熟畜肉中金黄色葡萄球菌污染标记物动态变化及与菌体生长关联分析
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摘要
为考察不同类型的熟猪肉、牛肉、羊肉对金黄色葡萄球菌污染标记物的影响,采用顶空固相微萃取/气相色谱质谱联用技术对三种熟肉进行本底值检测,并对人工染菌的三种熟肉进行标记物动态分析和细菌生长数量变化测定。结果表明,在采集的大量样本中,三种熟肉基质都检出一定强度的3-甲基丁醛和3-甲基丁酸信号,作为熟畜肉的本底值。对于受金黄色葡萄球菌污染的三种熟肉,3-甲基丁醛信号均在第12 h时(1～3×10~7 CFU/g)出现显著增长(p<0.05),强度增长到最大值后迅速下降;3-甲基丁酸信号均在第14 h时(0.7～2×10~8CFU/g)出现显著增长(p<0.05),在整个培养过程中强度不断上升。对于三种熟肉,3-甲基丁醛(0～18 h)和3-甲基丁酸(0～24 h)的信号强度分别与金黄色葡萄球菌生长量之间均具有极显著(p<0.01)的相关性。结论:金黄色葡萄球菌生长量分别达到10~7、10~8 CFU/g时开始检测到3-甲基丁醛和3-甲基丁酸的增长,之后在一定的培养周期内这两个物质稳定存在并与菌体生长显著相关。研究结果为建立畜肉中金黄色葡萄球菌污染标记物检测技术奠定了前期基础。
To explore the influences of different meats on the volatile markers of Staphylococcus aureus contaminating on cooked meat,headspace-solid phase micro-extraction/gas chromatography-mass spectrometry(HS-SPME-GC/MS)was employed to measure the background values of three cooked meats(pork,beef and mutton)and production regularities of volatile markers of S. aureus growing on meats. The growth of S. aureus was determined by enumerating bacterial cells. The results showed that all of three cooked meat were detected both 3-methyl-butanal and 3-methyl-butanoic acid signals,so that the contamination of S. aureus on meat could be judged by the detection value obvious above meat background values. For S. aureus growing on these three cooked meats,the 3-methyl-butanal signal increased significantly(p<0.05)at the 12 th hour(corresponding to 1～3×10~7CFU/g),then it showed an increase up to the maximum with a sharp decrease. The 3-methyl-butanoic acid signal increased significantly(p<0.05)at the 14 th hour(corresponding to 0.7～2×10~8 CFU/g),then increased continuously during the whole incubation process. And the correlation between 3-methyl-butanal intensities and bacterial counts of S. aureus were extremly significant during 0～18 h,also the correlation between 3-methyl-butanoci acid intensities and bacterial counts of S. aureus were extremly significant during 0～24 h(p<0.01). Therefore,the increasing of 3-methyl-butanal and 3-methyl-butanoic acid was detected while the bacterial cells of S. aureus reached up to 10~7 and 10~8 CFU/g respectively. Subsequently,the two volatile markers were stable and significantly correlative with bacterial counts of S. aureus during a certain period of incubation. These results provided foundations for the establishment of the detection of S. aureus contaminating on meat based on volatile markers.
To explore the influences of different meats on the volatile markers of Staphylococcus aureus contaminating on cooked meat,headspace-solid phase micro-extraction/gas chromatography-mass spectrometry(HS-SPME-GC/MS)was employed to measure the background values of three cooked meats(pork,beef and mutton)and production regularities of volatile markers of S. aureus growing on meats. The growth of S. aureus was determined by enumerating bacterial cells. The results showed that all of three cooked meat were detected both 3-methyl-butanal and 3-methyl-butanoic acid signals,so that the contamination of S. aureus on meat could be judged by the detection value obvious above meat background values. For S. aureus growing on these three cooked meats,the 3-methyl-butanal signal increased significantly(p<0.05)at the 12 th hour(corresponding to 1～3×10~7CFU/g),then it showed an increase up to the maximum with a sharp decrease. The 3-methyl-butanoic acid signal increased significantly(p<0.05)at the 14 th hour(corresponding to 0.7～2×10~8 CFU/g),then increased continuously during the whole incubation process. And the correlation between 3-methyl-butanal intensities and bacterial counts of S. aureus were extremly significant during 0～18 h,also the correlation between 3-methyl-butanoci acid intensities and bacterial counts of S. aureus were extremly significant during 0～24 h(p<0.01). Therefore,the increasing of 3-methyl-butanal and 3-methyl-butanoic acid was detected while the bacterial cells of S. aureus reached up to 10~7 and 10~8 CFU/g respectively. Subsequently,the two volatile markers were stable and significantly correlative with bacterial counts of S. aureus during a certain period of incubation. These results provided foundations for the establishment of the detection of S. aureus contaminating on meat based on volatile markers.
