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OCURRENCE OF PHOTOBACTERIUM DAMSELAE IN BIJUPIRÁ
(RACHYCENTRON CANADUM), NORTHERN COAST OF SÃO PAULO STATE,
NAPOLEÃO, S.R.1*; MOURINO, J.L.2; MORAES, J.R.E.3, SILVA, A.C.1;
SHIMADA, M.T.4; MELLO, H.4; KERBER, C.5; MORAES, F.R3.
* Adress: Via de Acesso Prof. Paulo Donato Castellane s/n - CEP: 14884-900 City: Jaboticabal - State: São Paulo - Country: Brasil - silvianapoleao@yahoo.com.br SUMMARY:
Fish photobacteriosis is a bacterial fish disease commonly called “pasteurellosis” or
“pseudotuberculosis” due to infection by halophilic Photobacterium damselae. The signs of
the disease in the chronic form showing typical whitish tubercles in the internal organs which
consist of bacterial accumulations. Strain CP1, originally isolated from a farm (without using
any antibiotics for disease control) in northern coast, São Paulo, Brazil, in June of 2011, was
used in this study. The strain was isolated from head kidney, spleen and/or liver of diseased
cobia Rachycentron canadum (weighing 0,3 kg) using Trypticase Soy Agar with 5% Sheep
Blood (Difco, supplemented with 1% NaCl; TSA1) and/or Brain Heart Infusion Agar (Difco,
supplemented with 1% NaCl; BHIA1) plates. On the basis of biochemical characteristics
comparisons with those of the type (ATCC 51736) strain, results from the API 20E (the two
strains exhibited identical characters) conformed to the species description of Photobacterium
damselae. The strain CP1 was susceptible to two of the 7 antibiotics tested: chloramphenicol
and enrofloxacin.
KEY-WORDS: Photobacterium damselae, marine fish, tanque-rede, Rachycentron canadum.
TEXT:
Fish photobacteriosis is a bacterial fish disease commonly called “pasteurellosis” or “pseudotuberculosis” due to infection by halophilic Photobacterium damselae. The signs of the disease in the chronic form showing typical whitish tubercles in the internal organs which consist of bacterial accumulations (MAGARINOS et al. 1996). The disease was first described in wild populations of white perch (Morone americanus) and striped bass (Morone saxatilis) in 1963, when a massive epizootic occurred in Chesapeake Bay (USA). Sniezsko and coworkers (1964) placed the microorganism isolated during this first epizootic within the genus Pasteurella on the basis of its morphological and biochemical properties. Later, Jansen and Surgalla (1968) examined the microorganism and found sufficient distinctions in physiological and serological characteristics from other species in the genus to propose the new species name Pasteurella piscicida. A few years later, pasteurellosis became a problem in yellowtail (Seriola quinqueradiata) and ayu (Plecoglosus altivelis) cultures in Japan (ROMALDE, 2001) Soon thereafter, the disease spread to other fish species in Japan, and _______________________ 1 Master Degree, Laboratory of Experimental and Comparative Pathology, Veterinary Pathology Department-FCAV/Unesp Jaboticabal/SP – Brazil 2 Researcher, UFSC – Federal University of Santa Catarina, Shrimp Marine Laboratory. 3 Prof. Dr. of Veterinary Pathology Department-FCAV/Unesp Jaboticabal/SP and Center of Aquaculture of Unesp. 4 Master Degree, Caunesp – Center of Aquaculture of Unesp. 5 Veterinary, Itapema Farm – SP. several epizootics were described in red seabream (Acanthopagrus schlegeli), red grouper (Epinephelus akaara), oval file fish( Navodan modestus), and hybrid striped bass [M. saxatilis (M. chrysops)]. Strain CP1, originally isolated from a farm (without using any antibiotics for disease control) in northern coast, São Paulo, Brazil, in June of 2011, was used in this study. The strain was isolated from head kidney, spleen and/or liver of diseased cobia Rachycentron canadum (weighing 0,3 kg) using Trypticase Soy Agar with 5% Sheep Blood (Difco, supplemented with 