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A method named sequence-specific capture of oligonucleotide probes (SCOPE) was developed for quantification of microbial rRNA molecules in a multiplex manner. In this method, a molecular weight cutoff membrane (MWCOM) was used for the separation of fluorescence-labeled oligonucleotide probes hybridized with rRNA from free unhybridized probes. To demonstrate proof of concept, probes targeting bacteria or archaea at different taxonomic levels were prepared and were hybridized with rRNAs. The hybridization stringency was controlled by adjusting reaction temperature and urea concentration in the mixture. Then, the mixture was filtered through the MWCOM. The rRNA and hybridized probes collected on the MWCOM were recovered and quantified using a spectrophotometer and fluorospectrometer, respectively. The method showed high accuracy in detecting specific microbial rRNA in a defined nucleic acid mixture. Furthermore, the method was capable of simultaneous detection and quantification of multiple target rRNAs in a sample with sensitivity up to a single-base mismatch. The SCOPE method was tested and benchmarked against reverse transcription-quantitative PCR (RT-qPCR) for the quantification of Bacteria , Archaea , and some key methanogens in anaerobic sludge samples. It was observed that the SCOPE method produced more reliable and coherent results. Thus, the SCOPE method allows simple and rapid detection and quantification of target microbial rRNAs for environmental microbial population analysis without any need for enzymatic reactions. IMPORTANCE Microorganisms play integral roles in the Earth's ecosystem. Microbial populations and their activities significantly affect the global nutrient cycles. Quantification of key microorganisms provides important information that is required to understand their roles in the environment. Sequence-based analysis of microbial population is a powerful tool, but it provides information only on relative abundance of microorganisms. Hence, the development of a simpler and quick method for the quantification of microorganisms is necessary. To address the shortcomings of a variety of molecular methods reported so far, we developed a simple, rapid, accurate, and multiplexed microbial rRNA quantification method to evaluate the abundance of specific microbial populations in complex ecosystems. This method demonstrated high specificity, reproducibility, and applicability to such samples. The method is useful for quantitative detection of particular microbial members in the environment.

The plastic film is extensively applied with limited recycling, leading to the long-run residue accumulation in soil, which offers a distinctive habitat for microorganisms, and creates a plastisphere. In this study, traditional low-density polyethylene (LDPE) plastic film and biodegradable polybutylene adipate terephthalate (PBAT) plastic film materials were selected to test their effects on soil microbial ecology. Based on high-throughput sequencing, compared to the soil environment, the alpha-diversity of bacterial communities in plastisphere was lower, and the abundance of Actinobacteria increased. Plastic film residues, as bacterial habitats, exhibited greater heterogeneity and harbor unique bacterial communities. The communities were distinguished between plastisphere and soil environment by means of a random-forest (RF) machine-learning model. Prominent distinctions emerged among bacterial functions between soil environment and plastisphere, especially regarding organics degradation. The neutral model and null model indicated that the constitution of bacterial communities was dominated by random processes except in LDPE plastisphere. The bacterial co-occurrence network of the plastisphere exhibited higher complexity and modularity. This study contributes to our comprehending of characteristics of plastisphere bacterial communities in soil environment and the associated ecological risks of plastic film residues accumulation., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

A pilot-scale experiment of natural rubber processing wastewater treatment was conducted using a combination system consisting of a two-stage up-flow anaerobic sludge blanket (UASB) and a down-flow hanging sponge (DHS) reactor for more than 10 months. The system achieved a chemical oxygen demand (COD) removal efficiency of 95.7% ± 1.3% at an organic loading rate of 0.8 kg COD/(m(3).d). Bacterial activity measurement of retained sludge from the UASB showed that sulfate-reducing bacteria (SRB), especially hydrogen-utilizing SRB, possessed high activity compared with methane-producing bacteria (MPB). Conversely, the acetate-utilizing activity of MPB was superior to SRB in the second stage of the reactor. The two-stage UASB-DHS system can reduce power consumption by 95% and excess sludge by 98%. In addition, it is possible to prevent emissions of greenhouse gases (GHG), such as methane, using this system. Furthermore, recovered methane from the two-stage UASB can completely cover the electricity needs for the operation of the two-stage UASB-DHS system, accounting for approximately 15% of the electricity used in the natural rubber manufacturing process.

Photo-crosslinked polymer beads were introduced into a laboratory activated-sludge unit containing municipal sewage sludge and the effect on nitrifying capacity was examined. The ammonia load started at a nitrogen-loading rate of 0.02 kg m(-3) day(-1) and was increased stepwise. It was found that the bead-containing unit could almost completely oxidize ammonia (over 95%) up to a nitrogen-loading rate of 0.216 kg m(-3) day(-1), whereas the maximum loading rate of the control unit (without polymer beads) was 0.096 kg m(-3) day(-1). The nitrifying potential of suspended and bead-associated organisms in the bead-containing unit was measured under different loading conditions. It was found that the bead-associated organisms exhibited high specific activities under high loading conditions and that the contribution of the bead-associated organisms to nitrification was greater than that of the suspended solids under these conditions. The bacterial population dynamics in the suspended solids and bead-associated organisms were analysed by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rRNA gene fragments and by fluorescence in situ hybridization with group-specific probes. Among the known nitrifying organisms, ammonia-oxidizing beta-proteobacteria and Nitrospira-related organisms were detected by these approaches. A comparison of the activity dynamics and population dynamics, however, suggested a possibility that other organisms may also have been involved in the nitrification process under high loading conditions.

In January 1997, the tanker Nakhodka sank in the Japan Sea, and more than 5000 tons of heavy oil leaked. The released oil contaminated more than 500 km of the coastline, and some still remained even by June 1999. To investigate the long-term influence of the Nakhodka oil spill on marine bacterial populations, sea water and residual oil were sampled from the oil-contaminated zones 10, 18, 22 and 29 months after the accident, and the bacterial populations in these samples were analysed by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments. The dominant DGGE bands were sequenced, and the sequences were compared with those in DNA sequence libraries. Most of the bacteria in the sea water samples were classified as the Cytophaga-Flavobacterium-Bacteroides phylum, alpha-Proteobacteria or cyanobacteria. The bacteria detected in the oil paste samples were different from those detected in the sea water samples; they were types related to hydrocarbon degraders, exemplified by strains closely related to Sphingomonas subarctica and Alcanivorax borkumensis. The sizes of the major bacterial populations in the oil paste samples ranged from 3.4 x 10(5) to 1.6 x 10(6) bacteria per gram of oil paste, these low numbers explaining the slow rate of natural attenuation.

Objective: Heterotrophic manganese (Mn)-oxidizing microorganisms responsible for biogenic manganese oxide (Bio-MnOx) production are fastidious. Their enrichment is not easily accomplished by merely adding a soluble organic substrate to non-sterile mixed cultures. The objective of this study was to evaluate polycaprolactone (PCL), an aliphatic polyester, as an effective solid organic substrate for the enrichment of marine Mn-oxidizing microorganisms., Results: We successfully obtained marine microbial enrichment with the capacity for dissolved Mn removal and MnOx production using PCL as a solid organic substrate. The removal of dissolved Mn by the Mn-oxidizing enrichment culture followed first-order kinetics with a rate constant of 0.014 h -1 . 16S rRNA gene amplicon sequencing analysis revealed that the Mn-oxidizing enrichment culture was highly dominated by operational taxonomic units related to the bacterial phyla Cyanobacteria, Planctomycetes, and Proteobacteria., Conclusions: Our data demonstrate that PCL can serve as a potential substrate to enrich Mn-oxidizing microorganisms with the ability to produce MnOx under marine conditions.

