Fungal diversity associated with Goa's tarballs: Insights from ITS region amplicon sequencing.

This study explores the fungal diversity associated with tarballs, weathered crude oil deposits, on Goa's tourist beaches. Despite tarball pollution being a longstanding issue in Goa state in India, comprehensive studies on associated fungi are scarce. Our research based on amplicon sequence analysis of fungal ITS region fills this gap, revealing a dominance of Aspergillus, particularly Aspergillus penicillioides, associated with tarballs from Vagator and Morjim beaches. Other notable species, including Aspergillus sydowii, Aspergillus carbonarius, and Trichoderma species, were identified, all with potential public health and ecosystem implications. A FUNGuild analysis was conducted to investigate the potential ecological roles of these fungi, revealing a diverse range of roles, including nutrient cycling, disease propagation, and symbiotic relationships. The study underscores the need for further research and monitoring, given the potential health risks and contribution of tarball-associated fungi to the bioremediation of crude oil-contaminated beaches.

Metabolism of 4-chloro-2-nitrophenol in a gram-positive bacterium, Exiguobacterium sp. PMA.

BACKGROUND: Chloronitrophenols (CNPs) are widely used in the synthesis of dyes, drugs and pesticides, and constitute a major group of environmental pollutants. 4-Chloro-2-nitrophenol (4C2NP) is an isomer of CNPs that has been detected in various industrial effluents. A number of physicochemical methods have been used for treatment of wastewater containing 4C2NP. These methods are not as effective as microbial degradation, however. RESULTS: A 4C2NP-degrading bacterium, Exiguobacterium sp. PMA, which uses 4C2NP as the sole carbon and energy source was isolated from a chemically-contaminated site in India. Exiguobacterium sp. PMA degraded 4C2NP with the release of stoichiometeric amounts of chloride and ammonium ions. The effects of different substrate concentrations and various inoculum sizes on degradation of 4C2NP were investigated. Exiguobacterium sp. PMA degraded 4C2NP up to a concentration of 0.6 mM. High performance liquid chromatography and gas chromatography-mass spectrometry identified 4-chloro-2-aminophenol (4C2AP) and 2-aminophenol (2AP) as possible metabolites of the 4C2NP degradation pathway. The crude extract of 4C2NP-induced PMA cells contained enzymatic activity for 4C2NP reductase and 4C2AP dehalogenase, suggesting the involvement of these enzymes in the degradation of 4C2NP. Microcosm studies using sterile and non-sterile soils spiked with 4C2NP were carried out to monitor the bioremediation potential of Exiguobacterium sp. PMA. The bioremediation of 4C2NP by Exiguobacterium sp. PMA was faster in non-sterilized soil than sterilized soil. CONCLUSIONS: Our studies indicate that Exiguobacterium sp. PMA may be useful for the bioremediation of 4C2NP-contaminated sites. This is the first report of (i) the formation of 2AP in the 4C2NP degradation pathway by any bacterium and (iii) the bioremediation of 4C2NP by any bacterium.

Metaxa: a software tool for automated detection and discrimination among ribosomal small subunit (12S/16S/18S) sequences of archaea, bacteria, eukaryotes, mitochondria, and chloroplasts in metagenomes and environmental sequencing datasets.

The ribosomal small subunit (SSU) rRNA gene has emerged as an important genetic marker for taxonomic identification in environmental sequencing datasets. In addition to being present in the nucleus of eukaryotes and the core genome of prokaryotes, the gene is also found in the mitochondria of eukaryotes and in the chloroplasts of photosynthetic eukaryotes. These three sets of genes are conceptually paralogous and should in most situations not be aligned and analyzed jointly. To identify the origin of SSU sequences in complex sequence datasets has hitherto been a time-consuming and largely manual undertaking. However, the present study introduces Metaxa ( http://microbiology.se/software/metaxa/ ), an automated software tool to extract full-length and partial SSU sequences from larger sequence datasets and assign them to an archaeal, bacterial, nuclear eukaryote, mitochondrial, or chloroplast origin. Using data from reference databases and from full-length organelle and organism genomes, we show that Metaxa detects and scores SSU sequences for origin with very low proportions of false positives and negatives. We believe that this tool will be useful in microbial and evolutionary ecology as well as in metagenomics.

Degradation of tarballs using associated bacterial consortia.

