Regulation Of RpoS Factor On The Growth Of Salmonella Typhimurium Under Environmental Stress

Salmonella Typhimurium,food-borne pathogen,is one of the main pathogens causing acute gastroenteritis.It is widely distributed in the natural world and has strong resistance to the external environment.The contamination of Salmonella has brought great economic loss to the food industry.Salmonella is able to adapt to the bad environment by its own regulation mechanism where RpoS plays a very important role.To study the role of RpoS on Salmonella Typhimurium during stationary phase under osmotic stress,the SL1344 and SL1344/(35)rpoS were treated in different environmental stresses to identify the role of RpoS in environmental stresses,and then selected the Salmonella under selective osmotic stress to further study the regulation mechanism of RpoS under osmotic stress by using RNA-seq.The results were as follows:1)SL1344 and SL1344/(35)rpoS were cultured in a single environment stress,the inhibitory effect was the strongest in 3.5% NaCl,the pH 4.0 was the second,and the 42℃ was the weakest.2)SL1344 and SL1344/(35)rpoS were stimulated in different concentrations of NaCl for different times,and then cultured in 3.5% NaCl.The concentration order of SL1344 in logarithmic phase at the same time when stimulated in different concentrations of NaCl for the same time was 5%(27)3.5%(27)0%,and SL1344 could grow in 3.5% NaCl after stimulation in different salt concentrations for different times;but the bacteria concentrations of SL1344/(35)rpoS stimulated in different salt concentrations for the same time was different from that of SL1344.When stimulated in 5%NaCl for different times,the lag phases of SL1344/(35)rpoS were much longer than that of SL1344,even unable to grow in the 5% NaCl.The growth trends of SL1344/(35)rpoS in 0% and 3.5% NaCl were similar,which is 60min(27)30min(27)3h.The results showed that RpoS played an important role in regulating osmotic stress with Salmonella Typhimurium.3)In order to study the regulation role of RpoS on Salmonella Typhimurium with pretreatment of pH 4.0 on Salmonella under osmotic stress,SL1344 and SL1344/(35)rpoS were cultured in 3.5% NaCl with or without pretreatment of pH4.0,and RNA-seq was used to identify different genes expressed under osmotic stress.According to the results of RNA-seq,genes related osmotic regulation of Salmonella Typhimurium were regulated by RpoS under osmotic stress,and genes negatively regulated were more than positively.Those genes were classified as carbohydrate metabolism(katE,otsA,otsB,glgC,glgX,sgrT,celA,etc.),amino acid metabolism(asnA,metA,metH,aroQ,etc.),transmembrane transport(sfbA,sfbB,yehZ,ugpB,artJ,chaA,etc.),transcription and regulation(yeaG,marB,marA,marR,yaiB,hilD,hilC,etc.),energy metabolism(arsA,arsB,arsC,osmC,etc.),and so on.The differential expressed genes could enhance the resistance to osmotic pressure of Salmonella Typhimurium.We also compared the growths of SL1344 and SL1344/(35)rpoS with or without the pH 4.0 pretreatment.The lag phase of SL1344/(35)rpoS was longer than that of the control,but there was only small difference showed in the results of RNA-seq.This result suggested that the effect of pH 4.0 pretreatment may be only effect the lag phase,can not last into the stationary phase.4)The differential RpoS-dependent genes involved in the main metabolic and regulatory processes under the osmotic pressure were selected for qPCR verification.The results were consistent with the expression trend of the differential genes in the results of RNA-seq,but the difference multiplier was not consistent,indicating that the results of RNA-seq was effective.The study of Salmonella Typhimurium under environmental stress and RNA-seq under osmotic stress laid the foundation for further study of the regulation mechanism of Salmonella under environmental stress.The results in this study could be used for prevention and control of Salmonella Typhimurium contamination in foods.