Aims: This study aimed to develop an efficient, cost-effective and eco-friendly bacterial consortium to degrade petroleum sludge., Methods and Results: Four bacterial strains belonging to genera Acinetobacter and Pseudomonas were selected to constitute three different consortia based on their initial concentration. The highest degradation rate (78%) of 1% (v/v) crude oil after 4 weeks of incubation was recorded when the concentration of biosurfactant (BS) producing isolate was high. Genes, such as alkB, almA, cyp153, pah-rhdGN, nah, phnAC and cat23 were detected using the polymerase chain reaction method and their induction levels were optimal at pH 7·0. A crude oil sludge was artificially constituted, and its bacterial composition was investigated using 16S rRNA gene amplicon sequencing. The results showed that the soil bacterial community was dominated by plant growth-promoting bacteria (PGPB) after crude oil treatment., Conclusions: Our findings indicate the decontamination of the crude oil contaminated soil was more effective in the presence of both the constituted consortium and PGPB compared to the presence of PGPB alone., Significance and Impact of the Study: This study showed that the PGPB (Taibaiella) present in petroleum uncontaminated soil can promote the soil decontamination. The addition of both efficient hydrocarbon-degrading and BS producing bacteria is also necessary to improve the decontamination., (© 2020 The Society for Applied Microbiology.)

In this study, continuous operation of a pilot-scale upflow anaerobic sludge blanket (UASB) reactor for sewage treatment was conducted for 630 days to investigate the physical and microbial characteristics of the retained sludge. The UASB reactor with a working volume of 20.2 m³ was operated at ambient temperature (16-29 °C) and seeded with digested sludge. After 180 days of operation, when the sewage temperature had dropped to 20 °C or lower, the removal efficiency of both total suspended solids (TSS) and total biochemical oxygen demand (BOD) deteriorated due to washout of retained sludge. At low temperature, the cellulose concentration of the UASB sludge increased owing to the rate limitation of the hydrolytic reaction of suspended solids in the sewage. However, after an improvement in sludge retention (settleability and concentration) in the UASB reactor, the process performance stabilized and gave sufficient results (68% of TSS removal, 75% of total BOD removal) at an hydraulic retention time (HRT) of 9.7 h. The methanogenic activity of the retained sludge significantly increased after day 246 due to the accumulation of Methanosaeta and Methanobacterium following the improvement in sludge retention in the UASB reactor. Acid-forming bacteria from phylum Bacteroidetes were detected at high frequency; thus, these bacteria may have an important role in suspended solids degradation. [ABSTRACT FROM AUTHOR]

The petrochemicals produced from refining oil have become a large part of human life, making oil a valuable and expensive commodity. As a non-renewable resource, extraction and transport efforts have intensified to keep up with the demand, increasing the occurrences of oil spills. Such accidents have devastating impacts on the environment, the health of organisms, and a country's economy like the Philippines, and thus, need to be resolved immediately. One way to deal with oil spills is through bioremediation but the process is still facing several challenges. For one, the toolbox for bioremediation is limited. About 79 genera of bacteria were observed to degrade oil but there are only a small number of bacterial species and/or strains that have been recognized as useful for bioremediation. Second is that most oil-degrading bacteria found have low oil degradation efficiencies. Another challenge is keeping the bacteria alive to carry out the process. Fortunately, progress has been made in solving these challenges. Researchers are now testing different consortia, including bacteria--bacteria, bacteria--fungi, bacteria--microalgae, that can complement each other such as biosurfactant-producing bacteria with different oil-degrading microorganisms or microalgae or fungi that enhance the growth of oil-degrading bacteria. A consortium like this improves the survivability of each microorganism and enhances the oil-degrading efficiency. Moreover, the search for additional oil-degrading and biosurfactant-producing bacteria and other microorganisms to add to the bioremediation toolbox has been improved with the emergence of high-throughput sequencing. Aside from microorganisms, seaweeds have shown potential for bioremediation. The seaweed Caulerpa prolifera has been demonstrated to degrade diesel up to a certain concentration with the help of the bacteria growing on its surface. Bioremediation has a long way to go, but recent developments have shown promise and it remains to be the cheapest, most environment-friendly, and most effective way of dealing with oil spills. [ABSTRACT FROM AUTHOR]

Tanjung Emas Port is the entry and exit point for trade commodities, both regional and international filled with many ships. This condition makes biofouling a very massive process in that place by various types of marine bacteria. The initial formation of a biofilm is relevant to bacterial diversity, colonization and adhesion. The objective of the study was assessing bacterial diversity in relation to with biofouling within Tanjung Mas Port Semarang, by using 16S rRNA eDNA metabarcoding Next-Generation Sequencing (NGS). Seawater samples from aquatic sites of Tanjung Mas harbor was used for DNA extraction and amplification of the 16S rRNA V-3-V4 hypervariable region, followed by sequencing and library construction of eDNA Metabarcoding. Sequence processing and analysis was performed in QIIME 2 and RStudio using DADA2 for advanced sequencing processing and Phyloseq. The results of this research showed that bacteria is the predominant taxon constituting 100% of the community. The taxon consists of Proteobacteria (49.38%), Bacteroidota (8.67%), and Firmicutes (8.88%). Alphaproteobacteria (20.92%) and Gammaproteobacteria (12.39%) dominate at the Class level, emphasizing their versatility and ecological influence. At the Order and Family levels reveals the prevalence of Rhodobacterales (10.04%), Chitinophagales (2.53%), Rhizobiales (3.61%), Rhodobacteraceae (5.67%), Saprospiraceae (3.63%), and Rhizobiaceae (2.15%). It was found that the unculturable taxa dominance in Tanjung Mas Port was 44.66%. These taxonomic entities contribute significantly to the taxonomic and functional diversity of the microbial community, influencing nutrient cycling, organic matter degradation, ecosystem stability and biofilm formation. [ABSTRACT FROM AUTHOR]

The oceans cover over 70% of our planet, hosting a biodiversity of tremendous wealth. Sponges are one of the major ecosystem engineers on the seafloor, providing a habitat for a wide variety of species to be considered a good source of bioactive compounds. In this study, a metataxonomic approach was employed to describe the bacterial communities of the sponges collected from Faro Lake (Sicily) and Porto Paone (Gulf of Naples). Morphological analysis and amplification of the conserved molecular markers, including 18S and 28S (RNA ribosomal genes), CO1 (mitochondrial cytochrome oxidase subunit 1), and ITS (internal transcribed spacer), allowed the identification of four sponges. Metataxonomic analysis of sponges revealed a large number of amplicon sequence variants (ASVs) belonging to the phyla Proteobacteria, Cloroflexi, Dadabacteria, and Poribacteria. In particular, Myxilla (Myxilla) rosacea and Clathria (Clathria) toxivaria displayed several classes such as Alphaproteobacteria, Dehalococcoidia, Gammaproteobacteria, Cyanobacteria, and Bacteroidia. On the other hand, the sponges Ircinia oros and Cacospongia mollior hosted bacteria belonging to the classes Dadabacteriia, Anaerolineae, Acidimicrobiia, Nitrospiria, and Poribacteria. Moreover, for the first time, the presence of Rhizobiaceae bacteria was revealed in the spongeM. (Myxilla) rosacea, which was mainly associated with soil and plants and involved in biological nitrogen fixation. [ABSTRACT FROM AUTHOR]

