Aquaculture Vol. 192 (2-4) pp. 101-110
Relationships between disease outbreak in cultured tiger shrimp (Penaeus monodon) and the composition of Vibrio communities in pond water and shrimp hepatopancreas during cultivation 

a Hung-Hung Sung
a Shi-Fang Hsu
a Chih-Kun Chen
b Yun-Yuan Ting
a Wei-Liang Chao

a Department of Microbiology, Soochow University, Shih Lin, , Taipei, Taiwan
b Tainan Fish Culture Branch, Taiwan Fisheries Research Institute, , Tainan, Taiwan

Abstract
Tiger shrimp (Penaeus monodon) larvae were first cultivated for 1 month in the same pond and then at postlarval 43 were transferred into three different culture ponds (E2, E4, and W4). Vibrio spp. from the pond water and shrimp hepatopancreas were isolated and identified using thiosulfate-citrate-bile salt-sucrose (TCBS) agar and fatty acid methyl esters (FAME) analysis. For the initial 60 days after transfer, the composition of the Vibrio community in the pond water remained fairly diverse, but subsequently decreases in species diversity were observed in all three ponds, suggesting that the culture system was under some kind of stress. However, no clear relationship between this reduction in species diversity, and the subsequent occurrence of disease to the changes in total viable and Vibrio counts were observed. Although the Vibrio community in pond E4 remained relatively diverse, in water samples taken from E2 and W4 between Day 73 and 89, the dominant species according to FAME were Vibrio cholerae and, apparently, Vibrio furnissii/Aeromonas spp., respectively. By Day 95, at which time all the shrimps, including those in pond E4, were dead, the microbial communities in all these ponds appeared to be dominated by Aeromonas spp. and V. furnissii. When thirty-eight of these Aeromonas spp. and V. furnissii isolates were randomly selected and re-identified using Biolog GN plates, the majority of them (58%) were reassigned either to Vibrio parahaemolyticus or the Vibrio harveyi/Vibrio carchariae group. Vibrios were only detected once in the hepatopancreas of shrimp from pond E2. In W4 shrimp, more than 104 CFU of vibrios/g were detected in samples taken on Day 62 and thereafter. The average weight of shrimp collected on Day 89 from ponds W4 and E2 was 4.91 and 7.96 g, respectively, which suggests that the presence of a large number of vibrios in the hepatopancreas may be associated with growth retardation in shrimps. The Biolog system identified 53.3% of the vibrios isolated from the hepatopancreas of shrimps collected from pond W4 on Day 62, as V. harveyi and 20% as V. parahaemolyticus. One month later, immediately before mass mortality of the shrimp, Biolog results suggested that the W4 hepatopancreatic Vibrio communities were dominated by V. parahaemolyticus.

Aquaculture Vol. 164 (1-4) pp. 359-366
Influence of vaccination on vibriosis resistance of the giant black tiger shrimp Penaeus monodon (Fabricius)

^a O.S.P. Teunissen
^b R. Faber
^a G.H.R. Booms
^b T. Latscha
^a J.H. Boon
^a , Wageningen Institute of Animal Science, Wageningen Agricultural University, P.O. Box 338, , 6700 AH Wageningen, Netherlands
^b , Intervet International, P.O. Box 31, , 5830 AA Boxmeer, Netherlands
Abstract:    The study aims to test the influence of vaccination with polyvalent vaccine prototypes on the vibriosis resistance of Penaeus monodon. Vaccinated P. monodon post-larvae were challenged with a virulent Vibrio alginolyticus strain 10, 20, and 30 days post-vaccination. Results showed that vaccination significantly enhances the resistance of shrimp to vibriosis. This effect decreased by age of the post-larvae, particularly for the glucan group. © 1998 Elsevier Science B.V. All rights reserved.

