Emerging fish pathogens Lactococcus petauri and L. garvieae in Nile tilapia (Oreochromis niloticus) farmed in Brazil.

Abstract

Lactococcosis in fish has been associated with Lactococcus garvieae and the recently described L. petauri. However, the relevance of these emerging fish pathogens to Nile tilapia still requires thorough understanding. This study investigated lactococcosis outbreaks in Nile tilapia on Brazilian farms and characterized the isolates through molecular identification of the bacterial species, multilocus sequence typing (MLST) analysis, virulence to Nile tilapia, and antimicrobial susceptibility. Lactococcosis outbreaks were monitored from 2019 to 2022 throughout Brazil. The outbreaks occurred mainly during warmer months, and co-infections were observed in four farms, whereas concurrent bacterial infections were identified in all farms. Since the sequence of the 16S rRNA was not capable of differentiating between L. petauri and L. garvieae, Lactococcus spp. isolates were identified at the species level using the gyrB gene sequence. In total, 30 isolates were classified as L. petauri and two as L. garvieae. All L. petauri isolates were grouped in ST24, except for one isolate which belonged to the newly described ST47. A new ST was also described for the L. garvieae isolates identified, ST46. Furthermore, L. petauri ST24 and ST47 were characterized as singletons, whereas L. garvieae ST46 was grouped with ST16 and ST17 and formed CC17. For the challenge trial, an L. petauri ST24 isolate was chosen considering that this MLST lineage was the most frequently observed. L. petauri was reisolated from challenged Nile tilapia, confirming the pathogenicity of this bacterium to Nile tilapia. The infection in the fish progressed very rapidly, and within 48 h post-challenge clinical signs and the first mortalities were observed. The estimated LD50 was 5.74 x 103 CFU 15 days post-challenge. Provisional epidemiological cutoff values were determined for L. petauri for six antimicrobial agents from different drug classes. All isolates were characterized as wild type (WT) for neomycin and oxytetracycline, whereas 96.67 % of the isolates were characterized as WT for amoxicillin, erythromycin, and florfenicol, and 83.33 % were WT for norfloxacin. Of the 14 outbreaks analyzed, 12 were caused by L. petauri and two by L. garvieae. The gyrB gene sequence was used to differentiate L. petauri from L. garvieae and allowed for the correct identification of these pathogens. Two MSLT lineages of L. petauri were identified and ST24 was observed in different regions of the country, illustrating a rapid expansion of this bacterial lineage. Highlights of the manuscriptO_LILactococcus petauri is pathogenic to Nile tilapia. C_LIO_LIThe MLST lineage most observed was L. petauri ST24, indicating its adaption to infect Nile tilapia. C_LIO_LIThe analysis of the gyrB gene sequence allowed for the correct identification of L. petauri and L. garvieae. C_LI