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Salmonella enterica infections continue to be a significant burden on public health worldwide. The ability of S. enterica to produce hydrogen sulfide (H2S) is an important phenotypic characteristic used to screen and identify Salmonella with selective medium; however, H2S-negative Salmonella have recently emerged. In this study, the H2S phenotype of Salmonella isolates was confirmed, and the selected isolates were subjected to antimicrobial susceptibility testing and molecular identification by multilocus sequence typing, pulsed-field gel electrophoresis, and clustered regularly interspaced short palindromic repeat (CRISPR) analysis. The phs genetic operon was also analyzed. A total of 160 S. enterica serovar Aberdeen isolates were detected between 2005 and 2013 in China. Of them, seven non-H2S-producing isolates were detected. Notably, four samples yielded four pairs of isolates with different H2S phenotypes, simultaneously. The data demonstrated that H2S-negative isolates were genetically closely related to H2S-positive isolates. Three new spacers (Abe1, Abe2, and Abe3) were identified in CRISPR locus 1 in four pairs of isolates with different H2S phenotypes from the same samples. Sequence analysis revealed a new nonsense mutation at position 208 in the phsA gene of all non-H2S-producing isolates. Additionally, we describe a new screening procedure to avoid H2S-negative Salmonella, which would normally be overlooked during laboratory and hospital screening. The prevalence of this pathogen may be underestimated; therefore, it is important to focus on improving surveillance of this organism to control its spread., Competing Interests: The authors have declared that no competing interests exist.

A clustered regularly interspaced short palindromic repeat (CRISPR) typing method has recently been developed and used for typing and subtyping of Salmonella spp., but it is complicated and labor intensive because it has to analyze all spacers in two CRISPR loci. Here, we developed a more convenient and efficient method, namely, CRISPR locus spacer pair typing (CLSPT), which only needs to analyze the two newly incorporated spacers adjoining the leader array in the two CRISPR loci. We analyzed a CRISPR array of 82 strains belonging to 21 Salmonella serovars isolated from humans in different areas of China by using this new method. We also retrieved the newly incorporated spacers in each CRISPR locus of 537 Salmonella isolates which have definite serotypes in the Pasteur Institute's CRISPR Database to evaluate this method. Our findings showed that this new CLSPT method presents a high level of consistency (kappa = 0.9872, Matthew's correlation coefficient = 0.9712) with the results of traditional serotyping, and thus, it can also be used to predict serotypes of Salmonella spp. Moreover, this new method has a considerable discriminatory power (discriminatory index [DI] = 0.8145), comparable to those of multilocus sequence typing (DI = 0.8088) and conventional CRISPR typing (DI = 0.8684). Because CLSPT only costs about $5 to $10 per isolate, it is a much cheaper and more attractive method for subtyping of Salmonella isolates. In conclusion, this new method will provide considerable advantages over other molecular subtyping methods, and it may become a valuable epidemiologic tool for the surveillance of Salmonella infections., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Salmonella enterica serovar Senftenberg is a common nontyphoidal Salmonella serotype which causes human Salmonella infections worldwide. In this study, 182 S. Senftenberg isolates, including 17 atypical non-hydrogen sulfide (H2S)-producing isolates, were detected in China from 2005 to 2011. The microbiological and genetic characteristics of the non-H2S-producing and selected H2S-producing isolates were determined by using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and clustered regularly interspaced short palindromic repeat (CRISPR) analysis. The phs operons were amplified and sequenced. The 17 non-H2S-producing and 36 H2S-producing isolates belonged to 7 sequence types (STs), including 3 new STs, ST1751, ST1757, and ST1758. Fourteen of the 17 non-H2S-producing isolates belonged to ST1751 and had very similar PFGE patterns. All 17 non-H2S-producing isolates had a nonsense mutation at position 1621 of phsA. H2S-producing and non-H2S-producing S. Senftenberg isolates were isolated from the same stool sample from three patients; isolates from the same patients displayed the same antimicrobial susceptibility, ST, and PFGE pattern but could be discriminated based on CRISPR spacers. Non-H2S-producing S. Senftenberg isolates belonging to ST1751 have been prevalent in Shanghai, China. It is possible that these emerging organisms will disseminate further, because they are difficult to detect. Thus, we should strengthen the surveillance for the spread of this atypical S. Senftenberg variant., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

