Isolation and biological characterization of a bacteriophage infecting Shigella flexneri

Abstract:

[Background] Shigella is a class of pathogenic bacteria that can cause diarrhea in humans and animals. As the problem of drug resistance is becoming more and more serious due to the abuse of antibiotics, alternative antimicrobial agents and treatment methods have become an urgent need. [Objective] To detect the pathogenicity of Shigella to broiler chickens, isolate and purify a bacteriophage (phage) that can lyse the strongly pathogenic Shigella, and study its biological characteristics. [Methods] Shigella was isolated from intestinal mucosa of diseased chickens. Healthy broiler chickens were used as animal models to test the drug resistance of highly pathogenic strains. The phage was isolated by using the host bacterium, and concentrated phage was purified by polyethylene glycol (PEG) precipitation. The morphological characteristics of the phage were observed by transmission electron microscopy (TEM). The double-layer plate method was used to determine the host spectrum, multiplicity of infection, one-step growth curve, pH, and thermal stability of the phage. [Results] This study has isolated 26 Shigella species, designated BDS1-BDS26. Among, BDS8 was the most pathogenic with multiple drug resistance and was identified as Shigella flexneri. Severe diarrhea and bloody stools were observed in the broiler chickens after feeding BDS8. Anatomical symptoms were mainly manifested as cardiac hypertrophy and obvious mesenteric haemorrhage, etc. Furthermore, the phage ΦDS8 was isolated from BDS8. TEM results showed that the head of ΦDS8 was icosahedral, with a diameter of 61±2 nm and a tail length of 165±2 nm. ΦDS8 belongs to the family of Siphoviridae. ΦDS8 could maintain high activity in the range of pH 4.0-10.0 and below 50℃. The infection cycle of ΦDS8 was about 120 min, including an incubation period of 75 min and an outbreak period of 45 min. The outbreak volume was 52 PFU/cell. [Conclusion] The phage ΦDS8 has host specificity and a better tolerance to general environments. This study provides a theoretical basis for phage therapy of shigellosis flexneri.