was obtained with strain OPN cultured under low-aeration conditions, reaching a value two-fold higher than that obtained from the parental strain OP (MM = 1,013 kDa) grown under the same conditions. Aeration conditions and the ptsN mutation influence the molecular mass of the PHB produced by A. vinelandii affecting in turn its physico-chemical properties.

Bionectria ochroleuca NOTL33—an endophytic fungus from Nothapodytes foetida producing antimicrobial and free radical scavenging metabolites

Abstract: Endophytic fungi are reported to produce diverse classes of secondary metabolites. This study investigated the antimicrobial and free radical scavenging activity of a foliar endophytic fungus from Nothapodytes foetida, a medium sized tree known to produce the antineoplastic compound camptothecin. The fungal isolate was identified as Bionectria ochroleuca based on the ITS rDNA analysis. The differences among endophytic, pathogenic and free living Bionectria ochroleuca were established by RNA secondary structure analysis. The metabolites showed a broad spectrum of antibacterial, antifungal and anti-dermatophytic activity. Minimum inhibitory concentration values of ethyl acetate extracts were in the range of 78–625 μg/mL against all test organisms, except for Pseudomonas aeruginosa (5 mg/mL). Antimicrobial components in the ethyl acetate extract were identified by GC-MS analysis. The isolate was also produced volatile antifungal compounds. A dose-dependent free radical quenching was observed in the ethyl acetate extract. This is the first report on Bionectria sp. as an endophyte of N. foetida. The results indicate that the B. ochroleuca NOTL33 isolate is a potential source of antimicrobial agents and could be used as an effective biofumigant.

Reduced nitrite and biogenic amine concentrations and improved flavor components of Chinese sauerkraut via co-culture of Lactobacillus plantarum and Zygosaccharomyces rouxii

Abstract: The aim of this study was to investigate the effect of inoculated fermentation on the quality of Chinese sauerkraut. To this end we studied a co-culture system consisting of Lactobacillus plantarum Shanghai brewing 1.08 and Zygosaccharomyces rouxii CGMCC 3791 during inoculated sauerkraut fermentation. The nitrite concentrations in pickled cabbage and radish inoculated with starter cultures of L. plantarum and Z. rouxii were significantly lower than those in the spontaneous fermentation system during the whole fermentation process. In addition, co-culture of L. plantarum and Z. rouxii during the production of sauerkraut decreased the formation of biogenic amines in the pickled vegetables. Using gas chromatography–mass spectrometry we also compared the levels of volatile compounds in inoculated and naturally fermented Chinese sauerkraut. Sixty compounds were identified, with the sauerkraut inoculated with starter cultures containing overall higher contents of volatile compounds, including acids, alcohols, esters, and phenols. The structure of the microbial community during the production of sauerkraut was studied using phospholipid fatty-acid (PLFA) analysis. This analysis revealed that the brine of inoculated sauerkraut contained significantly higher contents of Gram-positive and fungal PLFAs and a lower content of Gram-negative PLFAs, suggesting that the improved quality of inoculated Chinese sauerkraut may be ascribed to the inhibition of the growth of Gram-positive during sauerkraut fermentation. These results may indicate a new strategy to enhance the quality of Chinese sauerkraut.