Horizontal Dissemination of Antimicrobial Resistance Determinants in Multiple Salmonella Serotypes following Isolation from the Commercial Swine Operation Environment after Manure Application

Abstract

The aim of this study was to characterize the plasmids carrying antimicrobial resistance (AMR) determinants in multiple Salmonella serotypes recovered from the commercial swine farm environment after manure application on land. Manure and soil samples were collected on day 0 before and after manure application on six farms in North Carolina, and sequential soil samples were recollected on days 7, 14, and 21 from the same plots. All environmental samples were processed for Salmonella, and their plasmid contents were further characterized. A total of 14 isolates including Salmonella enterica serotypes Johannesburg (n ϭ 2), Ohio (n ϭ 2), Rissen (n ϭ 1), Typhimurium var5Ϫ (n ϭ 5), Worthington (n ϭ 3), and 4,12:i:Ϫ (n ϭ 1), representing different farms, were selected for plasmid analysis. Antimicrobial susceptibility testing was done by broth microdilution against a panel of 14 antimicrobials on the 14 confirmed transconjugants after conjugation assays. The plasmids were isolated by modified alkaline lysis, and PCRs were performed on purified plasmid DNA to identify the AMR determinants and the plasmid replicon types. The plasmids were sequenced for further analysis and to compare profiles and create phylogenetic trees. A class 1 integron with an ANT(2Љ)-Ia-aadA2 cassette was detected in the 50-kb IncN plasmids identified in S. Worthington isolates. We identified 100-kb and 90-kb IncI1 plasmids in S. Johannesburg and S. Rissen isolates carrying the bla CMY-2 and tet(A) genes, respectively. An identical 95-kb IncF plasmid was widely disseminated among the different serotypes and across different farms. Our study provides evidence on the importance of horizontal dissemination of resistance determinants through plasmids of multiple Salmonella serotypes distributed across commercial swine farms after manure application. IMPORTANCEThe horizontal gene transfer of antimicrobial resistance (AMR) determinants located on plasmids is considered to be the main reason for the rapid proliferation and spread of drug resistance. The deposition of manure generated in swine production systems into the environment is identified as a potential source of AMR dissemination. In this study, AMR gene-carrying plasmids were detected in multiple Salmonella serotypes across different commercial swine farms in North Carolina. The plasmid profiles were characterized based on Salmonella serotype donors and incompatibility (Inc) groups. We found that different Inc plasmids showed evidence of AMR gene transfer in multiple Salmonella serotypes. We detected an identical 95-kb plasmid that was widely distributed across swine farms in North Carolina. These conjugable resistance plasmids were able to persist on land after swine ma-

Whole genome sequencing analysis of multiple Salmonella serovars provides insights into phylogenetic relatedness, antimicrobial resistance, and virulence markers across humans, food animals and agriculture environmental sources

“…Plasmids were observed specific to Salmonella serovar that was very similar to the AMR genes (Table 3 ). This is in accordance to our previous study that the plasmid profiles were correlated to Salmonella serovar and incompatibility (Inc) groups [ 13 ]. The IncF (both FI and FII) family found across the different Salmonella serovars in our study is known to be a well-adapted and commonly distributed plasmid among members of the Enterobacterceae family [ 53 , 54 ].…”

“…The IncF (both FI and FII) family found across the different Salmonella serovars in our study is known to be a well-adapted and commonly distributed plasmid among members of the Enterobacterceae family [ 53 , 54 ]. Although our data cannot fully explain the transmission of AMR determinants among various species, they are in line with previous studies that reported on the role of animals and environment as important sources of multiple AMR genes as well as plasmids, and that transmission can occur by horizontal gene transfer [ 13 , 55 , 56 ].…”

“…The selection pressure on Salmonella is created by antimicrobial use in human health and food animal production leading to development and potential spread of antimicrobial resistance [ 8 – 11 ]. Our previous studies reported the persistence and dissemination of multiple resistant Salmonella serovars along with their determinants in the environment of commercial swine operation due to the manure application on land [ 12 , 13 ].…”

“…The Centers for Disease Control and Prevention (CDC) reported that the incidence of human Salmonella infections caused by monophasic 4,[5],12:i:-, which is in the top 4 of the most frequently reported Salmonella serovars, continue to rise while the incidence of the other serovars is decreasing [ 16 ]. The increase in the incidence of this serovar in human cases is paralleled by a similar increase in swine and environmental detection of this serovar variant [ 12 , 13 , 18 ]. However, there are gaps that still exist in our understanding of the temporal and spatial connection of resistant Salmonella transmission within humans, animals, and the environment sources.…”

Shifts in microbial community, pathogenicity‐related genes and antibiotic resistance genes during dairy manure piled up

“…Eventually, humans could become the ultimate receptors of ARGs. Previous studies have also confirmed that HGT of ARGs could impact food safety by the enrichment of ARGs in the pasture or in the composting process of pig, cow and poultry manure (Soucy et al , 2015; Baker et al , 2016; Pornsukarom and Thakur, 2017).…”