Secondary Metabolites with α-Glucosidase Inhibitory Activity from Mangrove Endophytic Fungus Talaromyces sp. CY-3.

Eight new compounds, including two sambutoxin derivatives (1-2), two highly oxygenated cyclopentenones (7-8), four highly oxygenated cyclohexenones (9-12), together with four known sambutoxin derivatives (3-6), were isolated from semimangrove endophytic fungus Talaromyces sp. CY-3, under the guidance of molecular networking. The structures of new isolates were elucidated by analysis of detailed spectroscopic data, ECD spectra, chemical hydrolysis, 13C NMR calculation, and DP4+ analysis. In bioassays, compounds 1-5 displayed better α-glucosidase inhibitory activity than the positive control 1-deoxynojirimycin (IC50 = 80.8 ± 0.3 µM), and the IC50 value was in the range of 12.6 ± 0.9 to 57.3 ± 1.3 µM.

Natural Wolbachia infection in field-collected Anopheles and other mosquito species from Malaysia.

BACKGROUND: The endosymbiont bacterium Wolbachia is maternally inherited and naturally infects some filarial nematodes and a diverse range of arthropods, including mosquito vectors responsible for disease transmission in humans. Previously, it has been found infecting most mosquito species but absent in Anopheles and Aedes aegypti. However, recently these two mosquito species were found to be naturally infected with Wolbachia. We report here the extent of Wolbachia infections in field-collected mosquitoes from Malaysia based on PCR amplification of the Wolbachia wsp and 16S rRNA genes. METHODS: The prevalence of Wolbachia in Culicinae mosquitoes was assessed via PCR with wsp primers. For some of the mosquitoes, in which the wsp primers failed to amplify a product, Wolbachia screening was performed using nested PCR targeting the 16S rRNA gene. Wolbachia sequences were aligned using Geneious 9.1.6 software, analyzed with BLAST, and the most similar sequences were downloaded. Phylogenetic analyses were carried out with MEGA 7.0 software. Graphs were drawn with GraphPad Prism 8.0 software. RESULTS: A total of 217 adult mosquitoes representing 26 mosquito species were screened. Of these, infections with Wolbachia were detected in 4 and 15 mosquito species using wsp and 16S rRNA primers, respectively. To our knowledge, this is the first time Wolbachia was detected using 16S rRNA gene amplification, in some Anopheles species (some infected with Plasmodium), Culex sinensis, Culex vishnui, Culex pseudovishnui, Mansonia bonneae and Mansonia annulifera. Phylogenetic analysis based on wsp revealed Wolbachia from most of the mosquitoes belonged to Wolbachia Supergroup B. Based on 16S rRNA phylogenetic analysis, the Wolbachia strain from Anopheles mosquitoes were more closely related to Wolbachia infecting Anopheles from Africa than from Myanmar. CONCLUSIONS: Wolbachia was found infecting Anopheles and other important disease vectors such as Mansonia. Since Wolbachia can affect its host by reducing the life span and provide resistance to pathogen infection, several studies have suggested it as a potential innovative tool for vector/vector-borne disease control. Therefore, it is important to carry out further studies on natural Wolbachia infection in vector mosquitoes' populations as well as their long-term effects in new hosts and pathogen suppression.

Cercospora beticola: The intoxicating lifestyle of the leaf spot pathogen of sugar beet.

