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猪粪便和粪坑中细菌的分离、特性鉴定及比较

Isolation, characterization and comparison of bacteria from swine faeces and manure storage pits.

作者信息

Cotta Michael A, Whitehead Terence R, Zeltwanger Rhonda L

机构信息

Fermentation Biotechnology Research Unit, National Center for Agricultural Utilization Research, USDA, Agricultural Research Service, Peoria, IL 61604, USA.

出版信息

Environ Microbiol. 2003 Sep;5(9):737-45. doi: 10.1046/j.1467-2920.2003.00467.x.

Abstract

Storage of swine manure is associated with the microbiological production of a variety of odorous chemicals including ammonia, organic acids and alcohols, and sulphides. Although largely the product of microbiological activity, little is known about the microorganisms present in swine manure. In order to gain a better understanding of the types and activities of the microorganisms present, representative strains of microorganisms were isolated from faeces and stored manure slurry, identified, and physiologically characterized. For swine manure slurry samples, total anaerobe colony counts were greatest when a non-selective, habitat simulating medium containing clarified swine manure slurry was used whereas the highest counts for faecal anaerobes were obtained on rumen fluid containing medium. Faecal and slurry samples were also plated onto the appropriate medium containing the antibiotics tetracycline, erythromycin and tylosin (10 micro g ml-1, individually) and the proportional counts of organisms capable of growing in the presence of these antibiotics determined. Randomly selected isolates from the highest dilutions were identified by 16 s rDNA sequence analysis, and selected physiological characteristics were determined. The results of these examinations indicate that the predominant culturable microorganisms from these environments are obligately anaerobic, low mol percentage G + C Gram positive bacteria (Firmicutes) who are members of Clostridial, Eubacterial, and Lactobacillus/Streptococcus phylogenetic groups. Isolates similar to Sporomusa and Flexibacter/Cytophaga/Bacteroides (CFB or Bacteroidetes) groups were also obtained. Although similar overall, faecal and slurry samples differed in bacterial composition. Manure slurry samples were dominated by organisms similar to Clostridium coccoides and Enterococcus species whereas the distribution of species present in faeces appeared much broader. Whereas most of the pure cultures could be assigned to known phylogenetic groupings, few could be identified as known species. Examination of some growth and physiological characteristics of faecal and slurry isolates showed these to be primarily carbohydrate fermenters, although some were able to ferment lactate and amino acids. When the ability of manure and faecal isolates to ferment protein, peptides and amino acids was examined, a relatively small percentage of these were able to do so and most of these fermented carbohydrates in addition to the amino acid sources provided. The predominant amino acid fermenters were most closely related to C. coccoides and C. botulinum, but representatives of the Bacteroides, Staphylococcus, Enterococcus and other phylogenetic groups were also found. The results reported here are compared with those obtained from clone libraries prepared from the same environmental samples.

摘要

猪粪的储存与多种有气味化学物质的微生物产生有关,这些化学物质包括氨、有机酸、醇类和硫化物。尽管这些物质很大程度上是微生物活动的产物,但对于猪粪中存在的微生物却知之甚少。为了更好地了解其中存在的微生物类型和活性,从粪便和储存的粪浆中分离出具有代表性的微生物菌株,进行鉴定并对其生理特性进行表征。对于猪粪浆样品,当使用含有澄清猪粪浆的非选择性、模拟生境培养基时,总厌氧菌菌落计数最高,而粪便厌氧菌的最高计数是在含有瘤胃液的培养基上获得的。粪便和粪浆样品也被接种到含有抗生素四环素、红霉素和泰乐菌素(各10微克/毫升)的合适培养基上,并测定能够在这些抗生素存在下生长的微生物的比例计数。通过16 s rDNA序列分析对从最高稀释度中随机选择的分离株进行鉴定,并测定选定的生理特性。这些检测结果表明,来自这些环境的主要可培养微生物是专性厌氧、低摩尔百分比G + C革兰氏阳性菌(厚壁菌门),它们是梭菌、真细菌和乳酸杆菌/链球菌系统发育组的成员。还获得了与芽孢八叠球菌和屈挠杆菌/噬细胞菌/拟杆菌(CFB或拟杆菌门)组相似的分离株。尽管总体相似,但粪便和粪浆样品的细菌组成有所不同。粪浆样品以类似于球形梭菌和肠球菌属的微生物为主,而粪便中存在的物种分布似乎更为广泛。虽然大多数纯培养物可以归入已知的系统发育分组,但很少能鉴定为已知物种。对粪便和粪浆分离株的一些生长和生理特性进行检测表明,它们主要是碳水化合物发酵菌,不过有些能够发酵乳酸和氨基酸。当检测粪肥和粪便分离株发酵蛋白质、肽和氨基酸的能力时,能够这样做的比例相对较小,而且其中大多数除了利用提供的氨基酸来源外,还能发酵碳水化合物。主要的氨基酸发酵菌与球形梭菌和肉毒梭菌关系最为密切,但也发现了拟杆菌、葡萄球菌、肠球菌和其他系统发育组的代表。本文报道的结果与从相同环境样品制备的克隆文库中获得的结果进行了比较。

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