Tarballs are semi-solid blobs of crude-oil formed in marine environment. Microbial degradation of tarballs is poorly understood, though there are indications that tarball-associated microbes can degrade recalcitrant hydrocarbons present in tarballs. In this study, 38 tarball-associated bacteria from Betul beach, Goa, India were initially screened for crude oil degradation. Based on preliminary studies and literature survey, four bacterial strains, Alcanivorax sp. Betul-O, Marinobacter sp. Betul-26, Pseudomonas sp. Betul-14, and Pseudomonas sp. Betul-M were selected for bacterial consortia preparation. Eleven bacterial consortia were prepared and studied for degradation of n-alkanes and polycyclic aromatic hydrocarbon compounds (PAHs) of tarballs based on gravimetric and GC-MS-MS analyses. The bacterial consortia depleted 53.69-97.78% and 22.78-61.98% of n-alkanes and PAH compounds, respectively, within 45 days. Bacterial consortium comprising Pseudomonas sp. Betul-14, Pseudomonas sp. Betul-M, and Alcanivorax sp. Betul-O exhibited promising tarball degradation abilities with 97.78% and 61.98% degradation of n-alkanes and PAH, respectively, within 45 days. Further research is required to obtain insights into degradation products and possible pathways involved.

High abundance of Vibrio in tarball-contaminated seawater from Vagator beach, Goa, India.

Tarballs are semi-solid remnants of crude oil and they are formed in marine environment after oil-spill incidents. They are composed of diverse hydrocarbons; some of which are recalcitrant in nature. Recent studies based on amplicon sequencing of 16S rRNA gene suggested that tarballs support hundreds of bacterial genera and provided insights into their role as hydrocarbon degraders and potential human pathogens. In this study, bacterial composition of tarball-contaminated seawater from Vagator beach, Goa, India was characterized by amplicon sequencing of V3-V4 regions of 16S rRNA gene. The DNA data revealed an unusual surge of Vibrio in sea-water contaminated with tarballs in May 2018 (16.16% OTUs), compared to tarball-free seawater samples collected in March 2018 (no detectable OTUs) and September 2018 (0.17% OTUs). Further studies are required to investigate if Vibrio species form biofilms on tarballs which may act as good reservoirs for their survival and transmission success.

Diversity of Bacterial Community in the Oxygen Minimum Zones of Arabian Sea and Bay of Bengal as Deduced by Illumina Sequencing.

The Indian Ocean harbors oxygen minimum zones (OMZs) in the Arabian Sea and Bay of Bengal, with dissolved oxygen < 20 µM, located at the mid-depths of the water column. Till date, high-throughput sequence-data on depth-wise distribution of prokaryotic communities have rarely been reported from these OMZs. The present study aimed to characterize the prokaryotic diversity inhabiting Arabian Sea Time Series (ASTS) and India's Idea 2 (II2) in the Arabian Sea, and Bay of Bengal Time Series (BoBTS) in the Bay of Bengal OMZs based on amplicon sequencing of 16S rRNA gene regions, along six sampled depths in the water column. High prokaryotic richness was observed in the Arabian Sea and Bay of Bengal samples. Operational taxonomic units (OTUs) in the range of 1249-3298 were identified, wherein, less prokaryotic diversity was observed at surface and within oxygen minimum depths. At phylum level, most OTUs were affiliated to Bacteroidetes, Chloroflexi, Cyanobacteria, Marinimicrobia, Planctomycetes, and Proteobacteria. Prokaryotic community differed between ASTS, II2 and BoBTS locations along varying physicochemical conditions. Predictive functional profiling of the bacterial communities suggested the involvement of abundant microbes in nitrogen and sulfur metabolism pathways. Bacterial isolates belonging to genera from the clades, δ-Proteobacteria and γ-Proteobacteria, described previously for their participation in biogeochemical cycling of N-and-S in the OMZs were reported from deoxygenated waters of both the basins. Bacteria involved in anammox such as Candidatus Scalindua were found to be relatively high at ASTS and II2 locations in the Arabian Sea. Further studies are required to ascertain the role of abundant bacteria along the dynamic oceanographic processes in the OMZs.

Metagenomic analysis of tarball-associated bacteria from Goa, India.

