Validation of crAssphage microbial source tracking markers and comparison with Bacteroidales markers for detection and quantification of faecal contaminations in surface water.

Human-specific faecal contamination has been affecting surface water and is a threat to both the environment and public health due to its potential co-occurrence with pathogens. Extended studies were conducted to detect and quantify faecal contamination using microbial source tracking (MST) markers targeting bacteria and viruses. The prototypical crAssphage, a presumed Bacteroides-infecting phage discovered in 2014, showed superior specificity to human faeces and high abundance in untreated sewage water.

The rat telemetry assay and venous catheter access buttons for use in cardiovascular safety pharmacology assessments - Surgical methods, refinements and colony maintenance.

Introduction: Rat telemetry is the assay of choice to assess the potential effects of novel drug candidates on cardiovascular parameters during early drug discovery. Telemetry device implantation can be combined with venous catheter and access button implantation when intravenous administration of the drug substance is required.

Methods: Rats (Sprague Dawley or Han Wistar) were implanted with telemetry devices for arterial blood pressure measurement using either direct aortic catheterisation (n = 131) or aortic catheterisation via the femoral artery (n = 17).

Tracing the fate of wastewater viruses reveals catchment-scale virome diversity and connectivity.

The discharge of wastewater-derived viruses in aquatic environments impacts catchment-scale virome composition. To explore this, we used viromic analysis of RNA and DNA virus-like particles to holistically track virus communities entering and leaving wastewater treatment plants and the connecting river catchment system and estuary. We reconstructed >40 000 partial viral genomes into 10 149 species-level groups, dominated by dsDNA and (+)ssRNA bacteriophages (Caudoviricetes and Leviviricetes) and a small number of genomes that could pose a risk to human health.

The Site-Specific Recombination System of the Bacteriophage Φ24.

Stx bacteriophages are members of the lambdoid group of phages and are responsible for Shiga toxin (Stx) production and the dissemination of Shiga toxin genes () across shigatoxigenic (STEC). These toxigenic bacteriophage hosts can cause life-threatening illnesses, and Stx is the virulence determinant responsible for the severe nature of infection with enterohemorrhagic , a subset of pathogenic STEC. Stx phages are temperate, and in the present study, the identification of what is actually required for Stx phage Φ24 and bacterial DNA recombination was tested using both and recombination assays.

Impact of Temperature, Nutrients and Heavy Metals on Bacterial Diversity and Ecosystem Functioning Studied by Freshwater Microcosms and High-Throughput DNA Sequencing.

Microbial communities are fundamental components in freshwater, and community shifts in ecosystem structure are indicative of changing environmental conditions. This study aimed at investigating the influence of key environmental parameters on bacterial diversity and ecosystem functioning (i.e.

Next-generation sequencing reveals fecal contamination and potentially pathogenic bacteria in a major inflow river of Taihu Lake.

Taihu Lake is one of the largest freshwater lakes in China and serves as an important source for drinking water. This lake is suffering from eutrophication, cyanobacterial blooms and fecal pollution, and the inflow Tiaoxi River is one of the main contributors. The goal here was to characterize the bacterial community structure of Tiaoxi River water by next-generation sequencing (NGS), paying attention to bacteria that are either fecal-associated or pathogenic, and to examine the relationship between environmental parameters and bacterial community structure.

Quantification of Microbial Source Tracking and Pathogenic Bacterial Markers in Water and Sediments of Tiaoxi River (Taihu Watershed).

Taihu Lake is one of the largest freshwater lakes in China, serving as an important source of drinking water; >60% of source water to this lake is provided by the Tiaoxi River. This river faces serious fecal contamination issues, and therefore, a comprehensive investigation to identify the sources of fecal contamination was carried out and is presented here. The performance of existing universal (BacUni and GenBac), human (HF183-Taqman, HF183-SYBR, BacHum, and Hum2), swine (Pig-2-Bac), ruminant (BacCow), and avian (AV4143 and GFD) associated microbial source tracking (MST) markers was evaluated prior to their application in this region.

Urbanization Impacts the Physicochemical Characteristics and Abundance of Fecal Markers and Bacterial Pathogens in Surface Water.

Urbanization is increasing worldwide and is happening at a rapid rate in China in line with economic development. Urbanization can lead to major changes in freshwater environments through multiple chemical and microbial contaminants. We assessed the impact of urbanization on physicochemical characteristics and microbial loading in canals in Suzhou, a city that has experienced rapid urbanization in recent decades.