Cercospora leaf spot, caused by the fungal pathogen Cercospora beticola, is the most destructive foliar disease of sugar beet worldwide. This review discusses C. beticola genetics, genomics, and biology and summarizes our current understanding of the molecular interactions that occur between C. beticola and its sugar beet host. We highlight the known virulence arsenal of C. beticola as well as its ability to overcome currently used disease management strategies. Finally, we discuss future prospects for the study and management of C. beticola infections in the context of newly employed molecular tools to uncover additional information regarding the biology of this pathogen. TAXONOMY: Cercospora beticola Sacc.; Kingdom Fungi, Phylum Ascomycota, Class Dothideomycetes, Order Capnodiales, Family Mycosphaerellaceae, Genus Cercospora. HOST RANGE: Well-known pathogen of sugar beet (Beta vulgaris subsp. vulgaris) and most species of the Beta genus. Reported as pathogenic on other members of the Chenopodiaceae (e.g., lamb's quarters, spinach) as well as members of the Acanthaceae (e.g., bear's breeches), Apiaceae (e.g., Apium), Asteraceae (e.g., chrysanthemum, lettuce, safflower), Brassicaceae (e.g., wild mustard), Malvaceae (e.g., Malva), Plumbaginaceae (e.g., Limonium), and Polygonaceae (e.g., broad-leaved dock) families. DISEASE SYMPTOMS: Leaves infected with C. beticola exhibit circular lesions that are coloured tan to grey in the centre and are often delimited by tan-brown to reddish-purple rings. As disease progresses, spots can coalesce to form larger necrotic areas, causing severely infected leaves to wither and die. At the centre of these spots are black spore-bearing structures (pseudostromata). Older leaves often show symptoms first and younger leaves become infected as the disease progresses. MANAGEMENT: Application of a mixture of fungicides with different modes of action is currently performed although elevated resistance has been documented in most employed fungicide classes. Breeding for high-yielding cultivars with improved host resistance is an ongoing effort and prudent cultural practices, such as crop rotation, weed host management, and cultivation to reduce infested residue levels, are widely used to manage disease. USEFUL WEBSITE: https://www.ncbi.nlm.nih.gov/genome/11237?genome_assembly_id=352037.

Antimicrobial Peptides from Lactobacillus plantarum UTNGt2 Prevent Harmful Bacteria Growth on Fresh Tomatoes.

In a previous study, the antimicrobial peptides extracted from Lactobacillus plantarum UTNGt2 of wild-type fruits of Theobroma grandiflorum (Amazon) were characterized. This study aimed to investigate the antimicrobial mechanisms of peptides in vitro and its protective effect on fresh tomatoes. The addition of partially purified Gt2 peptides to the E. coli suspension cells at the exponential (OD605 = 0.7) growth phase resulted in a decrease with 1.67 (log10) order of magnitude compared to the control without peptide. A marginal event (< 1 log10 difference) was recorded against Salmonella, while no effect was observed when combined with EDTA, suggesting that the presence of a chelating agent interfered with the antimicrobial activity. The Gt2 peptides disrupted the membrane of E. coli, causing the release of ß-galactosidase and leakage of DNA/RNA molecules followed by cell death, revealing a bacteriolytic mode of action. The tomatoes fruits coated with Gt2 peptides showed growth inhibition of the artificially inoculated Salmonella cocktail, demonstrating their preservative potential.

Optimization of plant hormonal balance by microorganisms prevents plant heavy metal accumulation.

Heavy metal contamination is a threat to global food safety. Reducing heavy metal uptake in plants is a promising way to make plants safer, yet breeding the right set of traits can be tedious. We test whether microorganisms are able to impact the plant's hormonal balance hereby helping to manage plant heavy metal uptake. We focus on ethylene, a plant hormone regulating plant stress tolerance and nutrition. We grew three phylogenetically distinct plants, Rumex palustris, Alcea aucheri and Arabidopsis thaliana, on a cadmium-spiked soil. Plants roots were coated with the bacterium Pseudomonas putida UW4, which degrades the precursor of ethylene, or an isogenic ACC deaminase-deficient mutant lacking this ability. We followed ethylene concentrations, plant growth and cadmium uptake. Wildtype bacteria reduced shoot cadmium concentration by up to 35% compared to the control, while the mutant increased cadmium concentration. This effect was linked to ethylene, which was consistently positively correlated with cadmium concentration. We therefore propose that bacteria modulating plant hormonal balance may offer new possibilities to improve specific aspects of plant phenotype, in the present context reducing heavy metal. They may thus pave the way for new strategies to improve food safety in a context of the widespread soil contamination.

Interspecies-cooperations of abutilon theophrasti with root colonizing microorganisms disarm BOA-OH allelochemicals.

A facultative, microbial micro-community colonizing roots of Abutilon theophrasti Medik. supports the plant in detoxifying hydroxylated benzoxazolinones. The root micro-community is composed of several fungi and bacteria with Actinomucor elegans as a dominant species. The yeast Papiliotrema baii and the bacterium Pantoea ananatis are actively involved in the detoxification of hydroxylated benzoxazolinones by generating H2O2. At the root surface, laccases, peroxidases and polyphenol oxidases cooperate for initiating polymerization reactions, whereby enzyme combinations seem to differ depending on the hydroxylation position of BOA-OHs. A glucosyltransferase, able to glucosylate the natural benzoxazolinone detoxification intermediates BOA-5- and BOA-6-OH, is thought to reduce oxidative overshoots by damping BOA-OH induced H2O2 generation. Due to this detoxification network, growth of Abutilon theophrasti seedlings is not suppressed by BOA-OHs. Polymer coats have no negative influence. Alternatively, quickly degradable 6-hydroxy-5-nitrobenzo[d]oxazol-2(3H)-one can be produced by the micro-community member Pantoea ananatis at the root surfaces. The results indicate that Abutilon theophrasti has evolved an efficient strategy by recruiting soil microorganisms with special abilities for different detoxification reactions which are variable and may be triggered by the allelochemical´s structure and by environmental conditions.