Aquaculture Vol. 164 (1-4) pp. 367-374
Bacterial flora in the hepatopancreas of pond-reared Penaeus monodon juveniles with luminous vibriosis

^a Eduardo M. Leaño
^a Celia R. Lavilla-Pitogo
^a Milagros G. Paner
^a , Aquaculture Department, Southeast Asian Fisheries Development Center, Tigbauan, , 5021 Iloilo, Philippines
Abstract:    Quantification and characterization of bacterial flora in the hepatopancreas (hp) of pond-reared Penaeus monodon juveniles affected with luminous bacteria were conducted in 1994 and 1995. Shrimp samples were taken from 23 grow-out ponds, 14 of which had disease outbreaks. Luminous bacterial (LB) load of the shrimps' hp with (mean=2.4×10¹ colony forming units (CFU)/hp) and without (mean=0.3×10¹ CFU/hp) disease outbreaks were comparable during the first 15 days of culture (DOC). During disease outbreaks at 18 to 32 DOC, however, LB load of affected shrimps (mean=9.0×10⁴ CFU/hp) were higher than healthy shrimps (mean=7.0×10¹ CFU/hp). At 50 to 60 DOC, levels of LB were comparable in older shrimps with or without disease. Total viable and presumptive Vibrio counts were also comparable in both shrimp samples from 1 to 60 DOC. Characterization of the 172 bacterial isolates collected showed that most (90.12%) were Vibrio species dominated by V. harveyi (27.91%), V. splendidus II (13.37%) and V. parahaemolyticus (10.46%). © 1998 Elsevier Science B.V. All rights reserved.

DAO 40:101-107 (2000)

Toxic factors of Vibrio strains pathogenic to shrimp

Cyrille Goarant^1,*, José Herlin¹, Raphaël Brizard¹, Anne-Laure Marteau¹, Claude Martin², Bernard Martin²

¹Laboratoire de Recherche Aquacole IFREMER en Nouvelle-Calédonie, Station d'Aquaculture de Saint Vincent, BP 2059, 98845 Nouméa Cedex, New Caledonia
²Laboratoire de Biologie Cellulaire, Université Française du Pacifique, Centre Universitaire de Polynésie Française, BP 6570, FAA'A Aéroport, Tahiti, French Polynesia

*E-mail: cgoarant@ifremer.fr
ABSTRACT:     Vibriosis is a major disease problem in shrimp aquaculture. 'Syndrome 93' is a seasonal juvenile vibriosis caused by Vibrio penaeicida which affects Litopenaeus stylirostris in grow-out ponds in New Caledonia. This study assessed the toxic activities of extracellular products (ECPs) from V. penaeicida, V. alginolyticus and V. nigripulchritudo using in vivo injections in healthy juvenile L. stylirostris (= Penaeus stylirostris) and in vitro assays on shrimp primary cell cultures and the fish cell line epithelioma papulosum cyprini (EPC). Toxic effects of ECPs were demonstrated for all pathogenic Vibrio strains tested both in vivo and in vitro, but for shrimp only; no effect was observed on the fish cell line. ECP toxicity for New Caledonian V. penaeicida was found only after cultivation at low temperature (20°C) and not at higher temperature (30°C). This points to the fact that 'Syndrome 93' episodes are triggered by temperature drops. The assays used here demonstrate the usefulness of primary shrimp cell cultures to study virulence mechanisms of shrimp pathogenic bacteria.

DAO 43:153-157 (2000) Abstract 
Use of prawn blood agar hemolysis to screen for bacteria pathogenic to cultured tiger prawns Penaeus monodon 
Chin-I Chang, Wen-Yu Liu*, Chung-Zen Shyu 
Department of Aquaculture, Taiwan Fisheries Research Institute, 199 Hou-Ih Rd, Keelung 202, Taiwan 
*Corresponding author. E-mail: wyliu@mail.tfrin.gov.tw 

ABSTRACT:     A newly developed prawn blood agar consisting of 1 ml of tiger prawn hemolymph in medium containing 200 ppm Rose Bengal was used to determine the hemolytic activity of 35 isolates of bacteria obtained from cultured tiger prawns Penaeus monodon and their rearing water. For comparison, the hemolytic activity of these isolates was also determined in sheep blood agar. Nine isolates (25.7% of total) showed different hemolytic reactions on prawn blood agar and sheep blood agar. From the 35 isolates, 8 with various hemolytic characteristics were selected and the relationship between the type of hemolytic activity and pathogenicity was determined and compared. Four isolates that showed hemolytic activity in prawn blood agar caused high mortality to cultured tiger prawns. By contrast, a significantly lower mortality rate was observed for tiger prawns injected with 4 isolates that did not exhibit hemolytic activity on prawn blood agar. Results further showed that mortality did not correlate with hemolytic activity determined using sheep blood agar. Prawn blood agar containing P. monodon hemocytes was faster and more accurate for determining prawn hemolytic activity of bacterial isolates.