Salmonella can cause serious foodborne diseases. We have developed a lateral flow immunoassay combined with recombinase polymerase amplification (LFD-RPA) for detection of Salmonella in food. The conserved fragment (fimY) was selected as the target gene. Under an optimal condition (37 °C, 10 min), the sensitivity was 12 colony-forming units (CFU)/mL in a pure culture. Testing with 16 non-Salmonella strains as controls revealed that LFD-RPA was specific to the fimY gene of Salmonella. The established assay could detect Salmonella at concentrations as low as 1.29 × 10² CFU/mL in artificially contaminated samples. This detection was at a slightly higher level than that for a pure bacterial culture. Combined with the test strip reader, the LFD-RPA is a feasible method for quantitative detection of Salmonella based on the test line intensity, which was the ratio for the test line and control line of the reflected light. The method could be a potential point-of-care test in limited resource areas and provides a new approach and technical support for the diagnosis of food safety.

Vibrio parahaemolyticus is the leading cause of seafood-borne bacterial poisoning in China and is a threat to human health worldwide. The aim of this study was to assess the antibiotic resistance profiles and distribution of heavy metal resistance of V. parahaemolyticus isolates from Penaeus vannamei from freshwater farms, seawater farms, and their corresponding markets in Zhejiang, China and to assess the relationship between multidrug resistance (MDR) and multi-heavy metal resistance (MHMR). Of the 360 P. vannamei samples that we tested, 90 (25.00%) were V. parahaemolyticus positive, but the occurrence of pathogenic isolates carrying the toxin genes tdh (4.44%) and trh (3.33%) was low. None of the tested isolates harbored both the tdh and trh genes. However, antibiotic resistance profiles varied among different sampling locations, levels of resistance to the antibiotics ampicillin (76.67%) and streptomycin (74.44%) were high overall, and MDR isolates were common (40.00% of all isolates). Heavy metal resistance patterns were similar among the different sampling locations. Overall, the majority of V. parahaemolyticus isolates displayed tolerance to Cd²⁺ (60.00%), and fewer were resistant to Cu²⁺ (40.00%), Zn²⁺ (38.89%), Ni²⁺ (24.44%), Cr³⁺ (14.44%), and Co²⁺ (8.89%). In addition, 34.44% (31/90) of isolates tested in this study were found to be MHMR. Using Pearson’s correlation analysis, MDR and MHMR were found to be positively correlated (P = 0.004; R = 0.759). The 18 V. parahaemolyticus isolates that were both MDR and MHMR represented 18 sequence types, of which 12 were novel to the PubMLST database, and displayed a high level of genetic diversity, suggesting that dissemination may be affected by mobile genetic elements via horizontal gene transfer. However, a low percentage of class 1 integrons without gene cassettes and no class 2 or 3 integrons were detected in the 18 MDR and MHMR isolates or in the 90 V. parahaemolyticus isolates overall. Thus, we suggest that future research focus on elucidating the mechanisms that lead to a high prevalence of resistance determinants in V. parahaemolyticus. The results of this study provide data that will support aquatic animal health management and food safety risk assessments in the aquaculture industry.

The worrying emergence of multiple resistance genes to last-resort antibiotics in food animals and human populations throughout the food chain and relevant environments has been increasingly reported worldwide. Enterobacteriaceae pathogens are considered the most common reservoirs of such antibiotic resistance genes (ARGs). Thus, a rapid, efficient and accurate detection method to simultaneously screen and monitor such ARGs in Enterobacteriaceae pathogens has become an urgent need. Our study developed a recombinase polymerase amplification (RPA) assay combined with a lateral flow dipstick (LFD) for simultaneously detecting predominant resistance genes to last-resort antibiotics of Enterobacteriaceae pathogens, including mcr-1, bla_NDM-1 and tet(X4). It is allowed to complete the entire process, including crude DNA extraction, amplification as well as reading, within 40 min at 37°C, and the detection limit is 10¹ copies/μl for mcr-1, bla_NDM-1 and tet(X4). Sensitivity analysis showed obvious association of color signals with the template concentrations of mcr-1, bla_NDM-1 and tet(X4) genes in Enterobacteriaceae pathogens using a test strip reader (R² = 0.9881, R² = 0.9745, and R² = 0.9807, respectively), allowing for quantitative detection using multiplex RPA-LFD assays. Therefore, the RPA-LFD assay can suitably help to detect multiple resistance genes to last-resort antibiotics in foodborne pathogens and has potential applications in the field.

Class I integrons have garnered significant attention due to pivotal roles in the dissemination of antimicrobial resistance genes (ARGs), which impose risks to public health and food safety. Here, the prevalence and characteristics of class I integrons in Escherichia coli isolates derived from food-producing animals and human patients were assessed. Of 721 E. coli isolates collected from human patients (113), pigs (298), and poultry (310), 93 (12.90%) carried the class I integrase gene (intI1). Multilocus sequence typing identified 39 sequence types from 93 intI1-postive isolates, including three novel types. Sequence analysis revealed that 59 classical class I integrons encompassed six distinct gene cassettes arrangements [dfrA17-aadA5, dfrA12-aadA2, dfrA1-aadA1, dfrA7, aac(6’)-Ib, and aadA1-aac(3)-VIa]. Six insertion sequences (IS1, IS6, IS21, IS91, IS110, and IS256) and one transposon (Tn3) were harbored in proximity to the integrons. A comparison with sequences retrieved from the National Center for Biotechnology Information database demonstrated that E. coli isolates with integron sequences were detected in various food-producing animals and human hosts in environmental niches across Asia, Europe, and North America. These findings indicate the potential risk of ARG transmission between food-producing animals and humans by bacteria populations and provide useful baseline data for monitoring of ARGs.