Molecular profiling and bioactive potential of an endophytic fungus Aspergillus sulphureus isolated from Sida acuta: a medicinal plant.

CONTEXT: Sida acuta Burm.f. (Malvaceae) extracts are reported to have applications against malaria, diuretic, antipyretic, nervous and urinary diseases. No fungal endophytes of S. acuta are reported. OBJECTIVE: Isolation, identification and evaluation of antibacterial, antioxidant, anticancer and haemolytic potential of fungal endophytes from the ethnomedcinal plant S. acuta. MATERIALS AND METHODS: Sida acuta stem segments were placed on PDA medium to isolate endophytic fungi. The fungus was identified by genomic DNA analysis and phylogenetic tree was constructed using ITS sequences (GenBank) to confirm species. The antibacterial efficacy of Aspergillus sulphureus MME12 ethyl acetate extract was tested against Gram-positive and Gram-negative pathogenic bacteria. DPPH free radical scavenging activity, anticancer and DNA fragmentation against EAC cells, and direct haemolytic activity (100-500 µg/mL) using human erythrocytes were determined. RESULTS AND DISCUSSION: The ethyl acetate extract of A. sulphureus (Fresen.) Wehmer (Trichocomaceae) demonstrated significant antibacterial potential against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Salmonella typhi compared to streptomycin. MIC against test pathogens was in the range of 15.6-62.5 µg/mL. The antioxidant results revealed significant RSA from 12.43% to 62.02% (IC50 = 350.4 µg/mL, p ≤ 0.05). MME12 offered considerable inhibition of EAC proliferation (23% to 84%, IC50 = 216.7 µg/mL, p ≤ 0.05) supported by DNA fragmentation studies. The extract also offered insignificant haemolysis (5.6%) compared to Triton X-100. CONCLUSIONS: A single endophytic fungus, A. sulphureus MME12 was isolated and identified using molecular profiling. The above-mentioned findings support the pharmacological application of A. sulphureus MME12 extract and demand for purification of the active principle(s).

Selection of Lactic Acid Bacteria with Probiotic Potential Isolated from the Fermentation Process of "Cupuaçu" (Theobroma grandiflorum).

In the present study, nine lactic acid bacteria isolated from the fermentation process of "cupuaçu" (Theobroma grandiflorum) were selected for probiotic use. In vitro (resistance to gastrointestinal environment, in vitro antagonism and co-aggregation with pathogens) and in vivo (intestinal colonization and ex vivo antagonism in germ-free mice, cumulative mortality, translocation to liver and spleen, histopathological examination of liver and ileum and mRNA cytokine gene expression during an experimental infection with S. Typhimurium) assays were used. Among the nine Lactobacillus strains isolated from the "cupuaçu" fermentation, L. plantarum 81 and L. plantarum 90 were selected as potential probiotics based on better results obtained in in vitro evaluations (production of diffusible inhibitory compounds and co-aggregation) as well as in vivo experiments (resistance to gastrointestinal environment, ex vivo antagonism, higher survival after enteropathogen challenge, lower hepatic translocation of enteropathogen, lower histopathological lesions in ileum and liver and anti-inflammatory pattern of immunological response). Concluding, L. plantarum 81 and L. plantarum 90 showed in vitro and in vivo capacities for probiotic use through different mechanisms of protection and its origin would allow an easier adaptation in an alimentary matrix for its administration.

Antibacterial compound produced by Pseudomonas aeruginosa strain UICC B-40, an endophytic bacterium isolated from Neesia altissima.