With the rapid emergence of plasmid-mediated colistin resistance gene mcr-1 , the increased resistance of Salmonella has attracted extensive attention. This study reports on 11 multidrug-resistant Salmonella enterica serovar Typhimurium strains harboring mcr-1 in China. They all presented resistance to colistin, and additionally, one that was isolated from a child's stool sample was also resistant to ceftriaxone and azithromycin. We screened 1454 strains of Salmonella for mcr-1 gene through PCR, and these strains are all preserved in our laboratory. Antimicrobial sensitivity analysis was carried out for the screened mcr-1 positive strains. Genetic polymorphism analysis of S. Typhimurium was performed by using the Pulsed-Field Gel Electrophoresis (PFGE). The plasmids harboring mcr-1 were identified by S1-PFGE and southern blotting. Plasmid conjugation assays were used to analyze the transferability of colistin resistance. The plasmids harboring mcr-1 were characterized by sequencing and bioinformatic analysis. Eleven S. Typhimurium strains harboring mcr-1 with colistin resistance (MICs 4μg/ml) were detected, which were isolated from children and pig offal in China. All of them were multidrug-resistant strains. PFGE results revealed that the strains isolated from different samples or locations have identical genotypes. S1-PFGE and southern blotting experiments showed that three plasmids of different sizes (33, 60, and 250 kb) all carried the mcr-1 gene. The plasmid conjugation assays revealed that Salmonella acquired mcr-1 harboring plasmids by horizontal transfer. Sequencing and plasmid type analysis revealed that these plasmids were types IncX4, IncI2, and IncHI2. Among them, IncX4 and IncI2 plasmids had extremely similar backbones and contained one resistant gene mcr-1. IncHI2 plasmid contained multiple resistant genes including bla CTX–M, oqxB , sul , aph , aadA , and bla TEM. We identified 11 mcr-1 harboring S. Typhimurium strains in China and described their characteristics. Our findings indicate that the mcr-1 gene can effectively spread among intestinal bacteria by horizontal transfer of three types of plasmids. Moreover, the IncHI2 plasmid can also mediate the transfer of other drug resistance genes. These results reveal that constant surveillance of mcr-1 harboring S Typhimurium is imperative to prevent the spread of colistin resistance. [ABSTRACT FROM AUTHOR]

Salmonella contamination of eggs and egg shells has been identified as a public health problem worldwide. Here, we reported an outbreak of severe gastrointestinal symptoms caused by Salmonella enterica serovar Enteritidis (S. enteritidis) in China. We evaluated the outbreak by using epidemiological surveys, routine laboratory testing methods, and whole genome sequencing (WGS). This outbreak occurred in a canteen in Beijing, during March 9–11, 2021, 225 of the 324 diners who have eaten at the canteen showed gastrointestinal symptoms. The outbreak had characteristical epidemiological and clinical features. It caused a very high attack rate (69.4%) in a short incubation time. All patients developed diarrhea and high fever, accompanied by abdominal pain (62.3%), nausea (50.4%), and vomiting (62.7%). The average frequency of diarrhea was 12.4 times/day, and the highest frequency of diarrhea was as high as 50 times/day. The average fever temperature was 39.4°C, and the highest fever temperature was 42°C. Twenty strains of S. enteritidis were recovered, including 19 from the patients samples, and one from remained egg fried rice. Antibiotic susceptibility test showed that the 20 outbreak strains all had the same resistance pattern. PFGE results demonstrated that all 20 strains bore completely identical bands. Phylogenetic analysis based on WGS revealed that all 20 outbreak strains were tightly clustered together. So the pathogenic source of this food poisoning incident may was contaminated egg fried rice. Resistance gene analysis showed that the outbreak strains are all multi-drug resistant strains. Virulence gene analysis indicated that these outbreak strains carried a large number of virulence genes, including 2 types of Salmonella pathogenicity islands (SPI-1 and SPI-2). Other important virulence genes were also carried by the outbreak strains, such as pef ABCD, rck and shd A. And the shd A gene was not in other strains located in the same evolutionary branch as the outbreak strain. We speculated that this is a significant reason for the serious symptoms of gastroenteritis in this outbreak. This outbreak caused by S. enteritidis suggested government should strengthen monitoring of the prevalence of outbreak clone strains, and take measures to mitigate the public health threat posed by contaminated eggs. [ABSTRACT FROM AUTHOR]

