PCR primer sets for the 16S rRNA gene of six phylogenetic groups of sulfate-reducing bacteria (SRB) were designed. Their application in conjunction with group-specific internal oligonucleotide probes was used to detect SRB DNA in samples of landfill leachate. Six generic/suprageneric groups could be differentiated: Desulfotomaculum; Desulfobulbus; Desulfobacterium; Desulfobacter; Desulfococcus–Desulfonema–Desulfosarcina; Desulfovibrio–Desulfomicrobium. The predicted specificities of the PCR primer and oligonucleotide probe combinations were confirmed with DNA from reference strains. In all cases, the PCR primers and probes were specific, the only exception being that the Desulfococcus–Desulfonema–Desulfosarcina (group 5) PCR primers were able to amplify DNA from Desulfobacterium (group 3) reference strains but these groups could nevertheless be differentiated with the internal oligonucleotide probes. The proliferation of SRB in landfill sites interferes with methanogenesis and waste stabilization, but relatively little is known about the composition of SRB populations in this environment. DNA was extracted from samples of landfill leachate from several municipal waste landfill sites and used as template in PCR reactions with SRB group-specific primer sets. Group-specific oligonucleotide probes were then used to confirm that the PCR products obtained contained the target SRB 16S rDNA. Both ‘direct’ and ‘nested’ PCR protocols were used to amplify SRB 16S rDNA from landfill leachates. Three of the six SRB groups could be detected using the ‘direct’ PCR approach (Desulfotomaculum, Desulfobacter and Desulfococcus–Desulfonema–Desulfosarcina). When ‘nested’ PCR was applied, an additional two groups could be detected (Desulfobulbus and Desulfovibrio–Desulfomicrobium). Only Desulfobacterium could not be detected in any leachate samples using either direct or nested PCR. The SRB-specific 16S rDNA primers and probes described here can be applied to investigations of SRB molecular ecology in general, and can be further developed for examining SRB population composition in relation to landfill site performance.

Taihu Lake is the third largest freshwater lake in China and serves as a drinking water source for ~30 million residents. Tiaoxi River is one of the main rivers connected to this lake and contributes >60% of the source water. Taihu Lake has been facing various environmental issues; therefore, it is important to study the water quality of its inflow rivers. This study aimed to evaluate the physico-chemical and microbiological characteristics of Tiaoxi River and to determine the spatial and seasonal variations in the water quality. Water samples were collected from 25 locations across the Tiaoxi River in three seasons in 2014–2015. Fourteen water quality parameters including multiple nutrients and indicator bacteria were assessed, and the data analyzed by multivariate statistical analyses. The physico-chemical analysis showed high levels (>1 mg/L) of total nitrogen (TN) in all locations for all seasons. Total phosphorus (TP), nitrite-N (NO₂-N), and ammonium-N (NH₄-N) exceeded the acceptable limits in some locations and fecal coliform counts were high (>250 CFU/100 mL) in 15 locations. Hierarchical cluster analysis showed that the sampling sites could be grouped into three clusters based on water quality, which were categorized as low, moderate, and high pollution areas. Principal component analysis (PCA) applied to the entire dataset identified four principal components which explained 83% of the variation; pH, conductivity, TP, and NO₃-N were found to be the key parameters responsible for variations in water quality. The overall results indicated that some of the sampling locations in the Tiaoxi River are heavily contaminated with pollutants from various sources which can be correlated with land use patterns and anthropogenic activities.

A verocytotoxigenic bacteriophage isolated from a strain of enterohemorrhagic Escherichia coli O157, into which a kanamycin resistance gene ( aph3 ) had been inserted to inactivate the verocytotoxin gene ( vt ₂ ), was used to infect Enterobacteriaceae strains. A number of Shigella and E. coli strains were susceptible to lysogenic infection, and a smooth E. coli isolate (O107) was also susceptible to lytic infection. The lysogenized strains included different smooth E. coli serotypes of both human and animal origin, indicating that this bacteriophage has a substantial capacity to disseminate verocytotoxin genes. A novel indirect plaque assay utilizing an E. coli recA441 mutant in which phage-infected cells can enter only the lytic cycle, enabling detection of all infective phage, was developed.

