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Feeding Cyprinus carpio with infectious materials mediates cyprinid herpesvirus 3 entry through infection of pharyngeal periodontal mucosa.

Fournier G, Boutier M, Stalin Raj V, Mast J, Parmentier E, Vanderwalle P, Peeters D, Lieffrig F, Farnir F, Gillet L, Vanderplasschen A - Vet. Res. (2012)

Bottom Line: Finally, we compared the disease induced by the two inoculation modes.They led to comparable clinical signs and mortality rate.The results of the present study suggest that, based on epidemiological conditions, CyHV-3 can enter carp either by skin or periodontal pharyngeal mucosal infection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Immunology-Vaccinology (B43b), Department of Infectious and Parasitic Diseases (B43b), Faculty of Veterinary Medicine, University of Liège, 4000 Liège, Belgium. a.vdplasschen@ulg.ac.be.

ABSTRACT
Cyprinid herpesvirus 3 (CyHV-3), also known as Koi herpesvirus, is the etiological agent of a mortal disease in common and koi carp. Recently, we investigated the entry of CyHV-3 in carp using bioluminescence imaging and a CyHV-3 recombinant strain expressing luciferase (LUC). We demonstrated that the skin is the major portal of entry after inoculation of carp by immersion in water containing CyHV-3. While this model of infection mimics some natural conditions in which infection takes place, other epidemiological conditions could favour entry of virus through the digestive tract. Here, we investigated whether ingestion of infectious materials mediates CyHV-3 entry through the digestive tract. Carp were fed with materials contaminated with the CyHV-3 LUC recombinant (oral contamination) or immersed in water containing the virus (contamination by immersion). Bioluminescence imaging analyses performed at different times post-infection led to the following observations: (i) the pharyngeal periodontal mucosa is the major portal of entry after oral contamination, while the skin is the major portal of entry after contamination by immersion. (ii) Both modes of inoculation led to the spreading of the infection to the various organs tested. However, the timing and the sequence in which some of the organs turned positive were different between the two modes of inoculation. Finally, we compared the disease induced by the two inoculation modes. They led to comparable clinical signs and mortality rate. The results of the present study suggest that, based on epidemiological conditions, CyHV-3 can enter carp either by skin or periodontal pharyngeal mucosal infection.

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In situ localization of LUC activity and detection of viral replication in carp periodontal pharyngeal mucosa. Carp weighing 100 g were fed with food pellets contaminated with the CyHV-3 LUC strain. At 2 dpi, carp were anesthetized, injected with luciferine, and euthanized immediately before dissection of the oropharyngeal cavity. Dissected fish were analysed for ex vivo bioluminescence (A). A fragment of periodontal pharyngeal mucosa emitting bioluminescence was harvested and processed for histological (B) and electron microscopy analysis (C and D). Panel C shows low magnification of the epithelium. Panel D shows one representative infected epithelial cell at higher magnification.
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Figure 3: In situ localization of LUC activity and detection of viral replication in carp periodontal pharyngeal mucosa. Carp weighing 100 g were fed with food pellets contaminated with the CyHV-3 LUC strain. At 2 dpi, carp were anesthetized, injected with luciferine, and euthanized immediately before dissection of the oropharyngeal cavity. Dissected fish were analysed for ex vivo bioluminescence (A). A fragment of periodontal pharyngeal mucosa emitting bioluminescence was harvested and processed for histological (B) and electron microscopy analysis (C and D). Panel C shows low magnification of the epithelium. Panel D shows one representative infected epithelial cell at higher magnification.

Mentions: In the present study, we investigated the role of carp digestive tract as a putative portal of entry for CyHV-3 using bioluminescence imaging. Carp were infected with the CyHV-3 LUC strain using two modes of inoculation: immersion in water containing the virus or feeding with contaminated materials (Figure 2). Fish were analysed by IVIS 24 and 48 h post-infection. Because photon emission is drastically attenuated in fish tissues [10], each fish was analysed in vivo lying on its right and then left side, and ex vivo after euthanasia and dissection. The results can be summarized as follows: in fish inoculated by immersion, 5 out of 6 fish analysed 1 day post-infection (dpi) expressed at least one focal source of light on the body surface. The signals were detected from various anatomic sites of the fish body, but principally on the fins. Analyses performed 2 dpi revealed that all of the fish had LUC signals on their surface (n = 6). In comparison to day 1, the number and the intensity of light focal sources detected 2 dpi increased in number and intensity. None of the fish inoculated by immersion expressed internal LUC signals neither at 1 dpi nor at 2 dpi. These observations confirmed our former results [10] demonstrating that the skin is the major portal of entry of CyHV-3 after inoculation by immersion in infectious water. Analysis of fish inoculated by ingestion of infectious materials led to unexpected results (Figure 2, oral inoculation mode). While none of the six fish analysed 1 dpi displayed LUC signals on the skin, one of them expressed LUC at the posterior part of the carp pharyngeal cavity. At 2 dpi, all of the analysed fish (n = 6) had intense light-emitting foci in the posterior part of the pharyngeal cavity. For 5 of the fish, no other LUC signal was detected elsewhere on or in the body (Figure 2). In addition to a strong pharyngeal signal, one single fish expressed a focal source of light on one branchial arch (data not shown). Because of the small size of the common carp used for this experiment, it was difficult to identify precisely the site of light emission within the pharyngeal cavity. Consequently this part of the experiment was repeated with larger carp (100 g, n = 5) (Figure 3).


