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. KEYWORDS E. coli, colistin resistance, mcr-1.3, plasmid P olymyxins (polymyxin B and colistin) are a last-resort treatment for infections caused by multidrug-resistant (MDR) Gram-negative bacteria (1). In veterinary use, colistin is administered with food in pig and poultry farming to prevent infections caused by pathogens (2). The mcr-1 gene, which confers plasmid-mediated colistin resistance to Enterobacteriaceae, was identified in an IncI2 plasmid from Escherichia coli and Klebsiella pneumoniae in China in 2016 (3). The mcr-1 gene found in E. coli (4), K. pneumoniae (5), and Salmonella spp. (6) has been proven to disseminate ubiquitously. The transmission of mcr-1-mediated colistin resistance between animals and human has been a threat to human health. It has also been demonstrated that the mcr-1 gene can coexist with bla CTX-M (5), bla NDM (7), and other resistance genes (4), which threatens a return of untreatable infections worldwide. Previous reports described the unique mcr-1 gene sequence compared with that of the originally published sequence (3), which indicates that mcr-1 is relatively conserved. Recently, a point mutation of A¡T at position 8 in mcr-1 was identified in K. pneumoniae (8). To investigate the epidemiology of mcr-1 and its variant, E. coli isolates collected from chickens nationwide in China were assessed.In total, 1,136 nonduplicate E. coli isolates were collected between 2010 and 2015 from sick chickens in 20 provinces and municipalities in China. All of these isolates were preliminarily screened on Mueller-Hinton agar medium with 2 g/ml colistin. Because the cooccurrence of mcr-1 with bla CTX-M may accelerate the transmission of resistance to colistin and cephalosporins, the mcr-1 (3) and bla CTX-M (9) genes were detected by PCR amplification of the isolates with resistance to colistin. The corresponding primers used to amplify the whole mcr-1 gene and parts of the ISApl1 element are listed in Table S1 in the supplemental material. For all of the positive PCR products of mcr-1, Sanger sequencing was performed (Tsingke Biological Technology, Chengdu, China) by using a DNA analyzer (Applied Biosystems, Life Technologies, Carlsbad, CA). We found a total of 58 (5.11%) mcr-1-positive isolates, including one isolate harboring the mcr-1 gene with mutations not found in the originally published gene sequence (3). MICs of colistin

Necrotic enteritis (NE) causes huge economic losses to the poultry industry. Probiotics are used as potential alternatives to antibiotics to prevent NE. It is known that Clostridium butyricum can act as a probiotic that can prevent infection. However, whether or not it exerts a beneficial effect on NE in chickens remains elusive. Therefore, we investigated the impact of C. butyricum on immune response and intestinal microbiota during the development of NE in chickens, including experimental stages with basal diets, high-fishmeal-supplementation diets, and Clostridium perfringens challenge. Chickens were divided into two groups from day 1 to day 20: one group had its diet supplemented with C. butyricum supplementation and one did not. At day 20, the chickens were divided into four groups: C. perfringens challenged and unchallenged chickens with and without C. butyricum supplementation. All groups were fed a basal diet for 13 days and thereafter a basal diet with 50% fishmeal from day 14 to 24. Chickens were infected with C. perfringens from day 21 to 23. At days 13, 20 and 24, samples were collected for analysis of the relative expression of immune response and intestinal mucosa barrier-related genes and intestinal microbes. The results show that C. butyricum can inhibit the increase in IL-17A gene expression and the reduction in Claudin-1 gene induced-expression caused by C. perfringens challenge. Moreover, C. butyricum was found to increase the expression of anti-inflammatory IL-10 in infected chickens. Although C. butyricum was found to have a significant beneficial effect on the structure of intestinal bacteria in the basal diet groups and decrease the abundance of C. perfringens in the gut, it did not significantly affect the occurrence of intestinal lesions and did not significantly correct the shift in gut bacterial composition post C. perfringens infection. In conclusion, although C. butyricum promotes the expression of anti-inflammatory and tight junction protein genes and inhibits pro-inflammatory genes in C. perfringens-challenged chickens, it is not adequate to improve the structure of intestinal microbiota in NE chickens. Therefore, more effective schemes of C. butyricum supplementation to prevent and treat NE in chickens need to be identified.

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.

Anthropogenic activities near urban rivers may have significantly increased the acquisition and dissemination of antibiotic resistance. In this study, we investigated the prevalence of colistin resistant strains in the Funan River in Chengdu, China. A total of 18 mcr-1-positive isolates (17 Escherichia coli and 1 Enterobacter cloacae) and 6 mcr-3-positive isolates (2 Aeromonas veronii and 4 Aeromonas hydrophila) were detected, while mcr-2, mcr-4 and mcr-5 genes were not detected in any isolates. To further explore the overall antibiotic resistance in the Funan River, water samples were assayed for the presence of 15 antibiotic resistance genes (ARGs) and class 1 integrons gene (intI1). Nine genes, sul1, sul2, intI1, aac(6′)-Ib-cr, blaCTX-M, tetM, ermB, qnrS, and aph(3′)-IIIa were found at high frequencies (70–100%) of the water samples. It is worth noting that mcr-1, blaKPC, blaNDM and vanA genes were also found in water samples, the genes that have been rarely reported in natural river systems. The absolute abundance of selected antibiotic resistance genes [sul1, aac(6′)-Ib-cr, ermB, blaCTX-M, mcr-1, and tetM] ranged from 0 to 6.0 (log10 GC/mL) in water samples, as determined by quantitative polymerase chain reaction (qPCR). The sul1, aac(6′)-Ib-cr, and ermB genes exhibited the highest absolute abundances, with 5.8, 5.8, and 6.0 log10 GC/mL, respectively. The absolute abundances of six antibiotic resistance genes were highest near a residential sewage outlet. The findings indicated that the discharge of resident sewage might contribute to the dissemination of antibiotic resistant genes in this urban river. The observed high levels of these genes reflect the serious degree of antibiotic resistant pollution in the Funan River, which might present a threat to public health.

No abstract