The textile industry generates significant amounts of wastewater containing high concentrations of azo dyes. An important point in the process of purification of azo dyes is their influence on the activated sludge (AS) in wastewater treatment plants. Azo dyes, such as amaranth, play the role of xenobiotics. This article seeks to answer the question of how organisms manage to respond to xenobiotics remains very important and open, i.e., how they will react to toxic conditions. The aim of this research was to study how these changes are expressed in terms of the different trophic levels of AS. In our experiment, it was found that the dominant trophic units are significantly changed due to the xenobiotic entering the system. The data reveal the significant development of the bacterial segment (genus Pseudomonas and azo-degrading bacteria) at times of large amaranth removal. In the most active phase of amaranth biodetoxification (48 h), the culturable bacteria of the genus Pseudomonas change by about 40%, while the azo-degrading bacteria change by about 2%. Fauna organisms have a sharp change in the dominant groups—from attached and crawling ciliates and testate amoebas to the mass development of small and large flagellates. This is of great importance because micro- and metafauna play an important role in the detoxification process by ingesting some of the xenobiotics. This role is expressed in the fact that after dying, macro-organisms release this xenobiotic in small portions so that it can then be effectively degraded by adapting to the amaranth biodegradation bacteria. In this study, it is clear that all these events lead to a decline in the quality of AS. But on the other hand, these allow AS to survive as a microbial community, and the fauna segment does not disappear completely. [ABSTRACT FROM AUTHOR]

A thermophilic UASB reactor was operated at 55 deg. C for greater than 470 days in order to investigate the effects of feed composition on the changes in microbial community structure where thermophilic granular sludge was used as the inoculum source. The feed compositions were changed with cultivation days; phase 1 (1-70 days), alcohol distillery wastewater; phase 2 (71-281 days), artificial acetate wastewater; phase 3 (282-474 days), artificial sucrose wastewater. During the first one month of each phase, the methanogenic activity and cell density of methanogens quantified by fluorescence in situ hybridization (FISH) drastically changed as a result of shift in feed composition. When artificial acetate wastewater was used as feed, retained granular sludge was partially disintegrated due to a decrease in the number of symbiotic bacterial community members: acetogens (acidogens) and hydrogenotrophic methanogens. In contrast, when the feed was shifted to sucrose (phase 3), granulation of biomass was promoted by a remarkable proliferation of the symbiotic community. The presence of hydrogen-utilizing methanogens and acetogens (acidogens) are shown to be effective for the enhancement of thermophilic granulation. The cell density of methanogens determined by FISH was strongly correlated with the methane-producing potential of the retained thermophilic granular sludge. [ABSTRACT FROM AUTHOR]

Marine oil spills are a significant concern worldwide, destroying the ecological environment and threatening the survival of marine life. Various oil-degrading bacteria have been widely reported in marine environments in response to marine oil pollution. However, little information is known about culturable oil-degrading bacteria in cold seep of the deep-sea environments, which are rich in hydrocarbons. This study enriched five oil-degrading consortia from sediments collected from the Haima cold seep areas of the South China Sea. Parvibaculum, Erythrobacter, Acinetobacter, Alcanivorax, Pseudomonas, Marinobacter, Halomonas, and Idiomarina were the dominant genera. Further results of bacterial growth and degradation ability tests indicated seven efficient alkane-degrading bacteria belonging to Acinetobacter, Alcanivorax, Kangiella, Limimaricola, Marinobacter, Flavobacterium, and Paracoccus, whose degradation rates were higher in crude oil (70.3-78.0%) than that in diesel oil (62.7-66.3%). From the view of carbon chain length, alkane degradation rates were medium chains > long chains > short chains. In addition, Kangiella aquimarina F7, Acinetobacter venetianus F1, Limimaricola variabilis F8, Marinobacter nauticus J5, Flavobacterium sediminis N3, and Paracoccus sediminilitoris N6 were first identified as oil-degrading bacteria from deep-sea environments. This study will provide insight into the bacterial community structures and oil-degrading bacterial diversity in the Haima cold seep areas, South China Sea, and offer bacterial resources to oil bioremediation applications. [ABSTRACT FROM AUTHOR]

Five strains of psychrotrophic hydrocarbon-oxidizing bacteria were isolated from the bottom sediments of Peter the Great Bay of the Sea of Japan. They were classified into the following species: Rhodococcus erythropolis, Rhodococcus sp., Sphingomonas sp., Pseudomonas sp., and Alcanivorax sp. All studied bacteria showed high oxidizing ability in relation to the decomposition of n-alkanes (C9–C27), phytane, pristane, and polycyclic aromatic hydrocarbons at 5 and 22°C. At the same time, the degradation of hydrocarbons was more intense at 5°C. Despite the different taxonomic affiliations of the obtained microorganisms, all strains primarily utilized short- (C9–C13) and long-chain (C21–27) alkanes, as well as polycyclic aromatic hydrocarbons. The highest hydrocarbon-oxidizing activity was shown by the strain Rhodococcus erythropolis AP_291. The latter utilized more than 50% of all hydrocarbons in the model mixture during the first week of the experiment at 5°C. [ABSTRACT FROM AUTHOR]

The present work provides the first knowledge about the diversity and functionality of the microorganisms occurring in oily bilge wastes, from the port of Mar del Plata, Argentina. We used a culture independent approach, based on massive sequencing of the 16S rRNA gene (Illumina), to analyze the microbiome structure from different oily bilge wastes. To gain insight into functional biomarkers for hydrocarbon (HC) degradation in these wastes, the occurrence of alkB genes in the degrading bacteria communities was also investigated trough PCR-DGGE method. The microbial compositions were determined in bilge waste oily phase (BWOP) samples from a deep-sea, a fishing ship, a dredge ship, a small coastal vessel, and an open pool, where bilge wastes from Mar del Plata's vessels are deposited. A core microbiome with OTUs classified as bacterial types associated with HC biodegradation was found across all samples. Seven highly predominant and stable bacterial types accounted for 82% of the relative composition determined in all the BWOP samples. These results indicate that oily bilge is a strong selector for bacteria having common taxonomical and functional features, independent of the vessel of collection. Alkane degradation marker gene alkB PCR-DGGE fingerprinting showed distinct diversity patterns among the samples. Our analysis showed that there is a well-defined bacterial assemblage able to withstand and thrive in the harsh conditions of the oily bilge wastes. Also, a well-established aliphatic biodegradation potential marker was detected, indicating that it could promptly be used for the generation of new bioremediation technologies. [ABSTRACT FROM AUTHOR]

Marine hydrocarbon-degrading bacteria play an important role in natural petroleum biodegradation processes and were initially associated with man-made oil spills or natural seeps. There is no full clarity though on what, in the absence of petroleum, their natural niches are. Few studies pointed at some marine microalgae that produce oleophilic compounds (alkanes, long-chain fatty acids, and alcohols) as potential natural hosts of these bacteria. We established Dansk crude oil-based enrichment cultures with photobioreactor-grown marine microalgae cultures Pavlova lutheri and Nannochloropsis oculata and analyzed the microbial succession using cultivation and SSU (16S) rRNA amplicon sequencing. We found that petroleum enforced a strong selection for members of Alpha- and Gamma-proteobacteria in both enrichment cultures with the prevalence of Alcanivorax and Marinobacter spp., well-known hydrocarbonoclastic bacteria. In total, 48 non-redundant bacterial strains were isolated and identified to represent genera Alcanivorax , Marinobacter , Thalassospira , Hyphomonas , Halomonas , Marinovum , Roseovarius , and Oleibacter , which were abundant in sequencing reads in both crude oil enrichments. Our assessment of public databases demonstrated some overlaps of geographical sites of isolation of Nannochloropsis and Pavlova with places of molecular detection and isolation of Alcanivorax and Marinobacter spp. Our study suggests that these globally important hydrocarbon-degrading bacteria are associated with P. lutheri and N. oculata. [ABSTRACT FROM AUTHOR]

