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Background and Aim: In several countries, two Entamoeba porcine species, Entamoeba suis and Entamoeba polecki (subtype 1 and 3), have been detected using molecular methods and identified pathogenicity associated with enteritis. However, globally, Entamoeba infection prevalence in pigs is extremely limited. This study aimed to coprologically and genetically examine pig parasites to estimate prevalence of Entamoeba in three pig farms in East Java, Indonesia. Materials and Methods: Hundred porcine fecal samples (Landrace) were collected from three East Javan farms in well-known swine industry regions. Fecal samples were examined under a microscope after sugar-flotation centrifugation, and molecular species and subtype identification were performed using polymerase chain reaction (PCR) and primer pairs targeting small-subunit ribosomal RNA. Results: Microscopy examinations identified parasites in 89/100 fecal samples; Entamoeba spp. cysts were the most frequent in these samples. Polymerase chain reaction showed that 58 samples were comprised of mixed Entamoeba suis and Entamoeba polecki, 22 E. suis alone, and nine E. polecki alone infections. Epolec F6-Epolec R6 primers successfully amplified E. polecki ST1-4 subtypes, while Epolecki 1-Epolecki 2 amplified only the E. polecki ST1 subtype. Entamoeba polecki ST1-specific primers successfully detected the ST1 subtype in 19/67 E. polecki positive samples. Conclusion: Entamoeba spp. prevalence in Indonesian pigs was previously shown to be high. On coprological examination of East Javan pigs, we detected high Entamoeba spp. levels, in which we genetically identified as E. suis (80.0%), E. polecki (67.0%), and E. polecki ST1 (19%).
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AIM: This study aimed to sequence the Cytochrome c oxidase (COX-1) gene sequence from mitochondrial DNA of Sarcoptes scabiei isolated from Lamongan goats and Mojokerto rabbits, align it with DNA isolated from Zi'gong rabbit (GenBank accession No. EU256389.1), and produce a phylogenetic analysis of S. scabiei COX-1 gene. MATERIALS AND METHODS: S. scabiei mites were obtained from goats and rabbits, and DNA was extracted using QIAamp DNA Mini Kit. The forward and reverse primer sequences were designed based on the DNA sequence of an S. scabiei COX-1 gene isolated from the Zi'gong rabbit (5'-TCTTAGGGGCTGGATTTAGTATG-3' and 5'-AGTTCCTCTACCAGTTCCAC-3', respectively). To confirm sequencing output, the sequence resulting from the reverse primer was inverted and aligned to the sequence from the forward primer using Clone Manager Professional Version 9 for Windows (Scientific & Educational Software; http://www.scied.com). This alignment was subsequently used to build a phylogenetic tree, using the Neighbor-Joining method, in the MEGA6 program (https://www.megasoftware.net/). RESULTS: Polymerase chain reaction (PCR) products from S. scabiei isolates from Lamongan goats and Mojokerto rabbits produced bands of around 290 bp with 2% agarose gel electrophoresis. Comparing the DNA sequences of the S. scabiei COX-1 gene with those isolated from Lamongan goats and Mojokerto rabbits showed 99% homology. CONCLUSION: PCR products of the S. scabiei COX-1 gene isolated from Lamongan goats and Mojokerto rabbits were around 290 bp long. The sequences had more than 99% homology. The sequences of the COX-1 gene of S. scabiei from Lamongan goats and Mojokerto rabbits were relatively close to the sequence of the gene in S. scabiei obtained from various hosts according to National Center for Biotechnology Information data.
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BACKGROUND: Scabies or mange is an infectious skin disease caused by the mite Sarcoptes scabiei. This skin disease affects various livestock such as goats, sheep, swine, cattle, other animals like dogs, cats, wild animals and also affect human. This research aimed to explore the protein in mites S. scabiei which has antigenic character and play roles in scabies pathogenesis in goats and rabbits. METHODS: S.scabiei mites were isolated from goats and rabbits, and characterized using SDS-PAGE. In addition the protein was also analysed using Western Blot assay. The isolation and identification were carried out in 2015 at the Parasitology Laboratory of Veterinary Medicine Faculty, Universitas Airlangga, Surabaya, Indonesia. RESULTS: The identification results using SDS-PAGE of mites S. scabiei var. caprae expressed 12 protein bands between 26,7 kDa and 205,8 kDa, continued by Western Blot showed 3 protein bands, after being reacted with blood serum from scabies infected goat, it could be identified antigenic protein with molecule weight 205.8 kDa, 57.3 kDa, and 43 kDa. While protein in mites S. scabiei var. cuniculi identified 9 protein bands between 24 kDa and 75 kDa by SDS-PAGE, and the Western Blot assay identified antigenic protein with molecule weight 62 kDa and 51 kDa. CONCLUSION: The antigenic protein of S. scabiei var. caprae and S. scabiei var. cuniculi showed that they are probably involved in the scabies pathogenesis in goats and rabbits.
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AIM: Scabies is one of the most important diseases in goats and caused by a complex hypersensitivity process that involves both humoral and cell-mediated immune responses. This phenomenon shows that the variety of Sarcoptes scabiei has different characteristics of specific antigenic protein or different immune-dominant. This research aims to detect the humoral and cellular immune response of rabbits which were immunized with the protein of S. scabiei var. caprae. MATERIALS AND METHODS: This research was done as follows, identification and collection of Sarcoptes scabiei var caprae from goat infected with scabies, separation of protein antigen from S. scabiei mites with ultrasonic sonicator, measurement of protein content with spectrophotometry, rabbit injection with 500 µg dose of antigen protein which was repeated 5 times (5x booster) every 2 weeks. Measurement of IgG titer using indirect ELISA, whereas to detect the expression of cellular immune response (TLR-9, CD4, and CD8) using Direct Immunofluorescence assay. RESULTS: Based on the statistical analysis, it showed that there was a significant enhancement (p<0.05) for optical density value or antibody titer and cellular immune response was shown by TLR-9, CD4, and CD8 expression in rabbit T lymphocytes which appear yellow to green fluorescent color using fluorescence microscope. The amount of fluorescence T lymphocytes showed a significant difference (p<0.05) between the control and various boosters. CONCLUSION: Antigenic protein of S. scabiei var. caprae contains ligands, which are involved in the pathogen-associated molecular pattern that has an ability to induce humoral and cellular immune response in rabbit. Specifically, that TLR-9 is not only involved in innate immunity but also in adaptive immunity and can be used as alternative adjuvant development research.