| Salmonella is a common foodborne pathogen causing salmonellosis of mammals,birds,reptiles,amphibians,etc.Human can also be infected through livestock and poultry products.The typing of Salmonella is very complicated.At present,it can be divided into nearly 2600 serotypes.Most of Salmonella can be pathogenic to humans and animals.The pathogenicity of different serotypes of Salmonella is pantropic and specific.Under the pressure of antibiotic selection,drug-resistant strains of Salmonella are increasing.It poses new challenges to public health.In this paper,the serotype,MLST typing,drug resistance and pathogenicity of 60 Salmonella isolates from animal origin in China during 1950 to 2009 were studied.It is hoped that the relationship among the typing,drug resistance and pathogenicity of Salmonella will be understood to provide basic information for detection,prevention and control of salmonellosis.The 60 strains of Salmonella from animal origin were collected in this study,including 33 isolates from chicken(55%),25 from pig(41.67%)and 2 from horse(3.33%).Among these strains,the Salmonella choleraesuis,Salmonella Enteritidis and Salmonella Pullorum were the main serotypes.Salmonella Typhimurium,Salmonella Abortusequi,Salmonella Tennessensis and Salmonella Indiana serotypes accounted for 10%(6/60),3.330%(2/60),3.33%(2/60)and 3.33%(2/60),respectively.In the 1950-1969.the serotypes were Salmonella Choleraesuis(12/20),Salmonella Typhimurium,(2/20),Salmonella Pullorum(4/20)and Salmonella Enteritidis(2/20).In 1970-1989,the serotypes were Salmonella Choleraesuis(7/20)and Salmonella Pullorum(7/20).The Salmonella Typhimurium(4/20)and Salmonella Abortusequi(2/20)were also isolated.In 1990-2009,the serotypes were Salmonella Enteritidis(14/20),Salmonella Tennessensis(2/20)and Salmonella Indiana(2/20).There were 9 ST types by MLST analysis of the 60 strains of Salmonella.The proportion of ST68,ST11 and ST92 were 30%(18/60),26.67%(16/60)and 15%(9/60),respectively.The rest of the strains were ST19,ST132,ST17,ST251,ST319,and ST139.MLST typing results correspond well with serotyping results.ST11 is Salmonella Enteritidis,ST17 is Salmonella Indiana,ST92 and ST132 are Salmonella Pullorum,ST319 is Salmonella Tennessensis,ST68 and ST139 are Salmonella Choleraesuis,ST19 is Salmonella Typhimurium,ST251 is Salmonella Abortusequi.In addition,MLST typing was slightly higher than serum typing method,the different ST types would be distinguished in the same serotype.This indicated that the phylogenetic evolution and phylogenetic relationship of different isolates within the same serotype could be further analyzed by MLST typing.In this study,the drug sensitivity test of Salmonella to ampicillin(AM),doxycycline(DOX),tetracycline(TE)and sulfanilidoxazole(SF)showed a bimodal distribution,and the distribution of drug resistant strains and sensitive strain was relatively concentrated;more than 30%of the strains were less sensitive to polymyxin(CL),and all strains were sensitive to meropenem(MEM).Amoxicillin/clavulanic acid(A/C),gentamicin(GM),spectinomycin(SPF),ofloxacin(OFL),enrofloxacin(ENR)and compound sulfamethoxazole(SXT)had better antibacterial effect,and the proportion of resistant strains was less than 5%.Ceftiofur(CEF),florfenicol(FFC)and ceftazidime(CAZ)were slightly less effective than the above antibiotics,11.67%,8.33%and 6.67%of the strains produced resistance respectively.According to the statistics of isolation time,only one strain of Salmonella was intermediary to DOX,and the others were sensitive to 15 antibiotics.In 19'60-1969,the sensitive strains to CAZ,GM,SPF and CL decreased,4 strains were intennediary to SPF and 2 strains were resistance to CL.Salmonella strains isolated in 1970-1989 were resistant to DOX,GM,SPF and CL,and the number of strains above the intermediate value was not significantly different from that in the 1960s;during 1990-1999,the resistance of Salmonella suddenly increased,more than 50%of the strains were resistant to AM,TE,SF and CL,and 60%of the strains were resistant to AM,TE,SF and CL.In 2000-2009,more than 50%of Salmonella strains were resistant to AM,SF,TE,DOX,CL and CEF,and resistant to 14 antibiotics except meropenem.23 strains of Salmonella were resistant to antibiotics,of which 5 strains were resistant to only one antibiotic,18 strains were resistant to two or more antibiotics,and 9 strains(50%)and 6 strains(33.33%)were resistant to four or five antibiotics,respectively.Even a super MDR isolates which could resistance 12 drug appeard in 2000s.There were 12 drug resistance spectrums the AM-TE-DOX-SF