Prevalence and genomic investigation of Salmonella isolates recovered from animal food-chain in Xinjiang, China

Highlights

• Salmonella was detected in 314 of the 2408 samples (13.04%).

• Six serovars and four serogroups were identified with dominance of Agona and serogroup O:4(B).

• 92.04%of studied isolates were multi-drug resistance (MDR).

• We reported the presence of cdtB gene encoding typhoid toxins in four S. Indiana isolates.

• We demonstrated the use of genomic framework for routine surveillance in China.

Abstract

Salmonella is a major foodborne pathogen worldwide, causing serious cases of morbidity and mortality due to the consumption of contaminated foods. Animal-borne foods were considered the main source of transferring Salmonella to humans; however, route surveillance by genomic platforms along the food-chain is limited in China. Here, we proceeded to the application of whole genome sequencing in the epidemiological analysis of Salmonella isolated along the food-chain in Xinjiang, China. A total of 2408 samples were collected from farms, slaughterhouses, and markets, and subjected to the isolation of Salmonella strains. 314 (13.04%) of the samples were positive for Salmonella. Phenotypic antimicrobial resistance was conducted by the broth dilution method using 14 antimicrobial agents belonging to ten classes for all 314 isolates. A selection of representative 103 isolates was subjected to whole-genome sequencing for understanding the Salmonella diversity, including serovars, antimicrobial and virulence genes, plasmid types, multi-locus sequence types, and allelic types. We found that S. Agona was the dominant serovar and O:4(B) was the dominant serogroup. The dominant genotype was ST13 and each serovar has a unique MLST pattern. Plasmids prediction reported Col(MGD2)_1 and Col(Ye4449)_1 as the dominant plasmids, in addition to the detection of IncFII(S)_1 and IncFIB(S)_1 carried by all S. Enteritidis isolates. Importantly, virulence genes prediction showed the presence of cdtB gene encoding typhoid toxins, spv genes, and pef gene cluster encoding fimbriae in the genomes of S. Indiana and S. Enteritidis. Phenotypic antimicrobial resistance identified 92.04% of the sampled isolates as multi-drug resistance (MDR), with high resistance to tetracycline (78.03%; 245/314), amoxicillin/ clavulanic acid (75.80%; 238/314), and ampicillin (70.70%; 222/314). Together, we firstly reported the prevalence of MDR Salmonella isolates harboring critical virulence factors transmission via animal-borne food-chain in Xinjiang, hence route surveillance by whole-genome sequencing platform could facilitate recognition and project early warning for the emerging MDR clones along the food-chain.

Introduction

Salmonella spp. are important foodborne pathogens worldwide contaminating a wide range of animal-borne products, including meat and meat products. Salmonella strains growing in animal farms frequently cross-contaminate animal carcasses during slaughtering operations before being transmitted to humans via the food-chain (Bonardi, 2017; Z. Jiang et al., 2019, Liu et al., 2020). In fact, previous studies have reported the high contamination of meat products with Salmonella strains (Ed-Dra et al., 2017, Yang et al., 2019). Therefore, the consumption of contaminated meat and meat products may lead to severe cases of salmonellosis, characterized by self-limited gastroenteritis in immunocompetent adults and invasive infections, particularly in immunocompromised patients (Elbediwi et al., 2020, Xu et al., 2020a, Yue, 2016).

Diarrheal and invasive non-typhoidal Salmonella infections were classified among the major foodborne diseases worldwide (Kirk et al., 2015). In fact, it is estimated that Salmonella strains were responsible for about 180 million (9%) of the diarrheal illnesses that occur globally each year, and caused about 298,000 deaths which represent 41% of all diarrheal disease-associated deaths (Besser, 2018). In the United States, it is estimated that non-typhoidal Salmonella were responsible for about one million cases of foodborne illnesses each year, with 19,586 hospitalizations and 378 deaths (Scallan et al., 2011). In the European Union, 91,662 confirmed human salmonellosis cases were reported during the year 2017, with an incidence rate of 19.7 cases per 100,000 populations (EFSA & ECDC, 2018). In China, Salmonella was classified among the major foodborne pathogens, causing n foodborne outbreaks (Paudyal et al., 2018, Zhou et al., 2020). Currently, Salmonella remains an important foodborne pathogen worldwide, presenting a major threat to public health with a significant economic burden (EFSA & ECDC, 2018, Kirk et al., 2015, Pan et al., 2019).

The use of antibiotics for treatment or prevention of bacterial infection in both human and animal fields has increased significantly during past decades. However, the overuse and misuse of antibiotics have led to the selection or accumulation of antimicrobial resistance in numerous bacterial species, especially those belonging to the Enterobacteriaceae family (Ed-Dra et al., 2019, Qiao et al., 2018, Sampaio and Gales, 2016). Consequently, the emergence of antibiotic-resistant bacteria has reached different corners of life, including humans, animals, environments, and food-chains (Elbediwi et al., 2019, Paudyal and Yue, 2019, Qiao et al., 2018). In fact, the World Health Organization classified antimicrobial resistance as a global public health crisis that must be managed with the utmost urgency (WHO, 2017). Importantly, surveillance data based on the epidemiological investigations demonstrated a dramatic increase in overall antimicrobial resistance of Salmonella strains since the early 1990s. This resistance was firstly linked to penta-resistant S. Typhimurium DT104 and then to the appearance of Salmonella strains resistant to the 3rd generation cephalosporin and quinolone (Eller et al., 2013; H. X. Jiang et al., 2014, Le Hello, 2014, Su et al., 2004). Recently, numerious studies reported the emergence of colistin resistance in different Salmonella serovars (Biswas et al., 2019, Elbediwi et al., 2020, Elbediwi et al., 2020, Lima et al., 2019). The high spreading of antimicrobial resistant Salmonella strains in food products is a serious problem for public health worldwide and the multidrug resistant strains originated from food products could be transmitted to humans leading to adverse outcomes.

Surveillance of Salmonella along the food-chain has a major concern for public health; in fact, conventional various labor intensive methods have been used to evaluate the antimicrobial resistance, virulence, plasmids profiles as well as the discrimination of isolates when an outbreak occurs (Ed-Dra et al., 2018, Murgia et al., 2015, Skyberg et al., 2006), which requires long-time and sometimes includes bias assessments for the choice of the particular method. In most of the cases, the results lose the value for a comparative purpose among relevant studies. Recently, the development in sequencing technologies stimulated the use of whole genome sequencing (WGS) as one-step approach for an unbiased understanding of serovars, antimicrobial resistance, virulence genes, plasmid types, multi-locus sequence types, and importantly, for an early recognition of emerging threats that could be responsible for severe humans and/or animal infections (Köser et al., 2014, Liu et al., 2020, Thomas et al., 2017, Viana et al., 2020, Xu et al., 2020a, Yu et al., 2020).

Xinjiang is considered the largest Chinese Autonomous Region with a superficial of about 1.6 million km². In Xinjiang, animal husbandry is considered the major activity of the local population, and animal-borne foods have taken a principal place in the local diet. In fact, animal-borne foods have been known as a major vehicle for foodborne pathogen transmission, including Salmonella. So far, studies investigated the prevalence of Salmonella in animal food-chain in Xinjiang are very limited (Yue et al., 2020). In this study, we investigated the prevalence and antimicrobial resistance of Salmonella isolates in animal farms, slaughterhouses, and food markets. Moreover, genomic sequences of 103 strains were conducted in order to produce a streamlined study of serovars, antimicrobial and virulence genes, plasmid types, multi-locus sequence types, and allelic types.