Viromic Analysis of Wastewater Input to a River Catchment Reveals a Diverse Assemblage of RNA Viruses.

Detection of viruses in the environment is heavily dependent on PCR-based approaches that require reference sequences for primer design. While this strategy can accurately detect known viruses, it will not find novel genotypes or emerging and invasive viral species. In this study, we investigated the use of viromics, i.

Lignocellulose-Degrading Microbial Communities in Landfill Sites Represent a Repository of Unexplored Biomass-Degrading Diversity.

The microbial conversion of lignocellulosic biomass for biofuel production represents a renewable alternative to fossil fuels. However, the discovery of new microbial enzymes with high activity is critical for improving biomass conversion processes. While attempts to identify superior lignocellulose-degrading enzymes have focused predominantly on the animal gut, biomass-degrading communities in landfill sites represent an unexplored resource of hydrolytic enzymes for biomass conversion.

Fifty important research questions in microbial ecology.

Microbial ecology provides insights into the ecological and evolutionary dynamics of microbial communities underpinning every ecosystem on Earth. Microbial communities can now be investigated in unprecedented detail, although there is still a wealth of open questions to be tackled. Here we identify 50 research questions of fundamental importance to the science or application of microbial ecology, with the intention of summarising the field and bringing focus to new research avenues.

Acute liver effects, disposition and metabolic fate of [C]-fenclozic acid following oral administration to normal and bile-cannulated male C57BL/6J mice.

The distribution, metabolism, excretion and hepatic effects of the human hepatotoxin fenclozic acid were investigated following single oral doses of 10 mg/kg to normal and bile duct-cannulated male C57BL/6J mice. Whole body autoradiography showed distribution into all tissues except the brain, with radioactivity still detectable in blood, kidney and liver at 72 h post-dose. Mice dosed with [C]-fenclozic acid showed acute centrilobular hepatocellular necrosis, but no other regions of the liver were affected.

Characterising the Canine Oral Microbiome by Direct Sequencing of Reverse-Transcribed rRNA Molecules.

PCR amplification and sequencing of phylogenetic markers, primarily Small Sub-Unit ribosomal RNA (SSU rRNA) genes, has been the paradigm for defining the taxonomic composition of microbiomes. However, 'universal' SSU rRNA gene PCR primer sets are likely to miss much of the diversity therein. We sequenced a library comprising purified and reverse-transcribed SSU rRNA (RT-SSU rRNA) molecules from the canine oral microbiome and compared it to a general bacterial 16S rRNA gene PCR amplicon library generated from the same biological sample.

Transcriptomic analysis of Shiga-toxigenic bacteriophage carriage reveals a profound regulatory effect on acid resistance in Escherichia coli.

Shiga-toxigenic bacteriophages are converting lambdoid phages that impart the ability to produce Shiga toxin to their hosts. Little is known about the function of most of the genes carried by these phages or the impact that lysogeny has on the Escherichia coli host. Here we use next-generation sequencing to compare the transcriptomes of E.

Anoxic Biodegradation of Isosaccharinic Acids at Alkaline pH by Natural Microbial Communities.

One design concept for the long-term management of the UK's intermediate level radioactive wastes (ILW) is disposal to a cementitious geological disposal facility (GDF). Under the alkaline (10.013.

Bacterio-plankton transformation of diazepam and 2-amino-5-chlorobenzophenone in river waters.

Benzodiazepines are a large class of commonly-prescribed drugs used to treat a variety of clinical disorders. They have been shown to produce ecological effects at environmental concentrations, making understanding their fate in aquatic environments very important. In this study, uptake and biotransformations by riverine bacterio-plankton of the benzodiazepine, diazepam, and 2-amino-5-chlorobenzophenone, ACB (a photo-degradation product of diazepam and several other benzodiazepines), were investigated using batch microcosm incubations.

Development and application of a method for the purification of free shigatoxigenic bacteriophage from environmental samples.

Shiga toxin producing Escherichia coli (STEC) strains are foodborne pathogens whose ability to produce Shiga toxin (Stx) is due to the integration of Stx-encoding lambdoid bacteriophage (Stx phage). Circulating, infective Stx phages are very difficult to isolate, purify and propagate such that there is no information on their genetic composition and properties. Here we describe a novel approach that exploits the phage's ability to infect their host and form a lysogen, thus enabling purification of Stx phages by a series of sequential lysogen isolation and induction steps.

