The aim of this study was to investigate the intergeneric transfer of vancomycin resistance gene vanA between probiotic enterococci in the fermentation progress of soybean meal and in the digestive tract of growing pigs. One vanA genotype vancomycin resistant E. faecium strain, Efm4, and one chloramphenicol-resistant E. faecalis strain, Efs2, were isolated from twenty-nine probiotic basis feed material / additive samples. For in vitro conjugation, Efm4 and Efs2 were used as starter to ferment soybean meal. For in vivo conjugation, thirty growing pigs were randomly assigned to five groups (n = 6), treated with a basic diet, or supplemented with 10% fermented soybean meal, 1% Efm4, 5% Efs2 or a combination of 1% Efm4 + 5% Efs2 for 7 d, respectively. Fecal samples of pigs in each group were collected daily for the isolation and dynamic analysis of Efm4, Efs2 and transconjugants. The sequence types (STs) of Efm4, Efs2 and transconjugants were analyzed by multilocus sequence typing (MLST). The vanA harboring plasmid in Efm4 and transconjugants was analyzed by S1-pulsed field gel electrophoresis (PFGE) and further verified by multiple alignments.The results showed that, in FSBM, transconjugants were detected 1 h after the fermentation, with a conjugation frequency of ~ 10- 3 transconjugants / recipient. Transconjugants proliferated with Efm4 and Efs2 in the first 8 h and maintained steadily for 10 d till the end of the experiment. Additionally, in vivo experiment showed that transcojugants were recovered in one of six pigs in both FSBM and Efm4 + Efs2 groups, with conjugation frequency of ~ 10- 5 and ~ 10- 4, respectively. MLST revealed the ST of Efm4, Efs2 and transconjugants was ST1014, ST69 and ST69, respectively. S1-PFGE confirmed the existence of the vanA-harboring, 142,988-bp plasmid, which was also a multi-drug resistant plasmid containing Tn1546-like transposon.The findings revealed the potential safety hazard existing in the commercial probiotic enterococci in China, because the horizontal transfer from farm to fork could potentially pose a safety risk to the public.
The emergence of the mobile tigecycline-resistance gene, tet(X4), poses a significant threat to public health. To investigate the prevalence and genetic characteristics of the tet(X4)-positive Escherichia coli in humans, 1101 human stool samples were collected from a tertiary class-A hospital in Beijing, China, in 2019. Eight E. coli isolates that were positive for tet(X4) were identified from clinical departments of oncology (n = 3), hepatology (n = 2), nephrology (n = 1), urology (n = 1), and general surgery (n = 1). They exhibited resistance to multiple antibiotics, including tigecycline, but remained susceptible to meropenem and polymyxin B. A phylogenetic analysis revealed that the clonal spread of four tet(X4)-positive E. coli from different periods of time or departments existed in this hospital, and three isolates were phylogenetically close to the tet(X4)-positive E. coli from animals and the environment. All tet(X4)-positive E. coli isolates contained the IncX1-plasmid replicon. Three isolates successfully transferred their tigecycline resistance to the recipient strain, C600, demonstrating that the plasmid-mediated horizontal gene transfer constitutes another critical mechanism for transmitting tet(X4). Notably, all tet(X4)-bearing plasmids identified in this study had a high similarity to several plasmids recovered from animal-derived strains. Our findings revealed the importance of both the clonal spread and horizontal gene transfer in the spread of tet(X4) within human clinics and between different sources.
Abstract Clostridium perfringens is often associated with foodborne diseases, posing significant public health risks. However, genomic investigation of C. perfringens isolates from the human population has been lacking. This study aims to fill this knowledge gap by examining the genomic characteristics of C. perfringens isolates from 699 individuals at a provincial hospital in China. We further conducted evolutionary and pan‐genomic analyses, incorporating isolates from humans and animals worldwide. The results reveal potential regional and transregional transmission of C. perfringens among individuals, along with the common transfer of small gene clusters during this process. Notably, the food poisoning‐associated toxin gene cpe was identified in a fusion plasmid for the first time in an isolate, indicating fusion of pCP13‐like and pCW3‐like plasmids and the potential for transfer of cpe across genetic backgrounds. Moreover, we observed that the genomic characteristics of C. perfringens correlate with host species, with specific toxin genes, such as pfoA and colA , potentially influencing host selectivity. Through this comprehensive genomic analysis, we provide novel insights into the fusion of pCW3‐like and pCP13‐like plasmids, the genetic location of cpe , the transmission dynamics of C. perfringens strains, and the relationship between toxin genes and host relevance. These findings expand our understanding of C. perfringens and its implications for public health.
Abstract Objectives The wide spread of tet(X4) gene orthologues in the environment, food, poultry and humans is causing serious tigecycline resistance. Consequently, developing a fast and universal method to detect tigecycline resistance has become increasingly important. Methods During 2019–2022, 116 Escherichia coli isolates were obtained from nine provinces in China. All isolates were tested for their susceptibility to antimicrobial agents by the microdilution broth method and for the tet(X4) gene by PCR. Ten tet(X4)-positive E. coli isolates were used to confirm certain conditions, including the optimal incubation time, the optimal concentration of tigecycline, and the cut-off of the relative growth (RG) value. Results The optimal time and concentration of tigecycline for separation of susceptible and resistant isolates was 2 h and 4 mg/L, and the RG cut-off value was 0.4. We validated whether the experiment was feasible using 116 isolates of E. coli. The method yielded a susceptibility of 94.9% (95% CI: 81.4%–99.1%) and a specificity of 96.1% (95% CI: 88.3%–99.0%). Conclusions This research has shown that this optical antimicrobial susceptibility testing method can rapidly differentiate between susceptible and resistant phenotypes in isolates of E. coli. In the same range as the current gold-standard methods, the clinical turnaround time is reduced from 48 h to 2.5 h. The above results suggest that the method has splendid specificity and operationality.
It is crucial to discover novel antimicrobial drugs to combat resistance. This study investigated the antibacterial properties of halicin (SU3327), an AI-identified anti-diabetic drug, against 13 kinds of common clinical pathogens of animal origin, including multidrug-resistant strains. Employing minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assessments, halicin demonstrated a broad-spectrum antibacterial effect. Time-killing assays revealed its concentration-dependent bactericidal activity against Escherichia coli ATCC 25922 (E. coli ATCC 25922), Staphylococcus aureus ATCC 29213 (S. aureus ATCC 29213), and Actinobacillus pleuropneumoniae S6 (APP S6) after 4 h of treatment at concentrations above the MIC. Halicin exhibited longer post-antibiotic effects (PAEs) and sub-MIC effects (PA-SMEs) for E. coli 25922, S. aureus 29213, and APP S6 compared to ceftiofur and ciprofloxacin, the commonly used veterinary antimicrobial agents, indicating sustained antibacterial action. Additionally, the results of consecutive passaging experiments over 40 d at sub-inhibitory concentrations showed that bacteria exhibited difficulty in developing resistance to halicin. Toxicology studies confirmed that halicin exhibited low acute toxicity, being non-mutagenic, non-reproductive-toxic, and non-genotoxic. Blood biochemical results suggested that halicin has no significant impact on hematological parameters, liver function, and kidney function. Furthermore, halicin effectively treated respiratory A. pleuropneumoniae infections in murine models. These results underscore the potential of halicin as a new antibacterial agent with applications against clinically relevant pathogens in veterinary medicine.
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