This study's aim was to determine the identity of antibacterial compounds produced by Pseudomonas aeruginosa strain UICC B-40 and describe the antibacterial compounds' mechanisms of action for damaging pathogenic bacteria cells. Isolation and identification of the compounds were carried out using thin layer chromatography (TLC), nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography mass spectrometry (LC-MS) analyses. Antibacterial activity was assayed via minimum inhibitory concentration (MIC) and the antibacterial compound mechanism was observed morphologically through scanning electron microscopy (SEM). This study successfully identified the (2E,5E)-phenyltetradeca-2,5-dienoate antibacterial compound (molecular weight 300 g/mol), composed of a phenolic ester, fatty acid and long chain of aliphatic group structures. MIC values for this compound were determined at 62.5 µg/ml against Staphylococcus aureus strain ATCC 25923. The mechanism of the compound involved breaking down the bacterial cell walls through the lysis process. The (2E,5E)-phenyltetradeca-2,5-dienoate compound exhibited inhibitory activity on the growth of Gram-positive bacteria.

Cultivable endophytic bacteria from heavy metal(loid)-tolerant plants.

To evaluate the interactions among endophytes, plants and heavy metal/arsenic contamination, root endophytic bacteria of Prosopis laevigata (Humb and Bonpl. ex Willd) and Sphaeralcea angustifolia grown in a heavy metal(loid)-contaminated zone in San Luis Potosi, Mexico, were isolated and characterized. Greater abundance and species richness were found in Prosopis than in Sphaeralcea and in the nutrient Pb-Zn-rich hill than in the poor nutrient and As-Cu-rich mine tailing. The 25 species identified among the 60 isolates formed three groups in the correspondence analysis, relating to Prosopis/hill (11 species), Prosopis/mine tailing (4 species) and Sphaeralcea/hill (4 species), with six species ungrouped. Most of the isolates showed high or extremely high resistance to arsenic, such as ≥100 mM for As(V) and ≥20 mM for As(III), in mineral medium. These results demonstrated that the abundance and community composition of root endophytic bacteria were strongly affected by the concentration and type of the heavy metals and metalloids (arsenic), as well as the plant species.

Abutilon theophrasti's defense against the allelochemical benzoxazolin-2(3H)-one: support by Actinomucor elegans.

Abutilon theophrasti Medik., previously found to be rather insensitive to benzoxazinoid containing rye mulch and the allelochemical benzoxazolin-2(3H)-one (BOA), can be associated with the zygomycete Actinomucor elegans, whereby the fungus colonizes the root relatively superficially and mainly in the maturation zone. The fungus mitigates necrosis of the cotyledons when seedlings are incubated with 2 mM BOA, in contrast to those that lack the fungus. In liquid cultures of the fungus, tryptophan was identified. The accumulation of tryptophan is increased in presence of BOA. This amino acid seems to be important in protecting Abutilon against BOA and its derivatives since it suppressed the accumulation of BOA derived, highly toxic 2-aminophen-oxazin-3-one (APO) in the medium and on the root surface during BOA incubations of Abutilon seedlings. Although A. elegans is insensitive to BOA and APO, the fungus is not able to protect the plant against harmful effects of APO, when seedlings are treated with the compound. Abutilon can detoxify BOA via BOA-6-OH glucosylation probably by a cell wall associated glucosyltransferase, but only low amounts of the product accumulate. Low tryptophan concentrations can contribute to a degradation of the toxic intermediate BOA-6-OH by Fenton reactions, whereby the amino acid is oxidized. One of the oxidation products was identified as 4(1H)-quinolinone, which is the core substructure of the quorum sensing molecule 2-heptyl-3-hydroxy-4-quinolone. The mutualistic association of Abutilon theophrasti with Actinomucor elegans is considered as opportunistic and facultative. Such plant-fungus associations depend rather likely on environmental conditions, such as the mode of fertilization.

Molecular characterization by amplified ribosomal DNA restriction analysis and antimicrobial potential of endophytic fungi isolated from Luehea divaricata (Malvaceae) against plant pathogenic fungi and pathogenic bacteria.

