Emergence of Salmonella Infantis carrying the pESI-like plasmid from eggs in egg grading and packing plants in Korea.

The plasmid of emerging S. Infantis (pESI) or pESI-like plasmid in Salmonella enterica Infantis are consistently reported in poultry and humans worldwide. However, there has been limited research on these plasmids of S. Infantis isolated from eggs. Therefore, this study aimed to analyze the prevalence and characteristics of S. Infantis carrying the pESI-like plasmid from eggs in egg grading and packing plants. In this study, the pESI-like plasmid was only detected in 18 (78.3%) of 23 S. Infantis isolates, and it was absent in the other 9 Salmonella serovars. In particular, S. Infantis isolates carrying the pESI-like plasmid showed the significantly higher resistance to ß-lactams, phenicols, cephams, aminoglycosides, quinolones, sulfonamides, and tetracyclines than Salmonella isolates without the pESI-like plasmid (p < 0.05). Moreover, all S. Infantis isolates carrying the pESI-like plasmid were identified as extended-spectrum ß-lactamase (ESBL) producer, harboring the blaCTX-M-65 and blaTEM-1 genes, and carried non-ß-lactamase resistance genes (ant(3'')-Ia, aph(4)-Ia, aac(3)-IVa, aph(3')-Ic, sul1, tetA, dfrA14, and floR) against five antimicrobial classes. However, all isolates without the pESI-like plasmid only carried the blaTEM-1 gene among the ß-lactamase genes, and either had no non-ß-lactamase resistance genes or harbored non-ß-lactamase resistance genes against one or two antimicrobial classes. Furthermore, all S. Infantis isolates carrying the pESI-like plasmid carried class 1 and 2 integrons and the aadA1 gene cassette, but none of the other isolates without the pESI-like plasmid harbored integrons. In particular, D87Y substitution in the gyrA gene and IncP replicon type were observed in all the S. Infantis isolates carrying the pESI-like plasmid but not in the S. Infantis isolates without the pESI-like plasmid. The distribution of pulsotypes between pESI-positive and pESI-negative S. Infantis isolates was clearly distinguished, but all S. Infantis isolates were classified as sequence type 32, regardless of whether they carried the pESI-like plasmid. This study is the first to report the characteristics of S. Infantis carrying the pESI-like plasmid isolated from eggs and can provide valuable information for formulating strategies to control the spread of Salmonella in the egg industry worldwide.

From farm to fork: Spread of a multidrug resistant Salmonella Infantis clone encoding blaCTX-M-1 on pESI-like plasmids in Central Italy.

Salmonella enterica subsp. enterica serovar Infantis (S. Infantis) is one of the "top five Salmonella serovars" of clinical significance in the European Union (EU). Antimicrobial resistant and extended spectrum ß-lactamase (ESBL) AmpC-producing S. Infantis have been described in food production systems and human clinical samples in Italy. Recently, an increase of MDR S. Infantis carrying blaCTX-M genes involved in 3rd generation cephalosporin resistance was noticed in the EU, including Italy, mainly due to the spread of S. Infantis harboring a pESI-like plasmid. The aim of this study was to investigate the occurrence of the S. Infantis pESI-like plasmid among antibiotic resistant S. Infantis strains isolated at different points of the food chain, and to provide a phylogenetic analysis to gain further insight on their transmission pathways from 'farm to fork'. MDR S. Infantis strains (n. 35) isolated from 2016 to 2021 at different stages of the food chain (animals, food, food-related environments, and humans) were investigated with in depth molecular characterization using real-time PCR, S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and whole genome sequencing (WGS). Our study reported the occurrence of S. Infantis strains harboring pESI-like plasmids, carrying blaCTX-M-1 genes, in Central Italy, at different sampling points along the food chain. Results confirmed the presence of a plasmid with a molecular size around 224-310 kb, thus consistent with the pESI-like, in 97 % of the 35 samples investigated. Two variants of S. Infantis pESI-like IncFIB(K)_1_Kpn3 were detected, one associated with the European clone carrying blaCTX-M-1 (21 isolates) and the other associated with U.S. isolates carrying blaCTX-M-65 (2 isolates, pESI-like U.S. variant). The majority was resistant to 3rd generation cephalosporins but none of the strains tested positive for the carbapenemase encoding genes. A total of 118 virulence genes were identified in isolates harboring the pESI-like plasmid. cgMLST and SNP-based analysis revealed the presence of one main cluster, composed by strains isolated from the environment, animals, food and humans. The results of this investigation underline the importance of phylogenetic studies to monitor and understand pathogen and AMR spread in a One Health approach.

Infection dynamics of Salmonella Infantis strains displaying different genetic backgrounds - with or without pESI-like plasmid - vary considerably.