引文
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[14]史辉,唐俊妮,陈娟,等.顶空固相微萃取分析金黄色葡萄球菌挥发性代谢产物的条件优化[J].食品科学,2015,36(12):185-190.
[15]徐薇薇,姚瑞基,袁维新,等.宁夏滩羊后腿肉营养评价及挥发性风味物质分析[J].肉类研究,2017,31(10):41-45.
[16]Takakura Y,Sakamoto T,Hirai S,et al. Characterization of the key aroma compounds in beef extract using aroma extract dilution analysis[J]. Meat Science,2014,97(1):27-31.
[17]Inagaki S,Amano Y,Kumazawa K. Identification and characterization of volatile components causing the characteristicflavor of Wagyu beef(Japanese black cattle)[J]. Journal of Agricultural and Food Chemistry,2017,65(39):8691-8695.
[18]Pulgar Del J S,Roldan M,Ruiz-Carrascal J. Volatile compounds profile of sous-vide cooked pork cheeks as affected by cooking conditions(vacuum packaging,temperature and time)[J].Molecules,2013,18(10):12538-12547.
[19]张迪雅,谢丹婷,李晔.应用电子鼻和GC-MS比较牛肉不同部位的挥发性物质组成[J].食品工业科技,2017,38(21):241-246.
[20]Hettinga K A,Van Valenberg H J,Lam T J,et al. The influence of incubation on the formation of volatile bacterial metabolites in mastitis milk[J]. Journal of Dairy Science,2009,92(10):4901-4905.
[21]Silcock P,Alothman M,Zardin E,et al. Microbially induced changes in the volatile constituents of fresh chilled pasteurized milk during storage[J]. Food Packaging and Shelf Life,2014,2(2):81-90.
[22]Hennekinne J A,De Buyser M L,Dragacci S. Staphylococcus aureus and its food poisoning toxins:characterization and outbreak investigation[J]. FEMS Microbiology Reviews,2012,36(4):815-836.
[23]侯巧娟,王锡昌,刘源,等.气味指纹技术在肉品品质及安全检测中的应用[J].食品科学,2011,32(7):327-331.
[24]金伟平,黄志强,刘群群,等.顶空固相微萃取-气质联用法分析单增李斯特菌污染冷藏牛肉的挥发性物质[J].食品科学,2012,33(2):243-247.
[25]Xu Y,Cheung W,Winder C L,et al. VOC-based metabolic profiling for food spoilage detection with the application to detecting Salmonella typhimurium contaminated pork[J]. Analytical and Bioanalytical Chemistry,2010,397(6):2439-2449.
[26]Xu Y,Cheung W,Winder C L,et al. Metabolic profiling of meat:Assessment of pork hygiene and contamination with Salmonella typhimurium[J]. Analyst,2011,136(3):508-514.
[2]鲁延迅,肖甜甜,宋军,等.凉菜与熟肉中金黄色葡萄球菌与大肠菌群的分离鉴定及污染分析[J].中国卫生检验杂志,2015,25(2):184-186.
[3]Young C P,O’Donoghue M M,Ho J,et al. High levels of Staphylococcus aureus contamination in Chinese style roast pork[J]. Foodborne Pathogens and Disease,2014,11(7):552-554.
[4]许振伟,韩奕奕,孟瑾,等.熟食肉制品中金黄色葡萄球菌风险评估基础研究[J].包装与食品机械,2012,30(5):40-43.
[5]Tait E,Perry J D,Stanforth S P,et al. Use of volatile compounds as a diagnostic tool for the detection of pathogenic bacteria[J]. Trends in Analytical Chemistry,2014,53:117-125.
[6]吴清平,李玉冬,张菊梅.常见食源性致病菌代谢组学研究进展[J].微生物学通报,2016,43(3):609-618.
