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Deep-sea environments can become contaminated with petroleum hydrocarbons. The effects of hydrostatic pressure (HP) in the deep sea on microbial oil degradation are poorly understood. Here, we performed long-term enrichments (100 days) from a natural cold seep while providing optimal conditions to sustain high hydrocarbon degradation rates. Through enrichments performed at increased HP and ambient pressure (AP) and by using control enrichments with marine broth, we demonstrated that both pressure and carbon source can have a big impact on the community structure. In contrast to previous studies, hydrocarbonoclastic operational taxonomic units (OTUs) remained dominant at both AP and increased HP, suggesting piezotolerance of these OTUs over the tested pressure range. Twenty-three isolates were obtained after isolation and dereplication. After recultivation at increased HP, an Alcanivorax sp. showed promising piezotolerance in axenic culture. Furthermore, preliminary co-cultivation tests indicated synergistic growth between some isolates, which shows promise for future synthetic community construction. Overall, more insights into the effect of increased HP on oil-degrading communities were obtained as well as several interesting isolates, e.g. a piezotolerant hydrocarbonoclastic bacterium for future deep-sea bioaugmentation investigation. [ABSTRACT FROM AUTHOR]

The development of high-throughput sequencing technologies has revolutionised the field of microbial ecology via 16S rRNA gene amplicon sequencing approaches. Clustering those amplicon sequencing reads into operational taxonomic units (OTUs) using a fixed cut-off is a commonly used approach to estimate microbial diversity. A 97% threshold was chosen with the intended purpose that resulting OTUs could be interpreted as a proxy for bacterial species. Our results show that the robustness of such a generalised cut-off is questionable when applied to short amplicons only covering one or two variable regions of the 16S rRNA gene. It will lead to biases in diversity metrics and makes it hard to compare results obtained with amplicons derived with different primer sets. The method introduced within this work takes into account the differential evolutional rates of taxonomic lineages in order to define a dynamic and taxonomic-dependent OTU clustering cut-off score. For a taxonomic family consisting of species showing high evolutionary conservation in the amplified variable regions, the cut-off will be more stringent than 97%. By taking into consideration the amplified variable regions and the taxonomic family when defining this cut-off, such a threshold will lead to more robust results and closer correspondence between OTUs and species. This approach has been implemented in a publicly available software package called DynamiC. [ABSTRACT FROM AUTHOR]

Butyrate-producing bacteria are promising probiotic candidates to target microbial dysbiosis in gastrointestinal disorders like inflammatory bowel diseases. Butyricicoccus pullicaecorum 25-3T, a butyrate-producing clostridial cluster IV strain, is such a candidate. Little is known about its abundance in the colon microbiota and its butyrogenic properties. We used the M-SHIMER® , an in vitro simulator for the human intestinal microbial ecosystem, to study the effect of supplementing a single dose of B. pullicaecorum 25-3T on lumen- and mucus-associated microbiota of eight individuals. Butyricicoccus pullicaecorum was more abundant in mucus-associated microbiota compared with lumen microbiota. Supplementation with a single dose of B. pullicaecorum 25-3T resulted in a temporary increase in B. pullicaecorum bacteria in lumen compartment of all individuals. In two cases, the responders, an increased butyrate production was observed as compared with the control. 16S rRNA gene amplicon sequencing revealed the microbiota of responders to be different as compared to non-responder microbiota. We can conclude that B. pullicaecorum 25-3T is a mucus-associated bacterium whose potency to stimulate butyrate production is characterized by a large interindividual variability in terms of composition of the receiving microbial community. [ABSTRACT FROM AUTHOR]

Although efforts have been made the past few years, knowledge on genomic and phenotypic diversity and occurrence of the denitrification ability in Gram-positive bacteria are still fragmentary. Many environmental monitoring approaches have used nir, nor, and nos genes as marker genes for detection of denitrification or denitrifying bacteria. However, primers used in these methods often fail to detect the genes in specific bacterial taxa, such as Gram-positive denitrifiers. In this study, novel primer sets specifically targeting nirK, qnorB, and nosZ genes of the Firmicute genus Geobacillus were developed by genomic mining and tested in parallel with commonly used primers on a set of phylogenetically closely related denitrifying geobacilli. Novel nirK and qnorB sequences were recovered from all strains tested, whereas nosZ was detected in part of the strain set, which was in agreement with observed phenotypes. Interspecies and modest intraspecies variations in amplified fragment length polymorphism ( AFLP) patterns were observed, verifying presence of genomic variation within the strain set. Our study shows that closely related Gram-positive denitrifiers may differ in denitrification phenotype and genotype. But foremost, novel primers targeting very divergent nirK, qnorB, and nosZ gene sequences of Gram-positive denitrifiers, are now available for cultivation-independent environmental surveys. [ABSTRACT FROM AUTHOR]

