Porcine sapeloviruses (PSVs) are widely distributed in pig populations; however, little information on their evolutionary history and the mechanisms driving their divergence is available. Therefore, in the present study, 241 fecal samples and 91 intestinal contents collected from pigs at 26 farms in Hunan, China, were tested for the presence of PSVs. The overall PSV positivity rate was 46.39 %, with a particularly high infection rate detected in nursery and fattening pigs. A total of 29 PSV strains (PSV-HuNs) were isolated, with these showing high genetic diversity based on phylogenetic and pairwise distance analyses of the capsid-protein gene sequences. Incongruence between phylognetic trees of the capsid-protein and 3CD regions indicated frequent recombination within the PSV-HuNs, and a putative recombinant hotspot near the 3' end of the P1 region was identified. Our results suggested that recombination played an important role in driving PSV genetic diversity and evolution.

Streptococcus suis is a zoonotic pathogen that harbors anti-oxidative stress genes, which have been reported to be associated with virulence. Serial passage has been widely used to obtain phenotypic variant strains to investigate the functions of important genes. In the present study, S. suis serotype 9 strain DN13 was serially passaged in mice 30 times. The virulence of a single colony from passage 10 (SS9-P10) was found to increase by at least 140-fold as indicated by LD50 values, and the increased virulence was stable for single colonies from passage 20 (SS0-P20) and 30 (SS0-P30). Compared to the parental strain, the mouse-adapted strains were more tolerant to oxidative and high temperature stress. Genome-wide analysis of nucleotide variations found that reverse mutations occurred in seven genes, as indicated by BLAST analysis. Three of the reverse mutation genes or their homologs in other bacteria were reported to be virulence-associated, including ideSsuis in S. suis, a homolog of malR of Streptococcus pneumoniae, and a homolog of the prepilin peptidase-encoding gene in Legionella pneumophila. However, these genes were not involved in the stress response. Another gene, srtR (stress response transcriptional regulator), encoding an XRE family transcriptional regulator, which had an internal stop in the parental strain, was functionally restored in the adapted strains. Further analysis of DN13 and SS9-P10-background srtR-knock-out and complementing strains supported the contribution of this gene to stress tolerance in vitro and virulence in mice. srtR and its homologs are widely distributed in Gram-positive bacteria including several important human pathogens such as Enterococcus faecium and Clostridioides difficile, indicating similar functions in these bacteria. Taken together, our study identified the first member of the XRE family of transcriptional regulators that is involved in stress tolerance and virulence. It also provides insight into the mechanism of enhanced virulence after serial passage in experimental animals.

Deinagkistrodon acutus is a venomous pit viper commonly used in traditional Chinese medicine; farming these snakes has become a major industry. In 2017, an outbreak of fatal diarrhoea among farm-raised D. acutus in Hunan Province caused the deaths of 5,600 snakes within 3 weeks. We isolated a brand-new sequence type of Aeromonas hydrophila (ST516) from lesions and confirmed that this bacterium was the causal agent of the outbreak. Snakes infected with the bacterium in the laboratory showed similar clinical symptoms to those of snakes in the original outbreak. We also tested bacterial virulence in Kunming mice to examine the likelihood of zoonosis. Isolates were pathogenic to mice, causing diarrhoea within 4 hr post-challenge, which indicates that the bacterium can potentially infect mammals. Environmental analysis showed that polluted spring water likely caused the diarrhoea in snakes. This study is the first to report on a large-scale outbreak of fatal diarrhoea in farm-raised snakes, originating in a pathogen that can infect mammals. These results should raise awareness regarding potential anthropozoonosis among poikilotherms, mammals, and humans; appropriate prevention or control methods should be developed.

Bacterial surface proteins can be good vaccine candidates. In the present study, we used polyclonal antibodies purified with intact Erysipelothrix rhusiopthiae to screen phage-displayed random dodecapeptide and loop-constrained heptapeptide libraries, which led to the identification of mimotopes. Homology search of the mimotope sequences against E. rhusiopthiae-encoded ORF sequences revealed 14 new antigens that may localize on the surface of E. rhusiopthiae. When these putative surface proteins were used to immunize mice, 9/11 antigens induced protective immunity. Thus, we have demonstrated that a combination of using the whole bacterial cells to purify antibodies and using the phage-displayed peptide libraries to determine the antigen specificities of the antibodies can lead to the discovery of novel bacterial surface antigens. This can be a general approach for identifying surface antigens for other bacterial species.

Environmental exposure is considered to be responsible for nontuberculous mycobacterial infections in humans. To facilitate the isolation of mycobacteria from soil, Middlebrook 7H10 agar was optimized as an enhanced selective medium by increasing the concentration of malachite green. A series of modified Middlebrook 7H10 agar media with malachite green concentrations ranging from 2.5 to 2500 mg/L was evaluated using 20 soil samples decontaminated with 3% sodium dodecyl sulfate plus 2% NaOH for 30 min. Among these modified Middlebrook 7H10 media, the medium with malachite green at a concentration of 250 mg/L, i.e., at the same concentration as in Löwenstein–Jensen medium, was the most effective in terms of the number of plates with mycobacterial growth. This medium was further evaluated with 116 soil samples. The results showed that 87.1% (101/116) of the samples produced mycobacterial growth, and 15 samples (12.9%) produced no mycobacterial growth. Of the plates inoculated with the soil samples, each in duplicate, 5.2% (12/232) showed late contamination. In total, 19 mycobacterial species were isolated, including seven (36.8%) rapidly growing mycobacteria and 12 (63.2%) slowly growing mycobacteria. Our results demonstrate that the modified Middlebrook 7H10 agar with 250 mg/L malachite green is useful for the primary isolation of nontuberculous mycobacteria from soil.Electronic supplementary materialThe online version of this article (doi:10.1186/s13568-017-0373-6) contains supplementary material, which is available to authorized users.