The emergence of extended-spectrum β-lactamase-producing Salmonella enterica serovar Enteritidis (ESBL-SE) in humans and foods has gained global attention. In particular, CTX-M-type ESBL-SE are increasingly being detected from various sample types. The aim of this study was to comprehensively analyze the epidemiology and characteristics of bla_CTX-M-55-carrying ESBL-SE isolates of clinical origin in Shanghai, China. A total of 292 S. Enteritidis isolates were recovered from the feces and blood of outpatients and inpatients between 2006 and 2014. Overall, there was a high frequency of cefotaxime-resistant isolates (97.3%), which was significantly higher (p < 0.01) than that of isolates resistant to the other tested antibiotics. All S. Enteritidis isolates exhibited resistance to ≥1 antibiotic, and 98.0% were multidrug resistant. A total of 233 isolates were identified as ESBL-SE, 166 of which were CTX-M type. Six subtypes of CTX-M-encoding genes were detected, among which bla_CTX-M-55 (91.6%, 152/166) was the most prevalent genotype. There was high genetic similarity among bla_CTX-M-55-positive ESBL-SE. The bla_CTX-M-55 gene in the ESBL-SE donor strains could be easily transferred into Enterobacteriaceae recipient strains. This study highlights that CTX-M-55 should be considered an important surveillance target in Shanghai, China. Cephalosporins, especially cefotaxime, must be used with caution in empirical treatment for Salmonella infections.

The aim of this study was to develop homogeneous and stable plasmid DNA reference materials for detecting the mechanisms of resistance to quinolones and fluoroquinolones in foodborne pathogens. The DNA fragments of 11 target genes associated with quinolone and fluoroquinolone resistance were artificially synthesized, inserted into plasmid vectors, and transferred into recipient cells. PCR and sequencing of DNA were performed to assess the genetic stability of the target DNA in recombinant Escherichia coli DH5α cells during subculturing for 15 generations. The limit of detection (LOD) of the target DNA was determined using PCR and real-time qualitative PCR (qPCR). The homogeneity and storage stability of plasmid DNA reference materials were evaluated in terms of plasmid DNA quantity, PCR-measured gene expression, and qPCR threshold cycle. All 11 target DNAs were successfully synthesized and inserted into vectors to obtain recombinant plasmids. No nucleotide mutations were identified in the target DNA being stably inherited and detectable in the corresponding plasmids during subculturing of recombinant strains. When the target DNA was assessed using PCR and qPCR, the LOD was ≤1.77 × 10⁵ and 3.26 × 10⁴ copies/μL, respectively. Further, when the reference materials were stored at 37 °C for 13 days, 4 °C for 90 days, and −20 °C for 300 days, each target DNA was detectable by PCR, and no mutations were found. Although the threshold cycle values of qPCR varied with storage time, they were above the LOD, and no significant differences were found in the quantity of each plasmid DNA at different timepoints. Further, the homogeneity and stability of the materials were highly consistent with the requirements of standard reference materials. To summarize, considering that our plasmid DNA reference materials conformed to standard requirements, they can be used to detect the mechanisms of quinolone and fluoroquinolone resistance in foodborne pathogens.