Biofilms represent a substantial problem in the food industry, with food spoilage, equipment failure, and public health aspects to consider. Besides, biofilms may be a hot spot for plasmid transfer, by which antibiotic resistance can be disseminated to potential foodborne pathogens. This study investigated biomass and plasmid transfer in dual-species ( Pseudomonas putida and Escherichia coli) biofilm models relevant to the food industry. Two different configurations (flow-through and drip-flow) and two different inoculation procedures (donor-recipient and recipient-donor) were tested. The drip-flow configuration integrated stainless steel coupons in the setup while the flow-through configuration included a glass flow cell and silicone tubing. The highest biomass density [10 log (cells cm-²)] was obtained in the silicone tubing when first the recipient strain was inoculated. High plasmid transfer ratios, up to 1/10 (transconjugants/total bacteria), were found. Depending on the order of inoculation, a difference in transfer efficiency between the biofilm models could be found. The ease by which the multiresistance plasmid was transferred highlights the importance of biofilms in the food industry as hot spots for the acquisition of multiresistance plasmids. This can impede the treatment of foodborne illnesses if pathogens acquire this multiresistance in or from the biofilm. [ABSTRACT FROM AUTHOR]

In drinking water (DW) and the distribution systems, bacterial growth and biofilm formation have to be controlled both for limiting taste or odour development and preventing clogging or biocorrosion problems. After a contamination with undesired bacteria, factors like nutrient availability and temperature will influence the survival of these invaders. Understanding the conditions enabling invaders to proliferate is essential for a holistic approach towards microbial risk assessment in DW. Pseudomonas putida was used as a model invader because this easy-growing bacterium can use a wide range of substrates. Invasion experiments in oligo- to eutrophic waters showed the requirement of both a carbon and phosphate source for survival of P. putida in DW. Addition of C, N and P enabled P. putida to grow in DW from 5.80 × 104 to 1.84 × 108 cells mL−1 and survive for at least 12 days. However, in surface water with similar nutrient concentrations, P. putida did not survive, indicating the concomitant importance of the present indigenous microbial community of the specific water sample. Either extensive carbon or phosphate limitation can be used in water treatment design in order to obtain a DW which is not susceptible for unwanted bacterial growth. [ABSTRACT FROM AUTHOR]

Bacillus cereus comprises a highly versatile group of bacteria, which are of particular interest because of their capacity to cause disease. Emetic food poisoning is caused by the toxin cereulide produced during the growth of emetic B. cereus in food, while diarrhoeal food poisoning is the result of enterotoxin production by viable vegetative B. cereus cells in the small intestine, probably in the mucus layer and/or attached to the host's intestinal epithelium. The numbers of B. cereus causing disease are highly variable, depending on diverse factors linked to the host (age, diet, physiology and immunology), bacteria (cellular form, toxin genes and expression) and food (nutritional composition and meal characteristics). Bacillus cereus group strains show impressive ecological diversity, ranging from their saprophytic life cycle in soil to symbiotic (commensal and mutualistic) lifestyles near plant roots and in guts of insects and mammals to various pathogenic ones in diverse insect and mammalian hosts. During all these different ecological lifestyles, their toxins play important roles ranging from providing competitive advantages within microbial communities to inhibition of specific pathogenic organisms for their host and accomplishment of infections by damaging their host's tissues. [ABSTRACT FROM AUTHOR]

