To identify a novel plasmid-mediated colistin resistance gene in Klebsiella pneumoniae isolated from chickens in China. WGS was used to identify a novel colistin resistance gene. The transferability of plasmids carrying mcr-7.1 was investigated by conjugation experiments. The expression of the mcr-7.1 gene was examined using an expression vector. A novel plasmid-mediated colistin resistance gene mcr-7.1, sharing 70% amino acid identity with the mcr-3 gene, was identified in three K. pneumoniae strains isolated from chickens in China. The mcr-7.1 gene was found in an IncI2-type plasmid (pSC20141012) that co-harboured the bla_CTX-M-55 gene in one isolate. pSC20141012 can be transferred from K. pneumoniae SC20141012 to Escherichia coli J53Az^r, exhibiting a ≥8-fold increase in colistin MIC compared with the recipient E. coli J53Az^r. We identified a novel plasmid-mediated colistin resistance gene named mcr-7.1 in K. pneumoniae in China. The prevalence of mcr-7.1 in various species of human and animal origin needs to be investigated immediately.

The mcr-1 gene was detected in 5.11% (58/1136) of Escherichia coli isolates of chicken origin from 13 provinces in China. A novel mcr-1 variant, named mcr-1.3 , encoding an Ile-to-Val functional variant of MCR-1 was identified in a sequence type 155 (ST155) strain. An mcr-1.3 -containing IncI2 plasmid, pHeN867 (60,757 bp), was identified. The transfer of pHeN867 led to a 32-fold increase in the MIC of colistin in the recipient, exhibiting an effect on colistin resistance that was similar to that of mcr-1 .

Background: Salmonella Enteritidis (S. Enteritidis) being one of the most prevalent foodborne pathogens worldwide poses a serious threat to public safety. Prevention of zoonotic infectious disease and controlling the risk of transmission of S. Enteriditidis critically requires the evolution of rapid and sensitive detection methods. The detection methods based on nucleic acid and conventional antibodies are fraught with limitations. Many of these limitations of the conventional antibodies can be circumvented using natural nanobodies which are endowed with characteristics, such as high affinity, thermal stability, easy production, especially higher diversity. This study aimed to select the special nanobodies against S. Enteriditidis for developing an improved nanobody-horseradish peroxidase-based sandwich ELISA to detect S. Enteritidis in the practical sample. The nanobody-horseradish peroxidase fusions can help in eliminating the use of secondary antibodies labeled with horseradish peroxidase, which can reduce the time of the experiment. Moreover, the novel sandwich ELISA developed in this study can be used to detect S. Enteriditidis specifically and rapidly with improved sensitivity. Results: This study screened four nanobodies from an immunized nanobody library, after four rounds of screening, using the phage display technology. Subsequently, the screened nanobodies were successfully expressed with the prokaryotic and eukaryotic expression systems, respectively. A sandwich ELISA employing the SE-Nb9 and horseradish peroxidase-Nb1 pair to capture and to detect S. Enteritidis, respectively, was developed and found to possess a detection limit of 5 × 10⁴ colony forming units (CFU)/mL. In the established immunoassay, the 8 h-enrichment enabled the detection of up to approximately 10 CFU/mL of S. Enteriditidis in milk samples. Furthermore, we investigated the colonization distribution of S. Enteriditidis in infected chicken using the established assay, showing that the S. Enteriditidis could subsist in almost all parts of the intestinal tract. These results were in agreement with the results obtained from the real-time PCR and plate culture. The liver was specifically identified to be colonized with quite a several S. Enteriditidis, indicating the risk of S. Enteriditidis infection outside of intestinal tract. Conclusions: This newly developed a sandwich ELISA that used the SE-Nb9 as capture antibody and horseradish peroxidase-Nb1 to detect S. Enteriditidis in the spike milk sample and to analyze the colonization distribution of S. Enteriditidis in the infected chicken. These results demonstrated that the developed assay is to be applicable for detecting S. Enteriditidis in the spiked milk in the rapid, specific, and sensitive way. Meanwhile, the developed assay can analyze the colonization distribution of S. Enteriditidis in the challenged chicken to indicate it as a promising tool for monitoring S. Enteriditidis in poultry products. Importantly, the SE-Nb1-vHRP as detection antibody can directly bind S. Enteritidis captured by SE-Nb9, reducing the use of commercial secondary antibodies and shortening the detection time. In short, the developed sandwich ELISA ushers great prospects for monitoring S. Enteritidis in food safety control and further commercial production. Graphic

Tibetan Chickens should have unique gastrointestinal microbiota because of their particular habitats. Thus, the aim of this study was to investigate the cecal microbiota of Tibetan Chickens from five typical high‐altitude regions of China. Lohmann egg‐laying hens (LMs) and Daheng broiler chickens (DHs) were chosen as controls. The cecal bacterial populations of Tibetan Chickens were surveyed by high‐throughput sequencing (HTS) of the bacterial 16S rRNA hypervariable region V3‐V4 (16S rRNAV3‐V4) combined with community‐fingerprinting analysis of the 16S rRNA gene based on polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE). The results revealed that the majority of cecal microbiota differed between the Tibetan Chicken and LM/DH. The microbial communities in the cecum were composed of 16 phyla, 28 classes, 36 orders, 57 families, 101 genera, and 189 species. Represented phyla were Bacteroidetes (>47%), Firmicutes (>18.8%), Spirochaetae (>0.3%), and Proteobacteria (>0.4%). Bacteroides and the RC9 gut group were the two most abundant genera. There were relatively more Christensenellaceae, Subdoligranulum, Spirochaeta, and Treponema in Tibetan Chickens, whereas there were more Phascolarctobacterium, Faecalibacterium, Megamonas, and Desulfovibrio in LMs and DHs. The cecal microbiota of Tibetan Chicken have slightly diverged due to exposure to different geographic environments. Differences in the intestinal bacterial communities of Tibetan Chicken and LM/DH were noted.

Sixteen different sequence types (STs) of Escherichia coli isolates from a commercial swine farm in China were confirmed to coharbor the carbapenem resistance gene bla _NDM-5 and the colistin resistance gene mcr-1 . Whole-genome sequencing revealed that bla _NDM-5 and mcr-1 were located on a 46-kb IncX3 plasmid and a 32-kb IncX4 plasmid, respectively. The two plasmids can transfer together with a low fitness cost, which might explain the presence of various STs of E. coli coharboring bla _NDM-5 and mcr-1 .