Laboratory Diagnosis of Porcine Rotavirus – Article

Rotaviruses that cause diarrhea in pigs are mainly divided into three categories: rotavirus A, rotavirus B and rotavirus C. Each type of virus is different from the others and requires specific tests designed for them. Diarrhea in pigs often involves co-infection with multiple rotaviruses.


Available tests:


Macroscopic pathology


  • Assess the walls of the small intestine for thinning, which may indicate disease.
  • Lesion site: jejunum and ileum.
  • advantage:

    • Fast and economical results
    • Applicable to all three groups of rotavirus

  • shortcoming:

    • Non-diagnostic – similar lesions may be seen in other diseases, especially coronavirus infections (transmissible gastroenteritis, porcine epidemic diarrhea, and porcine delta coronavirus)
    • Fresh tissue (<15 minutes) is required as breakdown can significantly change the appearance of the small intestine
    • Does not distinguish between different groups of rotaviruses


Villous atrophy caused by rotavirus. Photos of swine diseases by Perlezhitnitskiy, DVM, MSpVM. Source: Dr. Carlos Pijoan https://creativecommons.org/licenses/by-nc/4.0/

Villous atrophy caused by rotavirus. Swine Disease Photography: Perlezhitnitskiy, DVM, MSpVM. Source: Dr. Carlos Pijoan https://creativecommons.org/licenses/by-nc/4.0/


Histopathology


  • Assess for the presence of tissue lesions (villous atrophy of the small intestine) that may strongly suggest the presence of disease.
  • Sample type: Tissue from jejunum and ileum.
  • advantage:

    • Confirmation of villous atrophy suggesting viral enteritis

  • shortcoming:

    • Intestinal tissue needs to be fixed in formalin within 15 minutes after the animal dies.Intestinal tissue breaks down rapidly


Immunohistochemistry (IHC)


  • Detects the presence of viral antigens.
  • Sample type: Tissue from jejunum and ileum.
  • advantage:

    • Confirmation: can confirm the presence of the virus and related lesions

  • shortcoming:

    • Currently only available for rotavirus A and rotavirus C (not for rotavirus B)


Polymerase chain reaction (PCR)


  • Detects the presence of specific viral nucleic acid (RNA) sequences
  • Sample types: Jejunal and ileal intestinal tissue, fecal swabs, feces.
  • advantage:

    • Multiplex PCR detects different groups of rotaviruses, including rotavirus A, rotavirus B, and rotavirus C
    • High sensitivity.
    • PCR quantification
    • Moderate cost

      • Stool or tissue samples can often be pooled to reduce cost and minimize loss of sensitivity (especially in terms of clinical relevance).

  • shortcoming:

    • High sensitivity – can confirm the presence of the virus, but not the disease (high prevalence of the virus in the environment)


Virus isolation


  • Isolation of live virus.
  • Sample types: Jejunal and ileal intestinal tissue, fecal swabs, feces.
  • advantage:

    • Isolating viruses for vaccine development (autologous vaccines)

  • shortcoming:

    • expensive
    • slow results
    • Currently only available for rotavirus A, not rotavirus B or rotavirus C
    • Often difficult to culture (many false negatives)
    • Sample handling from field to laboratory may affect virus survival


Enzyme-linked immunosorbent assay (ELISA)


  • Detects the presence of antibodies.
  • Sample type: serum.
  • advantage:

    • Animals remain positive for weeks
    • Negative farms can be identified

  • shortcoming:

    • High prevalence of the virus in the environment results in widespread exposure of many/most pigs to the virus
    • Specific testing is required for each rotavirus group
    • There is no cross-protection between different rotavirus groups.


Table 1. Summary of applicable tests for different rotavirus groups.











Rotavirus A Rotavirus B Rotavirus C Does it allow you to differentiate between different groups?
Macroscopic pathology Yes Yes Yes No
Histopathology Yes Yes Yes No
Immunohistochemistry (IHC) Yes No Yes Yes
Polymerase chain reaction (PCR) Yes Yes Yes Yes
Virus isolation Yes No No No
Enzyme-linked immunosorbent assay (ELISA) Yes No No No


Interpretation of results:


Macroscopic pathology


  • Positive: Preliminary confirmation of the presence of intestinal thinning.
  • Negative: Early or mild cases may not show extensive intestinal pathology.


Histopathology


  • Positive: The disease is confirmed, but the cause of the disease is not confirmed.
  • Negative: Negative or undetectable lesions if the wrong sample is tested or if the test takes too long after infection.


Immunohistochemistry


  • Positive: Virus is present at the injured site.
  • Negative: If the test is done a long time after infection or if the virus concentration is too low (it is only present for a short period of time), it is negative or no virus is detected.


polymerase chain reaction


  • Positive: Confirmation of the presence of different rotavirus groups (A, B and C).
  • Negative: Negative if the test is done long after infection or if the virus is not detected due to incorrect sample selection or handling.


Virus isolation


  • Positive: Confirmation that the virus is present, but no disease is present.
  • Negative: Negative, the virus cannot be detected if the test is done long after infection, or it cannot grow due to other contamination or mishandling (it is present but difficult to culture). Does not detect rotavirus B or rotavirus C.


Enzyme-linked immunosorbent assay


  • Positive: Previous exposure to vaccine or wild virus (>2-4 weeks). There is no correlation between positivity and disease.
  • Negative:

    • Negative for vaccine or wild virus
    • Infection too recent to detect


Application scenarios:


Diarrhea in newborn piglets (acute or chronic)


  • Fecal samples were collected from 10 or more untreated pigs with diarrhea and PCR assayed in groups of 5 for rotavirus A, rotavirus B, and rotavirus C.
  • Necropsy 1-3 recently deceased pigs or euthanize pigs suffering from diarrhea. Macroscopic assessment of intestinal wall thinning in the jejunum and ileum. Samples were collected and fixed in formalin for histopathology and IHC.


Diarrhea in newly weaned piglets (acute or chronic)


  • Fecal samples were collected from 10 or more untreated pigs with diarrhea and PCR assayed in groups of 5 for rotavirus A, rotavirus B, and rotavirus C.
  • Necropsy 1-3 recently deceased pigs or euthanize pigs suffering from diarrhea. Macroscopic assessment of intestinal wall thinning in the jejunum and ileum. Samples were collected and fixed in formalin for histopathology and IHC.

Source link

Leave a Comment