Life in hypersaline environments is typically limited by bioenergetic constraints. Microbial activity at the thermodynamic edge, such as the anaerobic oxidation of methane ( AOM) coupled to sulphate reduction ( SR), is thus unlikely to thrive in these environments. In this study, carbon and sulphur cycling was investigated in the extremely hypersaline cold seep sediments of Mercator mud volcano. AOM activity was partially inhibited but still present at salinity levels of 292 g L−1 ( c. eightfold sea water concentration) with rates of 2.3 nmol cm−3 day−1 and was even detectable under saturated conditions. Methane and evaporite-derived sulphate comigrated in the ascending geofluids, which, in combination with a partial activity inhibition, resulted in AOM activity being spread over unusually wide depth intervals. Up to 79% of total cells in the AOM zone were identified by fluorescence in situ hybridization ( FISH) as anaerobic methanotrophs of the ANME-1. Most ANME-1 cells formed monospecific chains without any attached partner. At all sites, AOM activity co-occurred with SR activity and sometimes significantly exceeded it. Possible causes of these unexpected results are discussed. This study demonstrates that in spite of a very low energy yield of AOM, microorganisms carrying this reaction can thrive in salinity up to halite saturation. [ABSTRACT FROM AUTHOR]

The mucus layer in the colon, acting as a barrier to prevent invasion of pathogens, is thinner and discontinuous in patients with ulcerative colitis ( UC). A recent developed in vitro dynamic gut model, the M- SHIME, was used to compare long-term colonization of the mucin layer by the microbiota from six healthy volunteers ( HV) and six UC patients and thus distinguish the mucin adhered from the luminal microbiota. Although under the same nutritional conditions, short-chain fatty acid production by the luminal communities from UC patients showed a tendency toward a lower butyrate production. A more in-depth community analysis of those microbial groups known to produce butyrate revealed that the diversity of the C lostridium coccoides/ E ubacterium rectale and C lostridium leptum group, and counts of F aecalibacterium prausnitzii were lower in the luminal fractions of the UC samples. Counts of R oseburia spp. were lower in the mucosal fractions of the UC samples. q PCR analysis for butyryl- Co A:acetate Co A transferase, responsible for butyrate production, displayed a lower abundance in both the luminal and mucosal fractions of the UC samples. The M- SHIME model revealed depletion in butyrate producing microbial communities not restricted to the luminal but also in the mucosal samples from UC patients compared to HV. [ABSTRACT FROM AUTHOR]

Poly-β-hydroxybutyrate ( PHB) is a natural polymer that can be depolymerized into water-soluble short-chain fatty acid monomers. These monomers can act as microbial control agents. In this study, the effects of partially replacing the diet of Siberian sturgeon fingerlings with 2% and 5% PHB were investigated. Replacing 2% of the diet with PHB improved weight gain, specific growth rate ( SGR) and survival in the sturgeon fingerlings during the 10-week experimental period. Community-level physiological profiling and PCR-denaturing gradient gel electrophoresis ( PCR- DGGE) were used to analyze the microbial community diversity and community organization in the sturgeon gastrointestinal tract. DGGE analysis revealed that PHB affected the intestinal microbial species richness and diversity. The highest species richness was observed with 2% PHB. DNA sequencing of the dominant bands in 2% and 5% PHB treatments revealed that PHB stimulated bacteria belonging to the genera B acillus and R uminococcaceae. Principal component analysis, Lorenz curves and the Shannon index of Biolog™ Ecoplate data revealed that aerobic metabolic potential of the bacterial community was different in the PHB-treated fishes as compared with the control situation. Overall, our results indicate that PHB act as microbial control agents and replacement of 2% of Siberian sturgeon fingerling diet with PHB has beneficial effects. [ABSTRACT FROM AUTHOR]

The use of poly-β-hydroxybutyrate (PHB) was shown to be successful in increasing the resistance of brine shrimp against pathogenic infections. In this study, we isolated for the first time PHB-degrading bacteria from a gastrointestinal environment. Pure strains of PHB-degrading bacteria were isolated from Siberian sturgeon, European sea bass and giant river prawn. The capability of selected isolates to degrade PHB was confirmed in at least two of three setups: (1) growth in minimal medium containing PHB as the sole carbon (C) source, (2) production of clearing zones on minimal agar containing PHB as the sole C source and (3) degradation of PHB (as determined by HPLC analysis) in 10% Luria–Bertani medium containing PHB. Challenge tests showed that the PHB-degrading activity of the selected isolates increased the survival of brine shrimp larvae challenged to a pathogenic Vibrio campbellii strain by a factor 2–3. Finally, one of the PHB-degrading isolates from sturgeon showed a double biocontrol effect because it was also able to inactivate acylhomoserine lactones, a type of quorum-sensing molecule that regulates the virulence of different pathogenic bacteria. Thus, the combined supplementation of a PHB-degrading bacterium and PHB as a synbioticum provides perspectives for improving the gastrointestinal health of aquatic animals. [ABSTRACT FROM AUTHOR]

