Susceptibility of Escherichia coli isolated from uteri of postpartum dairy cows to antibiotic and environmental bacteriophages. Part I: Isolation and lytic activity estimation of bacteriophages
Susceptibility of Escherichia coli isolated from uteri of postpartum dairy cows to antibiotic and environmental bacteriophages. Part I: Isolation and lytic activity estimation of bacteriophages
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DOI：
10.3168/jds.2009-2298
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发表时间：
2010-01-01
影响因子：
3.5
通讯作者：
Machado, V. S.
中科院分区：
文献类型：
--
作者：
Bicalho, R. C.;Santos, T. M. A.;Machado, V. S.
本研究以产后荷斯坦奶牛子宫内分离的大肠杆菌为宿主，从2个大型商业奶牛场的环境样品中分离噬菌体。从商业奶牛场的粪便系统中分离出11种噬菌体制剂，并对其体外抗菌活性进行了表征。此外，对5头奶牛分离的57株子宫大肠杆菌进行了系统进化分组。在不同的感染多重度（multiple of infections， MOI）下，用不同的噬菌体制剂接种来自子宫的每个大肠杆菌宿主，以确定最小抑制MOI。采用微孔板技术评估单剂量（MOI = 10(2)）噬菌体对所有57株大肠杆菌分离株生长曲线的影响。此外，通过噬菌体纯化、DNA提取、酶切和琼脂糖凝胶电泳来评估不同噬菌体制剂内部和之间的遗传多样性。基于三重PCR的系统进化分组表明，所有分离株均属于系统群B1。在相当低的MOI条件下，细菌的生长被完全抑制，单剂量（MOI = 102）噬菌体制剂对57株大肠杆菌的生长曲线的影响表明，所有噬菌体制剂都显著降低了分离株的生长速度。噬菌体制剂1230-10的抑菌活性最高，能完全抑制71.7% （n = 57）的菌株的生长。在MOI = 102时，1230-10、6375-10、2540-4和6547-2噬菌体制剂的联合作用谱最广，在600 nm处完全抑制了噬菌体的生长（最终光密度）
The objective of this study was to isolate bacteriophages from environmental samples of 2 large commercial dairy farms using Escherichia coli isolated from the uteri of postpartum Holstein dairy cows as hosts. A total of 11 bacteriophage preparations were isolated from manure systems of commercial dairy farms and characterized for in vitro antimicrobial activity. In addition, a total of 57 E. coli uterine isolates from 5 dairy cows were phylogenetically grouped by triplex PCR. Each E. coli bacterial host from the uterus was inoculated with their respective bacteriophage preparation at several different multiplicities of infections (MOI) to determine minimum inhibitory MOI. The effect of a single dose (MOI = 10(2)) of bacteriophage on the growth curve of all 57 E. coli isolates was assessed using a microplate technique. Furthermore, genetic diversity within and between the different bacteriophage preparations was assessed by bacteriophage purification followed by DNA extraction, restriction, and agarose gel electrophoresis. Phylogenetic grouping based on triplex PCR showed that all isolates of E. coli belonged to phylogroup B1. Bacterial growth was completely inhibited at considerably low MOI, and the effect of a single dose (MOI = 102) of bacteriophage preparations on the growth curve of all 57 E. coli isolates showed that all bacteriophage preparations significantly decreased the growth rate of the isolates. Bacteriophage preparation 1230-10 had the greatest antimicrobial activity and completely inhibited the growth of 71.7% (n = 57) of the isolates. The combined action of bacteriophage preparations 1230-10, 6375-10, 2540-4, and 6547-2, each at MOI = 102, had the broadest spectrum of action and completely inhibited the growth (final optical density at 600 nm