Vinasse is a by-product from the ethanol industry and can be used for methane production through anaerobic digestion process driven by microbial consortia. The microorganisms involved must be known to obtain an ideal methane production. The present work evaluated the production of methane and byproducts from different ratios substrate/biomass (S0/X0), using sludge from an effluent treatment of the vegetable oil industry as inoculum in media containing vinasse. Also, the microbial community of the best methane production bioassay was characterized by high-throughput DNA sequencing. The following chemical parameters were evaluated: methanogenic activity, chemical oxygen demand, carbohydrate consumption, and production of volatile fatty acids. The highest methane production occurred at S0/X0 ratio of 1.5, which produced 59.78 mmol CH4 L−1. A great variety of microorganism genera was identified by high-throughput DNA sequencing, showing differences in the microbial consortia of the initial and final sampling times. At the final sampling point, the classes Bacteroidia (Porphyromonadaceae—OTU genera unknown 42.26% and Bacteroides genus 10.58%) and the class Betaproteobacteria (Proteobacteria-Comamonadaceae OTU) were identified as the dominant bacteria. The most abundant archaeal genera in the bioassay were Methanosaeta, Methanomassiliicoccaceae OTU vadinCA11, and Methanobacterium. The identification of the microorganisms of consortia involved in anaerobic digestion can collaborate on technologies to increase methane production through microbial isolation, bioaugmentation, and co-cultures. [ABSTRACT FROM AUTHOR]

This study examined impacts of Macondo MC252 oil from the Deepwater Horizon spill on oil-degrading bacteria in fresh and oligohaline marshes dominated by the common reed Phragmites australis (Cav.) Trin. ex Steud. and correlated microbial changes to soil variables. We hypothesized that indigenous oil-degrading bacteria and soil characteristics in the marshes effectively respond to Macondo oil. We tested this hypothesis with a greenhouse mesocosm study. Weathered and emulsified Macondo oil was applied to P. australis sods at different rates (0, 4, 8, 12, and 16 L m−2). Populations of oil degrading bacteria containing alkane monooxygenase and PAH-ring hydroxylating dioxygenase alpha subunit (PAH-RHDα) genes, the expression of these genes, soil respiration rate, residual total petroleum hydrocarbon, redox potential (Eh), as well as dissolved organic carbon (DOC) and its aromaticity index of SUVA254 were measured in soils 2 months after oil addition. Oil exposure generally increased oil-degrading bacterial populations, in particular Gram-positive (GP) PAH-degraders, soil respiration rate, DOC concentration, and aromaticity of DOC, but decreased Eh values. GP PAH-RHDα genes accounted for approximately 98% of total detected genes, and expression of these genes increased by a factor of 3 to 5 at various oil dosages. Both abundance and expression of GP PAH-RHDα genes significantly correlated to SUVA254 (P < 0.05). The study revealed that indigenous oil-degrading bacteria effectively responded to weathered Macondo oil in the P. australis marsh soil wherein GP bacteria with PAH-RHDα genes played a major role in biodegradation of PAH-enriched petroleum hydrocarbons. [ABSTRACT FROM AUTHOR]

Field data from the first several days after an oil spill is rare but crucial for our understanding of a spill’s impact on marine microbiota given their short generation times. Field data collected within days of the Texas City “Y” oil spill showed that exposure to crude oil can rapidly imbalance populations of marine microbiota, which leads to the proliferation of more resistant organisms. Vibrionales bacteria were up to 48 times higher than background concentrations at the most impacted sites and populations of the dinoflagellate Prorocentrum texanum increased significantly as well. Laboratory microcosm experiments with a natural plankton community showed that P. texanum grew significantly faster under oiled conditions but monocultures of P. texanum did not. Additional laboratory experiments with natural communities from Tampa Bay, Florida showed similar results although a different species dominated, P. minimum. In both cases, tolerance to the presence of crude oil was enhanced by higher sensitivity of grazers led to a release from grazing pressure and allows Prorocentrum species to dominate after an oil spill. The results suggest careful monitoring for Vibrionales and Prorocentrum during future spills would be beneficial given the potential implications to human health. [ABSTRACT FROM AUTHOR]

Variations on hydrophobicity were monitored in two marine obligate hydrocarbonoclastic bacteria: Alcanivorax borkumensis SK2T and Thalassolituus oleivoras MIL-1T. These strains were inoculated, separately in ONR7a mineral medium with different concentration of sodium acetate. During 10 days measurements of cellular abundance and cellular hydrophobicity (capacity to adhere at polystyrene) were carried out. Data obtained revealed that the natures of carbon source and growth phase are important factors in the regulation of adhesion of bacteria to the surfaces. Moreover Alcanivorax showed a major capability to colonize polystyrene respect on Thalassolituus in the experimented conditions. The understanding of capacity to adhesion of these bacteria for utilization of hydrophobic compounds is fundamental for their potential use in the mitigation of oil spills phenomenon. [ABSTRACT FROM AUTHOR]

Background Synthetic microplastics (⩽5-mm fragments) are emerging environmental contaminants that have been found to accumulate within coastal marine sediments worldwide. The ecological impacts and fate of microplastic debris are only beginning to be revealed, with previous research into these topics having primarily focused on higher organisms and/or pelagic environments. Despite recent research into plastic-associated microorganisms in seawater, the microbial colonization of microplastics in benthic habitats has not been studied. Therefore, we employed a 14-day microcosm experiment to investigate bacterial colonization of lowdensity polyethylene (LDPE) microplastics within three types of coastal marine sediment from Spurn Point, Humber Estuary, U.K. Results Bacterial attachment onto LDPE within sediments was demonstrated by scanning electron microscopy and catalyzed reporter deposition fluorescence in situ hybridisation (CARDFISH). Log-fold increases in the abundance of 16S rRNA genes from LDPE-associated bacteria occurred within 7 days with 16S rRNA gene numbers on LDPE surfaces differing significantly across sediment types, as shown by quantitative PCR. Terminal-restriction fragment length polymorphism (T-RFLP) analysis demonstrated rapid selection of LDPEassociated bacterial assemblages whose structure and composition differed significantly from those in surrounding sediments. Additionally, T-RFLP analysis revealed successional convergence of the LDPE-associated communities from the different sediments over the 14- day experiment. Sequencing of cloned 16S rRNA genes demonstrated that these communities were dominated after 14 days by the genera Arcobacter and Colwellia (totalling 84-93% of sequences). Attachment by Colwellia spp. onto LDPE within sediments was confirmed by CARD-FISH. Conclusions These results demonstrate that bacteria within coastal marine sediments can rapidly colonize LDPE microplastics, with evidence for the successional formation of plastisphere-specific bacterial assemblages. Although the taxonomic compositions of these assemblages are likely to differ between marine sediments and the water column, both Arcobacter and Colwellia spp. have previously been affiliated with the degradation of hydrocarbon contaminants within low-temperature marine environments. Since hydrocarbon-degrading bacteria have also been discovered on plastic fragments in seawater, our data suggest that recruitment of hydrocarbonoclastic bacteria on microplastics is likely to represent a shared feature between both benthic and pelagic marine habitats. [ABSTRACT FROM AUTHOR]

Atomic force microscope techniques and multi-staining fluorescence microscopy were employed to study the steps in drinking water biofilm formation. During the formation of a conditioning layer, surface hydrophobic forces increased and the range of characteristic hydrophobic forces diversified with time, becoming progressively complex in macromolecular composition, which in return triggered irreversible cellular adhesion. AFM visualization of 1 to 8 week drinking water biofilms showed a spatially discontinuous and heterogeneous distribution comprising an extensive network of filamentous fungi in which biofilm aggregates were embedded. The elastic modulus of 40-day-old biofilms ranged from 200 to 9000 kPa, and the biofilm deposits with a height >0.5 μm had an elastic modulus <600 kPa, suggesting that the drinking water biofilms were composed of a soft top layer and a basal layer with significantly higher elastic modulus values falling in the range of fungal elasticity. [ABSTRACT FROM PUBLISHER]

