Isolation and identification of Klebsiella pneumoniae from Tibetan pigs and preparation of inactivated vaccine with Tibetan medicine adjuvant.

This study investigated the prevalence of Klebsiella pneumoniae (KP) in Tibetan pigs. Nasal swabs and lung tissue samples were collected, from which hypervirulent Klebsiella pneumoniae (hvKP) were isolated through culture, identification, serotyping, and virulence gene detection. These strains subsequently underwent antimicrobial susceptibility testing and pathogenicity assessment. Using a hvKP as the antigen, three types of vaccines were prepared: an inactivated vaccine, an inactivated vaccine adjuvanted with Tibetan astragalus polysaccharide, and an inactivated vaccine with a conventional adjuvant. Mouse immunization experiments were conducted to evaluate immunogenic potency, survival rates post-challenge, and antibody production, thereby comparing the protective efficacy of the different vaccine formulations.The results of this experiment showed that 54 strains of KP were isolated from nasal swabs and lungs of Tibetan pigs; They were typed and found to contain two highly virulent serovars (K20 and K54);The virulence genes carried by the two serovars of KP were found to contain hvKP characteristic virulence genes in the virulence gene assay, which is consistent with the hvKP characteristic.Both serovars of KP exhibited limited drug resistance but still belonged to multidrug-resistant bacteria. In this study, mouse challenge experiments confirmed that these strains possess significant pathogenicity and lethality toward healthy mice, meeting the criteria for use as immunogenic antigens The results of sterility test and animal safety test proved that the final concentration of the inactivator was 0.3% and had no obvious adverse reaction to individual animals; the results of immune response test showed that the three kinds of inactivated vaccines had no adverse reaction to mice, and the antibody potency of inactivated vaccine with adjuvant of Tibetan astragalus polysaccharide was higher than that of inactivated vaccine of KP in the immune potency test, and the antibody potency of inactivated vaccine with conventional adjuvant was the highest, which proved that Tibetan astragalus polysaccharide could be used as a vaccine to increase the immune antibody potency. This proves that Tibetan astragalus polysaccharide can be used as a vaccine adjuvant to increase the antibody potency of immunity. In a murine model, the inactivated vaccine elicited complete protection (100% survival) against a lethal challenge with the homologous K20 and K54 strains under the tested conditions. Furthermore, bacterial re-isolation experiments confirmed that deceased mice in control groups were positive for KP, while all immunized mice survived and cleared the infection. In summary, the prevalent KP of porcine origin in the plateau Linzhi area is mainly prevalent in K20 and K54. These strains carry the virulence genes of hvKP, are pathogenic and lethal, and belong to the high-virulence KP. The inactivated vaccine prepared against these two serovars has been shown to have significant protective efficacy, and the incorporation of Tibetan astragalus polysaccharide as a vaccine adjuvant has been demonstrated to enhance the protective efficacy of the vaccine. This provides a valuable foundation for the development of subsequent preventive and control measures against Klebsiella pneumoniae.