肉制品加工中脂肪对甲型副伤寒沙门氏菌rpoE的应激特性
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摘要
目的:为了解甲型副伤寒沙门氏菌缺陷株和野生株在肉制品加工过程不同介质中的热应激响应特征,并进一步探究肉制品加工中脂肪对甲型副伤寒沙门氏菌rpoE表达的影响。方法:以55,63,72℃对接种108CFU/g甲型副伤寒沙门氏菌缺陷株△rpoE和野生株在不同介质下进行热处理,测定处理后样品中甲型副伤寒沙门氏菌的残存数量,比较缺陷株和野生株在不同介质下的失活情况,对野生株和缺陷株在不同介质下的耐热情况进行分析,然后以Real-time PCR法检测野生株在近似巴氏杀菌条件下不同介质中热处理后的残存菌体样本的rpoE基因的表达水平,比较脂肪对rpoE表达水平的影响。结果:缺陷株和野生株D值之间存在显著性差异(P<0.05),缺陷株的D值都明显低于野生株,相同温度下缺陷株和野生株分别在肥瘦1∶1和纯瘦肉介质之间的D值都存在显著性差异(P<0.05),肥瘦介质中菌株更耐热,野生株在肥瘦1∶1和纯瘦肉介质的近似巴氏杀菌条件下,表达上调水平分别为3.53,3.29倍,且不存在显著性差异(P>0.05)。结论:肉制品在巴氏杀菌近似条件下,甲型副伤寒沙门氏菌通过提高rpoE的表达水平来增加甲型副伤寒沙门氏菌的耐热性。脂肪对菌株的耐热性起重要作用,并不会通过调节rpoE的表达来提高耐热性。
Objective: In order to understand the heat stress responses characteristics of Salmonella paratyphi A mutant and wild-type strain in different medium during the meat processing, and to further explore the effect of fat on the expression of rpoE gene of Salmonella paratyphi A in meat processing. Methods: In this study, the first, the vaccinated108 CFU/g Salmonella paratyphi A mutant strains and wild strains in different medium, then subjected to thermal treatment. The surviving cell number was counted on selective media, the inactivation of mutant strains and wild strains were compared and analyzed in meat products processing. The next real-time PCR method was used to detect the expression level of rpoE gene of wild strain under approximative conditions of meat products pasteurization in different medium. The effect of the fat on the expression level of rpoE was compared. Results: The results showed that D values of the wild and the mutant strains were a significant difference(P<0.05), and the D values of the mutant strains were significantly lower than the wild strains. Meanwhile, the D values of the mutant strains and wild-type strain has a significant difference(P<0.05) between the medium of sausages lean percentage 50% and 100%, bacterial strain in medium fat percentage 50% more heat resistant. The up-regulated expression level of rpoE gene in the medium of sausages lean percentage50% and 100% were 3.53 and 3.29 times respectively, and there was no significant difference(P >0.05). Conclusion:Salmonella paratyphi A increased the expression levels of rpoE to improve its thermal resistance. Fat plays an important role in the thermal resistance of the bacterial strains, it does not improve the thermal resistance by regulating rpoE expression level.
Objective: In order to understand the heat stress responses characteristics of Salmonella paratyphi A mutant and wild-type strain in different medium during the meat processing, and to further explore the effect of fat on the expression of rpoE gene of Salmonella paratyphi A in meat processing. Methods: In this study, the first, the vaccinated108 CFU/g Salmonella paratyphi A mutant strains and wild strains in different medium, then subjected to thermal treatment. The surviving cell number was counted on selective media, the inactivation of mutant strains and wild strains were compared and analyzed in meat products processing. The next real-time PCR method was used to detect the expression level of rpoE gene of wild strain under approximative conditions of meat products pasteurization in different medium. The effect of the fat on the expression level of rpoE was compared. Results: The results showed that D values of the wild and the mutant strains were a significant difference(P<0.05), and the D values of the mutant strains were significantly lower than the wild strains. Meanwhile, the D values of the mutant strains and wild-type strain has a significant difference(P<0.05) between the medium of sausages lean percentage 50% and 100%, bacterial strain in medium fat percentage 50% more heat resistant. The up-regulated expression level of rpoE gene in the medium of sausages lean percentage50% and 100% were 3.53 and 3.29 times respectively, and there was no significant difference(P >0.05). Conclusion:Salmonella paratyphi A increased the expression levels of rpoE to improve its thermal resistance. Fat plays an important role in the thermal resistance of the bacterial strains, it does not improve the thermal resistance by regulating rpoE expression level.
