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首页> 外文期刊>Poultry Science >Development of a quantitative real-time polymerase chain reaction assay to target a novel group of ammonia-producing bacteria found in poultry litter.
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Development of a quantitative real-time polymerase chain reaction assay to target a novel group of ammonia-producing bacteria found in poultry litter.

机译：实时定量聚合酶链反应测定法的开发，以靶向在家禽垫料中发现的一组新的产氨细菌。

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Ammonia production in poultry houses has serious implications for flock health and performance, nutrient value of poultry litter, and energy costs for running poultry operations. In poultry litter, the conversion of organic N (uric acid and urea) to NH(4)-N is a microbially mediated process. The urease enzyme is responsible for the final step in the conversion of urea to NH(4)-N. Cloning and analysis of 168 urease sequences from extracted genomic DNA from poultry litter samples revealed the presence of a novel, dominant group of ureolytic microbes (representing 90% of the urease clone library). Specific primers and a probe were designed to target this novel poultry litter urease producer (PLUP) group, and a new quantitative real-time PCR assay was developed. The assay allowed for the detection of 10(2) copies of target urease sequences per PCR reaction (approximately 1 x 10(4) cells per gram of poultry litter), and the reaction was linear over 8 orders of magnitude. Our PLUP group was present only in poultry litter and was not present in environmental samples from diverse agriculutural settings. This novel PLUP group represented between 0.1 to 3.1% of the total microbial populations (6.0 x 10(6) to 2.4 x 10(8) PLUP cells per gram of litter) from diverse poultry litter types. The PLUP cell concentrations were directly correlated to the total cell concentrations in the poultry litter and were found to be influenced by the physical parameters of the litters (bedding material, moisture content, pH), as well as the NH(4)-N content of the litters, based on principal component analysis. Chemical parameters (organic N, total N, total C) were not found to be influential in the concentrations of our PLUP group in the diverse poultry litters Future applications of this assay could include determining the efficacy of current NH(4)-N-reducing litter amendments or in designing more efficient treatment protocols.
机译：家禽舍中的氨生产对鸡群的健康和生产性能，家禽垫料的营养价值以及经营家禽的能源成本产生了严重影响。在家禽垫料中，有机氮（尿酸和尿素）向NH（4）-N的转化是微生物介导的过程。脲酶是尿素向NH（4）-N转化的最后一步。从家禽垫料样品中提取的基因组DNA的168个尿素酶序列的克隆和分析表明，存在一个新的占优势的尿素分解微生物群（占尿素酶克隆文库的90％）。设计特异性引物和探针来靶向该新型家禽垃圾脲酶生产者（PLUP）组，并开发了一种新的定量实时PCR分析方法。该测定法允许每个PCR反应检测10（2）份目标脲酶序列（每克家禽垃圾约1 x 10（4）个细胞），并且该反应在8个数量级上呈线性。我们的PLUP组仅出现在家禽垫料中，而没有出现在来自各种农业环境的环境样本中。这个新颖的PLUP组代表各种家禽垫料类型的总微生物种群（每克垫料6.0 x 10（6）到2.4 x 10（8）个PLUP细胞）的0.1％至3.1％。 PLUP细胞浓度与家禽垫料中的总细胞浓度直接相关，并且发现受垫料的物理参数（垫料，水分，pH）以及NH（4）-N含量的影响基于主成分分析的垃圾量。化学参数（有机氮，总氮，总碳）在不同家禽垫料中对我们的PLUP组的浓度没有影响，该测定法的未来应用可能包括确定当前减少NH（4）-N的功效乱抛垃圾或设计更有效的处理方案。

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