Luehea divaricata is an important plant in popular medicine; it is used for its depurative, anti-inflammatory, and other therapeutic activities. We evaluated the antimicrobial activity of endophytic fungi isolated from leaves of L. divaricata against phytopathogens and pathogenic bacteria, and characterized the isolates based on amplified ribosomal DNA restriction analysis (ARDRA). The in vitro antagonistic activity of these endophytes against the phytopathogen Alternaria alternata was assayed by dual culture technique. Based on this evaluation of antimicrobial activity, we extracted secondary metabolites from nine endophytic fungi by partitioning in ethyl acetate and methanol. These were tested against the phytopathogens A. alternata, Colletotrichum sp and Moniliophthora perniciosa, and against the human pathogenic bacteria Escherichia coli and Staphylococcus aureus. Molecular characterization by ARDRA technique was used for phylogenetic analysis, based on comparison with sequences in GenBank. The endophytes had varied effects on A. alternata. One isolate produced an inhibition halo against M. perniciosa and against E. coli. This antibiosis activity indicates a role in the protection of the plant against microbial pathogens in nature, with potential for pharmaceutical and agricultural applications. Based on ARDRA, the 13 isolates were grouped. We found three different haplotypes of Phomopsis sp, showing interspecific variability. It appears that examination of the microbial community associated with medicinal plants of tropical regions has potential as a useful strategy to look for species with biotechnological applications.

Root and arbuscular mycorrhizal mycelial interactions with soil microorganisms in lowland tropical forest.

Tropical forests have high rates of soil carbon cycling, but little information is available on how roots, arbuscular mycorrhizal fungi (AMF), and free-living microorganisms interact and influence organic matter mineralization in these ecosystems. We used mesh ingrowth cores and isotopic tracers in phospholipid fatty acid biomarkers to investigate the effects of roots and AMF mycelia on (1) microbial community composition, microbial carbon utilization, and hydrolytic enzyme activities for large, potted tropical trees and (2) enzyme activities and litter mass loss in a lowland tropical forest. Under the tropical tree, plant-derived carbon was incorporated predominantly into bacterial groups in both rhizosphere and AMF-only soils. Gram-positive bacteria incorporated additional soil-derived carbon in rhizosphere soils, which also contained the highest microbial biomass. For hydrolytic enzymes, ß-glucosidase and N-acetyl ß-glucosaminidase activities were highest in rhizosphere soils, while phosphomonoesterase activity was highest in AMF-only soil. In the forest, leaf litter mass loss was increased by the presence of roots, but not by the presence of AMF mycelia only. Root-microbial interactions influenced organic matter cycling, with evidence for rhizosphere priming and accelerated leaf litter decomposition in the presence of roots. Although AMF mycelia alone did not stimulate organic matter mineralization, they were a conduit of carbon to other soil microorganisms.

Impact of weed control on arbuscular mycorrhizal fungi in a tropical agroecosystem: a long-term experiment.

Cover crop species represent an affordable and effective weed control method in agroecosystems; nonetheless, the effect of its use on arbuscular mycorrhizal fungi (AMF) has been scantily studied. The goal of this study was to determine root colonization levels and AMF species richness in the rhizosphere of maize plants and weed species growing under different cover crop and weed control regimes in a long-term experiment. The treatment levels used were (1) cover of Mucuna deeringian (Muc), (2) "mulch" of Leucaena leucocephala (Leu), (3) "mulch" of Lysiloma latisiliquum (Lys), (4) herbicide (Her), (5) manual weeding (CD), (6) no weeding (SD), and (7) no maize and no weeding (B). A total of 18 species of AMF belonging to eight genera (Acaulospora, Ambispora, Claroideoglomus, Funneliformis, Glomus, Rhizophagus, Sclerocystis, and Scutellospora) were identified from trap cultures. Muc and Lys treatments had a positive impact on AMF species richness (11 and seven species, respectively), while Leu and B treatments on the other hand gave the lowest richness values (six species each). AMF colonization levels in roots of maize and weeds differed significantly between treatment levels. Overall, the use of cover crop species had a positive impact on AMF species richness as well as on the percentage of root colonized by AMF. These findings have important implications for the management of traditional agroecosystems and show that the use of cover crop species for weed control can result in a more diverse AMF community which should potentially increase crop production in the long run.

Expression of Xylella fastidiosa fimbrial and afimbrial proteins during biofilm formation.

