Abstract
Diagnostic methods were used to identify and quantify Myxobolus cerebralis, a myxozoan parasite of salmonid fish. In this study, 7-week-old, pathogen-free rainbow trout (Oncorhynchus mykiss) were experimentally infected with M. cerebralis and at 7 months postinfection were evaluated with 5 diagnostic assays: 1) pepsin-trypsin digest (PTD) to detect and enumerate spores found in cranial cartilage, 2) 2 different histopathology grading scales that provide a numerical score for severity of microscopic lesions in the head, 3) a conventional single-round polymerase chain reaction (PCR), 4) a nested PCR assay, and 5) a newly developed quantitative real-time TaqMan PCR. There were no significant differences (P > 0.05) among the 5 diagnostic assays in distinguishing between experimentally infected and uninfected control fish. The 2 histopathology grading scales were highly correlated (P < 0.001) for assessment of microscopic lesion severity. Quantification of parasite levels in cranial tissues using PTD and real-time TaqMan PCR was significantly correlated r = 0.540 (P < 0.001). Lastly, 104 copies of the 18S rDNA gene are present in the M. cerebralis genome, a feature that makes this gene an excellent target for PCR-based diagnostic assays. Also, 2 copies of the insulin growth factor-I gene are found in the rainbow trout genome, whose detection can serve both as an internal quality control for amplifiable DNA and as a basis to quantify pathogen genome equivalents present in quantitative PCR assays.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 202-211 |
| Number of pages | 10 |
| Journal | Journal of Veterinary Diagnostic Investigation |
| Volume | 16 |
| Issue number | 3 |
| DOIs | |
| State | Published - Jan 1 2004 |
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ASJC Scopus subject areas
- veterinary(all)
Cite this
Evaluation of five diagnostic methods for the detection and quantification of Myxobolus cerebralis. / Kelley, Garry O.; Zagmutt-Vergara, Francisco J.; Leutenegger, Christian M.; Myklebust, Karin A.; Adkison, Mark A.; McDowell, Terry S.; Marty, Gary D.; Kahler, Alex L.; Bush, Arla L.; Gardner, Ian; Hedrick, Ronald.
In: Journal of Veterinary Diagnostic Investigation, Vol. 16, No. 3, 01.01.2004, p. 202-211.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Evaluation of five diagnostic methods for the detection and quantification of Myxobolus cerebralis
AU - Kelley, Garry O.
AU - Zagmutt-Vergara, Francisco J.
AU - Leutenegger, Christian M.
AU - Myklebust, Karin A.
AU - Adkison, Mark A.
AU - McDowell, Terry S.
AU - Marty, Gary D.
AU - Kahler, Alex L.
AU - Bush, Arla L.
AU - Gardner, Ian
AU - Hedrick, Ronald
PY - 2004/1/1
Y1 - 2004/1/1
N2 - Diagnostic methods were used to identify and quantify Myxobolus cerebralis, a myxozoan parasite of salmonid fish. In this study, 7-week-old, pathogen-free rainbow trout (Oncorhynchus mykiss) were experimentally infected with M. cerebralis and at 7 months postinfection were evaluated with 5 diagnostic assays: 1) pepsin-trypsin digest (PTD) to detect and enumerate spores found in cranial cartilage, 2) 2 different histopathology grading scales that provide a numerical score for severity of microscopic lesions in the head, 3) a conventional single-round polymerase chain reaction (PCR), 4) a nested PCR assay, and 5) a newly developed quantitative real-time TaqMan PCR. There were no significant differences (P > 0.05) among the 5 diagnostic assays in distinguishing between experimentally infected and uninfected control fish. The 2 histopathology grading scales were highly correlated (P < 0.001) for assessment of microscopic lesion severity. Quantification of parasite levels in cranial tissues using PTD and real-time TaqMan PCR was significantly correlated r = 0.540 (P < 0.001). Lastly, 104 copies of the 18S rDNA gene are present in the M. cerebralis genome, a feature that makes this gene an excellent target for PCR-based diagnostic assays. Also, 2 copies of the insulin growth factor-I gene are found in the rainbow trout genome, whose detection can serve both as an internal quality control for amplifiable DNA and as a basis to quantify pathogen genome equivalents present in quantitative PCR assays.
AB - Diagnostic methods were used to identify and quantify Myxobolus cerebralis, a myxozoan parasite of salmonid fish. In this study, 7-week-old, pathogen-free rainbow trout (Oncorhynchus mykiss) were experimentally infected with M. cerebralis and at 7 months postinfection were evaluated with 5 diagnostic assays: 1) pepsin-trypsin digest (PTD) to detect and enumerate spores found in cranial cartilage, 2) 2 different histopathology grading scales that provide a numerical score for severity of microscopic lesions in the head, 3) a conventional single-round polymerase chain reaction (PCR), 4) a nested PCR assay, and 5) a newly developed quantitative real-time TaqMan PCR. There were no significant differences (P > 0.05) among the 5 diagnostic assays in distinguishing between experimentally infected and uninfected control fish. The 2 histopathology grading scales were highly correlated (P < 0.001) for assessment of microscopic lesion severity. Quantification of parasite levels in cranial tissues using PTD and real-time TaqMan PCR was significantly correlated r = 0.540 (P < 0.001). Lastly, 104 copies of the 18S rDNA gene are present in the M. cerebralis genome, a feature that makes this gene an excellent target for PCR-based diagnostic assays. Also, 2 copies of the insulin growth factor-I gene are found in the rainbow trout genome, whose detection can serve both as an internal quality control for amplifiable DNA and as a basis to quantify pathogen genome equivalents present in quantitative PCR assays.
UR - http://www.scopus.com/inward/record.url?scp=2442621210&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=2442621210&partnerID=8YFLogxK
U2 - 10.1177/104063870401600305
DO - 10.1177/104063870401600305
M3 - Article
C2 - 15152834
AN - SCOPUS:2442621210
VL - 16
SP - 202
EP - 211
JO - Journal of Veterinary Diagnostic Investigation
JF - Journal of Veterinary Diagnostic Investigation
SN - 1040-6387
IS - 3
ER -