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Diversity of the microbial community in the Zharkent geothermal hot spring, located in the southeastern region of Kazakhstan, was assessed using both culture-dependent and -independent approaches. Shotgun metagenomic sequencing of DNA extracted from the spring water yielded 11,061,725 high-quality sequence reads, totaling >1,67 Gb of nucleotide sequences. Furthermore, water samples were enriched in nutrient broth at varying high temperatures, and colonies isolated by being streaked onto nutrient agar. Finally, DNA extraction and amplification, as well as sequencing and phylogenetic analysis, were conducted. Bacteria constituted more than 99.97% of the total prokaryotic abundance, with Archaea contributing only an extremely small component; Firmicutes, Proteobacteria, and Actinobacteria dominated the community. At genus level, Firmicutes reads affiliated with Desmospora, Parageobacillus, Paenibacillus, and Brevibacillus, accounting for more than 60% of total prokaryotic abundance. Eight morphologically distinct, aerobic, endospore-forming thermophilic bacteria were recovered; isolates differed significantly in substrate utilization patterns, as well as their production of thermophilic, extracellular, hydrolytic enzymes for degradation of starch, lipids, cellulose, and protein. Five strains could degrade all four macromolecular types at temperatures ranging from 55 to 75 °C. Phylogenetic analyses based on 16S rRNA gene sequences placed all isolates into the genus Geobacillus with some of them possibly representing novel species. The results indicate that this hot spring represents a rich source of novel thermophilic bacteria and potentially useful thermostable enzymes., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The impact of the heavy metal contamination and acidity on the bacterial community was studied in samples collected from the Akhtala copper mine tailing using molecular approaches. The bacterial community structure analysis by PCR-DGGE fingerprinting revealed an abundance of Firmicutes, Acidobacteria, and Proteobacteria in different layers of the Akhtala tailing. 454 pyrotag sequence analyses revealed that a significant part of the sequences originated from Proteobacteria (49%) and Bacteroidetes (43%). Bacterial taxa are distributed also in phyla Saccharibacteria (2%), Verrucomicrobia (1.5%), Gammatimonadetes (1%), and some minor additional bacterial groups. The main primary producers in the Akhtala tailing appear to be obligate autotrophic Thiobacillus and Sulfuritalea species. Representatives of Lutibacter and Lysobacter genera are the most abundant acid-tolerant heterotrophs in the studied tailing. The presence of a large number of yet-uncultivated species emphasizes the importance of the future exploration of the tailing as an important source of novel bacteria.

The environmental Escherichia albertii strain DM104, which cross-reacts serologically with Shigella dysenteriae was assessed for pathogenic properties, immunogenicity, and protective efficacy in different animal models to evaluate it as a vaccine candidate against S. dysenteriae, which causes the severe disease, shigellosis. The DM104 isolate was found to be non-invasive and did not produce any entero- or cyto-toxins. The strain also showed negative results in the mouse lethal activity assay. The non-pathogenic DM104 strain gave, however, a high protective efficacy as an ocularly administered vaccine in the guinea pig eye model against S. dysenteriae type 4 challenge. It also induced a high titer of serum IgG against S. dysenteriae type 4 whole cell lysate and lipopolysaccharide. Taken together, all these results indicate a good potential for the use of the DM104 as a live vaccine candidate against shigellosis.

An environmental freshwater bacterial isolate, DM104, appearing as Shigella-like colonies on selective agar plates was found to show strong and specific serological cross-reactivity with Shigella dysenteriae type 4. Biochemical identification according to the analytical profile index, molecular serotyping by restriction of the amplified O-antigen gene cluster (rfb-RFLP), together with phylogenetic analysis of the 16S rRNA gene and multi-locus sequence analysis, identified the isolate as Escherichia albertii. rfb-RFLP of DM104, revealed a profile different from that of S. dysenteriae type 4. However, western blot analysis of extracted lipopolysaccharides demonstrated strong cross-reactivity with S. dysenteriae type 4 using specific monovalent antisera and a lipopolysaccharide gel banding profile similar to that of S. dysenteriae type 4. The observed O-antigen cross-reaction between an E. albertii isolate and S. dysenteriae extends our knowledge of the extent of O-antigen cross-reaction within the Escherichia/Shigella group of organisms, and offers the possibility of using DM104 and similar cross-reacting strains as shigellosis vaccine candidates.

A culture-independent study of the bacterial diversity in Lake Dhanmondi, located in the central region of Dhaka city, Bangladesh, was carried out using deep sequence analysis of 16S rRNA gene PCR amplicons. The results revealed the presence of a group of bacteria, termed LD11, phylogenetically unrelated to any previously cultivated bacteria at the phylum level. LD11 sequences comprised about 1.7 % of the total sequence reads after quality assessment. LD11 appears to constitute a novel division with a deep evolutionary lineage apparently branching between the Chloroflexi and Thermi-Deinococci phyla. Sequence similarity with molecular data from freshwater environments indicates that LD11 represents a widespread and novel clade of freshwater bacteria for which no cultivated representatives are yet available.

Little is known about the distribution, survival, and transmission of Shigella in environmental surface waters. To gain more insight into the environmental biology of Shigella we isolated five bacterial strains serotyped as Shigella flexneri 2b from a freshwater lake in Bangladesh using a modified nutrient broth supplemented with nucleic acid bases. The biochemical properties of the isolates, including inability to ferment lactose and a negative lysine decarboxylase test, indicated common physiological characteristics with Shigella, but differed significantly from that of standard clinical strains. The isolates possessed the ipaH virulence gene and a megaplasmid, but lacked other Shigella-related virulence marker genes. Genetic fingerprinting and sequence analysis of housekeeping genes confirmed the strains as S. flexneri isolates. An apparent clonal origin of strains recovered with a one-year interval indicates a strong environmental selection pressure on Shigella for persistence in the freshwater environment. The lack of a complete set of virulence genes as well as uncommon biochemical properties suggest that these strains might represent a group of non-invasive and atypical environmental Shigella variants, with the potential for further elucidation of the survival mechanism, diversity, and emergence of virulent Shigella in tropical freshwater environments.

Using protocols designed for the isolation of Shigella from environmental freshwater samples from different regions of Bangladesh, 11 bacterial strains giving rise to Shigella-like colonies on selective agar plates and showing serological cross-reaction with Shigella-specific antisera were isolated. Phylogenetic analyses revealed that three of the isolates were most closely related to Escherichia coli, four to Enterobacter sp., two to Stenotrophomonas, and two isolates belonged to the Gram-positive genus Aerococcus. The isolates cross-reacted with six different serotypes of Shigella and were, in each case, highly type-specific. Two of the isolates belonging to the Enterobacter and Escherichia genera gave extremely strong cross-reactivity with Shigella dysenteriae and Shigella boydii antisera, respectively. The Aerococcus isolates gave relatively weak but significant cross-reactions with S. dysenteriae. Western blot analysis revealed that a number of antigens from the isolates cross-react with Shigella spp. The results indicate that important Shigella spp. surface antigens are shared by a number of environmental bacteria, which have implications for the use of serological methods in attempts for the detection and recovery of Shigella from aquatic environments.