Evaluation of a reliable non-invasive molecular test for the diagnosis of the causative agent of acute hepatopancreatic necrosis disease of shrimp

Jee Eun Han, Feng-Jyu Tang-Nelson, Patharapol Piamsomboon, Carlos R Pantoja-Morales

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Acute hepatopancreatic necrosis disease (AHPND, also known as early mortality syndrome, EMS) has caused substantial mortality, up to 100%, in populations of penaeid shrimp cultured in SE Asia and in Latin America. The disease is caused by the bacterium Vibrio parahaemolyticus, which secretes binary toxins (PirAvp and PirBvp) resulting in the deterioration of the hepatopancreas tissue of infected shrimp. Diagnosis, screening, and monitoring of AHPND in shrimp populations involve sacrificing individuals to obtain tissue samples. This sampling method is undesirable when applied to valuable populations of broodstock. Here, we evaluated a non-invasive diagnostic method based on shrimp fecal samples that are analyzed by PCR. Small groups of Pacific white shrimp Penaeus vannamei were exposed to low levels of AHPND-bacteria and their feces were collected prior to any mortality observed (in the bioassays #1 and #2). Two protocols were evaluated. In one, DNA extracted from the fecal samples was directly analyzed by PCR. In the other, the fecal samples were cultured in TSB+ for 6 h to enrich the bacterial populations, then the enriched bacterial broth was used for PCR analyses. Our results showed that the presence of V. parahaemolyticus could be detected both in fecal DNA samples and in the enriched bacterial broth, but the bands from the bacterial broth showed stronger amplification than the DNA; 12 strong positive in the enriched bacterial broth, but only 7 strong positive in the fecal DNA samples. Also, the AHPND bacteria present in the feces is infectious, determined by a bioassay of feeding specific pathogen free indicator shrimp with AHPND-feces (in the bioassay #3), and this proves that the AHPND can be transmitted through a fecal-oral route.

Original languageEnglish (US)
Pages (from-to)58-61
Number of pages4
JournalAquaculture Reports
Volume5
DOIs
StatePublished - Feb 1 2017

Fingerprint

necrosis
shrimp
feces
DNA
bioassay
mortality
bacterium
testing
Vibrio parahaemolyticus
bioassays
sampling
bacteria
toxin
amplification
pathogen
breeding stock
Penaeidae
Litopenaeus vannamei
Latin America
hepatopancreas

Keywords

  • Acute hepatopancreatic necrosis disease (AHPND)
  • Aquaculture
  • Early Mortality Syndrome (EMS)Early mortality syndrome (EMS)
  • Fecal samples
  • Feces
  • PCR
  • Shrimp
  • Vibrio parahaemolyticus

ASJC Scopus subject areas

  • Aquatic Science
  • Animal Science and Zoology

Cite this

@article{3a3730aaa8f74a1690c9ac6ae961a637,
title = "Evaluation of a reliable non-invasive molecular test for the diagnosis of the causative agent of acute hepatopancreatic necrosis disease of shrimp",
abstract = "Acute hepatopancreatic necrosis disease (AHPND, also known as early mortality syndrome, EMS) has caused substantial mortality, up to 100{\%}, in populations of penaeid shrimp cultured in SE Asia and in Latin America. The disease is caused by the bacterium Vibrio parahaemolyticus, which secretes binary toxins (PirAvp and PirBvp) resulting in the deterioration of the hepatopancreas tissue of infected shrimp. Diagnosis, screening, and monitoring of AHPND in shrimp populations involve sacrificing individuals to obtain tissue samples. This sampling method is undesirable when applied to valuable populations of broodstock. Here, we evaluated a non-invasive diagnostic method based on shrimp fecal samples that are analyzed by PCR. Small groups of Pacific white shrimp Penaeus vannamei were exposed to low levels of AHPND-bacteria and their feces were collected prior to any mortality observed (in the bioassays #1 and #2). Two protocols were evaluated. In one, DNA extracted from the fecal samples was directly analyzed by PCR. In the other, the fecal samples were cultured in TSB+ for 6 h to enrich the bacterial populations, then the enriched bacterial broth was used for PCR analyses. Our results showed that the presence of V. parahaemolyticus could be detected both in fecal DNA samples and in the enriched bacterial broth, but the bands from the bacterial broth showed stronger amplification than the DNA; 12 strong positive in the enriched bacterial broth, but only 7 strong positive in the fecal DNA samples. Also, the AHPND bacteria present in the feces is infectious, determined by a bioassay of feeding specific pathogen free indicator shrimp with AHPND-feces (in the bioassay #3), and this proves that the AHPND can be transmitted through a fecal-oral route.",
keywords = "Acute hepatopancreatic necrosis disease (AHPND), Aquaculture, Early Mortality Syndrome (EMS)Early mortality syndrome (EMS), Fecal samples, Feces, PCR, Shrimp, Vibrio parahaemolyticus",
author = "Han, {Jee Eun} and Feng-Jyu Tang-Nelson and Patharapol Piamsomboon and Pantoja-Morales, {Carlos R}",
year = "2017",
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doi = "10.1016/j.aqrep.2016.12.004",
language = "English (US)",
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pages = "58--61",
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TY - JOUR