[7]Lemfack M C,Gohlke B O,Toguem S M T,et al. A database of microbial volatiles[J]. Nucleic Acids Research,2018,46(1):D1261-D1265.
[8]陈娟,史辉,王琼,等.细菌挥发性代谢产物的类型、检测技术及应用[J].微生物学杂志,2015,35(1):89-94.
[9]Boots A W,Smolinska A,Van Berkel J J,et al. Identification of microorganisms based on headspace analysis of volatile organic compounds by gas chromatography-mass spectrometry[J]. Journal of Breath Research,2014,8(2):27106.
[10]Ratiu I A,Bocos-Bintintan V,Patrut A,et al. Discrimination of bacteria by rapid sensing their metabolic volatiles using an aspiration-type ion mobility spectrometer(a-IMS)and gas chromatography-mass spectrometry(GC-MS)[J]. Analytica Chimica Acta,2017,982:209-217.
[11]Filipiak W,Sponring A,Baur M M,et al. Molecular analysis of volatile metabolites released specifically by Staphylococcus aureus and Pseudomonas aeruginosa[J]. BMC Microbiology,2012,12(1):113.
[12]Bos L D,Sterk P J,Schultz M J. Volatile metabolites of pathogens:A systematic review[J]. Plos Pathogens,2013,9(5):210-216.
[13]Chen J,Tang J,Shi H,et al. Characteristics of volatile organic compounds produced from five pathogenic bacteria by headspace-solid phase micro-extraction-gas chromatography/mass spectrometry[J]. Journal of Basic Microbiology,2017,57(3):228-237.
[14]史辉,唐俊妮,陈娟,等.顶空固相微萃取分析金黄色葡萄球菌挥发性代谢产物的条件优化[J].食品科学,2015,36(12):185-190.
[15]徐薇薇,姚瑞基,袁维新,等.宁夏滩羊后腿肉营养评价及挥发性风味物质分析[J].肉类研究,2017,31(10):41-45.
[16]Takakura Y,Sakamoto T,Hirai S,et al. Characterization of the key aroma compounds in beef extract using aroma extract dilution analysis[J]. Meat Science,2014,97(1):27-31.
[17]Inagaki S,Amano Y,Kumazawa K. Identification and characterization of volatile components causing the characteristicflavor of Wagyu beef(Japanese black cattle)[J]. Journal of Agricultural and Food Chemistry,2017,65(39):8691-8695.
[18]Pulgar Del J S,Roldan M,Ruiz-Carrascal J. Volatile compounds profile of sous-vide cooked pork cheeks as affected by cooking conditions(vacuum packaging,temperature and time)[J].Molecules,2013,18(10):12538-12547.
[19]张迪雅,谢丹婷,李晔.应用电子鼻和GC-MS比较牛肉不同部位的挥发性物质组成[J].食品工业科技,2017,38(21):241-246.
[20]Hettinga K A,Van Valenberg H J,Lam T J,et al. The influence of incubation on the formation of volatile bacterial metabolites in mastitis milk[J]. Journal of Dairy Science,2009,92(10):4901-4905.
[21]Silcock P,Alothman M,Zardin E,et al. Microbially induced changes in the volatile constituents of fresh chilled pasteurized milk during storage[J]. Food Packaging and Shelf Life,2014,2(2):81-90.
[22]Hennekinne J A,De Buyser M L,Dragacci S. Staphylococcus aureus and its food poisoning toxins:characterization and outbreak investigation[J]. FEMS Microbiology Reviews,2012,36(4):815-836.
[23]侯巧娟,王锡昌,刘源,等.气味指纹技术在肉品品质及安全检测中的应用[J].食品科学,2011,32(7):327-331.
[24]金伟平,黄志强,刘群群,等.顶空固相微萃取-气质联用法分析单增李斯特菌污染冷藏牛肉的挥发性物质[J].食品科学,2012,33(2):243-247.
[25]Xu Y,Cheung W,Winder C L,et al. VOC-based metabolic profiling for food spoilage detection with the application to detecting Salmonella typhimurium contaminated pork[J]. Analytical and Bioanalytical Chemistry,2010,397(6):2439-2449.
[26]Xu Y,Cheung W,Winder C L,et al. Metabolic profiling of meat:Assessment of pork hygiene and contamination with Salmonella typhimurium[J]. Analyst,2011,136(3):508-514.