The beaches of Goa state in India are frequently polluted with tarballs, specifically during pre-monsoon and monsoon seasons. Tarballs contain hydrocarbons, including polycyclic aromatic hydrocarbons, which pose significant environmental risks. Microbes associated with tarballs reportedly possess capabilities to degrade toxic hydrocarbons present in tarballs. In this study, bacterial diversity associated with tarballs from Vagator and Morjim beaches of north Goa was analysed based on V3-V4 regions of 16S rRNA gene sequenced using Illumina Miseq Platform. The Proteobacterial members were dominant in both Vagator (≥85.5%) and Morjim (≥94.0%) samples. Many of the identified taxa have been previously reported as hydrocarbon degraders (e.g. Halomonas, Marinobacter) or possible human pathogens (e.g. Acinetobacter, Klebsiella, Rhodococcus, Staphylococcus, Vibrio). This is the first study reported on a metagenomic analysis of bacteria associated with tarballs from Goa.

Diversity and extracellular enzyme activities of heterotrophic bacteria from sediments of the Central Indian Ocean Basin.

Sedimentary bacteria play a role in polymetallic nodule formation and growth. There are, however, limited reports on bacterial diversity in nodule-rich areas of the Central Indian Ocean Basin (CIOB). In this study, bacterial abundance in thirteen sediment cores collected from the CIOB was enumerated, followed by phylogenetic characterisation and, screening of select heterotrophic bacteria for extracellular enzyme activities. Total bacterial counts (TBC) were in the order of 107 cells g-1; there was a significant difference (p > 0.05) among the cores but not within the sub-sections of the cores. The retrievable heterotrophic counts ranged from non-detectable to 5.33 × 105 g-1; the heterotrophic bacteria clustered within the phyla Firmicutes, Proteobacteria and Actinobacteria. Bacillus was the most abundant genus. The extracellular enzyme activities were in the order: amylase > lipase > protease > phosphatase > Dnase > urease. Major findings are compared with previous studies from the CIOB and other areas.

Phylogenetic characterization of culturable bacteria and fungi associated with tarballs from Betul beach, Goa, India.

Tarballs are semisolid blobs of crude oil, normally formed due to weathering of crude-oil in the sea after any kind of oil spills. Microorganisms are believed to thrive on hydrocarbon-rich tarballs and possibly assist in biodegradation. The taxonomy of ecologically and economically important tarball-associated microbes, however, needs improvement as DNA-based identification and phylogenetic characterization have been scarcely incorporated into it. In this study, bacteria and fungi associated with tarballs from touristic Betul beach in Goa, India were isolated, followed by phylogenetic analyses of 16S rRNA gene and the ITS sequence-data to decipher their clustering patterns with closely-related taxa. The gene-sequence analyses identified phylogenetically diverse 20 bacterial genera belonging to the phyla Proteobacteria (14), Actinobacteria (3), Firmicutes (2) and Bacteroidetes (1), and 8 fungal genera belonging to the classes Eurotiomycetes (6), Sordariomycetes (1) and Leotiomycetes (1) associated with the Betul tarball samples. Future studies employing a polyphasic approach, including multigene sequence-data, are needed for species-level identification of culturable tarball-associated microbes. This paper also discusses potentials of tarball-associated microbes to degrade hydrocarbons.

Diversity of bacteria and fungi associated with tarballs: Recent developments and future prospects.

Tarballs are formed by weathering of crude oil in marine environment. They are transported from open ocean to the shores by sea currents and waves. Tarball pollution is a major concern to global marine ecosystem. Microbes such as bacteria and fungi are known to be associated with tarballs. They presumably play an important role in tarball degradation and some are potential human and animal pathogens. This paper highlights the recent studies on tarball-associated bacteria and fungi. Future perspectives on diversity, ecology and possible applications of tarball-associated microbes in bioremediation of beached tarballs have been discussed.

Molecular phylogeny of Neodevriesia, with two new species and several new combinations.

To identify Cladosporium-like isolates associated with marine algae, phylogenetic analyses were conducted using a combined data set of the nuc rDNA internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2 = ITS), 28S gene, and partial RNA polymerase II second largest subunit (RPB2). These isolates were confirmed as belonging to Neodevriesia (Neodevriesiaceae), a genus recently segregated from Cladosporium. Two new species, Neodevriesia cladophorae and N. grateloupiae, are described on the basis of morphological and phylogenetic distinctions, and this is the first report of Neodevriesia species from the marine environment. Devriesia fici, D. fraserae, and D. sardiniae are transferred to Neodevriesia based on the phylogenetic affinities and morphological characters of their ex-type specimens.

The ITS-based phylogeny of fungi associated with tarballs.

