Multiplex PCR for rapid diagnosis and differentiation of pox and pox-like diseases in dromedary Camels.

BACKGROUND: Pox and pox-like diseases of camels are a group of exanthematous skin conditions that have become increasingly important economically. Three distinct viruses may cause them: camelpox virus (CMLV), camel parapox virus (CPPV) and camelus dromedary papilloma virus (CdPV). These diseases are often difficult to differentiate based on clinical presentation in disease outbreaks. Molecular methods such as PCR targeting species-specific genes have been developed and used to identify these diseases, but not simultaneously in a single tube. Recently, multiplex PCR has gained reputation as a convenient diagnostic method with cost-and timesaving benefits. METHODS AND RESULTS: In the present communication, we describe the development, optimization and validation of a multiplex PCR assay able to detect simultaneously the genome of the three viruses in one single test allowing for rapid and efficient molecular diagnosis. The assay was developed based on the evaluation and combination of published and new primer sets and was validated with viral genomic DNA extracted from known virus strains (n = 14) and DNA extracted from homogenized clinical skin specimens (n = 86). The assay detects correctly the target pathogens by amplification of targeted genes, even in case of co-infection. The method showed high sensitivity, and the specificity was confirmed by PCR-product sequencing. CONCLUSION: This assay provide rapid, sensitive and specific method for identifying three important viruses in specimens collected from dromedary camels with varying clinical presentations.

Phylogenetic analysis of eight sudanese camel contagious ecthyma viruses based on B2L gene sequence.

BACKGROUND: Camel contagious ecthyma (CCE) is an important viral disease of camelids caused by a poxvirus of the genus parapoxvirus (PPV) of the family Poxviridae. The disease has been reported in west and east of the Sudan causing economical losses. However, the PPVs that cause the disease in camels of the Sudan have not yet subjected to genetic characterization. At present, the PPV that cause CCE cannot be properly classified because only few isolates that have been genetically analyzed. METHODS AND RESULTS: PCR was used to amplify the B2L gene of the PPV directly from clinical specimens collected from dromedary camels affected with contagious ecthyma in the Sudan between 1993 and 2013. PCR products were sequenced and subjected to genetic analysis. The results provided evidence for close relationships and genetic variation of the camel PPV (CPPV) represented by the circulation of both Pseudocowpox virus (PCPV) and Orf virus (ORFV) strains among dromedary camels in the Sudan. Based on the B2L gene sequence the available CPPV isolates can be divided into two genetic clades or lineages; the Asian lineage represented by isolates from Saudi Arabia, Bahrain and India and the African lineage comprising isolates from the Sudan. CONCLUSION: The camel parapoxvirus is genetically diverse involving predominantly viruses close to PCPV in addition to ORFVs, and can be divided into two genetically distant lineages. Based on sequences of the B2L gene it is not possible to suggest that the viruses that cause CCE form a monophylogenetic group or species within the PPV phylogeny.