Poultry Microbiological Safety Research Unit, R.B. Russell Agricultural Research Center, Agricultural Research Service, USDA, 950 College Station Road, Athens, GA 30605, USA.
Poult Sci. 2013 Feb;92(2):526-33. doi: 10.3382/ps.2012-02708.
There has been a resurgent interest in the use of bacteriophages or their gene products to control bacterial pathogens as alternatives to currently used antibiotics. Clostridium perfringens is a gram-positive, spore-forming anaerobic bacterium that plays a significant role in human foodborne disease as well as non-foodborne human, animal, and avian diseases. Countries that have complied with the ban on antimicrobial growth promoters in feeds have reported increased incidences of C. perfringens-associated diseases in poultry. To address these issues, new antimicrobial agents, putative lysins encoded by the genomes of bacteriophages, are being identified in our laboratory. Poultry intestinal material, soil, sewage, and poultry processing drainage water were screened for virulent bacteriophages that could lyse C. perfringens and produce clear plaques in spot assays. Bacteriophages were isolated that had long noncontractile tails, members of the family Siphoviridae, and with short noncontractile tails, members of the family Podoviridae. Several bacteriophage genes were identified that encoded N-acetylmuramoyl-l-alanine amidases, lysozyme-endopeptidases, and a zinc carboxypeptidase domain that has not been previously reported in viral genomes. Putative phage lysin genes (ply) were cloned and expressed in Escherichia coli. The recombinant lysins were amidases capable of lysing both parental phage host strains of C. perfringens as well as other strains of the bacterium in spot and turbidity reduction assays, but did not lyse any clostridia beyond the species. Consequently, bacteriophage gene products could eventually be used to target bacterial pathogens, such as C. perfringens via a species-specific strategy, to control animal and human diseases without having deleterious effects on beneficial probiotic bacteria.
人们对利用噬菌体或其基因产物来控制细菌病原体重新产生了兴趣，将其作为目前使用的抗生素的替代品。产气荚膜梭菌是一种革兰氏阳性、形成孢子的厌氧细菌，在人类食源性疾病以及非食源性人类、动物和禽类疾病中起着重要作用。那些遵守饲料中禁止使用抗菌生长促进剂的国家报告称，家禽中与产气荚膜梭菌相关的疾病发病率有所增加。为了解决这些问题，我们实验室正在鉴定新的抗菌剂，即噬菌体基因组编码的潜在溶菌酶。对家禽肠道材料、土壤、污水和家禽加工排水进行了筛选，以寻找能够裂解产气荚膜梭菌并在点分析中产生清晰斑块的烈性噬菌体。分离出具有长非收缩性尾巴的噬菌体，属于肌尾噬菌体科，以及具有短非收缩性尾巴的噬菌体，属于短尾噬菌体科。鉴定出几种噬菌体基因，这些基因编码 N-乙酰基胞壁酰-L-丙氨酸酰胺酶、溶菌酶内肽酶和锌羧肽酶结构域，这些结构域以前在病毒基因组中没有报道过。克隆和表达了推定的噬菌体溶菌酶基因 (ply) 在大肠杆菌中。重组溶菌酶是酰胺酶，能够裂解产气荚膜梭菌的两个亲本噬菌体宿主菌株以及斑点和浊度降低测定中的其他菌株，但不能裂解除该菌以外的任何梭菌。因此，噬菌体基因产物最终可以通过一种特定于物种的策略用于靶向细菌病原体，如产气荚膜梭菌，以控制动物和人类疾病，而不会对有益的益生菌产生有害影响。