Food-borne infections with Salmonella are among the most common causes of human diseases worldwide, and infections with the serovar Infantis are becoming increasingly important. So far, diverse phenotypes and genotypes of S. Infantis have been reported. Therefore, the present study aimed to investigate the infection dynamics of two different S. Infantis strains in broilers. For this purpose, 15 birds were infected on day 2 of life with 108 CFU/ml of a pESI+ or a pESI- S. Infantis strain, respectively. Ten uninfected birds served as in-contact birds to monitor transmission. In both groups, an increase of infection was observed from 7 days of age onwards, reaching its peak at 28 days. However, the pESI+ strain proved significantly more virulent being re-isolated from most cloacal swabs and organs by direct plating. In contrast, the pESI- strain could be re-isolated from cloacal swabs and caeca only when enrichment was applied. Although the excretion of this strain was limited, the transmission level to in-contact birds was similar to the pESI+ strain. Differences in infection dynamics were also reflected in the antibody response: whereas the pESI+ strain provoked a significant increase in antibodies, antibody levels following infection with the pESI- strain remained in the range of negative control birds. The actual findings provide for the first time evidence of S. Infantis strain-specific infectivity in broilers and confirm previous observations in the field regarding differences in persistence on farms and resistance against disinfectants.

Mega-plasmid found worldwide confers multiple antimicrobial resistance in Salmonella Infantis of broiler origin in Russia.

Plasmids which are the mobile part of the bacterial genome can acquire and carry over genes conferring antimicrobial resistance, thus contributing to rapid adaptation of bacterial community to human-defined environment. In 2014, Israeli scientists have reported a large conjugative mega-plasmid pESI (plasmid for emerging S. Infantis) that provides multiple drug resistance (MDR) of Salmonella Infantis isolated from broilers. Later, very similar pESI-like plasmids have been found in Salmonella isolated from poultry in the United States, Italy, Switzerland, Hungary, and Japan. Here we report detection of pESI-like plasmids in Salmonella Infantis isolated from chicken food products in Russia. Whole genome sequencing of three MDR isolates revealed pESI-like plasmids in all three cases. These plasmids have such typical pESI features as a locus for siderophore yersiniabactin, a cluster of IncI1 conjugative genes, a cluster of type IV pilus genes, and three toxin-antitoxin modules. The pESI-like plasmids carry from two to five resistance genes in each isolate. In total, we observed six antimicrobial resistance genes associated with pESI-like plasmids (aadA1, blaCTX-M-14, dfrA14, sul1, tetA/tetR, tetM). Besides plasmid genes of antimicrobial resistance, all three MDR isolates of S. Infantis harbor a mutation in chromosomal gene gyrA (p.S83Y or p.D87Y) that is associated with resistance to fluoroquinolones. In addition, we performed a comparative bioinformatics meta-analysis of 25 pESI-like plasmids hosted by S. Infantis from the USA, Europe, Latin America, Israel, and Japan. This analysis identified a 173 kB sequence that is common for all pESI-like plasmids and carries virulence operons and toxin-antitoxin modules.

Emergence of Multidrug-Resistant Salmonella enterica Subspecies enterica Serovar Infantis of Multilocus Sequence Type 2283 in German Broiler Farms.

During the last decade, Salmonella enterica subspecies enterica serovar Infantis (S. Infantis) has become more prevalent across Europe with an increased capability to persist in broiler farms. In this study, we aimed to identify potential genetic causes for the increased emergence and longer persistence of S. Infantis in German poultry farms by high-throughput-sequencing. Broiler derived S. Infantis strains from two decades, the 1990s (n = 12) and the 2010s (n = 18), were examined phenotypically and genotypically to detect potential differences responsible for increased prevalence and persistence. S. Infantis organisms were characterized by serotyping and determining antimicrobial susceptibility using the microdilution method. Genotypic characteristics were analyzed by whole genome sequencing (WGS) to detect antimicrobial resistance and virulence genes as well as plasmids. To detect possible clonal relatedness within S. Infantis organisms, 17 accessible genomes from previous studies about emergent S. Infantis were downloaded and analyzed using complete genome sequence of SI119944 from Israel as reference. In contrast to the broiler derived antibiotic-sensitive S. Infantis strains from the 1990s, the majority of strains from the 2010s (15 out of 18) revealed a multidrug-resistance (MDR) phenotype that encodes for at least three antimicrobials families: aminoglycosides [ant(3")-Ia], sulfonamides (sul1), and tetracyclines [tet(A)]. Moreover, these MDR strains carry a virulence gene pattern missing in strains from the 1990s. It includes genes encoding for fimbriae clusters, the yersiniabactin siderophore, mercury and disinfectants resistance and toxin/antitoxin complexes. In depth genomic analysis confirmed that the 15 MDR strains from the 2010s carry a pESI-like megaplasmid with resistance and virulence gene patterns detected in the emerged S. Infantis strain SI119944 from Israel and clones inside and outside Europe. Genotyping analysis revealed two sequence types (STs) among the resistant strains from the 2010s, ST2283 (n = 13) and ST32 (n = 2). The sensitive strains from the 1990s, belong to sequence type ST32 (n = 10) and ST1032 (n = 2). Therefore, this study confirms the emergence of a MDR S. Infantis pESI-like clone of ST2283 in German broiler farms with presumably high tendency of dissemination. Further studies on the epidemiology and control of S. Infantis in broilers are needed to prevent the transfer from poultry into the human food chain.