Characterization and virulence potential of phenotypically diverse Aeromonas veronii isolates recovered from moribund freshwater ornamental fishes of Kerala, India

Abstract

In the present study, we investigated the involvement of Aeromonas spp. in eliciting disease outbreaks in freshwater ornamental fishes across the state of Kerala, India. We investigated three incidences of disease, in which the moribund fishes exhibited clinical signs such as haemorrhagic septicemia (in gouramy, Trichogaster sp.), dropsy (in Oscar, Astronotus ocellatus) and tail rot/fin rot (in gold fish, Carassius carassius). Pure cultures (n = 20 from each fish; 60 in total) of Aeromonas spp. were recovered from the abdominal fluid as well as from internal organs of affected fishes, although they could not be identified to species level because of the variations in their phenotypic characters. The molecular fingerprinting of the isolates using Enterobacterial Repetitive Intergenic Consensus PCR proved the genetic diversity of the isolates from the three sites. The phylogenetic trees constructed using concatenated sequences (using 16S rRNA, gyrA, gyrB and rpoD genes) indicated that they were related to Aeromonas veronii. They exhibited marked cytotoxic and haemolytic activity, which were responsible for the pathogenic potential of the isolates. The isolates possessed multiple virulence genes such as enterotoxins (act and alt), haemolytic toxins (aerA and hlyA), genes involved in type III secretion system (ascV, aexT and ascF-ascG), glycerophospholipid-cholesterol acyltransferase (gcat) and a type IV pilus (tapA) gene, as determined by PCR. Virulence of representative isolates to goldfish was also tested, and we found LD(50) values of 10(4.07)-10(5.35) cfu/fish. Furthermore, the organisms could be recovered as pure cultures from the lesions as well as from the internal organs.

Aeromonas hydrophilaandAeromonas veroniicause motileAeromonassepticaemia in the cultured Chinese sucker,Myxocyprinus asiaticus

“…Bacterial identification should be carried out step by step, with colony morphology description followed by a physiological and biochemistry characteristic test and molecular analysis. These steps are important for the classification of bacterial phenotype, serotype and genotype (Martinez‐Murcia et al, ; Sreedharan, Philip, & Singh, ; Wei, Puah, Jin, Puthucheary, & Chua, ). Ma and Yu () provided nine critical reactions for bacterial phenotype identification, namely, arginine dihydrolase (+), lysine decarboxylase (+), glucose fermentation (+), esculin/salicin (+), mannitol/ketole (+), saccharose (+) and cellobiose (−).…”

“…Bacterial identification should be carried out step by step, with colony morphology description followed by a physiological and biochemistry characteristic test and molecular analysis. These steps are important for the classification of bacterial phenotype, serotype and genotype (Martinez-Murcia et al, 2011;Sreedharan, Philip, & Singh, 2013;Wei, Puah, Jin, Puthucheary, & Chua, 2015). Ma and Yu (1992) (Pidiyar, 2002).…”

Characterization and antimicrobial susceptibility of motile aeromonads isolated from freshwater ornamental fish showing signs of septicaemia

“…In a study done to investigate the causative organisms behind incidences of bacterial disease (n = 23) among a number of different species of ornamental fish in Sri Lanka, Hettiarachchi & Cheong (1994) found A. hydrophila to be the most dominant species, associated with 18 incidences (78.26%). In contrast, Sreedharan et al (2013) investigated 3 incidences of disease (1 each in gourami, goldfish and oscar), in Kerala, India, and isolated A. veronii as the causative organism in all 3 cases. A comparatively higher occurrence of A. veronii over A. hydrophila has been reported among aeromonads isolated from diseased eels in the Republic of Korea (Yi et al 2013) and diseased freshwater aquaculture fish from China (Hu et al 2012).…”

Evidence of fish and human pathogens associated with doctor fish (Garra rufa, Heckel, 1843) used for cosmetic treatment

“…This scenario suggests that the most probable cause of mortality outbreaks comprised the association between stress conditions (movement, captivity) and bacterial co‐infections. In particular, the poor management of fish debilitated by mycobacterial infection may have triggered a bacterial septicaemia due to opportunistic bacteria such as A. veronii and S. putrefacien s. These agents have in fact been associated with fish haemorrhagic septicaemia outbreaks (Jagoda et al, ; Lee et al, ; Sreedharan et al, ). A. veronii especially resulted in the most frequent Aeromonas spp.…”