Complete sequencing of the Xylella fastidiosa genome revealed characteristics that have not been described previously for a phytopathogen. One characteristic of this genome was the abundance of genes encoding proteins with adhesion functions related to biofilm formation, an essential step for colonization of a plant host or an insect vector. We examined four of the proteins belonging to this class encoded by genes in the genome of X. fastidiosa: the PilA2 and PilC fimbrial proteins, which are components of the type IV pili, and XadA1 and XadA2, which are afimbrial adhesins. Polyclonal antibodies were raised against these four proteins, and their behavior during biofilm development was assessed by Western blotting and immunofluorescence assays. In addition, immunogold electron microscopy was used to detect these proteins in bacteria present in xylem vessels of three different hosts (citrus, periwinkle, and hibiscus). We verified that these proteins are present in X. fastidiosa biofilms but have differential regulation since the amounts varied temporally during biofilm formation, as well as spatially within the biofilms. The proteins were also detected in bacteria colonizing the xylem vessels of infected plants.

Evaluation of foliar fungal endophyte diversity and colonization of medicinal plant Luehea divaricata (Martius et Zuccarini).

Endophyte microorganisms are organisms that live inside plants without causing any apparent damage to their hosts. Since all plants exhibit endophyte microorganisms, it is believed that mutual association is of great importance in nature. Luehea divaricata (Martius & Zuccarini), known popularly in Brazil as agoita-cavalo, is a big-sized tree with a wide distribution in the country that possesses medicinal qualities for: dysentery, leucorrhea, rheumatism, blennorrhoea, tumors, bronchitis, and depuration. This research aims at isolating and molecularly characterizing fungi isolates from L. divaricata by sequence analysis of ITS1-5.8S-ITS2 rDNA. Further, the colonization of endophyte in the host plant by Light and Scanning Electron Microscopy will also be investigated. Whereas, genera Alternaria, Cochliobolus, Diaporthe, Epicoccum, Guignardia, Phoma, and Phomopsis, were identified; rDNA sequence analysis revealed intra-species variability among endophyte isolates of the genus Phomopsis sp. Light and Scanning Electron Microscopy techniques showed the presence of endophyte fungi inside L. divaricata leaves, inhabiting inter- and intra-cellular spaces. These types of extensive colonization and dissemination were reported throughout all the leaf parts in palisade parenchyma, esclerenchyma, spongy parenchyma, adaxial epidermis, and vascular bundle indicating colonization of endophytes in múltiple structural sub-niches in the host plant.

Evaluation of foliar fungal endophyte diversity and colonization of medicinal plant Luehea divaricata (Martius et Zuccarini)

Endophyte microorganisms are organisms that live inside plants without causing any apparent damage to their hosts. Since all plants exhibit endophyte microorganisms, it is believed that mutual association is of great importance in nature. Luehea divaricata (Martius & Zuccarini), known popularly in Brazil as agoita-cavalo, is a big-sized tree with a wide distribution in the country that possesses medicinal qualities for: dysentery, leucorrhea, rheumatism, blennorrhoea, tumors, bronchitis, and depuration. This research aims at isolating and molecularly characterizing fungi isolates from L. divaricata by sequence analysis of ITS1-5.8S-ITS2 rDNA. Further, the colonization of endophyte in the host plant by Light and Scanning Electron Microscopy will also be investigated. Whereas, genera Alternaria, Cochliobolus, Diaporthe, Epicoccum, Guignardia, Phoma, and Phomopsis, were identified; rDNA sequence analysis revealed intra-species variability among endophyte isolates of the genus Phomopsis sp. Light and Scanning Electron Microscopy techniques showed the presence of endophyte fungi inside L. divaricata leaves, inhabiting inter- and intra-cellular spaces. These types of extensive colonization and dissemination were reported throughout all the leaf parts in palisade parenchyma, esclerenchyma, spongy parenchyma, adaxial epidermis, and vascular bundle indicating colonization of endophytes in múltiple structural sub-niches in the host plant.

Bacterial population structure of the jute-retting environment.