Comparative genomics of Shiga toxin encoding bacteriophages.

Background: Stx bacteriophages are responsible for driving the dissemination of Stx toxin genes (stx) across their bacterial host range. Lysogens carrying Stx phages can cause severe, life-threatening disease and Stx toxin is an integral virulence factor. The Stx-bacteriophage vB_EcoP-24B, commonly referred to as Ф24B, is capable of multiply infecting a single bacterial host cell at a high frequency, with secondary infection increasing the rate at which subsequent bacteriophage infections can occur.

Metagenomic approaches to the discovery of cellulases.

Most of the microorganisms responsible for nutrient cycling in the environment have yet to be cultivated, and this could include those species responsible for the degradation of cellulose. Known cellulases are well defined at the protein sequence level, but gene variants are difficult to amplify from environmental DNA. The identification of novel cellulase genes independent of DNA amplification is made possible by adopting a direct metagenome sequencing approach to provide genes that can be cloned, expressed, and characterized prior to potential exploitation, all in the absence of any information on the species from which they originated.

Methods for the isolation of cellulose-degrading microorganisms.

The biodegradation of lignocellulose, the most abundant organic material in the biosphere, is a feature of many aerobic, facultatively anaerobic and obligately anaerobic bacteria and fungi. Despite widely recognized difficulties in the isolation and cultivation of individual microbial species from complex microbial populations and environments, significant progress has been made in recovering cellulolytic taxa from a range of ecological niches including the human, herbivore, and termite gut, and terrestrial, aquatic, and managed environments. Knowledge of cellulose-degrading microbial taxa is of significant importance with respect to nutrition, biodegradation, biotechnology, and the carbon-cycle, providing insights into the metabolism, physiology, and functional enzyme systems of the cellulolytic bacteria and fungi that are responsible for the largest flow of carbon in the biosphere.

Identification of genes expressed in cultures of E. coli lysogens carrying the Shiga toxin-encoding prophage Φ24B.

Background: Shigatoxigenic E. coli are a global and emerging health concern. Shiga toxin, Stx, is encoded on the genome of temperate, lambdoid Stx phages.

Importance of Micromonospora spp. as colonizers of cellulose in freshwater lakes as demonstrated by quantitative reverse transcriptase PCR of 16S rRNA.

The relative abundance of micromonosporas in the bacterial communities inhabiting cellulose baits, water columns, and sediments of two freshwater lakes was determined by quantitative PCR (qPCR) of reverse-transcribed 16S rRNA. Micromonospora spp. were shown to be significant members of the active bacterial population colonizing cellulosic substrates in the lake sediment, and their increased prevalence with greater depth was confirmed by enumeration of CFU.

The microbial ecology of anaerobic cellulose degradation in municipal waste landfill sites: evidence of a role for fibrobacters.

Cellulose is reputedly the most abundant organic polymer in the biosphere, yet despite the fundamental role of cellulolytic microorganisms in global carbon cycling and as potential sources of novel enzymes for biotechnology, their identity and ecology is not well established. Cellulose is a major component of landfill waste and its degradation is therefore a key feature of the anaerobic microbial decomposition process. Here, we targeted a number of taxa containing known cellulolytic anaerobes (members of the bacterial genus Fibrobacter, lineages of Clostridium clusters I, III, IV and XIV, and anaerobic fungi of the Neocallimastigales) in landfill leachate and colonized cellulose 'baits' via PCR and quantitative PCR (qPCR).

The Fibrobacteres: an important phylum of cellulose-degrading bacteria.

The phylum Fibrobacteres currently comprises one formal genus, Fibrobacter, and two cultured species, Fibrobacter succinogenes and Fibrobacter intestinalis, that are recognised as major bacterial degraders of lignocellulosic material in the herbivore gut. Historically, members of the genus Fibrobacter were thought to only occupy mammalian intestinal tracts. However, recent 16S rRNA gene-targeted molecular approaches have demonstrated that novel centres of variation within the genus Fibrobacter are present in landfill sites and freshwater lakes, and their relative abundance suggests a potential role for fibrobacters in cellulose degradation beyond the herbivore gut.