For more than 100 years it was believed that bacteria were the only group responsible for the oxidation of ammonia. However, recently, a new strain of archaea bearing a putative ammonia monooxygenase subunit A ( amoA) gene and able to oxidize ammonia was isolated from a marine aquarium tank. Ammonia-oxidizing archaea (AOA) were subsequently discovered in many ecosystems of varied characteristics and even found as the predominant causal organisms in some environments. Here, we summarize the current knowledge on the environmental conditions related to the presence of AOA and discuss the possible site-related properties. Considering these data, we deduct the possible niches of AOA based on pH, sulfide and phosphate levels. It is proposed that the AOA might be important actors within the nitrogen cycle in low-nutrient, low-pH, and sulfide-containing environments. [ABSTRACT FROM AUTHOR]

The emission of hydrogen sulfide into the atmosphere of sewer systems induces the biological production of sulfuric acid, causing severe concrete corrosion. As a possible preventive solution, a microbial consortium of nitrate-reducing, sulfide-oxidizing bacteria (NR-SOB) was enriched in a continuously stirred tank reactor in order to develop a biological technique for the removal of dissolved sulfide. The consortium, dominated by Arcobacter sp., was capable of removing 99% of sulfide. Stable isotope fractioning of the sulfide indicated that the oxidation was a biological process. The capacity of the NR-SOB consortium for rapid removal of sulfide was demonstrated by using it as an inoculum in synthetic and real sewage. Removal rates up to 52 mg sulfide-S g VSS−1 h−1 were achieved, to our knowledge the highest removal rate reported so far for freshwater species in the absence of molecular oxygen. Further long-term incubation experiments revealed the capacity of the bacteria to oxidize sulfide without the presence of nitrate, suggesting that an oxidized redox reserve is present in the culture. [ABSTRACT FROM AUTHOR]

Three bacterial enrichment cultures (ECs) were isolated from the digestive tract of Pacific white shrimp Penaeus vannamei, by growing the shrimp microbial communities in a mixture of N-acyl homoserine lactone (AHL) molecules. The ECs, characterized by denaturing gradient gel electrophoresis analysis and subsequent rRNA sequencing, degraded AHL molecules in the degradation assays. Apparently, the resting cells of the ECs also degraded one of the three types of quorum-sensing signal molecules produced by Vibrio harveyi in vitro [i.e. harveyi autoinducer 1 (HAI-1)]. The most efficient AHL-degrading ECs, EC5, was tested in Brachionus experiments. EC5 degraded the V. harveyi HAI-1 autoinducer in vivo, neutralizing the negative effect of V. harveyi autoinducer 2 (AI-2) mutant, in which only the HAI-1- and CAI-1-mediated components of the quorum-sensing system are functional on the growth of Brachionus. This suggests that EC5 interferes with HAI-1-regulated metabolism in V. harveyi. These AHL-degrading ECs need to be tested in other aquatic systems for their probiotic properties, preferably in combination with specific AI-2-degrading bacteria. [ABSTRACT FROM AUTHOR]

A poly-β-hydroxybutyrate (PHB)-accumulating enrichment culture was obtained using activated sludge from a polyphosphate-accumulating reactor as inoculum. PHB accumulated by the enrichment culture significantly enhanced the survival of Artemia nauplii, infected with the virulent pathogen Vibrio campbellii LMG 21363. A strain was isolated from the enrichment culture, based on its ability to accumulate PHB, and 16S rRNA gene sequencing of the isolate revealed 99% sequence similarity to Brachymonas denitrificans AS-P1. The isolate, named PHB2, showed good PHB-accumulating activity (up to 32% of the cell dry weight). PHB accumulated by isolate PHB2 was able to protect Artemia completely from the V. campbellii strain. Our data indicate that PHB-accumulating bacteria, such as B. denitrificans PHB2, could be used as an an effective and economically interesting alternative strategy to control infections in aquaculture. [ABSTRACT FROM AUTHOR]