In many areas of the world, polybrominated diphenyl ethers are ubiquitous due to their use as fire retardants. It is known that the hydrophobic characteristics of PBDEs cause them to sink in soil and sediment, yet their effect on microbes within the soil is not well understood. In this study, soil was treated with 1, 10, and 100 mg kg BDE 209 for up to 45 days. Treatment effects on soil enzymatic activities for urease and catalase were evaluated. The impact on the microbe community structure was estimated using denaturing gradient gel electrophoresis after polymerase chain reaction amplification of total genomic DNA, using bacterial variable V3 region targeted primers. The effects on the soil microbial community size and major bacterial groups were evaluated using fluorescence in situ hybridization analysis. Forty-five days after the addition of BDE 209, urease activity was suppressed by BDE 209, even at a concentration of 1 mg kg. Catalase activity increased in the samples containing lower concentrations of BDE 209, but was suppressed in samples containing higher concentrations. The bacterial community also varied in response to the addition of BDE 209, and the variation of community composition differed among treatments. In addition, α, β and γ subclass proteobacteria decreased in the group of 100 mg kg BDE 209 spiked soil after 45 days of treatment. Throughout the experiment, no BDE 209 degradation was detected under darkness. These observations demonstrated that BDE 209 in soil, although of low bioavailability, had an adverse impact on the structure and function of the soil microbial community and microbial processes. [ABSTRACT FROM AUTHOR]

The ability of native bacteria to utilize diesel fuel as the sole carbon and energy source was investigated in this research. Ten bacterial strains were isolated from the oil refinery field in Tehran, Iran. Two biodegradation experiments were performed in low and high (500 and 10000 ppm, respectively) concentration of diesel fuel for 15 days. Only two isolates were able to efficiently degrade the petroleum hydrocarbons in the first test and degraded 86.67 % and, 80.60 % of diesel fuel, respectively. The secondary experiment was performed to investigate the toxicity effect of diesel fuel at high concentration (10000 ppm). Only one strain was capable to degrade 85.20 % of diesel fuel at the same time (15 days). Phenotype and phylogeny analysis of this strain was characterized and identified as dieseldegrading bacteria, based on gram staining, biochemical tests, 16S rRNA gene sequence analysis. These results indicate that this new strain was Bacillus sp. and could be considered as Bacillus Cereus with 98 % 16 S rRNA gene sequence similarity. The results indicate that native strains have great potential for in situ remediation of diesel-contaminated soils in oil refinery sites. [ABSTRACT FROM AUTHOR]

The microbial consortium involved in anaerobic digestion has not yet been precisely characterized and this process remains a ‘black box’ with limited efficiency. In this study, seven anaerobic sludge digesters were selected based on technology, type of sludge, process and water quality. The prokaryotic community of these digesters was examined by constructing and analysing a total of 9890 16S rRNA gene clones. Libraries were constructed using primers specific for the Bacteria and Archaea domains for each digester, respectively. After phylogenetic affiliation, the libraries were compared using statistical tools to determine the similarities or differences among the seven digesters. Results show that the prokaryotic community of an anaerobic digester is composed of phylotypes commonly found in all anaerobic digesters sampled and also of specific phylotypes. The Archaea community is represented by an equilibrium among a restricted number of operational taxonomic units (OTUs). These OTUs are affiliated with Methanosarcinales, Methanomicrobiales and Arc I phylogenetic groups. Statistical analysis revealed that the Bacteria community can be described as a three component model: one-third making up a core group of phylotypes common to most of the digesters, one-third are phylotypes shared among a few digesters and another one-third are specific phylotypes. The core group is composed of only six OTUs affiliated with Chloroflexi, Betaproteobacteria, Bacteroidetes and Synergistetes. Its role in anaerobic degradation appears critical to investigate. This comparison of anaerobic digester populations is a first step towards a future understanding of the relationship among biodiversity, operating conditions and digester efficiency.The ISME Journal (2009) 3, 700–714; doi:10.1038/ismej.2009.2; published online 26 February 2009 [ABSTRACT FROM AUTHOR]

Aims: Characterization and quantification of microbial community in dimethyl phthalate (DMP)-degrading anaerobic sludge using molecular techniques. Methods and Results: An enriched anaerobic sludge effectively degrading over 99% of dimethyl phthalate in an upflow anaerobic sludge blanket (UASB) reactor for 530 days was characterized and quantified by 16S rRNA-based molecular methods. A total of 78 Bacteria clones were classified into 22 operational taxonomic units (OTUs) in nine divisions, including Firmicutes, Proteobacteria, Chloroflexi, Thermotogae, Bacteroidetes/Chlorobi, Spirochaetes, Acidobacteria and two candidate divisions. The two most abundant OTUs were likely responsible, respectively, for the de-esterification of DMP and the subsequent phthalate degradation. The outer layer of the granule was dominated by Bacteria; whereas the interior was by Archaea, of which 89 ± 5% were acetoclastic Methanosaetaceae and 11 ± 5% hydrogenotrophic Methanomicrobiales. Conclusions: Twenty-two Bacteria OTUs in DMP-degrading anaerobic sludge distributed in nine divisions. The two most abundant OTUs were likely responsible respectively for the de-esterification of DMP and the subsequent phthalate degradation. Layered granular microstructure of DMP-degrading anaerobic sludge suggested that the rate of DMP de-esterification is faster than its inward diffusion rate. Significance and Impact of the Study: This work is the first study to characterize and quantify the microbial community in the anaerobic phthalic ester degrading sludge from the anaerobic reactor. [ABSTRACT FROM AUTHOR]

A lab-scale expanded granular sludge bed (EGSB) reactor was seeded with granular sludge and operated to investigate the influence of temperature decrease on both process performance and the microbial community structure of the granular sludge. Synthetic wastewater containing sucrose and volatile fatty acids was used as feed. The EGSB reactor was brought online at a starting temperature of 15°C and was reduced stepwise to a final temperature of 5°C. The reactor exhibited sufficient COD removal efficiency between 10°C and 15°C. However at 5°C serious deterioration of process performance was observed. The methane-producing activity of the retained sludge increased when it was compared to the activity of the seed sludge (day 0) during 10°C to 15°C operation. When hydrogen fed, sludge showed much higher methanogenic activity as compared with seed sludge activity at test temperatures of 15°C and 20°C on day 196 of reactor operation. At this time, proliferation of the genus Methanospirillum in the retained sludge was observed and a decrease in Methanobacterium species was also measured. Throughout the experiment, the genus Methanosaeta was detected in abundance and the community structure of the Domain Bacteria was stably maintained. The sugar-degrading acid-forming bacteria, Lactococcus and Anaerovibrio were detected in the retained sludge throughout the experiment as well and the propionate-degrading acetogen Syntrophobacter fumaroxidans was also detected, although its population size decreased at 5°C. [ABSTRACT FROM AUTHOR]

Rod-shaped Alphaproteobacteria possessing long prosthecae-like appendages have been detected abundantly in biofilms attaching onto anode graphite of cellulose-fed microbial fuel cells (MFCs). To identify their ecological roles, the present study isolated a corresponding bacterium (strain Mfc52) by direct plating of a biofilm suspension onto a solid medium containing glucose and ferric ion. Phylogenetic analysis revealed that this strain is deeply branched in the class Alphaproteobacteria and may represent a novel order. Strain Mfc52 fermented sugars and produced lactate, acetate, and fumarate, whereas ferric ion stimulated the growth on glucose. When an MFC was inoculated with this strain and supplemented with glucose, it fermented glucose and generated electricity by oxidizing organic acids produced from glucose. Electron micrographs showed that a fraction of cells in a liquid culture had prosthecae-like appendages that were abundantly observed in anode biofilm. These observations suggest that the bacterial population represented by strain Mfc52 shared an important niche in the cellulose-fed MFC, where it generated electricity by oxidizing intermediate metabolites from cellulose degradation. [ABSTRACT FROM AUTHOR]

