Genomic Epidemiology Of Salmonella Enterica Serovar Newport

Salmonella enterica is one of the major foodborne pathogens and is a major threat to global health and economic development.Salmonellosis is caused by various Salmonella serotypes that differ significantly in their ability to infect different hosts and to colonize or surivive in dynamic niches.Salmonella enterica.subsp.enterica serovar Newport(S.Newport)is one of the top 3 serotypes for Salmonella infection in the United States.Cattle is generally considered as the primary host of infection or carrier.The incidence of S.Newport infection has increased in the United States in recent years while in the European Union S.Newport infection has remained stable over the past few decades.In general,S.Newport disseminates through human consumption of contaminated animal food,such as beef,pork,poultry and milk or non-animal sources,such as vegetables and other fresh produce.However recent outbreaks have been attributed to various other sources,such as fresh produce,seafood,irrigation water and soil.In addition,S.Newport has developed a special capacity for long-term survival in the environment such as fertilizers.Particularly,it has the ability to colonize and infect the plant.It is worth noting that S.Newport can colonize,invade plant tissues during food processing.The dynamic prevalance of S.Newport can serve as a source of persistence,posing the reservoir for human infections.Only few investigations have focused on different routes of transmission,particularly in the context of the transmission of antimicrobial-resistant Salmonella to human.Moreover,the major ecology,antimicrobial resistance and disease burden of S.Newport in China are largely unknown.Whole genome sequencing data,multilocus sequence analysis(MLST)and multilocus enzyme electrophoresis(MLEE)analysis confirmed S.Newport as a serotype of multiple phylogenetic origins.MLST was used to analyze 384 S.Newport isolates from different hosts and geographic habitats,and three evolutionary clades were identified,including human clade I and non-human clade ?(usually from avian,livestock and reptile)whose antimicrobial resistance profiles were different.And there was a large niche difference between them.They were likely to show host preference.However,few studies have focused on dissemination in food chain,and analysis of population structure and host preference through global S.Newport genomes remains unclear as far as we learned.Therefore,in present study,we attempt to:(1)obtain the pattern of antimicrobial resistance of S.Newport from different sources;(2)analyze the basic rule of S.Newport evolution through MLST;(3)obtain the genomic epidemiological characteristics of S.Newport of Chinese human origin;(4)dissect the genetic evolution and antimicrobial resistance mechanism of Globally sampled S.Newport;(5)explore the potential mechanism of host preference of S.Newport at genome-wide level.?.Clustering of American S.Newport by antimicrobial-resistance pattern during 1996?2015.Characterization of transmission routes of Salmonella among various food-animal reservoirs and their antibiogram is crucial for appropriate intervention and medical treatment.Here,we analyzed 3,728 Salmonella Newport isolates collected from various food-animals,retail meats and humans in the United States between 1996 and 2015,based on their minimum inhibitory concentration(MIC)towards 27 antimicrobials.Random Forest and Hierarchical Clustering statistic was used to group the isolates according to their MICs.Classification and Regression Tree(CART)analysis was used to identify the appropriate antibiotic and its cut-off value between human-and animal-population.Two distinct populations were revealed based on the MICs of individual strain by both methods,with the animal population having significantly higher MICs which correlates to antibiotic-resistance(AR)phenotype.Only?9.7%(267/2763)human isolates could be attributed to food-animal originS.Furthermore,the isolates of animal origin had less diverse antibiogram than human isolates(P<0.001),suggesting multiple sources involved in human infection.CART identified trimethoprim-sulfamethoxazole to be the best classifier for differentiating the animal and human isolates.Additionally,two typical AR patterns,MDR-Amp(Multi-drug resistant Ampicillin)and Tet-SDR(Tetracycline single-drug resistant)dominant in bovine-or turkey-population,were identified,indicating that distinct food-animal sources could be involved in human infections.The AR analysis suggested fluoroquinolones(i.e.ciprofloxacin),but not extended-spectrum cephalosporins(i.e.ceftriaxone,cefoxitin),is the adaptive choice for empirical therapy.Antibiotic-resistant S.Newport from humans has multiple origins,with distinct food-animal-borne route contributing to a significant proportion of heterogeneous isolates.?.Resolve population structure by MLST typing.Salmonella Newport,with phylogenetic diversity feature,contributes to significant public health concerns.Our previous study suggested that S.Newport from multiple animal-borne routes,with distinct antibiotic resistant pattern,might transmit to human.However,their genetic information was lacking.Therefore,this study investigated the relationship between each other among the hosts,sources,genotype and antibiotic resistance in S.Newport.We used the multilocus sequence typing(MLST)in conjunction with minimum inhibitory concentration of 16 antimicrobials of globally sampled 1842 S.Newport strains,including 282 newly contributed Chinese strains,to evaluate this association.Our analysis reveals that sequence types(STs)are significantly associated with different host sources,including livestock(ST45),birds(ST5),contaminated water and soil(ST118),reptiles(ST46)and seafood(ST31).Importantly,ST45 contained most of(344/553)the multi-drug resistance(MDR)strains,which were believed to be responsible for human MDR bacterial