Tarballs, the remnants of crude oil which change into semi-solid phase due to various weathering processes in the sea, are rich in hydrocarbons, including toxic and almost non-degradable hydrocarbons. Certain microorganisms such as fungi are known to utilize hydrocarbons present in tarballs as sole source of carbon for nutrition. Previous studies have reported 53 fungal taxa associated with tarballs. There is apparently no gene sequence-data available for the published taxa so as to verify the fungal identification using modern taxonomic tools. The objective of the present study is to isolate fungi from tarballs collected from Candolim beach in Goa, India and investigate their phylogenetic diversity based on 5.8S rRNA gene and the flanking internal transcribed spacer regions (ITS) sequence analysis. In the ITS-based NJ tree, eight tarball-associated fungal isolates clustered with 3 clades of Dothideomycetes and 2 clades of Saccharomycetes. To the best of our knowledge, this is the first study that has employed ITS-based phylogeny to characterize the fungal diversity associated with tarballs. Further studies are warranted to investigate the role of the tarball-associated fungi in degradation of recalcitrant hydrocarbons present in tarballs and the role of tarballs as carriers of human pathogenic fungi.

Genome sequence, comparative analysis, and evolutionary insights into chitinases of entomopathogenic fungus Hirsutella thompsonii.

Hirsutella thompsonii (Ht) is a fungal pathogen of acarines and the primary cause of epizootics among mites. The draft genomes of two isolates of Ht (MTCC 3556: Ht3, 34.6 Mb and MTCC 6686: Ht6, 34.7 Mb) are presented and compared with the genomes of Beauveria bassiana (Bb) ARSEF 2860 and Ophiocordyceps sinensis (Os) CO18. Comparative analysis of carbohydrate active enzymes, pathogen-host interaction genes, metabolism-associated genes, and genes involved in biosynthesis of secondary metabolites in the four genomes was carried out. Reduction in gene family sizes in Ht3 and Os as compared with Ht6 and Bb is observed. Analysis of the mating type genes in Ht reveals the presence of MAT idiomorphs which is suggestive of cryptic sexual traits in Ht. We further identify and classify putative chitinases that may function as virulence factors in fungal entomopathogens due to their role in degradation of arthropod cuticle.

Stimulatory effects of arsenic-tolerant soil fungi on plant growth promotion and soil properties.

Fifteen fungi were obtained from arsenic-contaminated agricultural fields in West Bengal, India and examined for their arsenic tolerance and removal ability in our previous study. Of these, the four best arsenic-remediating isolates were tested for plant growth promotion effects on rice and pea in the present study. A greenhouse-based pot experiment was conducted using soil inocula of individual fungi. The results indicated a significant (P<0.05) increase in plant growth and improvement of soil properties in inoculated soils compared to the control. A significant increase in plant growth was recorded in treated soils and varied from 16-293%. Soil chemical and enzymatic properties varied from 20-222% and 34-760%, respectively, in inoculated soil. Plants inoculated with inocula of Westerdykella and Trichoderma showed better stimulatory effects on plant growth and soil nutrient availability than Rhizopus and Lasiodiplodia. These fungi improved soil nutrient content and enhanced plant growth. These fungi may be used as bioinoculants for plant growth promotion and improved soil properties in arsenic-contaminated agricultural soils.

Ribosomal and RPB2 DNA sequence analyses suggest that Sporidesmium and morphologically similar genera are polyphyletic.

Sporidesmium and morphologically similar dematiaceous, hyphomycetous genera are characterised by holoblastic phragmoconidia produced on proliferating or non-proliferating conidiophores. They include a number of asexual (anamorphic) genera taxonomically segregated from Sporidesmium sensu lato and are similar in having schizolytic conidial secession. The taxonomy of these ubiquitous asexual fungi and their affinities with known Ascomycetes are, however, still obscure. This study incorporates a phylogenetic investigation, based on the LSU nu-rDNA and RNA polymerase II second largest subunit (RPB2) gene sequence, to assess the possible familial placement of Ellisembia, Linkosia, Repetophragma, Sporidesmiella, Sporidesmium and Stanjehughesia, and justify whether anamorphic characters are proper phylogenetic indicators. Phylogenies provide conclusive evidence to suggest that Sporidesmium is not monophyletic and species are phylogenetically distributed in two major ascomycete classes, Dothideomycetes and Sordariomycetes. Morphologies currently used in their classification have undergone convergent evolution and are not phylogenetically reliable. The possible teleomorphic affinities of these anamorphic genera are discussed in light of morphology and molecular data. As these anamorphs, in most cases, are the sole known morph of the holomorph, it is proposed that in the absence of or failure to detect their teleomorphic phase, the anamorph names should be used for the holomorph.