T1 - Evaluation of a reliable non-invasive molecular test for the diagnosis of the causative agent of acute hepatopancreatic necrosis disease of shrimp

AU - Han, Jee Eun

AU - Tang-Nelson, Feng-Jyu

AU - Piamsomboon, Patharapol

AU - Pantoja-Morales, Carlos R

PY - 2017/2/1

Y1 - 2017/2/1

N2 - Acute hepatopancreatic necrosis disease (AHPND, also known as early mortality syndrome, EMS) has caused substantial mortality, up to 100%, in populations of penaeid shrimp cultured in SE Asia and in Latin America. The disease is caused by the bacterium Vibrio parahaemolyticus, which secretes binary toxins (PirAvp and PirBvp) resulting in the deterioration of the hepatopancreas tissue of infected shrimp. Diagnosis, screening, and monitoring of AHPND in shrimp populations involve sacrificing individuals to obtain tissue samples. This sampling method is undesirable when applied to valuable populations of broodstock. Here, we evaluated a non-invasive diagnostic method based on shrimp fecal samples that are analyzed by PCR. Small groups of Pacific white shrimp Penaeus vannamei were exposed to low levels of AHPND-bacteria and their feces were collected prior to any mortality observed (in the bioassays #1 and #2). Two protocols were evaluated. In one, DNA extracted from the fecal samples was directly analyzed by PCR. In the other, the fecal samples were cultured in TSB+ for 6 h to enrich the bacterial populations, then the enriched bacterial broth was used for PCR analyses. Our results showed that the presence of V. parahaemolyticus could be detected both in fecal DNA samples and in the enriched bacterial broth, but the bands from the bacterial broth showed stronger amplification than the DNA; 12 strong positive in the enriched bacterial broth, but only 7 strong positive in the fecal DNA samples. Also, the AHPND bacteria present in the feces is infectious, determined by a bioassay of feeding specific pathogen free indicator shrimp with AHPND-feces (in the bioassay #3), and this proves that the AHPND can be transmitted through a fecal-oral route.

AB - Acute hepatopancreatic necrosis disease (AHPND, also known as early mortality syndrome, EMS) has caused substantial mortality, up to 100%, in populations of penaeid shrimp cultured in SE Asia and in Latin America. The disease is caused by the bacterium Vibrio parahaemolyticus, which secretes binary toxins (PirAvp and PirBvp) resulting in the deterioration of the hepatopancreas tissue of infected shrimp. Diagnosis, screening, and monitoring of AHPND in shrimp populations involve sacrificing individuals to obtain tissue samples. This sampling method is undesirable when applied to valuable populations of broodstock. Here, we evaluated a non-invasive diagnostic method based on shrimp fecal samples that are analyzed by PCR. Small groups of Pacific white shrimp Penaeus vannamei were exposed to low levels of AHPND-bacteria and their feces were collected prior to any mortality observed (in the bioassays #1 and #2). Two protocols were evaluated. In one, DNA extracted from the fecal samples was directly analyzed by PCR. In the other, the fecal samples were cultured in TSB+ for 6 h to enrich the bacterial populations, then the enriched bacterial broth was used for PCR analyses. Our results showed that the presence of V. parahaemolyticus could be detected both in fecal DNA samples and in the enriched bacterial broth, but the bands from the bacterial broth showed stronger amplification than the DNA; 12 strong positive in the enriched bacterial broth, but only 7 strong positive in the fecal DNA samples. Also, the AHPND bacteria present in the feces is infectious, determined by a bioassay of feeding specific pathogen free indicator shrimp with AHPND-feces (in the bioassay #3), and this proves that the AHPND can be transmitted through a fecal-oral route.

KW - Acute hepatopancreatic necrosis disease (AHPND)

KW - Aquaculture

KW - Early Mortality Syndrome (EMS)Early mortality syndrome (EMS)

KW - Fecal samples

KW - Feces

KW - PCR

KW - Shrimp

KW - Vibrio parahaemolyticus

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U2 - 10.1016/j.aqrep.2016.12.004

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JO - Aquaculture Reports

JF - Aquaculture Reports

SN - 2352-5134

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