Scabies is a human skin disease due to the burrowing ectoparasite var. resulting in intense itching and inflammation and manifesting as a skin allergy. Because of insufficient mite material and lack of in vitro propagation system for antigen preparation, scabies is a challenging disease to develop serological diagnostics. For allergen characterization, full-length tropomyosin (Sar s 10) was cloned, expressed in pET-15b, and assessed for reactivity with IgE antibodies from human sera. IgE binding was observed to Sar s 10 with sera collected from subjects with ordinary scabies, house dust mite (HDM)-positive and naive subjects and a diagnostic sensitivity of< 30% was observed. paramyosin (Sar s 11) was cloned, and expressed in pET-28a in three overlapping fragments designated Sspara1, Sspara2, and Sspara3. IgE and IgG binding was observed to Sspara2 and Sspara3 antigens with sera collected from ordinary scabies, and HDM-positive subjects, but no binding was observed with sera collected from naive subjects. Sspara2 displayed excellent diagnostic potential with 98% sensitivity and 90% specificity observed for IgE binding and 70% sensitivity for IgG. In contrast, the diagnostic sensitivity of Sspara3 was 84% for IgE binding and 40% for IgG binding. In combination, Sspara2 and Sspara3 provided an IgE sensitivity of 94%. This study shows that IgE binding to Sspara2 and Sspara3 is a highly sensitive method for diagnosis of scabies infestation in clinical practice. The developed enzyme-linked immunosorbent assay helps direct future development of a specific diagnostic tool for scabies.
Objective: The coding of astigmatid mites based on their morphological and developmental characteristics often leads to uncertainty in the results. The ribosomal internal transcribed spacer (ITS-2) region, being highly conserved in eukaryotes is commonly employed as a barcode for identification of mite species. The present study was an attempt to characterize the gene sequences of astigmatid mites i.e. Sarcoptes scabiei (S. scabiei), Dermatophagoides farinae (D. farinae) using ITS-2 as a genetic marker. Place and Duration of Study: The study was conducted at Department of Dermatology, Military Hospital (MH), Rawalpindi from September 2012 to October 2013. Materials and Methods: In order to characterize relationship of astigmatid mites, the ITS-2 marker was successfully amplified and sequenced. The resulting ITS-2 gene sequences were aligned using Clustal W. MEGA 7 was used to construct phylogenetic tree of the aligned sequence. Results: The phylogenetic tree showed an overall genetic distance of 0.53 indicating close genetic relationship among astigmatid mite species. Pairwise distance was calculated for the ITS-2 gene and low genetic diversity values were observed within S. scabiei and D. farinae that range from 0.003-0.008 and 0.006-0.038 respectively. Conclusion: The study supports the view that the ITS-2 region can be used to identify morphologically difficult astigmatid mites but is not useful in characterization of different species based on the geographical distribution. This study has important implication in our understanding of the epidemiology of S. scabiei and D. farinae and development of control strategies in human transmission.
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