Paenibacillus is a spore-forming bacterial genus that is frequently isolated from fluid milk and is proposed to play a role in spoilage. To characterize the genetic and phenotypic diversity of Paenibacillus spp., we first used rpoB allelic typing data for a preexisting collection of 1,228 Paenibacillus species isolates collected from raw and processed milk, milk products, and dairy environmental sources. Whole-genome sequencing (WGS) and average nucleotide identity by BLAST (ANIb) analyses performed for a subset of 58 isolates representing unique and overrepresented rpoB allelic types in the collection revealed that these isolates represent 21 different Paenibacillus spp., with P. odorifer being the predominant species. Further genomic characterization of P. odorifer isolates identified two distinct phylogenetic clades, clades A and B, which showed significant overrepresentation of 172 and 164 ortholog clusters and 94 and 52 gene ontology (GO) terms, respectively. While nitrogen fixation genes were found in both clades, multiple genes associated with nitrate and nitrite reduction were overrepresented in clade A isolates; additional phenotypic testing demonstrated that nitrate reduction is specific to isolates in clade A. Hidden Markov models detected 9 to 10 different classes of cold shock-associated genetic elements in all P. odorifer isolates. Phenotypic testing revealed that all isolates tested here can grow in skim milk broth at 6°C, suggesting that psychrotolerance is conserved in P. odorifer Overall, our data suggest that Paenibacillus spp. isolated from milk in the United States represent broad genetic diversity, which may provide challenges for targeted-control strategies aimed at reducing fluid milk spoilage.IMPORTANCE Although Paenibacillus species isolates are frequently isolated from pasteurized fluid milk, the link between the genetic diversity and phenotypic characteristics of these isolates was not well understood, especially as some Bacillales isolated from milk are unable to grow at refrigeration temperatures. Our data demonstrate that Paenibacillus spp. isolated from fluid milk represent tremendous interspecies diversity, with P. odorifer being the predominant Paenibacillus sp. isolated. Furthermore, genetic and phenotypic data support that P. odorifer is well suited to transition from a soil-dwelling environment, where nitrogen fixation (and other nitrate/nitrite reduction pathways present only in clade A) may facilitate growth, to fluid milk, where its multiple cold shock-associated adaptations enable it to grow at refrigeration temperatures throughout the storage of milk. Therefore, efforts to reduce bacterial contamination of milk will require a systematic approach to reduce P. odorifer contamination of raw milk.

中文翻译：

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香菇（Paenibacillus odorifer）是在美国从牛奶中分离出的主要香菇物种，它显示了耐心理的遗传和表型保守性，但是进化枝相关的氮代谢途径差异。

芽孢杆菌是一种形成孢子的细菌属，通常从液态奶中分离出来，并被认为在腐败中起作用。为了表征Paenibacillus spp。的遗传和表型多样性，我们首先使用rpoB等位基因分型数据收集了从生乳和加工乳，奶制品和乳制品环境来源收集的1,228个Paenibacillus菌种的现有集合。对58个分离株的子集进行全基因组测序（WGS）和通过BLAST（ANIb）进行的平均核苷酸同一性分析，这些分离株代表了该集合中独特和过度表达的rpoB等位基因类型，显示这些分离株代表21种不同的Paenibacillus spp。，其中有P. odorifer主要物种。P. odorifer分离株的进一步基因组学鉴定鉴定了两个不同的系统发育进化枝，进化枝A和B，分别显示了172和164个直系同源簇以及94和52个基因本体论（GO）词的显着过表达。尽管在两个进化枝中都发现了固氮基因，但是在进化枝A分离物中，与硝酸盐和亚硝酸盐还原相关的多个基因却过分表达。额外的表型测试表明，硝酸盐的还原对进化枝A中的分离株具有特异性。隐马尔可夫模型在所有P. odorifer分离株中检测到9至10种与冷休克相关的遗传元素。表型测试表明，此处测试的所有分离株均可以在6°C的脱脂牛奶肉汤中生长，这表明香菇中的耐精神病菌是保守的。总体而言，我们的数据表明，Paenibacillus spp。在美国，从牛奶中分离出来代表广泛的遗传多样性，重要提示：尽管经常从巴氏灭菌的液态奶中分离出Paenibacillus菌种，但对这些分离株的遗传多样性和表型特征之间的联系仍知之甚少，尤其是一些芽孢杆菌。从牛奶中分离出来的物质在冷藏温度下无法生长。我们的数据表明，Paenibacillus spp。从流质奶中分离出代表巨大的种间多样性，其中P. odorifer是主要的芽孢杆菌。孤立。此外，遗传学和表型数据也证明，体育香菇非常适合从土壤居住环境过渡，在土壤环境中，固氮（以及仅在枝条A中存在的其他硝酸盐/亚硝酸盐还原途径）可以促进向液态奶的生长，多种与冷冲击相关的适应措施使它在整个牛奶存储过程中都能在冷藏温度下生长。因此，为减少牛奶中的细菌污染而做出的努力将需要一种系统的方法，以减少原奶中的臭味假单胞菌污染。