An environmentally representative stagnant-water model was developed to monitor the growth dynamics of Legionella pneumophila. This model was evaluated for three distinct water treatments: untreated tap water, heat-treated tap water, and heat-treated tap water supplemented with Pseudomonas putida, a known biofilm-forming bacterium. Bringing heat-treated tap water after subsequent cooling into contact with a densely formed untreated biofilm was found to promote the number of L. pneumophila by 4 log units within the biofilm, while the use of untreated water only sustained the L. pneumophila levels. Subsequent colonization of the water phase by L. pneumophila was noticed in the heat-treated stagnant-water models, with concentrations as high as 1 × 1010 mip gene copies L−1 stagnant water. Denaturing gradient gel electrophoresis in combination with clustering analysis of the prokaryotic community in the water phase and in the biofilm phase suggests that the different water treatments induced different communities. Moreover, boosts of L. pneumophila arising from heat treatment of water were accompanied by shifts to a more diverse eukaryotic community. Stimulated growth of L. pneumophila after heating of the water may explain the rapid recolonization of L. pneumophila in water systems. These results highlight the need for additional or alternative measures to heat treatment of water in order to prevent or abate potential outbreaks of L. pneumophila. [ABSTRACT FROM AUTHOR]

In most natural environments as well as in engineered environments, such as wastewater treatment plants, ammonia-oxidizing bacteria (AOB) experience fluctuating substrate concentrations. Several physiological traits, such as low maintenance energy demand and decay rate, cell-to-cell communication, cell mobility, stable enzymes and RNAs, could allow AOB to maintain themselves under unfavourable circumstances. This review examines whether AOB possess such traits and how these traits might offer advantages over competing organisms such as heterotrophic bacteria during periods of starvation. In addition, within the AOB groups, differences exist in adaptation to and competitiveness under conditions of high or low ammonia or oxygen concentrations. Because these findings are of importance with regard to the ecology and activity of AOB in natural and engineered environments, concluding remarks are directed towards future research objectives that may clarify unanswered questions, thereby contributing to the general knowledge of the ecology and activity of ammonia oxidizers. [ABSTRACT FROM AUTHOR]

An evolutionary algorithm was used to determine the optimal combination of parameters for transconjugant formation. As a model system, a gfp tagged TOL plasmid pWW0 was chosen to examine transfer from Pseudomonas putida to Escherichia coli. A comparison of flow cytometry results with plating and microscopy showed that the majority of transconjugants were not culturable. The transconjugant ratio therefore was determined by flow cytometry. The evolutionary algorithm showed that the optimal conditions were obtained at 28°C and at the highest nutrient concentrations. This work demonstrates that evolutionary algorithms can be used to find optimal parameter interactions in environmental microbiology. [ABSTRACT FROM AUTHOR]

Abstract: The prebiotic potential of native chicory inulin was assessed in the Simulator of the Human Intestinal Microbial Ecosystem (SHIME) by monitoring microbial community from the colon compartments, its metabolic activity and community structure. Inulin addition selected for a higher short chain fatty acid production with shifts towards propionic and butyric acid. Conventional culture-based techniques and PCR-denaturing gradient gel electrophoresis analysis showed no remarkable changes in the overall microbial community from the colon compartments of the SHIME, whereas selective effects were seen for lactic acid bacteria. Quantitative PCR with bifidobacteria-specific primers revealed a significant increase with more than 1 log CFU ml−1 from the proximal to distal colon, in contrast to culture-based techniques, which only showed a minor bifidogenic effect in the ascending colon. Our results indicate that inulin purports prebiotic effects from the proximal to distal colon and that real-time PCR is a more precise technique to detect differences in bifidobacterial populations whereas conventional culturing techniques are much more variable. [Copyright &y& Elsevier]