Background: Salmonella enterica subsp. enterica serovar Typhimurium infections continue to be a significant public health threat worldwide. The aim of this study was to investigate antibiotic resistance among 147 S. Typhimurium isolates collected from patients in Henan, China from 2006 to 2015.Methods: 147 S. Typhimurium isolates were collected from March 2006 to November 2015 in Henan Province, China. Antimicrobial susceptibility testing was performed, and the resistant genes of ciprofloxacin, cephalosporins (ceftriaxone and cefoxitin) and azithromycin were detected and sequenced. Clonal relationships were assessed by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE).Results: Of the 147 isolates, 91.1% were multidrug resistant (MDR), with 4.1% being resistant to all antibiotic classes tested. Of concern, 13 MDR isolates were co-resistant to the first-line treatments cephalosporins and ciprofloxacin, while three were also resistant to azithromycin. Seven PFGE patterns were identified among the 13 isolates. All of the isolates could be assigned to one of four main groups, with a similarity value of 89%. MLST assigned the 147 isolates into five STs, including two dominant STs (ST19 and ST34). Of the 43 ciprofloxacin-resistant isolates, 39 carried double gyrA mutations (Ser83Phe, Asp87Asn/Tyr/Gly) and a single parC (Ser80Arg) mutation, including 1 isolate with four mutations (gyrA: Ser83Phe, Asp87Gly; parC: Ser80Arg; parE: Ser458Pro). In addition, 12 isolates not only carried mutations in gyrA and parC but also had at least one plasmid-mediated quinolone resistance (PMQR) gene. Among the 32 cephalosporin-resistant isolates, the most common extended-spectrum β-lactamase (ESBL) gene was blaOXA-1, followed by blaCTX-M, blaTEM-1, and blaCMY-2. Moreover, the mphA gene was identified in 5 of the 15 azithromycin-resistant isolates. Four MDR isolates contained ESBL and PMQR genes, and one of them also carried mphA in addition.Conclusion: The high level of antibiotic resistance observed in S. Typhimurium poses a great danger to public health, so continuous surveillance of changes in antibiotic resistance is necessary. [ABSTRACT FROM AUTHOR]

Salmonella enterica infections continue to be a significant burden on public health worldwide. The ability of S. enterica to produce hydrogen sulfide (H2S) is an important phenotypic characteristic used to screen and identify Salmonella with selective medium; however, H2S-negative Salmonella have recently emerged. In this study, the H2S phenotype of Salmonella isolates was confirmed, and the selected isolates were subjected to antimicrobial susceptibility testing and molecular identification by multilocus sequence typing, pulsed-field gel electrophoresis, and clustered regularly interspaced short palindromic repeat (CRISPR) analysis. The phs genetic operon was also analyzed. A total of 160 S. enterica serovar Aberdeen isolates were detected between 2005 and 2013 in China. Of them, seven non-H2S-producing isolates were detected. Notably, four samples yielded four pairs of isolates with different H2S phenotypes, simultaneously. The data demonstrated that H2S-negative isolates were genetically closely related to H2S-positive isolates. Three new spacers (Abe1, Abe2, and Abe3) were identified in CRISPR locus 1 in four pairs of isolates with different H2S phenotypes from the same samples. Sequence analysis revealed a new nonsense mutation at position 208 in the phsA gene of all non-H2S-producing isolates. Additionally, we describe a new screening procedure to avoid H2S-negative Salmonella, which would normally be overlooked during laboratory and hospital screening. The prevalence of this pathogen may be underestimated; therefore, it is important to focus on improving surveillance of this organism to control its spread. [ABSTRACT FROM AUTHOR]