Isolation and characterization of multidrug-resistant Proteus mirabilis from diarrheic sika deer fawns in Northeast China

Abstract

Diarrhea in sika deer fawns is a multifactorial disease that severely affects the development of deer farming in China. In the present study, we focused on Proteus mirabilis isolates recovered from diarrheic sika deer fawns in Heilongjiang, Jilin, and Liaoning provinces from 2022 to 2024. A total of 75  P. mirabilis isolates were identified using colony morphology and Gram staining together with 16S rRNA gene sequencing, type-strain-inclusive phylogenetic analysis, and atpD gene-targeted PCR as a supplementary molecular assay. Virulence genes and antimicrobial resistance genes were detected by PCR, antimicrobial susceptibility was determined using the broth microdilution method, and biofilm formation was evaluated by crystal violet staining. The pathogenic potential of representative isolates was further assessed using murine oral gavage and intraperitoneal challenge models, followed by observation of clinical signs and histopathological examination of the heart, liver, spleen, lung, and kidney. The isolates carried multiple virulence and antimicrobial resistance genes, exhibited multidrug-resistant phenotypes, and showed moderate to strong biofilm-forming capacity. Oral gavage with representative isolates induced obvious diarrhea in mice within two days, while intraperitoneal challenge caused clinical signs, mortality, and histopathological lesions in multiple organs. Whole-genome sequencing of the principal challenge strain Y24N1546 yielded a single circular chromosome of 4,073,229 bp. Genome-wide comparison with the P. mirabilis type strain ATCC 29906 T yielded an average nucleotide identity (ANI) of 98.94% and a digital DNA-DNA hybridization (dDDH) value of 93.4%; independent TYGS analysis also assigned Y24N1546 to P. mirabilis . Overall, these findings support P. mirabilis as an important bacterial pathogen associated with diarrheic sika deer fawns in Northeast China. Because fawn diarrhea is multifactorial, the etiological contribution of P. mirabilis should be interpreted together with other host, environmental, and management-related factors.