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首页> 外文期刊>Applied Microbiology >Rapid Quantification of Viable Campylobacter Bacteria on Chicken Carcasses, Using Real-Time PCR and Propidium Monoazide Treatment, as a Tool for Quantitative Risk Assessment
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Rapid Quantification of Viable Campylobacter Bacteria on Chicken Carcasses, Using Real-Time PCR and Propidium Monoazide Treatment, as a Tool for Quantitative Risk Assessment

机译：实时PCR和单叠氮化丙锭处理技术作为定量风险评估的工具，可快速定量鸡尸体上的活弯曲菌

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A number of intervention strategies against Campylobacter -contaminated poultry focus on postslaughter reduction of the number of cells, emphasizing the need for rapid and reliable quantitative detection of only viable Campylobacter bacteria. We present a new and rapid quantitative approach to the enumeration of food-borne Campylobacter bacteria that combines real-time quantitative PCR (Q-PCR) with simple propidium monoazide (PMA) sample treatment. In less than 3 h, this method generates a signal from only viable and viable but nonculturable (VBNC) Campylobacter bacteria with an intact membrane. The method's performance was evaluated by assessing the contributions to variability by individual chicken carcass rinse matrices, species of Campylobacter , and differences in efficiency of DNA extraction with differing cell inputs. The method was compared with culture-based enumeration on 50 naturally infected chickens. The cell contents correlated with cycle threshold ( C_(T) ) values ( R ~(2) = 0.993), with a quantification range of 1 × 10~(2) to 1 × 10~(7) CFU/ml. The correlation between the Campylobacter counts obtained by PMA-PCR and culture on naturally contaminated chickens was high ( R ~(2) = 0.844). The amplification efficiency of the Q-PCR method was not affected by the chicken rinse matrix or by the species of Campylobacter . No Q-PCR signals were obtained from artificially inoculated chicken rinse when PMA sample treatment was applied. In conclusion, this study presents a rapid tool for producing reliable quantitative data on viable Campylobacter bacteria in chicken carcass rinse. The proposed method does not detect DNA from dead Campylobacter bacteria but recognizes the infectious potential of the VBNC state and is thereby able to assess the effect of control strategies and provide trustworthy data for risk assessment.
机译：针对弯曲杆菌感染的家禽的许多干预策略都集中在屠宰后减少细胞数量，强调仅对快速存活的弯曲杆菌细菌进行快速，可靠的定量检测的必要性。我们提出了一种新的，快速定量的食源弯曲杆菌细菌计数方法，该方法将实时定量PCR（Q-PCR）与简单的单叠氮化丙锭（PMA）样品处理相结合。在不到3小时内，此方法仅从具有完整膜的存活和存活但不可培养的弯曲杆菌（VBNC）产生信号。通过评估各个鸡car体漂洗基质对可变性的贡献，弯曲杆菌的种类以及不同细胞输入下DNA提取效率的差异来评估该方法的性能。将该方法与基于培养的枚举对50只自然感染的鸡进行了比较。细胞含量与循环阈值（C_（T））值（R〜（2）= 0.993）相关，定量范围为1×10〜（2）至1×10〜（7）CFU / ml。通过PMA-PCR获得的弯曲杆菌计数与自然污染鸡的培养之间的相关性很高（R〜（2）= 0.844）。 Q-PCR方法的扩增效率不受鸡肉漂洗基质或弯曲杆菌属物种的影响。当应用PMA样品处理时，从人工接种的鸡漂洗液中未获得Q-PCR信号。总之，这项研究提供了一种快速的工具，可用于产生关于鸡car体冲洗液中活弯曲菌的可靠定量数据。所提出的方法不能检测到死去的弯曲杆菌细菌的DNA，但是可以识别VBNC状态的感染潜力，因此能够评估控制策略的效果并为风险评估提供值得信赖的数据。

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