infections.Chinese isolates were detected to form two unique lineages of avian(ST808 group)and freshwater animal(ST2364 group)origin.Taken together,genotyping information of S.Newport could serve to improve Salmonella source-originated diagnostics and guide better selection of antibiotic therapy against Salmonella infections.?.Genomic epidemiology of S.Newport of Chinese human origin during 1991?2018Salmonella Newport infections are gradually on the rise in China from the last decade.For human infections,S.Newport has been ranked among the top five serovars responsible for persistent infections,globally.A total of 290 S.Newport strains with their relevant clinical metadata were analyzed,and the strains were subjected to whole-genome sequence analysis.Among these,62.4%(n=181)were from diarrheic patients and 28.9%(n=84)were from asymptomatic individuals(including adults and youngsters)while 8.6%(n=25)were from cases of persistent diarrhea in infants(28%,n=7)and toddlers(72%,n=18).The association between the sequence types(STs)and the variations in the clinical presentation was statistically significant(P=0.0432),with ST46 causing diarrhea or representing asymptomatic patients and ST31 or ST68 causing persistent diarrhea.Genomic analysis revealed that the highest proportion of the isolates(98.5%,n=279),primarily from patients with or without diarrhea rather than from asymptomatic individuals,carried antimicrobial resistance determinants corresponding to the aminoglycosides and beta-lactams,highlighting the need for cautionary usage of antimicrobials in such patients.These findings also suggest that cases of nontyphoidal Salmonella infection with symptoms of acute diarrhea or persistent diarrhea caused by S.Newport should be handled with caution,due to the high chance of development of an antimicrobial resistance phenotype that might lead to therapeutic failures.Together,S.Newport ST31 and ST46,which have the highest frequency of carriage of MDR,are potentially responsible for antimicrobial resistant diarrhea/persistent diarrhea in infants and children,while adult humans are more likely to be asymptomatic carriers of the S.Newport strains.?.Analysis of genetic evolution,antimicrobial resistance mechanism and host preference by global genomesGlobal 1560 S.Newport isolates could be divided into four clades,C-??C-?.ST45(28.42%),ST118(19.56%),ST46(13.60%)are the major ones.The main sources were human(42.05%),livestock(21.03%),cold-blooded animal(8.08%).One main lineage,ST45-WBM(warm-blooded mammal)of C-?,which mainly consisted of livestock strains,generally carried IncA/C2 plasmid with a large number of antimicrobial resistance genes.ST 166 and ST31 also carried a few plasmids with some antimicrobial-resistant genes.S.Newport contained 9 classes of antimicrobial-resistant genes among which aminoglycosides and tetracycline ranked top 2.Apart from rifampicin which mainly came from cold-blooded animal(CBA)and human,the other classes mainly came from WBM and avian.BEAST spatio-temporal analysis indicated that the molecular clock rate of S.Newport was at moderately high level among other Salmonella serotypes or species and C-I had the earliest differentiation time point with the lowest molecular clock rate while C-? had the highest.Analysis of main lineage ST45-WBM of C-?,suggested that the MDR IncA/C2 plasmid was the main driver of the emergence of ST45 among livestocks.After choosing representative strains of main STs and testing their phenotypes,it was verified that there was difference among the three main lineages ST45-WBM,ST46-CBA,ST118&5-Human in virulence to mice model.It was also confirmed that there was difference in host preference by infection tests to zebrafish and C.elegans model and in the abilities of biofilm formation and swarming motility.Genome-wide heatmap suggested a recombination characteristic of S.Newport and some genes that assisted lineages for host adaptation.Through the mining of three main lineages' genomes,it was found that their specific genes that assisted their host-adaptation were mainly categoried in nutrients metabolism&transport,and genetic material operation.Virulence gene scanning results showed the total number of virulence genes and indicated it was negatively correlated with the total number of resistance genes.Salmonella pathogenicity island(SPI)and mobile genetic element scanning results indicated that ST45-WBM lineage was characteristic of four specific transposons and one type I integron while ST118&5-human lineage contained a specific SPI.The detection results of recombination events indicated that S.Newport was the serotype with the widest genomic recombination region,the fastest recombination rate and the highest average number of events per strain among isolated Salmonella in the last hundred years.Among them,the reticulate evolution phenomenon was prominent in ST45-WBM.XerC₃ was its specific recombinase,which may support genetic diversity and dynamic prevalence.In conclusion,this study reveled a general phenomnon of antimicrobial-resistance of S.Newport,S.Newport group clustering by antimicrobial-resistance pattern,the best antimicrobial classifier,genotypes,ecological distribution and main route of transmission to human.This study also dissected the epidemiological features of the genomes of strains from Chinese human origin and formulate a global picture regarding population structure,phylogenomic evolution history,the antimicrobial-resistant genetic deteminants,the host preference,evolutionary pattern with lineage-specific molecular targets of S.Newport.This study pointed out that IncA/C2 plasmid could be a major driver for the emergence of ST45 among livestocks,and newly detected recombinase and nutrition metabolic transport component may be involved in mediating host adaptation.The gained knowledge in this study provides a new avenue for further investigation of dynamic Salmonella behavior and evolutionary history.