Plasmid pC1 of Delftia acidovorans CA28, encoding the oxidative deamination of 3-chloroaniline (3-CA), was tagged with a mini-Tn5 transposon containing the gfp gene. The labelled plasmid, designated pC1gfp, was subsequently transferred to Pseudomonas putida UWC3 and the plasmid transfer from this donor to the bacterial community in activated sludge was studied. Mating experiments were performed on both Luria–Bertani agar plates and directly in liquid activated sludge. Green fluorescent colonies appearing on mineral medium containing 3-CA as sole nitrogen source were picked up and verified to be true pC1gfp-harbouring transconjugants. REP- and BOX-PCR genomic fingerprinting revealed a large diversity in the transconjugant collection, indicating the occurrence of multiple plasmid transfer events. Remarkably, Luria–Bertani agar plate matings yielded a different set of transconjugants compared to plasmid transfers directly in liquid activated sludge. From the plate matings, mainly Aeromonas sp. was isolated, whereas D. acidovorans strains were predominant in the transconjugant collection from liquid activated sludge. The majority of the isolates showed 3-CA deamination, probably to chlorocatechol, but several pC1gfp-harbouring transconjugants performed a rapid and complete 3-CA degradation. [Copyright &y& Elsevier]

Genes that encode the degradation of both naturally occurring and xenobiotic organic compounds are often located on plasmids, transposons or other mobile and/or integrative elements. The list of published reports of such mobile genetic elements (MGEs) keeps growing as researchers continue to isolate and characterize new degrading bacteria and their corresponding degradative genes. There is also growing evidence that horizontal exchange of catabolic (degradative) genes among bacteria in microbial communities plays an important role in the evolution of catabolic pathways. Around 10 years ago the hypothesis was raised that we might be able to accelerate this natural gene exchange and pathway construction by introducing and subsequently spreading degradative genes, located on MGEs, into well established, competitive indigenous microbial populations as a means of bioaugmentation of polluted soils and waters. During the last decade, only a few reports on successful MGE- mediated bioaugmentation have been published. After summarizing the diversity of degradative MGEs, this review presents an overview of studies that have monitored the transfer of degradative genes in soil microcosms and in activated sludge and other wastewater treatment reactors, with emphasis on those that have clearly shown a direct effect of gene transfer on accelerated biodegradation. A few successful cases suggest that the strategy could indeed work under specific conditions, such as when the in situ degradation potential is absent and the pollutant degrading transconjugants can grow and become numerically dominant populations in the bacterial community. Further studies in this area are obviously needed to improve our current knowledge on the efficiency of gene dissemination as a tool in bioremediation. [Copyright &y& Elsevier]

The diversity of bacterial groups of activated sludge samples that received wastewater from four different types of industry was investigated by a nested PCR–DGGE (denaturing gradient gel electrophoresis) approach. Specific 16S rRNA primers were chosen for large bacterial groups (Bacteria and α-Proteobacteria in particular), which dominate activated sludge communities, as well as for actinomycetes, ammonium oxidisers and methanotrophs (types I and II). In addition primers for the new Acidobacterium kingdom were used to observe their community structure in activated sludge. After this first PCR amplification, a second PCR with bacterial primers yielded 16S rRNA gene fragments that were subsequently separated by DGGE, thus generating ‘group-specific DGGE patterns’. The community structure and diversity of the bacterial groups from the different samples was further analysed using different techniques, such as statistical analysis and Shannon diversity index evaluation of the band patterns. By combining the seven DGGE gels, cluster analysis, multidimensional scaling and principal component analysis clearly clustered two of the four activated sludge types separately. It was shown that the combination of molecular and statistical methods can be very useful to differentiate microbial communities. [Copyright &y& Elsevier]

Biofilm formation on 316L stainless steel was investigated in a pilotscale flow-through system fed with brackish surface water using an alternating flow/stagnation/flow regime. Microbial community analysis by denaturing gradient gel electrophoresis and sequencing revealed the presence of complex microbial ecosystems consisting of, amongst others, Leptothrix-related manganese-oxidizing bacteria in the adjacent water, and sulfur-oxidizing, sulfate-reducing and slime-producing bacteria in the biofilm. Selective plating of the biofilm indicated the presence of high levels of manganese-oxidizing microorganisms, while microscopic and chemical analyses of the biofilm confirmed the presence of filamentous manganese-precipitating microorganisms, most probably Leptothrix species. Strong accumulation of iron and manganese occurred in the biofilm relative to the adjacent water. No evidence of selective colonization of the steel surface or biocorrosion was found over the experimental period. The overall results of this study highlight the potential formation of complex microbial biofilm communities in flow-through systems thriving on minor concentrations of manganese. [Copyright &y& Elsevier]