引文
[1]CHAU T T,CAMPBELL J I,GALINDO C M,et al.Antimicrobial drug resistance of Salmonella enterica serovar typhi in asia and molecular mechanism of reduced susceptibility to the fluoroquinolones[J].Antimicrobial Agents&Chemotherapy,2008,51(12):4315-4323.
[2]HAN G,CHEN N,MA Z.Assessment of the uncertainty in thermal food processing decisions based on microbial safety objectives[J].Journal of Food Engineering,2011,102(3):247-256.
[3]HEIR E,HOLCK A L,OMER M K,et al.Effects of post-processing treatments on sensory quality and Shiga toxigenic Escherichia coli,reductions in dryfermented sausages[J].Meat Science,2013,94(1):47-54.
[4]DUCIC M,KLISARA N,MARKOV S,et al.The fate and pasteurization-based inactivation of Escherichia coli O157,Salmonella,Typhimurium and Listeria monocytogenes,in dry,fermented sausages[J].Food Control,2016,59:400-406.
[5]BUSBY S,EBRIGHT R H.Promoter structure,promoter recognition,and transcription activation in prokaryotes[J].Cell,1994,79(5):743-746.
[6]SPECTOR M P,KENYON W J.Resistance and survival strategies of Salmonella enterica,to environmental stresses[J].Food Research International,2012,45(2):455-481.
[7]NONAKA G,BLANKSCHIEN M,HERMAN C,et al.Regulon and promoter analysis of the E.coli heat-shock factor,σ32,reveals a multifaceted cellular response to heat stress[J].Genes&Development,2006,20(13):1776-1789.
[8]GUISBERT E,YURA T,RHODIUS V A,et al Convergence of molecular,modeling,and systems approaches for an understanding of the Escherichia coli heat shock response[J].Microbiology&Molecular Biology Reviews Mmbr,2008,72(3):545-554.
[9]RICHTER K,HASLBECK M,BUCHNER J.The heat shock response:Life on the verge of death[J].Molecular Cell,2010,40(2):253-266.
[10]YANG Y,WEI J K,ZHENG Q,et al.Growth temperature alters Salmonella,Enteritidis heat/acid resistance,membrane lipid composition and stress/virulence related gene expression[J].International Journal of Food Microbiology,2013,172C(1):102-109.
[11]ALISDAIR M M,MARK R,COGAN T A,et al Role of the alternative sigma factors sigmaE and sigmaS in survival of Salmonella enterica serovar Typhimurium during starvation,refrigeration and os motic shock[J].Microbiology,2007,153(Pt 1):263-269.
[12]KENYON W J,SAYERS D G,HUMPHREYS S,et al.The starvation-stress response of Salmonella enterica serovar Typhimurium requires sigma(E)-,but not CpxR-regulated extracytoplasmic functions[J].Microbiology,2002,148(Pt 1):113-122.
[13]THONGBOONKERD V,VANAPORN M,SONGTAWEEN,et al.Altered Proteome in Burkholderia pseudomallei rpoE operon knockout mutant:Insights into mechanisms of rpoE operon in stress tolerance,survival,and virulence[J].Journal of Proteome Re search,2007,6(4):1334-1341.
[14]TESTERMAN T L,VAZQUEZTORRES A,XU Y,et al.The alternative sigma factor sigmaE controls antioxidant defences required for Salmonella virulence and stationary-phase survival[J].Molecular Microbiology,2002,43(3):771-782.
[15]BANG I S,FRYE J G,MCCLELLAND M,et al Alternative sigma factor interactions in Salmonella:Sigma and sigma promote antioxidant defences by enhancing sigma levels[J].Molecular Microbiology,2005,56(3):811-823.
[16]HANSEN N H,RIEMANN H.Factors affecting the heat resistance of nonsporing organisms[J].Journal of Applied Microbiology,1963,26(3):314-333.
[17]AHMED N M,CONNER D E,HUFFMAN D LHeat-resistance of Escherichia coli O157:H7 in meat and poultry as affected by product composition[J].Journal of Food Science,1995,60(3):606-610.
[18]VEERAMUTHU G J,PRICE J F,DAVIS C E,et al.Thermal inactivation of Escherichia coli O157:H7,Salmonella senftenberg,and enzymes with potential as time-temperature indicators in ground turkey thigh meat[J].Journal of Food Protection,1998,61(2):171-175.
[19]HEIN I,FLEKNA G,KRASSNIG M,et al.Realtime PCR for the detection of Salmonella spp.in food:An alternative approach to a conventional PCR system suggested by the FOOD-PCR project[J].Journal of Microbiological Methods,2006,66(3):538-547.