Jute is one of the most versatile bast fibers obtained through the process of retting, which is a result of decomposition of stalks by the indigenous microflora. However, bacterial communities associated with the retting of jute are not well characterized. To investigate the presence of microorganisms during the process of jute retting, full-cycle rRNA approach was followed, and two 16S rRNA gene libraries, from jute-retting locations of Krishnanagar and Barrackpore, were constructed. Phylotypes affiliating to seven bacterial divisions were identified in both libraries. The bulk of clones came from Proteobacteria ( approximately 37, 41%) and a comparatively smaller proportion of clones from the divisions-Firmicutes ( approximately 11, 12%), Cytophaga-Flexibacter-Bacteroidetes group (CFB; approximately 9, 7%), Verrucomicrobia ( approximately 6, 5%), Acidobacteria ( approximately 4, 5%), Chlorobiales ( approximately 5, 5%), and Actinobacteria ( approximately 4, 2%) were identified. Percent coverage value and diversity estimations of phylotype richness, Shannon-Weiner index, and evenness confirmed the diverse nature of both the libraries. Evaluation of the retting waters by whole cell rRNA-targeted flourescent in situ hybridization, as detected by domain- and group-specific probes, we observed a considerable dominance of the beta-Proteobacteria (25.9%) along with the CFB group (24.4%). In addition, 32 bacterial species were isolated on culture media from the two retting environments and identified by 16S rDNA analysis, confirming the presence of phyla, Proteobacteria ( approximately 47%), Firmicutes ( approximately 22%), CFB group ( approximately 19%), and Actinobacteria ( approximately 13%) in the retting niche. Thus, our study presents the first quantification of the dominant and diverse bacterial phylotypes in the retting ponds, which will further help in improving the retting efficiency, and hence the fiber quality.

Distribution of mosquito (Diptera: Culicidae) species and Wolbachia (Rickettsiales: Rickettsiaceae) infections during the bird immigration season in Pathumthani province, central Thailand.

Mosquito distribution in the immigration bird-nested area, Pathumthani province, was investigated from August to December in 2006. Mosquitoes were collected by using CO2-baited Centers for Disease Control light traps in which dry ice was used as a source of CO2 to attract mosquitoes. Six traps were operated from 4 p.m. until 7 a.m. on each study day. Four genera, which were Anopheles, Armigeres, Culex, and Mansonia with 14 species of mosquitoes were collected. Culex gelidus (13.94-59.41%) and Culex tritaeniorhynchus (32.87-70.30%) were most collected species in this area for every month. Other two species with moderate distribution in this area were Anopheles barbirostris (0.76-3.30%) and Mansonia uniformis (1.55-11.36%). Polymerase Chain Reactions were performed for testing Wolbachia infection in Cx. gelidus and Cx. tritaeniorhynchus only. Fifty-four percent (15/28 pools) of Cx. gelidus and none (0/20 pools) of Cx. tritaeniorhynchus were positive for Wolbachia infection. Wolbachia infection in other mosquito species collected in this and other areas need to be investigated to understand species and geographic variation of Wolbachia infection in mosquitoes in nature.

Muscodor albus E-6, an endophyte of Guazuma ulmifolia making volatile antibiotics: isolation, characterization and experimental establishment in the host plant.

Muscodor albus is an endophytic fungus, represented by a number of isolates from tropical tree and vine species in several of the world's rainforests, that produces volatile organic compounds (VOCs) with antibiotic activity. A new isolate, E-6, of this organism, with unusual biochemical and biological properties, has been obtained from the branches of a mature Guazuma ulmifolia (Sterculiaceae) tree growing in a dry tropical forest in SW Ecuador. This unique organism produces many VOCs not previously observed in other M. albus isolates, including butanoic acid, 2-methyl-; butanoic acid, 3-methyl-; 2-butenal, 2-methyl-; butanoic acid, 3-methylbutyl ester; 3-buten-1-ol, 3-methyl; guaiol; 1-octene, 3-ethyl-; formamide, N-(1-methylpropyl); and certain azulene and naphthalene derivatives. Some compounds usually seen in other M. albus isolates also appeared in the VOCs of isolate E-6, including caryophyllene; phenylethyl alcohol; acetic acid, 2-phenylethyl ester; bulnesene; and various propanoic acid, 2-methyl- derivatives. The biological activity of the VOCs of E-6 appears different from the original isolate of this fungus, CZ-620, since a Gram-positive bacterium was killed, and Sclerotinia sclerotiorum and Rhizoctonia solani were not. Scanning electron micrographs of the mycelium of isolate E-6 showed substantial intertwining of the hyphal strands. These strands seemed to be held together by an extracellular matrix accounting for the strong mat-like nature of the mycelium, which easily lifts off the agar surface upon transfer, unlike any other isolate of this fungus. The ITS-5.8S rDNA partial sequence data showed 99 % similarity to the original M. albus strain CZ-620. For the first time, successful establishment of M. albus into its natural host, followed by recovery of the fungus, was accomplished in seedlings of G. ulmifolia. Overall, isolates of M. albus, including E-6, have chemical, biological and structural characteristics that make them potentially useful in medicine, agricultural and industrial applications.