The salinity of industrial wastewater evaporation ponds was artificially increased from 3–7% to 12–16% (w/v), in an attempt to reduce the activity of sulfate-reducing bacteria (SRB) and subsequent emission of H2S. To investigate the changes in bacterial diversity in general, and SRB in particular, following this salination, two sets of universal primers targeting the 16S rRNA gene and the functional apsA [adenosine-5′-phosphosulfate (APS) reductase α-subunit] gene of SRB were used. Phylogenetic analysis indicated that Proteobacteria was the most dominant phylum both before and after salination (with 52% and 68%, respectively), whereas Firmicutes was the second most dominant phylum before (39%) and after (19%) salination. Sequences belonging to Bacteroidetes, Spirochaetes and Actinobacteria were also found. Several groups of SRB from Proteobacteria and Firmicutes were also found to inhabit this saline environment. Comparison of bacterial diversity before and after salination of the ponds revealed both a shift in community composition and an increase in microbial diversity following salination. The share of SRB in the 16S rRNA gene was reduced following salination, consistent with the reduction of H2S emissions. However, the community composition, as shown by apsA gene analysis, was not markedly affected. [ABSTRACT FROM AUTHOR]

This article discusses a study which examined the distribution of indigenous oil-degrading bacteria in the coastal seawater of Toyama Bay, Japan. The study analyzed surface seawater samples with or without oil film in fishing by denaturing gradient gel electrophoresis (DGGE) of the polymerase chain reaction (PCR)-amplified V3 region of bacterial 16S recombinant DNA (rDNA). It revealed that several DGGE bands clearly detected only in samples with oil film corresponded to Cyanobacteria. Moreover, the study cultured surface seawater samples with oil film in two different liquid culture media, a marine broth and an NSW medium.

Like Bacteria, Archaea occur in a wide variety of environments, only some of which can be considered ‘extreme’. We compare archaeal diversity, as represented by 173 16S rRNA gene libraries described in published reports, to bacterial diversity in 79 libraries from the same source environments. An objective assessment indicated that 114 archaeal libraries and 45 bacterial libraries were large enough to yield stable estimates of total phylotype richness. Archaeal libraries were seldom as large or diverse as bacterial libraries from the same environments. However, a relatively larger proportion of libraries were large enough to effectively capture rare as well as dominant phylotypes in archaeal communities. In contrast to bacterial libraries, the number of phylotypes did not correlate with library size; thus, ‘larger’ may not necessarily be ‘better’ for determining diversity in archaeal libraries. Differences in diversity suggest possible differences in ecological roles of Archaea and Bacteria; however, information is lacking on relative abundances and metabolic activities within the sampled communities, as well as the possible existence of microhabitats. The significance of phylogenetic diversity as opposed to functional diversity remains unclear, and should be a high priority for continuing research. [ABSTRACT FROM AUTHOR]

A field experiment was performed to monitor changes in exogenous bacteria and to investigate the diversity of indigenous bacteria during a field trial of microbial enhanced oil recovery (MEOR). Two wells (26–195 and 27–221) were injected with three exogenous strains and then closed to allow for microbial growth and metabolism. After a waiting period, the pumps were restarted and the samples were collected. The bacterial populations of these samples were analyzed by denaturing gradient gel electrophoresis (DGGE) with PCR-amplified 16S rRNA fragments. DGGE profiles indicated that the exogenous strains were retrieved in the production water samples and indigenous strains could also be detected. After the pumps were restarted, average oil yield increased to 1.58 and 4.52 tons per day in wells 26–195 and 27–221, respectively, compared with almost no oil output before the injection of exogenous bacteria. Exogenous bacteria and indigenous bacteria contributed together to the increased oil output. Sequence analysis of the DGGE bands revealed that Proteobacteria were a major component of the predominant bacteria in both wells. Changes in the bacteria population in the reservoirs during MEOR process were monitored by molecular analysis of the 16S rRNA gene sequence. DGGE analysis was a successful approach to investigate the changes in microorganisms used for enhancing oil recovery. The feasibility of MEOR technology in the petroleum industry was also demonstrated. [ABSTRACT FROM AUTHOR]

BTX (benzene, toluene and xylene) degrading bacteria were isolated from Pacific Ocean sediment and nearshore surface water. In the seawater near a ferry dock, degrading bacteria of a relatively wide diversity were detected, including species of Pseudomonas, Rhodococcus, Exiguobacterium and Bacillus; while species of Bacillus only have been detected from the deep-sea sediment. Most of the isolates showed degradation to more than one compound. Generally better growth was obtained with p-xylene and ethylbenzene than with the other two. All the bacteria could tolerate and grow with the compounds at 5–20% (v/v). Both benzene and toluene degradation related genes had been successfully PCR cloned from the isolates of nearshore water, the detected benzene dioxygenase gene was identical among all the species and close to its soil counterpart. However, they were not detected in all the isolates from deep sea. Results in this report suggested that BTX degrading bacteria widely spread in marine environments and they might be of potentials in biotreatment of BTEX in saline environments. [ABSTRACT FROM AUTHOR]

Background: Microbial fuel cells (MFCs) are devices that exploit microorganisms to generate electric power from organic matter. Despite the development of efficient MFC reactors, the microbiology of electricity generation remains to be sufficiently understood. Results: A laboratory-scale two-chamber microbial fuel cell (MFC) was inoculated with rice paddy field soil and fed cellulose as the carbon and energy source. Electricity-generating microorganisms were enriched by subculturing biofilms that attached onto anode electrodes. An electric current of 0.2 mA was generated from the first enrichment culture, and ratios of the major metabolites (e.g., electric current, methane and acetate) became stable after the forth enrichment. In order to investigate the electrogenic microbial community in the anode biofilm, it was morphologically analyzed by electron microscopy, and community members were phylogenetically identified by 16S rRNA gene clone-library analyses. Electron microscopy revealed that filamentous cells and rodshaped cells with prosthecae-like filamentous appendages were abundantly present in the biofilm. Filamentous cells and appendages were interconnected via thin filaments. The clone library analyses frequently detected phylotypes affiliated with Clostridiales, Chloroflexi, Rhizobiales and Methanobacterium. Fluorescence in-situ hybridization revealed that the Rhizobiales population represented rod-shaped cells with filamentous appendages and constituted over 30% of the total population. Conclusion: Bacteria affiliated with the Rhizobiales constituted the major population in the cellulose-fed MFC and exhibited unique morphology with filamentous appendages. They are considered to play important roles in the cellulose-degrading electrogenic community. [ABSTRACT FROM AUTHOR]

Today, biomass covers about 10% of the world’s primary energy demand. Against a backdrop of rising crude oil prices, depletion of resources, political instability in producing countries and environmental challenges, besides efficiency and intelligent use, only biomass has the potential to replace the supply of an energy hungry civilisation. Plant biomass is an abundant and renewable source of energy-rich carbohydrates which can be efficiently converted by microbes into biofuels, of which, only bioethanol is produced on an industrial scale today. Biomethane is produced on a large scale, but is not yet utilised for transportation. Biobutanol is on the agenda of several companies and may be used in the near future as a supplement for gasoline, diesel and kerosene, as well as contributing to the partially biological production of butyl-t-butylether, BTBE as does bioethanol today with ETBE. Biohydrogen, biomethanol and microbially made biodiesel still require further development. This paper reviews microbially made biofuels which have potential to replace our present day fuels, either alone, by blending, or by chemical conversion. It also summarises the history of biofuels and provides insight into the actual production in various countries, reviewing their policies and adaptivity to the energy challenges of foreseeable future. [ABSTRACT FROM AUTHOR]