[20]MUJAHID S,PECHAN T,WANG C.Protein expression by Listeria monocytogenes grown on a RTE-meat matrix[J].International Journal of Food Microbiology,2008,128(2):203-211.
[2]HAN G,CHEN N,MA Z.Assessment of the uncertainty in thermal food processing decisions based on microbial safety objectives[J].Journal of Food Engineering,2011,102(3):247-256.
[3]HEIR E,HOLCK A L,OMER M K,et al.Effects of post-processing treatments on sensory quality and Shiga toxigenic Escherichia coli,reductions in dryfermented sausages[J].Meat Science,2013,94(1):47-54.
[4]DUCIC M,KLISARA N,MARKOV S,et al.The fate and pasteurization-based inactivation of Escherichia coli O157,Salmonella,Typhimurium and Listeria monocytogenes,in dry,fermented sausages[J].Food Control,2016,59:400-406.
[5]BUSBY S,EBRIGHT R H.Promoter structure,promoter recognition,and transcription activation in prokaryotes[J].Cell,1994,79(5):743-746.
[6]SPECTOR M P,KENYON W J.Resistance and survival strategies of Salmonella enterica,to environmental stresses[J].Food Research International,2012,45(2):455-481.
[7]NONAKA G,BLANKSCHIEN M,HERMAN C,et al.Regulon and promoter analysis of the E.coli heat-shock factor,σ32,reveals a multifaceted cellular response to heat stress[J].Genes&Development,2006,20(13):1776-1789.
[8]GUISBERT E,YURA T,RHODIUS V A,et al Convergence of molecular,modeling,and systems approaches for an understanding of the Escherichia coli heat shock response[J].Microbiology&Molecular Biology Reviews Mmbr,2008,72(3):545-554.
[9]RICHTER K,HASLBECK M,BUCHNER J.The heat shock response:Life on the verge of death[J].Molecular Cell,2010,40(2):253-266.
[10]YANG Y,WEI J K,ZHENG Q,et al.Growth temperature alters Salmonella,Enteritidis heat/acid resistance,membrane lipid composition and stress/virulence related gene expression[J].International Journal of Food Microbiology,2013,172C(1):102-109.
[11]ALISDAIR M M,MARK R,COGAN T A,et al Role of the alternative sigma factors sigmaE and sigmaS in survival of Salmonella enterica serovar Typhimurium during starvation,refrigeration and os motic shock[J].Microbiology,2007,153(Pt 1):263-269.
[12]KENYON W J,SAYERS D G,HUMPHREYS S,et al.The starvation-stress response of Salmonella enterica serovar Typhimurium requires sigma(E)-,but not CpxR-regulated extracytoplasmic functions[J].Microbiology,2002,148(Pt 1):113-122.
[13]THONGBOONKERD V,VANAPORN M,SONGTAWEEN,et al.Altered Proteome in Burkholderia pseudomallei rpoE operon knockout mutant:Insights into mechanisms of rpoE operon in stress tolerance,survival,and virulence[J].Journal of Proteome Re search,2007,6(4):1334-1341.
[14]TESTERMAN T L,VAZQUEZTORRES A,XU Y,et al.The alternative sigma factor sigmaE controls antioxidant defences required for Salmonella virulence and stationary-phase survival[J].Molecular Microbiology,2002,43(3):771-782.
[15]BANG I S,FRYE J G,MCCLELLAND M,et al Alternative sigma factor interactions in Salmonella:Sigma and sigma promote antioxidant defences by enhancing sigma levels[J].Molecular Microbiology,2005,56(3):811-823.
[16]HANSEN N H,RIEMANN H.Factors affecting the heat resistance of nonsporing organisms[J].Journal of Applied Microbiology,1963,26(3):314-333.
[17]AHMED N M,CONNER D E,HUFFMAN D LHeat-resistance of Escherichia coli O157:H7 in meat and poultry as affected by product composition[J].Journal of Food Science,1995,60(3):606-610.
[18]VEERAMUTHU G J,PRICE J F,DAVIS C E,et al.Thermal inactivation of Escherichia coli O157:H7,Salmonella senftenberg,and enzymes with potential as time-temperature indicators in ground turkey thigh meat[J].Journal of Food Protection,1998,61(2):171-175.
[19]HEIN I,FLEKNA G,KRASSNIG M,et al.Realtime PCR for the detection of Salmonella spp.in food:An alternative approach to a conventional PCR system suggested by the FOOD-PCR project[J].Journal of Microbiological Methods,2006,66(3):538-547.
[20]MUJAHID S,PECHAN T,WANG C.Protein expression by Listeria monocytogenes grown on a RTE-meat matrix[J].International Journal of Food Microbiology,2008,128(2):203-211.