1% NaCl; TSA1) and/or Brain Heart Infusion Agar (Difco, supplemented with 1% NaCl; BHIA1) plates. The pure cultures strains were stored in tryptic soy broth glicerol (+1% NaCl) at –20 °C. The strain was characterized and identified by using standard morphological, physiological and biochemical plate and tube tests, API 20E kits (ATB system, BioMerieux SA, Marcy-I’Etoile, France). The test were performed in triplicate at 30 °C and incubated for 24 h. The identification profiles were obtained by comparing the phenotypic results to the APIweb software on line database (Biomeriux®, France). Stock cultures of strain CP1 were grown on BHIA1 for 24 h at 28 °C. The bacteria were suspended in sterile phosphate buffered saline [PBS: 0.8775% (w/v) NaCl, 0.02% (w/v) KCl, 0.02% (w/v) KH2PO4, 0.115 % (w/v) Na2HPO4, 10% (v/v) glycerol, pH 7.2] and diluted to a turbidity equivalent to a MacFarland No. 0.5 standard solution (0.5 ml BaSO4 +99.5 ml 0.36 N HCl). The bacterial suspension (0.1 ml) was spread onto Mueller-Hinton agar (Difco) and antibiotic discs then added (KONEMAN et al. 1988). The antibiotic discs used in the assay included chloramphenicol, ciprofloxacin, doxycycline hydrochloride, enrofloxacin, norfloxacin, oxytetracycline and tetracycline. The plates were incubated at 35 °C for 24h and the inhibition of the bacteria by the antibiotics scored (KONEMAN et al. 1988). Only one kind of colonies, termed the strain CP1, was isolated from the kidney and/or liver of diseased cobia (R. canadum, weighing 0,3 kg) during the outbreak of serious mortality in June of 2011 in Ilhabela, São Paulo, Brazil. The moribund cobia displayed dark coloration and whitish granulomatous deposits observed on internal organs including kidney, liver. Haemorrhaging was not observed on the viscera or the walls of the peritoneal cavity. The strain was a Gram-negative, bipolar rod, non-motile, oxidase- and catalase-positive, and fermentative. The strain grow on thiosulfate citrate bile salt sucrose (TCBS; Difco) agar. On the basis of biochemical characteristics comparisons with those of the type (ATCC 51736) strain, results from the API 20E (the two strains exhibited identical characters) conformed to the species description of Photobacterium damselae. The strain CP1 was susceptible to two of the 7 antibiotics tested: chloramphenicol and enrofloxacin. This result had already been reported in R. canadum in Taiwan (LIU et al., 2003) and in Brazil (Costa et al., 2011), showing a common problem in farms of R. canadum around the world. References

COSTA, F.; LEAL, C.; FIGUEIREDO, H. 2011. Photobacterium dansalae infection in cobia
(Rachycentron canadum) fingerlings in a Brazilian hatchery. Abstract of Congress WAS
Aquaculture - Natal, Brazil.
JANSSEN, W. A. and SURGALLA, M. J., 1968. Morphology, physiology, and serology of a
Pasteurella species pathogenic for white perch (Roccus americanus). J. Bacteriol., 96, 1606–
1610.
KONEMAN, E. W., ALLEN, S. D., DOWELL, V. R. JR,, JANDA, W. M., SOMMERS, H.
M. and WINN, W. C. (Editors), 1988. Diagnostic Microbiology, 3rd edn. Philadelphia: J.B.
Lippincott.
LIU, PING-CHUNG; LIN, JI-YANG and LEE, KUO-KAU. 2003. Virulence of Photobacterium damselae subsp. Piscicida in cultured cobia Rachycentron canadum. J. Basic Microbiol. 43(6), 499–507 MAGARINOS, B., TORANZO, A. E. and ROMALDE, J. L. 1996. Phenotypic and pathobiological characteristics of Pasteurella piscicida. Annu. Rev. Fish Dis., 6, 41–64. ROMALDE, J.L. 2002. Photobacterium damselae subsp. piscicida: an integrated view of a bacterial fish pathogen. Int Microbiol 5: 3–9. SNIEZSKO, S. F., BULLOCK, G. L., HOLLIS, E. and BOONE, J. G., 1964. Pasteurella sp. from an Epizootic of white perch (Roccus americanus) in Chesapeake Bay tidewater area. J. Bacteriol., 88, 1814–1815.

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