We used catalysed reported deposition – fluorescence in situ hybridization (CARD-FISH) to analyse changes in the abundance of the bacterial groups Alphaproteobacteria, Gammaproteobacteria and Bacteroidetes, and of hydrocarbon-degrading Cycloclasticus bacteria in mesocosms that had received polycyclic aromatic hydrocarbons (PAHs) additions. The effects of PAHs were assessed under four contrasting hydrographic conditions in the coastal upwelling system of the Rías Baixas: winter mixing, spring bloom, summer stratification and autumn upwelling. We used realistic additions of water soluble PAHs (approximately 20–30 μg l−1 equivalent of chrysene), but during the winter period we also investigated the effect of higher PAHs concentrations (10–80 μg l−1 chrysene) on the bacterial community using microcosms. The most significant change observed was a significant reduction (68 ± 5%) in the relative abundance of Alphaproteobacteria. The magnitude of the response of Cycloclasticus bacteria (positive with probe CYPU829) to PAHs additions varied depending on the initial environmental conditions, and on the initial concentration of added PAHs. Our results clearly show that bacteria of the C ycloclasticus group play a major role in low molecular weight PAHs biodegradation in this planktonic ecosystem. Their response was stronger in colder waters, when their background abundance was also higher. During the warm periods, the response of Cycloclasticus was limited, possibly due to both, a lower bioavailability of PAHs caused by abiotic factors (solar radiation, temperature), and by inorganic nutrient limitation of bacterial growth. [ABSTRACT FROM AUTHOR]

A fatal bulking phenomenon was found to occur occasionally in the methanogenic granular sludge of a mesophilic (35–40°C), full-scale upflow anaerobic sludge blanket (UASB) reactor treating organic wastewater discharged from a sugar manufacturing factory. A vast number of filamentous cells were observed in the bulking sludge that were morphologically distinct from the previously recognized anaerobic bulking agent Anaerolinea thermophila. 16S rRNA gene-based analyses of the microbial populations in the bulking sludge revealed that the dominant filamentous organisms were members of proposed candidate bacterial phylum, KSB3. Fluorescence in situ hybridization (FISH) analysis of the healthy sludge granules showed that the KSB3 filaments were the dominant granule surface population suggesting that they are fundamental constituents of the sludge granules and that they occasionally overgrow in the reactor, possibly triggering the filamentous bulking. We surveyed 10 additional mesophilic and thermophilic anaerobic sludges for the presence and diversity of KSB3 populations. Bacteria closely related to the characterized KSB3 filaments were present in two types of mesophilically grown UASB sludge granules treating actual wastewater discharged from sugar-processing industries.The ISME Journal (2007) 1, 246–255; doi:10.1038/ismej.2007.28; published online 31 May 2007 [ABSTRACT FROM AUTHOR]

The bacterial and archaeal community structure was examined in two methanogenic anaerobic digestion processes degrading organic household waste at mesophilic (37°C) and thermophilic (55°C) temperatures. Analysis of bacterial clone libraries revealed a predominance of Bacteroidetes (34% of total clones) and Chloroflexi (27%) at the mesophilic temperature. In contrast, in the thermophilic clone library, the major group of clones were affiliated with Thermotogae (61%). Within the domain Archaea, the phyla Euryarchaeota and Crenarchaeota were both represented, the latter only at the mesophilic temperature. The dominating archaeons grouped with Methanospirillum and Methanosarcina species at the mesophilic and thermophilic temperature, respectively. Generally, there was a higher frequency of different sequences at the lower temperature, suggesting a higher diversity compared to the community present at the thermophilic temperature. Furthermore, it was not only the species richness that was affected by temperature, but also the phylogenetic distribution of the microbial populations. [ABSTRACT FROM AUTHOR]

A bacterial strain capable of utilizing n-alkanes with chain lengths ranging from decane (C10H22) to tetracontane (C40H82) as a sole carbon source was isolated using a system for screening microorganisms able to grow on paraffin (mixed long-chain n-alkanes). The isolate, identified according to its 16S rRNA sequence as Acinetobacter venetianus, was designated A. venetianus 6A2. Two DNA fragments encoding parts of AlkB-type alkane hydroxylase homologues, designated alkMa and alkMb, were polymerase chain reaction-amplified from the genome of A. venetianus 6A2. To study the roles of these two alkM paralogues in n-alkane utilization in A. venetianus 6A2, we constructed alkMa, alkMb, and alkMa/ alkMb disruption mutants. Studies on the growth patterns of the disruption mutants using n-alkanes with different chain lengths as sole carbon source demonstrated central roles for the alkMa and alkMb genes in utilization of C10 to C18 n-alkanes. Comparative analysis of these patterns also suggested different substrate preferences for AlkMa and AlkMb in n-alkane utilization. Because both single and double mutants were able to grow on n-alkanes with chain lengths of C20 and longer, we concluded that yet another enzyme(s) for the utilization of these n-alkanes must exist in A. venetianus 6A2. [ABSTRACT FROM AUTHOR]

Lipids (characterized as oils, greases, fats and long-chain fatty acids) are important organic components of wastewater. Their amount, for example, in municipal wastewater is approximately 30–40% of the total chemical oxygen demand. The concern over the behavior of lipids in biological treatment systems has led to many studies, which have evaluated their removal, but still the exact behavior of lipids in these processes is not well understood. In this review, we discuss the current knowledge of how lipids/fatty acids affect both aerobic and anaerobic processes and specific methods that have been used in an attempt to enhance their removal from wastewater. Overall, the literature shows that lipids/fatty acids are readily removed by biological treatment methods, inhibitory to microbial growth as well as the cause of foaming, growth of filamentous bacteria and floc flotation. [ABSTRACT FROM AUTHOR]

Methanosarcina species with a high maximum specific growth rate (µmax) and high half-saturation coefficient (Ks) and Methanosaeta species with a low µmax and low Ks are the only known aceticlastic methanogens. Because of Methanosaeta's low Ks. the low acetate concentrations in conventional, mesophilic anaerobic digestion yield Methanosaeta dominance. However, Methanosarcina absorbs increases in acetate more efficiently and thus promotes more stable digestion. This paper tests the hypothesis that decreasing digester feeding frequencies can increase Methanosarcina predominance. Two acetate-fed reactors were established at a 17-day solids retention time. One reactor was fed hourly, and one was fed once daily. Microscopic and molecular methods were used to verify that the hourly fed reactor enriched for Methanosaeta, while the dally fed reactor enriched for Methanosarcina. Growth and substrate-use kinetics were measured for each reactor. A digester overload condition was simulated, and the Methanosarcina-enriched reactor was found to perform belier than the Methanosaeta-enriched reactor. These findings indicate that Methanosarcina dominance can be achieved with infrequent feedings, leading to more stable digestion. [ABSTRACT FROM AUTHOR]

The effects of naphthalene on microbial communities in the bottom boundary layer of the Delaware Bay estuary were investigated in microcosms using denaturing gradient gel electrophoresis (DGGE) and fluorescent in situ hybridization (FISH) with oligonucleotide probes. Three days after the addition of naphthalene, rates of bacterial production and naphthalene mineralization were higher than in no-addition controls and than in cases where glucose was added. Analyses using both DGGE and FISH indicated that the bacterial community changed in response to the addition of naphthalene. FISH data indicated that a few major phylogenetic groups increased in response to the glucose addition and especially to the naphthalene addition. DGGE also demonstrated differences in community composition among treatments, with four phylotypes being unique to naphthalene-amended treatments and three of these having 16S rRNA genes similar to known hydrocarbon degraders. The bacterial community in the naphthalene-amended treatment was distinct from the communities in the glucose-amended treatment and in the no-addition control. These data suggest that polycyclic aromatic hydrocarbons may have large effects on microbial community structure in estuaries and probably on microbially mediated biogeochemical processes. [ABSTRACT FROM AUTHOR]

The article examines the microbial diversity in the groundwater samples using a culture- and cloning-independent technique in New Zealand. The researchers investigated 18 distinct terminal restriction fragment length polymorphism (T-RFLP) peak profiles were generated from groundwater samples. Based on the results, differences in the peak profiles appeared to be related to groundwater chemistry, especially to concentrations of iron and manganese.

