Arq. Bras. Med. Vet. Zootec., v.60, n.1, p.

275-277, 2008

Communication

[Comunicação]

Isolation and characterizaton of Edwardsiella tarda from pacu Myleus micans

[Isolamento e caracterização de Edwardsiella tarda em pacu Myleus micans]

L.C. Lima1, A.A. Fernandes1, A.A.P. Costa2, F.O. Velasco2, R.C. Leite1, J.L. Hackett3

1
Escola de Veterinária - UFMG
Caixa Postal 567
30123-970 – Belo Horizonte, MG
2
Aluno de graduação - EV-UFMG – Belo Horizonte, MG
3
Fisheries and Environmental Management Services – Victoria, BC, Canada

In January 2005, significant fish mortality A minimum of five alive and moribund fish were
occurred in several species in the São Francisco collected from the main zones of mortality. After
river which extended along 2006. The mortality clinical examination, kidney samples were
in fish stocks in this Brazilian river basin is of aseptically collected, streaked onto blood agar
great concern since fishery is the main source of and on brain heart infusion agar (BHI). The
economic income for the local communities and plates were brought to the laboratory for colonies
many of the affected species are of great identification. Sub samples were grown onto
economic value. In order to elucidate the causes BHI and blood agars and pure individualized
of the fish kill, a series of physical, chemical, colonies were streaked onto McConkey (MC)
toxicological, and microbiological analysis were and Salmonela-Shigella (SS) agars and incubated
carried out. Among the bacteria found in the at 30oC, for 24-48h. Et-LIS antimicrobial
species sampled, it was isolated Edwardsiella susceptibility patterns were determined by the
tarda from pacu Myleus micans. disk diffusion method using Muller-Hinton agar.
The following drugs were tested: erythromycin,
E. tarda, a member of the family ampicillin, penicillin, tetracycline, enrofloxacin,
Enterobacteriaceae, is the causative agent of sulfazotrin, neomycin, amoxillin, streptomycin,
septicemia in a variety of fish species, being ceftiofur, gentamycin, and florfenicol. Virulence
frequently isolated from Japanese flounder characteristics of Et-LIS were assessed by
Paralichthys olivaceous (Miwa and Nobuhiro, challenging two tropical species, tilapia
2000), eels Anguilla japonica, and catfish Oreochromis spp. and carp Cyprinus carpio. The
Ictalurus punctatus (Inglis et al., 2001). In fish were inoculated with 2ml of bacterial
Brazil, E. tarda was isolated from farmed solution prepared according pattern 2 of
rainbow trout Oncorhynchus mykiss McFarland standards. The individuals were kept
(Alexandrino et al., 1999). Although it is without feeding and isolated in aquaria until die.
commonly classified as opportunistic, E. tarda is The control group was injected with 2ml of
considered a serious pathogen of fish. This saline solution and sacrificed to permit
bacterium is also important due its zoonotic comparisons with the challenged individuals.
aspects. Infected fish processed for human Portions of kidneys, liver, and celomatic
consumption is a source of gastroenteritis and exudates were aseptically removed and streaked
meningitis (Janda and Abbot, 1993; Inglis et al., in SS agar to confirm the infection of E. tarda.
2001; Muratori et al. 2001).


Recebido em 23 de fevereiro de 2007

Aceito em 18 de dezembro de 2007
E-mail: [email protected]
 Lima et al.

On BHI agar plates, Et-LIS colonies were small characterizations, in triplicate, permitted the
(1mm diameter), transparent, and punctate. identification of the isolate as Edwardsiella tarda
Colonies on SS agar varied from 1-3mm (Table 1). To confirm the identification by
diameter and showed small black centers to conventional methods, a rapid diagnostic kit
predominantly black colonies indicating H2S (Bactray system)1 was also used. Run in
production. While presumptive identification duplicate, the kit identified E. tarda at 99.9%
revealed small Gram negative rods, biochemical confidence level.

Table 1. Biochemical profile of Edwardsiella tarda, Et-LIS, isolated from kidney of pacu Myleus micans
Characteristic Characteristic
Gram stain - Glucose +
Catalase + Arabinose -
Cytochrome oxidase - Lactose -
Voges-Proskauker - Trehalose -
Lysine descarboxylase + Inositol -
Ornithin descarboxylase + Salicin -
Citrate, Simmons -
Oxidative/Fermentative +/+ Fructose +
Sulphide + Sucrose -
Indole + Adonitol -
Motility + Dulcitol -
Esculin hydrolysis - Triple Sugar Iron agar * K/AG
Urease - Growth in BHI with 3% NaCl +
K= alkaline or no reaction, A= acid production, G= gas production.

In antimicrobial susceptibility assay, Et-LIS of E. tarda infection may slightly differ from fish
tested sensitive to all the 12 drugs used, except species to species. Experimentally infected,
for erythromycin (resistant) and penicillin channel catfish exhibited hemorrhages in the
(intermediate). According to the literature, E. visceral organs, however none of the usual
tarda is sensitive to a wide variety of external signs found among fish with natural
antimicrobials (Janda and Abbot, 1993) and, infections were noted in experimentally
therefore, nearly any approved drug can be used inoculated fish (Inglis et al., 2000). Together
to combat the infection. In challenged fish, Et- with other genus like Aeromonas, isolated from
LIS pure cultures were recovered from celomatic additional fish species sampled with the infected
exudates and from kidney. Tilapias challenged pacu, it is suggested that E. tarda might have had
were found dead within 24h after exposure, a role in the mortality occurred in the São
while the carps lived for 72h. Both species Francisco river system. Although Et-LIS showed
showed similar infection signs like reddening of to be susceptible to a relatively great range of
fins, opercula, and anal region. Also, tilapias and drugs, its isolate infection can be fatal to other
carps showed corneal opacity, body ulcers, tropical fish species. Et-LIS also seems to be
abdominal swelling, a generalized internal more fulminating to common carp than tilapia.
hyperemia and organs enlarged and hemorrhagic.
Similar symptoms were found in infected Keywords: fish, pacu, Myleus micans,
rainbow trout reared in intensive flow-through Edwardsiella tarda, Sao Francisco basin
systems (Alexandrino et al., 1999). Clinical signs


RESUMO

Investigaram-se as causas da mortalidade de peixes ocorrida em janeiro de 2005 na bacia do Rio São
Francisco, Brasil. Edwardsiella tarda foi isolada dos rins de pacu Myleus micans. O isolado, denominado
Et-LIS, caracterizado por bastonetes Gram negativos móveis, foi identificado por testes bioquímicos e
confirmado pelo kit comercial Bactray. A susceptibilidade a 10 drogas das 12 testadas foi determinada

1
Laborclin – Pinhais, PR ,Brazil.

276 Arq. Bras. Med. Vet. Zootec., v.60, n.1, p.275-277, 2008
 Isolation and characterization…

pelo método de difusão de discos, enquanto as características de virulência foram avaliadas mediante
inoculação experimental em Cyprinus carpio e em Oreochromis spp. Ambas as espécies desafiadas
apresentaram sinais compatíveis com infecção por E. tarda. As tilápias (Oreochromis spp.) morreram
48h após a inoculação, enquanto as carpas (Cyprinus carpio) sobreviveram por 72h. Este é o primeiro
relato da ocorrência de E. tarda em pacu.

Palavras-chave: peixe, pacu, Myleus micans, Edwardsiella tarda, bacia do São Francisco

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