Phage and application thereof
A technology of phage and Escherichia coli, applied in the direction of phage, virus/phage, application, etc., can solve the problem of insufficient products and achieve the effect of avoiding evolution and the generation of drug-resistant strains
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Embodiment 1
[0021] Example 1: Isolation and purification of enterotoxigenic Escherichia coli ETEC-K88ac phage
[0022] 1. Isolation of phage:
[0023] In July 2016, in a medium-sized farm in the suburbs of Hengyang, severe diarrhea occurred in multiple pens of piglets within 2 months, and the disease could not be controlled after antibiotic treatment. The mortality rate was as high as 20%. After isolation, a highly toxic strain of Enterotoxigenic Escherichia coli E.coli-HuNHY19 was deposited on April 24, 2019 in the General Microbiology Center of the China Committee for the Collection of Microorganisms (Address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing), and its preservation number It is CGMCC No.17622, named as Escherichia coli (Escherichia coli). After typing and identification, E.coli-HuNHY19 was O8: K91, K88ac, indicating that the main prevalent pathogenic Escherichia coli was enterotoxigenic Escherichia coli K88ac.
[0024] In this regard, sewage samples were ...
Embodiment 2
[0042] 1. Temperature Sensitivity of Phage-HY19
[0043] Take 600 µL for a titer of 10 8 The phage liquid of pfu / mL was placed in 5 sterile EP tubes, respectively, at 50°C, 60°C, 70°C, 80°C, and 90°C for 20 min, 40 min, and 60 min, and each temperature was set for 3 replicates , after the action time was over, samples were taken, immediately placed in a water bath to cool down, and then the titer of the phage was determined, and the experiment was repeated 3 times. The temperature sensitivity of Phage-HY19 as image 3 As shown, the activity of phage Phage-HY19 did not change significantly after being treated at 50-70°C for 1 hour; after being treated at 80°C for 40 minutes, all phages were inactivated; after being treated at 90°C for 20 minutes, all phages were inactivated.
[0044] Determination of the host spectrum of phage Phage-HY19
[0045] 2.1 Test material:
[0046] E.coli-HuNHY19 (preservation number is CGMCC No.17622, O8: K91, K88ac, self-isolated and identified...
Embodiment 3
[0052] Embodiment 3: Activity test of bacteriophage Phage-HY19
[0053] 3.1 Toxicity test
[0054] Take a total of 20 5-week-old female experimental Kunming mice, and randomly divide them into two groups, each with 10 mice, wherein group 1 is the test group, and the oral concentration is 10 9 Pfu / ml phage culture solution 2mL, group 2 was the control group, the same amount of PBS was orally administered for 14 days, the mice were killed by neck dislocation, and the changes of viscera, digestive tract and mucous membrane were observed. The results showed that the phage Phage-HY19 had no effect on the daily behavior of the mice, and there was no abnormality in the anatomical examination.
[0055] In vitro bactericidal activity
[0056] Escherichia coli E.coli-HuNHY19 (preservation number: CGMCC No. 17622), CVCC1524 (O8: K91, K88ac) cultured overnight was 1.0 mL each (the bacteria content was 1×10 9 cfu / mL) mixed and OD it with LB 600nm The value is adjusted to 1.0 (the bacte...
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