Antimicrobial Resistance and Genomic Characteristics of Escherichia coli Strains Isolated from the Poultry Industry in Henan Province, China.

Escherichia coli (E. coli) is an important foodborne pathogen and a biomarker for monitoring antimicrobial resistance. Investigating the prevalence of E. coli in the poultry industry holds great importance, particularly in Henan province, a major poultry-producing region in China. Here, we investigated the antimicrobial resistance (AMR) phenotypes of E. coli strains obtained from the poultry industry in Henan, China. A total of 344 E. coli strains were isolated from 638 samples collected from seven farms, three slaughterhouses, and ten terminal markets. Approximately 96.4%, 81.7%, and 52.5% of the isolates from the farms, slaughterhouses, and terminal markets exhibited multidrug resistance. Whole-genome sequencing was performed on 169 strains to reveal their genomic characteristics. The sequence type (ST) analysis revealed that ST10 and ST156 were the most frequent types within the poultry supply chain, whereas ST10 and ST162 were commonly found across the farms, slaughterhouses, and terminal markets. Fourteen ST10 E. coli strains belonged to phylogenetic group A, while fifteen ST165 and six ST162 E. coli strains belonged to phylogenetic group B1. In addition, several antimicrobial resistance genes and virulence factor genes were identified. The blaNDM-5 gene mediated carbapenem resistance in two E. coli strains, while mcr-1-mediated colistin resistance was detected in nine E. coli strains. Phylogenetic group A exhibited fewer virulence genes compared to other groups of E. coli. Plasmid replicons, such as IncFIB (AP001918), IncX1, IncFIC (FII), and IncFII (pHN7A8), were frequently observed. These findings provide valuable insights into the current AMR profiles of E. coli strains isolated from the poultry industry in Central China and highlight the need to implement good manufacturing practices and reduce antibiotic usage to mitigate potential risks associated with E. coli.

Antimicrobial resistance and population genomics of multidrug-resistant Escherichia coli in pig farms in mainland China.

The expanding use of antimicrobials in livestock is an important contributor to the worldwide rapid increase in antimicrobial resistance (AMR). However, large-scale studies on AMR in livestock remain scarce. Here, we report findings from surveillance of E. coli AMR in pig farms in China in 2018-2019. We isolated E. coli in 1,871 samples from pigs and their breeding environments, and found AMR in E. coli in all provinces in mainland China. We detected multidrug-resistance in 91% isolates and found resistance to last-resort drugs including colistin, carbapenems and tigecycline. We also identified a heterogeneous group of O-serogroups and sequence types among the multidrug-resistant isolates. These isolates harbored multiple resistance genes, virulence factor-encoding genes, and putative plasmids. Our data will help to understand the current AMR profiles of pigs and provide a reference for AMR control policy formulation for livestock in China.

Prevalence and Molecular Characterization of Antimicrobial-Resistant Escherichia coli in Pig Farms, Slaughterhouses, and Terminal Markets in Henan Province of China.

Escherichia coli is an important foodborne pathogen and also plays key roles in dissemination of antimicrobial resistance genes (ARGs). However, current data on the prevalence of antimicrobial-resistant E. coli at different nodes of the pork supplying chain are still limited. Herein, we investigated drug-resistant phenotypes and molecular characteristics of E. coli strains isolated from different pig farms, slaughterhouses, and terminal markets in the Henan Province of China. A total of 191 (70.74%), 140 (35.09%), and 77 (30.20%) E. coli strains were isolated from 270, 399, and 255 samples collected from pig farms, slaughterhouses, and retailing markets, respectively. Antimicrobial susceptibility testing revealed that these 408 strains showed severe antimicrobial resistance profiles. Approximately 93.19% (178/191), 66.43% (93/140), and 67.53% (52/77) of the isolates from farms, slaughterhouses, and terminal markets were resistant to three of the nine antibiotic classes tested, respectively. Multilocus sequence typing showed that sequence types (STs) 10 and ST101 were commonly identified among the isolates from farms, slaughterhouses, and terminal markets. Isolates belonging to these two STs carried multiple ARGs, conferring resistance to the antibiotics tested. Two important ARGs with great public health concerns (mcr-1 and blaNDM-1) were found from these two STs. Isolates belonging to these two STs also carried several virulence factor-encoding genes, including astA, tsh, and traT, which might contribute to the pathogenesis of these isolates. The wide prevalence and distribution of these two STs in different nodes of pork supplying chain might represent a big public health threat and should receive more attention.

Characteristics of a Colistin-Resistant Escherichia coli ST695 Harboring the Chromosomally-Encoded mcr-1 Gene.

Enterobacteriaceae having chromosomally-encoded mcr-1 is rarely reported. In this study, we recovered a chromosomal mcr-1 carrying Escherichia coli, designated HeN100, from the feces of a diarrheal pig in China. Antimicrobial susceptibility testing showed that HeN100 was resistant to three aminoglycosides, twelve ß-lactams including three carbapenems, one phenicol, two tetracyclines, two fluoroquinolones, nitrofurantoin, and colistin tested. Oxford Nanopore MinION sequencing revealed that the complete genomes of the multidrug resistant (MDR) HeN100 consisted of a single circular chromosome and five circular plasmids. Bioinformatical analysis determined HeN100 as ST695 and it contained many acquired resistance genes responsible for its MDR phenotypes, including colistin resistance mcr-1 and the carbapenem resistance blaNDM-1, and most of these genes were located on plasmids. However, the mcr-1 was found on the chromosome, and it was located between an IS30-like element ISApl1 and a PAP2-like encoding gene. These three genes consisted of an "ISApl1-mcr-1-orf" segment and inserted in high AT-rich regions. Finally, we found the blaNDM-1 was carried on an IncFII type conjugative plasmid. The conjugation frequency of this plasmid was 7.61 ± 2.11 × 10-5 per recipient, and its conjugation conferred resistance to carbapenems and other ß-lactams, as well as aminoglycosides. The spread of this mcr-1/blaNDM-1-carrying E. coli ST695 represents a great concern of public health.

[Establishment and identification of human cervical cancer cell line C33A stably expressing human papillomavirus type 58 E6E7 fusion gene].

The study was performed to construct a human cervical cancer cell line C33A which can stably express HPV58E6E7 fusion gene. Firstly, C33A cells were transfected with the recombinant lentivirus LV-HPV58E6E7 which contained HPV58E6E7 fusion gene, and the stably transfected cells (LV-HPV58E6E7/C33A) were screened out by flow cytometry. MTT was used to observe the growth of LV-HPV58E6E7/C33A cells and flow cytometry was carried out to detect the cell cycle. LV-HPV58E6E7/C33A cells were inoculated into the left armpits of nude mice. Then, the transcription and expression of HPV58E6E7 fusion gene was detected by qRT-PCR and Western blot, respectively. The results showed that HPV58E6E7 fusion gene can promote the proliferation of C33A cells. HPV58E6E7 fusion gene can be stably transcripted and expressed in vaccinated nude mice. The conclusion indicated that we successfully established a cervical cancer cell line LV-HPV58E6E7/C33A which can stably express HPV58E6E7 fusion gene. This cell line will provide an antigen cell line for the immune effect detection of HPV58 therapeutic vaccine.