The aim of this study was to find indicators of petroleum biodegradation in granitic groundwater. Both pristine and contaminated groundwaters from boreholes around petroleum storage vaults located approximately 40 m below the surface in granite and with storage capacities of up to 120,000 m 3 were sampled. Total numbers of microorganisms, “most probable numbers” (MPN) of anaerobic bacteria, and chemical indications of microbial activity were determined in the groundwater. Hydrocarbon contaminants and metabolites were detected using gas chromatography-mass spectrometry (GC-MS). In contaminated groundwater, the total number of microorganisms was 2–4 × 10 6 ml -1 , which was significantly higher than the 6 × 10 4 ml -1 found in pristine groundwater. This microbial abundance was also reflected in the MPN analysis. Up to 7 × 10 4 nitrate-, 2 × 10 3 iron-, and 3 × 10 4 sulfate-reducing bacteria were detected in contaminated groundwaters. In such groundwaters, depletion of anaerobic electron acceptors and detection of reduced species could be established. We also proposed using a high alkalinity/hardness of water quota (A/H quota) as an indicator of microbial activity. In contaminated groundwaters the A/H quota averaged 2.8, while in pristine groundwater the same was only 1.3. Moreover, the presence of 20 oxidized petroleum hydrocarbons, i.e., putative metabolites of which 9 were strictly intracellular, was detected in the contaminated groundwaters. Phylogenetic neighbor-joining analysis of 16S rRNA genes provided information about the bacterial communities. The bacteria in contaminated groundwater were found to be strikingly similar to bacteria in other hydrocarbon-contaminated environments. [ABSTRACT FROM AUTHOR]

To investigate the biomass and phylogenetic diversity of the microbial community inhabiting the deep aquifer of the Great Artesian Basin (GAB), geothermal groundwater gushing out from the aquifer was sampled and analyzed. Microbial cells in the groundwater were stained with acridine orange and directly counted by epifluorescence microscopy. Microbial cells were present at a density of 108–109 cells per liter of groundwater. Archaeal and bacterial small-subunit rRNA genes (rDNAs) were amplified by PCR with Archaea- and Bacteria-specific primer sets, and clone libraries were constructed separately. A total of 59 clones were analyzed in archaeal and bacterial 16S rDNA libraries, respectively. The archaeal 16S rDNA clones were divided into nine operated taxonomic units (OTUs) by restriction fragment length polymorphism. These OTUs were closely related to the methanogenic genera Methanospirillum and Methanosaeta, the heterotrophic genus Thermoplasma, or miscellaneous crenarchaeota group. More than one-half of the archaeal clones (59% of total 59 clones) were placed beside phylogenetic clusters of methanogens. The majority of the methanogen-related clones (83%) was closely related to a group of hydrogenotrophic methanogens (genus Methanospirillum). The bacterial OTUs branched into seven phylogenetic clusters related to hydrogen-oxidizing thermophiles in the genera Hydrogenobacter and Hydrogenophilus, a sulfate-reducing thermophile in the genus Thermodesulfovibrio, chemoheterotropic bacteria in the genera Thermus and Aquaspirillum, or the candidate division OP10. Clones closely related to the thermophilic hydrogen-oxidizers in the genera Hydrogenobacter and Hydrogenophilus were dominant in the bacterial clone library (37% of a total of 59 clones). The dominancy of hydrogen-users strongly suggested that H2 plays an important role as a primary substrate in the microbial ecosystem of this deep geothermal aquifer. [ABSTRACT FROM AUTHOR]

Among human faecal bacteria, many members of theClostridium leptumsubgroup are fibrolytic and butyrate producing microorganisms thereby contributing to processes important to colonic health. Yet this phylogenetic subgroup remains poorly described to date. To improve detection and description of members of theC. leptumsubgroup, the Clep 866 group probe was developed. Its association with probes targeting theClostridium viridecluster (Cvir 1414) andEubacterium desmolansspecies (Edes 635) allowed for the first time the detection of all members found in this phylogenetic group in human faecal microbiota. A species-specific probe was also designed to detect members of theRuminococcus callidusspecies (Rcal 733). The design of signature regions was based on alignment of 16S rRNA sequences isolated from faeces of five healthy adults. Furthermore, an oligonucleotide competitor strategy was developed in order to improve the specificity of the probes formerly validated or designed in this study. The oligonucleotide probes were tested using a collection of target and non-target strains using FISH combined with flow cytometry. These new probes were added to a panel of 18 phylogenetic probes selected to describe faecal microbiota composition in 21 human faeces of healthy adults.Clostridium leptumsubgroup represented 22% of the total faecal bacteria and codominated with members ofClostridium coccoidesgroup. The clusterFaecalibacterium prausnitziiwas the dominant component of theC. leptumsubgroup and 20% of the latter subgroup remained unidentified at the species level. [ABSTRACT FROM AUTHOR]

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The diversity of Archaea in anaerobic digesters was characterized by strand conformation polymorphism (SSCP) analysis and the sequencing of 16S rDNA genes. The 44 digesters sampled, located in eight different countries, treated effluents from agriculture, the food processing and petro-chemical industries, pulp and paper plant, breweries, slaughterhouses and municipal waste. All the existing processes were represented among the samples (fixed-film, fluidized bed, stirred-tank, UASB, sequential batch reactor, lagoon). Single strand conformation polymorphism analysis targeting the V3 region of 16S rDNA revealed between four to six distinct archaeal peaks per digester. The diversity of dominant Archaea in the 44 digesters was estimated as 23 different 16S rDNA sequences. Cloning of archaeal 16S rRNA genes from 11 distinct total genomic DNA, screening of clones by SSCP and the sequencing of 170 of them made it possible to characterize these SSCP peaks. All the sequences retrieved were members of the Euryarchaeaota subdomain. Furthermore, most of the sequences retrieved were very close to already known and cultivated strains or to environmental clones. The most frequent archaeal sequences were close to Methanosaeta concilii and to a 16S rDNA clone vadinDC06 located in the Methanobacterium clade (84% and 73% of digesters respectively). The other sequences were members of the Methanobacteriales and the Methanomicrobiales families. Only one sequence was far from any sequence of the database and it could be grouped with several sequences of environmental clones. Each digester harboured between two to nine archaeal sequences with only one of them corresponding to a putative acetate-utilizing species. Furthermore, the process in the digesters appeared to play a part in the distribution of archaeal diversity. [ABSTRACT FROM AUTHOR]

The marine γ-Proteobacterium Alcanivorax borkumensis is highly specialized in the assimilation of aliphatic hydrocarbons, and makes up a large part of the biomass in oil-polluted marine environments. In addition to the previously identified alkane hydroxylase AlkB1, a second alkane hydroxylase (AlkB2) showing 65% identity to the Pseudomonas aeruginosa AlkB2 alkane hydroxylase was identified. Unlike alkB1, alkB2 is not flanked by genes involved in alkane metabolism. Heterologous expression of the A. borkumensis AP1 alkB1 and alkB2 genes showed that they encode functional alkane hydroxylases with substrate ranges similar to those of their P. putida and P. aeruginosa homologues. The transcription initiation sites and levels of the alkB1, alkB2 and alkS mRNA transcripts were determined. Expression of both alkB1 and alkB2 was induced by alkanes, but transcripts corresponding to alkB1 were much more abundant than those of alkB2. An inverted repeat similar to the binding site for the P. putida GPo1 transcriptional activator AlkS was present upstream of the promoters for alkB1 and alkB2, although that of alkB2 was less well conserved, and only the transcriptional fusion of promoter PalkB1 to the reporter gene lacZ efficiently responded to n-octane. Contrary to what has been found for the P. putida GPo1 alkane degradation pathway, expression of the A. borkumensis AP1 alkS gene was not induced by alkanes, and an AlkS binding site was not present upstream of the promoter for alkS. This indicates that, in spite of the clear similarities, the A. borkumensis alk-genes are regulated by a strategy different from that of the P. putida GPo1 alk genes. [ABSTRACT FROM AUTHOR]