was the dominant drug resistance spectrum,accounting for 22.22%.Only one drug resistance spectrum was found in 50%of Salmonella.According to the isolation time,the main resistance spectrum was AM-TE-SF-CL and AM-TE-DOX-SF in 1990s,AM-TE-DOX-SF-CL and AM-TE-DOX-SF in 2000s.A total of 12 Salmonella strains contained AM-TE-SF resistance combinations,accounting for 66.67%of all multidrug resistant strains.It would be a fixed drug resistance spectrum.14 pairs of primers were used to detect sulfonamides,beta-lactams,aminoglycosides,chloramphenicols,tetracyclines and quinolones,respectively.Among them,sul2,blaTEM and let(A)were found to be the most common resistant genes,accounting for 76.19%,71.43%and 61.90%of the tested samples,23.81%for(?)ac(6)-1b and floR,19.05%for aacC4 and tet(G),blaOXA was 14.29%,blaCMY,blaPSE,blaSHV,catA1,qnr A and qnrB were not detected,blaOXA,aacC4,aac(6)-1b,floR and let(A)were detected in two multidrug resistant strains which resistance more than 11 kinds of drug.This genome combination should be the main reason for super-resistant clones.In this study,ICR mice were selected as experimental animals,and a unified animal model was established to compare the pathogenicity of Salmonella from different animal sources and reduce the differences between experimental animals.According to the results,60 strains of Salmonella were mainly concentrated in 107 CFU,accounting for 41.67%of the total strains.In 1960-1989,ID50 of the pathogenic bacteria was concentrated in 108 CFU,accounting for 50%of the isolates in this period.The most virulent strains were isolated from 2000 to 2009.The median of ID50 was 1.41 ×107.The virulence of Salmonella isolated from 1960 to 1969 was the weakest.The median was 12.28 ×107.Comparing the standard deviation of Salmonella strains in different ages,it was found that the fluctuation amplitude of the standard deviation was large,indicating that the pathogenicity of Salmonella strains in the same age was significantly different.Studies on the pathogenicity of Salmonella showed that the strongest was Salmonella Indiana with ID50 of 106,followed by Salmonella Typhimurium and Salmonella Enteritidis with ID50 of 107,Salmonella choleraesuis,Salmonella Pullorum,Salmonella Abortusequi and Salmonella Tennessensis with ID50 of 108.Compared with different strains of same serotype,the ID50 of Salmonella Typhimurium,Salmonella Enteritidis,Salmonella Tennessensis and Salmonella Indiana were similar,and the standard deviation(M ± SD)was less than 5.Salmonella choleraesuis,Salmonella Pullorum and Salmonella Abortusequi were very different.The standard deviation is about 10.The relationship between pathogenicity and ST type of Salmonella was analyzed.The pathogenicity of ST139 was the strongest,with ID50 of 4.4 ×106,followed by ST17 strain with ID50 of 9×106,ST19,ST11 and ST92 strains with ID50 of 107,ST68,ST132,ST251 and ST319 of 108.The standard deviations of pathogenicity of ST19,ST11,ST132,ST17 and ST319 were all less than 5,and the differences of pathogenicity among strains were small.The standard deviations of ST68,ST92 and ST251 were about 10.The above results showed that the pathogenicity of ST19,ST11,ST132,ST17 and ST319 were correlated with serotype/ST.The pathogenicity of the homozygous strains was not related to the pathogenicity,but only to the type specificity.There were 22 drug resistant strains studied,of which one super-resistant strain(12 kinds of resistant strains)was the most pathogenic with ID50 of 7 ×105,and the other multiresistant strains(drug resistant species>2)ID50 were Med in 107.One double-resistant strain with ID50 of 1.2 ×109,and the other strains were only resistant to one antibiotic with ID50 of 108.From the general trend,we can infer that the increase of Salmonella resistance will have a positive effect on pathogenicity.In summary,the following conclusions were drawn:(1)The 60 strains of Salmonella from animal origin in this study were typed into 7 serotypes and 9 ST types.Comparing the results of serotyping with MLST typing,most of the ST types could correspond to serotypes.The results of Salmonella typing were significantly correlated with the age of isolation.(2)The isolation rate of strains resistant to AM,TE,DOX,SF and CL was higher,forming an AM-TE-SF drug resistance spectrum(isolation rate was 66.67%).Detecting drug resistance gene,blaTEM,sul2 and let(A)were in high rate.(3)The pathogenicity of Salmonella is a type-specific characteristic,which is only related to the serotype/ST of Salmonella. |
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