Virulence factors and molecular characteristics of Shigella flexneri isolated from calves with diarrhea.

BACKGROUND: The natural hosts of Shigella are typically humans and other primates, but it has been shown that the host range of Shigella has expanded to many animals. Although Shigella is becoming a major threat to animals, there is limited information on the genetic background of local strains. The purpose of this study was to assess the presence of virulence factors and the molecular characteristics of S. flexneri isolated from calves with diarrhea. RESULTS: Fifty-four S. flexneri isolates from Gansun, Shanxi, Qinghai, Xinjiang and Tibet obtained during 2014 to 2016 possessed four typical biochemical characteristics of Shigella. The prevalences of ipaH, virA, ipaBCD, ial, sen, set1A, set1B and stx were 100 %, 100 %, 77.78 %, 79.63 %, 48.15 %, 48.15 and 0 %, respectively. Multilocus variable number tandem repeat analysis (MLVA) based on 8 variable number of tandem repeat (VNTR) loci discriminated the isolates into 39 different MLVA types (MTs), pulsed field gel electrophoresis (PFGE) based on NotI digestion divided the 54 isolates into 31 PFGE types (PTs), and multilocus sequence typing (MLST) based on 15 housekeeping genes differentiated the isolates into 7 MLST sequence types (STs). CONCLUSIONS: The findings from this study enrich our knowledge of the molecular characteristics of S. flexneri collected from calves with diarrhea, which will be important for addressing clinical and epidemiological issues regarding shigellosis.

Epidemic and molecular characterization of fluoroquinolone-resistant Shigella dysenteriae 1 isolates from calves with diarrhea.

BACKGROUND: The widespread distribution of antimicrobial-resistant Shigella has become a recurrent challenge in many parts of the developing world. Previous studies indicate that the host of Shigella has expanded from humans to animals. This study aimed to investigate the prevalence of fluoroquinolone resistance and associated molecular characterization of S. dysenteriae 1 isolated from calves. RESULTS: All 38 unduplicated S. dysenteriae 1 isolates were collected from calves in Gansu Province from October 2014 to December 2016. According to MLST and PFGE analysis, these isolates were separated into 4 and 28 genotypes, respectively. The most common STs identified were ST228 (34.21%, 13/38) and ST229 (39.47%, 15/38), which were first found in the present study. All isolates harbored virulence genes, and the incidence of the seven virulence genes were ipaH (100%), ipaBCD (92.11%), stx (73.68%), ial (57.89%), sen (28.95%), set1A and set1B (0%). According to the results of antimicrobial susceptibilities, 76.32% (29/38) were resistant to fluoroquinolone and showed multidrug resistance. In a study on the polymorphism of quinolone resistance-determining region (QRDR) of gyrA/B and parC/E genes, we identified two mutations in gyrA (Ser83 → Leu and Asp87 → Asn) and parC (Ser80 → Ile and Ser83 → Leu), respectively. Among them, 55.17% (16/29) of resistant strains had the gyrA point mutations (Ser83 → Leu) and parC point mutation (Ser83 → Leu). Moreover, 41.38% (12/29) of isolates had all five point mutations of gyrA and parC. In addition, the prevalence of the plasmid-mediated quinolone resistance (PMQR) determinant genes was also investigated. All 29 fluoroquinolone-resistant isolates were positive for the aac (6')-Ib-cr gene but negative for qepA, except for SD001. In addition, only 6 (20.69%, 6/29) isolates harbored the qnr gene, including two with qnrB (6.90%, 2/29) and four with qnrS (13.79%, 4/29). CONCLUSION: Given the increased common emergence of multidrug resistant isolates, uninterrupted surveillance will be necessary to understand the actual epidemic burden and control this infection.

Exploring the Prevalence and Distribution Patterns of Antibiotic Resistance Genes in Bovine Gut Microbiota Using a Metagenomic Approach.

Antibiotic resistance genes (ARGs) have become recognized contaminants and pose a high public health risk. The animal gut microbiota is a reservoir of ARGs, but the knowledge of the origin and dissemination of ARGs remains unclear. In this study, we provide a comprehensive profile of ARGs and mobile genetic elements in the gut microbiota from 30 bovines to study the impact of modern antibiotics on resistance. A total of 42 ARG types were detected by annotating the metagenomic sequencing data from Comprehensive Antibiotic Resistance Database (CARD). We found that the diversity and abundance of ARGs in individual yaks were significantly lower than those in dairy and beef cattle (p < 0.0001). The results of heat map and single nucleotide polymorphism clustering suggest that ARGs from dairy and beef cattle are more similar, whereas those from yaks cluster separately. The long-term use of antibiotics may contribute to this difference, suggesting that antibiotic consumption is the main cause of ARG prevalence. Furthermore, abundant insertions were also found in this study, signifying a strong potential for horizontal transfer of ARGs among microbes, especially pathogens.

Epidemic characterization and molecular genotyping of Shigella flexneri isolated from calves with diarrhea in Northwest China.

BACKGROUND: The widespread presence of antibiotics resistance genes in pathogenic bacteria can cause enormous problems. Food animals are one of the main reservoirs of intestinal pathogens that pose a potential risk to human. Analyzing the epidemiological characteristics and resistance patterns of Shigella flexneri in calves is necessary for animal and human health. METHODS AND RESULTS: A total of 54 Shigella flexneri isolates, including six serotypes (1a, 2a, 2b, 4a, 6 and Xv), were collected from 837 fecal samples obtained from 2014 to 2016. We performed pulsed-field gel electrophoresis (PFGE) and applied the restriction enzyme NotI to analyze the genetic relatedness among the 54 isolates and to categorize them into 31 reproducible and unique PFGE patterns. According to the results of antimicrobial susceptibility tests, all 26 Shigella flexneri 2a serotypes were resistant to cephalosporin and/or fluoroquinolones. The genes blaTEM-1 , blaOXA-1 , and blaCTX-M-14 were detected in 19 cephalosporin-resistant S. flexneri 2a isolates. Among 14 fluoroquinolone-resistant isolates, the aac(6')-Ib-cr gene was largely present in each strain, followed by qnrS (5). Only one ciprofloxacin-resistant isolate harbored the qepA gene. Sequencing the quinolone resistance determining regions (QRDRs) of the fluoroquinolone-resistant isolates revealed two point mutations in gyrA (S83 L, D87N/Y) and a single point mutation in parC (S80I). Interestingly, two gyrA (D87N/Y) strains were resistant to ciprofloxacin. CONCLUSIONS: The current study enhances our knowledge of Shigella in cattle, although continual surveillance is necessary for the control of shigellosis. The high level of cephalosporin and/or fluoroquinolone resistance in Shigella warns us of a potential risk to human and animal health.