Feeding Cyprinus carpio with infectious materials mediates cyprinid herpesvirus 3 entry through infection of pharyngeal periodontal mucosa.

Fournier G, Boutier M, Stalin Raj V, Mast J, Parmentier E, Vanderwalle P, Peeters D, Lieffrig F, Farnir F, Gillet L, Vanderplasschen A - Vet. Res. (2012)

In situ localization of LUC activity and detection of viral replication in carp periodontal pharyngeal mucosa. Carp weighing 100 g were fed with food pellets contaminated with the CyHV-3 LUC strain. At 2 dpi, carp were anesthetized, injected with luciferine, and euthanized immediately before dissection of the oropharyngeal cavity. Dissected fish were analysed for ex vivo bioluminescence (A). A fragment of periodontal pharyngeal mucosa emitting bioluminescence was harvested and processed for histological (B) and electron microscopy analysis (C and D). Panel C shows low magnification of the epithelium. Panel D shows one representative infected epithelial cell at higher magnification.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3295659&req=5

Figure 3: In situ localization of LUC activity and detection of viral replication in carp periodontal pharyngeal mucosa. Carp weighing 100 g were fed with food pellets contaminated with the CyHV-3 LUC strain. At 2 dpi, carp were anesthetized, injected with luciferine, and euthanized immediately before dissection of the oropharyngeal cavity. Dissected fish were analysed for ex vivo bioluminescence (A). A fragment of periodontal pharyngeal mucosa emitting bioluminescence was harvested and processed for histological (B) and electron microscopy analysis (C and D). Panel C shows low magnification of the epithelium. Panel D shows one representative infected epithelial cell at higher magnification.
Mentions: In the present study, we investigated the role of carp digestive tract as a putative portal of entry for CyHV-3 using bioluminescence imaging. Carp were infected with the CyHV-3 LUC strain using two modes of inoculation: immersion in water containing the virus or feeding with contaminated materials (Figure 2). Fish were analysed by IVIS 24 and 48 h post-infection. Because photon emission is drastically attenuated in fish tissues [10], each fish was analysed in vivo lying on its right and then left side, and ex vivo after euthanasia and dissection. The results can be summarized as follows: in fish inoculated by immersion, 5 out of 6 fish analysed 1 day post-infection (dpi) expressed at least one focal source of light on the body surface. The signals were detected from various anatomic sites of the fish body, but principally on the fins. Analyses performed 2 dpi revealed that all of the fish had LUC signals on their surface (n = 6). In comparison to day 1, the number and the intensity of light focal sources detected 2 dpi increased in number and intensity. None of the fish inoculated by immersion expressed internal LUC signals neither at 1 dpi nor at 2 dpi. These observations confirmed our former results [10] demonstrating that the skin is the major portal of entry of CyHV-3 after inoculation by immersion in infectious water. Analysis of fish inoculated by ingestion of infectious materials led to unexpected results (Figure 2, oral inoculation mode). While none of the six fish analysed 1 dpi displayed LUC signals on the skin, one of them expressed LUC at the posterior part of the carp pharyngeal cavity. At 2 dpi, all of the analysed fish (n = 6) had intense light-emitting foci in the posterior part of the pharyngeal cavity. For 5 of the fish, no other LUC signal was detected elsewhere on or in the body (Figure 2). In addition to a strong pharyngeal signal, one single fish expressed a focal source of light on one branchial arch (data not shown). Because of the small size of the common carp used for this experiment, it was difficult to identify precisely the site of light emission within the pharyngeal cavity. Consequently this part of the experiment was repeated with larger carp (100 g, n = 5) (Figure 3).

Bottom Line: Finally, we compared the disease induced by the two inoculation modes.They led to comparable clinical signs and mortality rate.The results of the present study suggest that, based on epidemiological conditions, CyHV-3 can enter carp either by skin or periodontal pharyngeal mucosal infection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Immunology-Vaccinology (B43b), Department of Infectious and Parasitic Diseases (B43b), Faculty of Veterinary Medicine, University of Liège, 4000 Liège, Belgium. a.vdplasschen@ulg.ac.be.

ABSTRACT
Cyprinid herpesvirus 3 (CyHV-3), also known as Koi herpesvirus, is the etiological agent of a mortal disease in common and koi carp. Recently, we investigated the entry of CyHV-3 in carp using bioluminescence imaging and a CyHV-3 recombinant strain expressing luciferase (LUC). We demonstrated that the skin is the major portal of entry after inoculation of carp by immersion in water containing CyHV-3. While this model of infection mimics some natural conditions in which infection takes place, other epidemiological conditions could favour entry of virus through the digestive tract. Here, we investigated whether ingestion of infectious materials mediates CyHV-3 entry through the digestive tract. Carp were fed with materials contaminated with the CyHV-3 LUC recombinant (oral contamination) or immersed in water containing the virus (contamination by immersion). Bioluminescence imaging analyses performed at different times post-infection led to the following observations: (i) the pharyngeal periodontal mucosa is the major portal of entry after oral contamination, while the skin is the major portal of entry after contamination by immersion. (ii) Both modes of inoculation led to the spreading of the infection to the various organs tested. However, the timing and the sequence in which some of the organs turned positive were different between the two modes of inoculation. Finally, we compared the disease induced by the two inoculation modes. They led to comparable clinical signs and mortality rate. The results of the present study suggest that, based on epidemiological conditions, CyHV-3 can enter carp either by skin or periodontal pharyngeal mucosal infection.

Show MeSH
Related in: MedlinePlus