RESUMEN
Streptococcus agalactiae and Francisella noatunensis subsp. orientalis (Fno) are important pathogens for farm-raised tilapia worldwide. There are no reports of coinfection caused by S. agalactiae and Fno in fish. This study aimed to determine the aetiology of atypical mortalities in a cage farm of Nile tilapia and to characterize the genetic diversity of the isolates. Fifty-two fish were sampled and subjected to parasitological and bacteriological examination. The S. agalactiae and Fno isolates were genotyped using MLST and REP-PCR, respectively. Whole-genome sequencing was performed to confirm the MLST results. Seven fish were shown coinfected by S. agalactiae and Fno. Chronic hypoxia and a reduction in the water temperature were determined as risk factors for coinfection. Fno isolates were shown clonally related in REP-PCR. The MLST analysis revealed that the S. agalactiae isolates from seven coinfected fish were negative for the glcK gene; however, these were determined to be members of clonal complex CC-552. This is the first description of coinfection by S. agalactiae and Fno in farm-raised Nile tilapia. The coinfection was predisposed by chronic hypoxia and was caused by the main genotypes of S. agalactiae and Fno reported in Brazil. Finally, a new S. agalactiae genotype with glcK gene partially deleted was described.
Asunto(s)
Cíclidos , Enfermedades de los Peces/mortalidad , Francisella/fisiología , Variación Genética , Infecciones por Bacterias Gramnegativas/veterinaria , Infecciones Estreptocócicas/veterinaria , Streptococcus agalactiae/fisiología , Animales , Acuicultura , Coinfección/microbiología , Coinfección/mortalidad , Coinfección/veterinaria , Enfermedades de los Peces/microbiología , Francisella/genética , Infecciones por Bacterias Gramnegativas/microbiología , Infecciones por Bacterias Gramnegativas/mortalidad , Secuencias Repetitivas Esparcidas , Secuencias Invertidas Repetidas , Tipificación de Secuencias Multilocus/veterinaria , Reacción en Cadena de la Polimerasa/veterinaria , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/mortalidad , Streptococcus agalactiae/genéticaRESUMEN
Despite the worldwide occurrence of Francisella noatunensis subsp. orientalis (Fno) infection in farmed tilapia, sensitivity and specificity estimates of commonly used diagnostic tests have not been reported. This study aimed to estimate the sensitivity and specificity of bacteriological culture and qPCR to detect Fno infection. We tested 559 fish, sampled from four farms with different epidemiological scenarios: (i) healthy fish in a hatchery free of Fno; (ii) targeted sampling of diseased fish with suggestive external clinical signs of francisellosis during an outbreak; (iii) convenience sampling of diseased and clinically healthy fish during an outbreak; and (iv) sampling of healthy fish in a cage farm without a history of outbreaks, but with francisellosis reported in other farms in the same reservoir. The qPCR had higher median sensitivity (range, 48.8-99.5%) than culture (range, 1.6-74.4%). Culture had a substantially lower median sensitivity (1.6%) than qPCR (48.8%) to detect Fno in carrier tilapia (farm 4). Median specificity estimates for both tests were >99.2%. The qPCR is the superior test for use in surveillance and monitoring programmes for francisellosis in farmed Nile tilapia, but both tests have high sensitivity and specificity which make them fit for use in the diagnosis of Fno outbreaks.
Asunto(s)
Cíclidos , Enfermedades de los Peces/microbiología , Francisella/clasificación , Infecciones por Bacterias Gramnegativas/veterinaria , Animales , Acuicultura , Brasil , ADN Bacteriano/genética , Brotes de Enfermedades/veterinaria , Enfermedades de los Peces/patología , Francisella/genética , Francisella/aislamiento & purificación , Infecciones por Bacterias Gramnegativas/microbiología , Infecciones por Bacterias Gramnegativas/patología , Reacción en Cadena en Tiempo Real de la Polimerasa , Sensibilidad y EspecificidadRESUMEN
Flavobacterium columnare is responsible for disease outbreaks in freshwater fish farms. Several Brazilian native fish have been commercially exploited or studied for aquaculture purposes, including Amazon catfish Leiarius marmoratus × Pseudoplatystoma fasciatum and pacamã Lophiosilurus alexandri. This study aimed to identify the aetiology of disease outbreaks in Amazon catfish and pacamã hatcheries and to address the genetic diversity of F. columnare isolates obtained from diseased fish. Two outbreaks in Amazon catfish and pacamã hatcheries took place in 2010 and 2011. Four F. columnare strains were isolated from these fish and identified by PCR. The disease was successfully reproduced under experimental conditions for both fish species, fulfilling Koch's postulates. The genomovar of these 4 isolates and of an additional 11 isolates from Nile tilapia Oreochromis niloticus was determined by 16S rRNA restriction fragment length polymorphism PCR. The genetic diversity was evaluated by phylogenetic analysis of the 16S rRNA gene and repetitive extragenic palindromic PCR (REP-PCR). Most isolates (n = 13) belonged to genomovar II; the remaining 2 isolates (both from Nile tilapia) were assigned to genomovar I. Phylogenetic analysis and REP-PCR were able to demonstrate intragenomovar diversity. This is the first report of columnaris in Brazilian native Amazon catfish and pacamã. The Brazilian F. columnare isolates showed moderate diversity, and REP-PCR was demonstrated to be a feasible method to evaluate genetic variability in this bacterium.