Microbiomic and transcriptomic insight into the pathogenesis of meningitis-like disease in cultured Pelophylax nigromaculata

Abstract In recent years, a deadly infectious disease named frog meningitis (or cataract and torticollis) raged the frog farms in China. Diseased frogs manifested torticollis, cataract, edema, and finally died. Yet, the molecular pathogenesis of this disease has not been studied. In this study, the etiologic factor of frog meningitis broken in a black-spotted frog farm was investigated by comparative microbiomics (brain, eyeball, and intestine). And the pathogenesis of two major types of symptomatic frogs (individuals with both torticollis and cataract and individuals with only torticollis) were investigated by histopathological section and comparative transcriptomics of brain, liver, and muscle. Our results indicated that Elizabethkingia bacteria had increased proportions in the brain and eyeball microbiomes (but not intestine microbiomes) of both types of symptomatic frogs, and E. miricola was dominant in the eyeball and brain microbiomes of most torticollis-cataract frogs. In the torticollis-only individuals, vascular congestion and reduced hepatic fat can be detected in their leptomeninges vessels and liver, respectively. While in the torticollis-cataract ones, we observed additional leukocyte extravasation in their leptomeninges vessel, and neurologic damage in their brain and spiral cord. Transcriptional analyses suggested that frogs could be well divided into asymptomatic, torticollis-only, and torticollis-cataract groups by the variation of their brain transcriptomes. The upregulated brain genes in symptomatic frogs were mainly enriched in immune-related pathways. And the Toll-like receptor and NOD-like receptor signaling pathways, inflammation, and adaptive humoral immunity were activated in response to E. miricola infection. Torticollis-only and torticollis-cataract individuals differed in the level of immune gene expression remarkably, and the emerge of cataract symptom indicated the onset of strong innate immune and inflammatory responses. Overall, our results suggested that E. miricola was likely the etiologic factor of meningitis emerged in this farm. In regards to pathological stages, the infection progressed to leptomeninges invasion in torticollis-only individuals, while it had caused meningitis in torticollis-cataract ones. Correspondingly, the torticollis symptom was likely due to leptomeninges infection, and the cataract was likely a concomitant symptom of meningitis due to disruption of vascular permeability during excessive inflammation. Our results provided a first molecular insight into the pathogenesis of frog meningitis.