Polysaccharides are an important source of organic carbon in the marine environment and degradation of the insoluble and globally abundant cellulose is a major component of the marine carbon cycle. Although a number of species of cultured bacteria are known to degrade crystalline cellulose, little is known of the polysaccharide hydrolases expressed by cellulose-degrading microbial communities, particularly in the marine environment. Next generation 454 Pyrosequencing was applied to analyze the microbial community that colonizes and degrades insoluble polysaccharides in situ in the Irish Sea. The bioinformatics tool MG-RAST was used to examine the randomly sampled data for taxonomic markers and functional genes, and showed that the community was dominated by members of the Gammaproteobacteria and Bacteroidetes. Furthermore, the identification of 211 gene sequences matched to a custom-made database comprising the members of nine glycoside hydrolase families revealed an extensive repertoire of functional genes predicted to be involved in cellulose utilization. This demonstrates that the use of an in situ cellulose baiting method yielded a marine microbial metagenome considerably enriched in functional genes involved in polysaccharide degradation. The research reported here is the first designed to specifically address the bacterial communities that colonize and degrade cellulose in the marine environment and to evaluate the glycoside hydrolase (cellulase and chitinase) gene repertoire of that community, in the absence of the biases associated with PCR-based molecular techniques.

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. Nine sampling locations covering three urban intensity classes (high, medium and low) in Suzhou were selected for field studies and three locations in Huangshan (natural reserve) were included as pristine control locations. Water samples were collected for physicochemical, microbiological and molecular analyses. Compared to medium and low urbanization sites, there were statistically significant higher levels of nutrients and total and thermotolerant coliforms (or fecal coliforms) in highly urbanized locations. The effect of urbanization was also apparent in the abundances of human-associated fecal markers and bacterial pathogens in water samples from highly urbanized locations. These results correlated well with land use types and anthropogenic activities at the sampling sites. The overall results indicate that urbanization negatively impacts water quality, providing high levels of nutrients and a microbial load that includes fecal markers and pathogens.

The autotrophic ammonia-oxidizing bacteria in a eutrophic freshwater lake were studied over a 12-month period. Numbers of ammonia oxidisers in the lakewater were small throughout the year, and tangential-flow concentration was required to obtain meaningful estimates of most probable numbers. Sediments from littoral and profundal sites supported comparatively large populations of these bacteria, and the nitrification potential was high, particularly in summer samples from the littoral sediment surface. In enrichment cultures, lakewater samples nitrified at low (0.67 mM) ammonium concentrations only whereas sediment samples exhibited nitrification at high (12.5 mM) ammonium concentrations also. Enrichments at low ammonium concentration did not nitrify when inoculated into high-ammonium medium, but the converse was not true. This suggests that the water column contains a population of ammonia oxidizers that is sensitive to high ammonium concentrations. The observation of nitrification at high ammonium concentration by isolates from some winter lakewater samples, identified as nitrosospiras by 16S rRNA probing, is consistent with the hypothesis that sediment ammonia oxidizers enter the water column at overturn. With only one exception, nested PCR amplification enabled the detection of Nitrosospira 16S rDNA in all samples, but Nitrosomonas ( N. europaea-eutropha lineage) 16S rDNA was never obtained. However, the latter were part of the sediment and water column communities, because their 16S rRNA could be detected by specific oligonucleotide probing of enrichment cultures. Furthermore, a specific PCR amplification regime for the Nitrosomonas europaea ammonia monooxygenase gene ( amoA ) yielded positive results when applied directly to sediment and lakewater samples. Patterns of Nitrosospira and Nitrosomonas detection by 16S rRNA oligonucleotide probing of sediment enrichment cultures were complex, but lakewater enrichments at low ammonium concentration were positive for nitrosomonads and not nitrosospiras. Analysis of enrichment cultures has therefore provided evidence for the existence of subpopulations within the lake ammonia-oxidizing community distinguishable on the basis of ammonium tolerance and possibly showing a seasonal distribution between the sediment and water column.