| Staphylococcus aureus(S.aureus)is an important zoonotic food-borne pathogens causing severe invasive infections because of its virulence factors.Staphylococcal enterotoxin O(SEO)is a member of the new types of SEs with superantigenic and emetic activity.It has been reported that some virulence factors of S.aureus such as?-Hemolysin(Hl?),toxic shock syndrome toxin-1(TSST-1)and Panton Valentine leukocidin(PVL)induced innate immune response,but SEO-induced host inflammatory response is still unclear.In additional,the increase in multi-drug resistant(MDR)strains of S.aureus is a growing concern.It has been reported that MRSA could be transmitted by feces.China is a major producer and consumer of pork.Once pig farms are contaminated by S.aureus,it will not only threaten human health but also cause significant economic losses.Based on,the purpose of this study was to investigate the prevalence and molecular characterization of pig farm S.aureus in Chongqing,and the host pro-inflammatory immune response against enterotoxins of S.aureus.1.Identification and antimicrobial resistance in S.aureus from pig farmsIn this study,swab samples were collected for isolation and identification of S.aureus from pig farms in six regions of Chongqing.A total of 724 swabs were collected from 12pig farms,68 isolates were S.aureus,57 of which were methicillin-resistant S.aureus(MRSA)based on the presence of the mec A gene.As for sampleing sources,the water,feed and air samples were seriously contaminated by S.aureus.The isolation rates were 13.6%(3/22),12.2%(5/41)and 30.0%(6/20),respectively.According to the different regions,the yongchuan(24%,12/50)and wanzhou(23.6%,41/174)were seriously contaminated by S.aureus.Subsequently,13 antimicrobial agents were used to test the antibiotic susceptibility of the 68 S.aureus isolates.It showed that high levels of resistance to antibiotics were found in these isolates.Resistance to penicillin,tetracycline,and erythromycin were observed in all 68 S.aureus isolates.Additionally,88.2%of these isolates were resistant to oxacillin,67.6%to chloramphenicol,22.1%to trimethoprim/sulphamethoxazole,and 10.3%to clindamycin.Low resistance rates to rifamycin(2.9%),gentamicin(1.5%),ciprofloxacin(1.5%),levofloxacin(1.5%),nitrofurantoin(0%),and teicoplanin(0%)were noted among the isolates.In conclusion,in some pig farm of Chongqing are contaminated to various degrees in the breeding processes.The resistance to antibiotics from pig farm-derived S.aureus isolates in Chongqing is constantly expanding.Therefore,it is necessary to strengthen the management of commonly used antibiotics in the clinic and advocate that cross-medication should be conducted.2.Prevalence and characterisation of integrons and enterotoxin genes in S.aureus isolates from pig farmsIntegrons are mobile DNA elements that allow for acquisition and dissemination of antibiotic resistance genes among clinical bacteria.However,limited information is available on integrons of S.aureus from pig farms.To investigate the prevalence of integrons and enterotoxin genes in multidrug resistant S.aureus isolates,the isolates from pig farm in Chongqing,China were examined for characterization of integrons and enterotoxin genes.All 68 isolates were carried integrons,of which 88.2%(60/68)harbored both class 1 and 2 integrons.In addition,85.3%(58/68)class 2 integron-positive isolates carried?-lactams gene(bla TEM-1),and 66.7%(40/60)class 1 integron-positive isolates carried aad A1c,aad A1 or dfr A1 gene.No class 3 integron was identified in the isolates tested.In addition,the enterotoxin genes of 68 pig farm derived S.aureus were detected by PCR.The results showed that 92.6%isolates(63/68)carried enterotoxin genes,of which the seg,seh,sem and seo genes with higher detection rates were 47.1%(32/68),82.4%(56/68),32.4%(22/68)and 54.4%(37/68),respectively.4.4%(3/68)isolates carried classic enterotoxin genes(sea-sed),and 92.6%isolates(63/68)carrying the new types of enterotoxin genes(seg-sep,ser).This is the first time to report the presence and characterization of class 1 and 2 integrons from pig farm-derived S.aureus isolates.Our results indicated that the integrons and enterotoxin genes are common among the pig farm-derived S.aureus isolates,and the strains mainly carrying new types of enterotoxin genes.For the significance of public health,the prevalence of integrons and their associated resistance genes in pig farm-derived S.aureus isolates should be paid adequate attention and surveillance.3.ATP facilitates Staphylococcal enterotoxin O induced neutrophil IL-1?secretion via NLRP3 inflammasome dependent pathwaysS.aureus is an important zoonotic food-borne pathogens causing severe invasive infections,such as sepsis,pneumonia,food poisoning,toxic shock syndrome,and autoimmune diseases.SEO is a new type of enterotoxins of S.aureus with superantigenic and emetic activity.It is still unclear about SEO-induced host inflammatory response.Therefore,the mechanism of SEO-induced IL-1?secretion in mice neutrophils was investigated in this study.Our results showed that recombinant SEO had superantigenic activity with high level of gamma interferon(IFN-?)production in mouse spleen cells and induced inflammatory cytokines expression including IL-1?,IL-1?,IL-6 and TNF-?in neutrophils under the action of ATP.In addition,SEO-induced IL-1?secretion was dependent on activation of Toll like receptor(TLR4),nuclear factor kappa B(NF-?B)and c-jun N-terminal kinase(JNK)signaling pathways.However,SEO-induced IL-1?secretion was abolished in neutrophils of NLRP3-/-mice compared with from WT mice,indicating that activation of NLRP3 inflammasome mediated IL-1?secretion during neutrophils stimulation with SEO under the action of ATP.Moreover,this process of SEO-induced IL-1?secretion was dependent on potassium(K+)efflux.Taken together,our study suggests that activation of TLR4/JNK/NLRP3 inflammasome signaling pathways mediates maturation and secretion of IL-1?and provides a new insight on S.aureus virulence factor-induced host immune response. |
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