Wisnu Nurcahyo
Bagian Klinik Hewan, Fakultas Kedokteran Hewan, Universitas Udayana, Bali

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Journal : Jurnal Veteriner

Molecular Detection of Toxoplasmosis Using Specific Primers P30, B1, and rDNA

Jurnal Veteriner Vol 13, No 1 (2012)
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Abstract

Study in order to develop molecular techniques using specific primers for the early diagnosis oftoxoplasmosis have been conducted. Detection of Toxoplasma gondii genome was performed usingpolymerase chain reaction (PCR) technique. The primers used in this study were rDNA, P30, and B1. ThePCR products were further run using gel electrophoresis (gel 1.5% – 2.0%) and the band was documented.Toxoplasma was detected at 500 bp and 600 bp using primer P30 and B1, respectively. Whereas usingprimer rDNA no band was observed. It was assumed that primer rDNA was not sensitive since the targetamplification was 88 bp.

Produksi dan Isolasi Protein Membran Stadium Bradizoit Toxoplasma gondii : Suatu Usaha untuk Mendapatkan Material Diagnostik dalam Mendiagnosa Toksoplasmo

Jurnal Veteriner Vol 10, No 3 (2009)
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A study was conducted to isolate the membrane protein of Toxoplasma gondii at bradyzoid stage forthe development of intradermal diagnostic test in livestock infected by the parasite. Toxoplasma wasinitially collected from meat of goat serologically positive to the parasite. The infected meat was then fedinto uninfected cat to obtain oocyst. The oocyst was inoculated into the stomach of mice to produce tachyzoitwhich in turn produce cyst in tissue known as bradyzoit . The membrane protein was then isolated from thebradyzoit. The protein was analysed by sodium dodecyl sulfate electrophoresis (SDS-PAGE). The dataobtained were presented descriptively. The protein concentration isolated from each mouse infected at thedose of 1x107 oocysts was 11.91 mg. Two protein bands specific for bradyzoit were identified at 97.72 kDaand 67.60 kDa.

Profil Protein Stadium Sporozoit Eimeria tenella Isolat Yogyakarta Melalui Analisis Protein SDS-PAGE (PROTEIN PROFILE OF THE SPOROZOITE OF Eimeria tenella ISOLATES FROM YOGYAKARTA USING SDS-PAGE PROTEIN ANALYSIS)

Jurnal Veteriner Vol 13, No 2 (2012)
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Coccidiosis is one of the important diseases in poultry industry. In Indonesia the morbidity of thedisease is between 80 to 90%. A rapid and prompt diagnosis would be one of the essential steps ineradication and control of the disease. The objective of this study is to determine the protein profile ofsporozoite of Eimeria tenella isolated in Yogyakarta using sodium dodecyl sulphate polyacrilamide gelelectrophoresis (SDS-PAGE) protein analysis. Protein analysis was performed in 12% polyacrilamide geland further electrophoresis at 100 volts and over-staining with Coomasie brilliant blue. The resultsshowed that the sporozoite of E. tenella isolated in Yogyakarta contained five proteins with molecularweights of 15, 26, 32, 80, and 91 kDa, respectively.

Penggunaan Protein Membran Stadium Bradizoit Toxoplasma gondii untuk Mendiagnosis Toksoplasmosis dengan Metode Intradermal Test

Jurnal Veteriner Vol 14, No 2 (2013)
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A research was conducted to find out an alternative diagnose in detecting toxoplasmosis in livestock/animal using intradermal test  from protein membrane of  T. gondii bradizoite stage. Local isolate ofmembrane T. gondii bradizoite stage  was used in the research. Ten of domestic sheep with the age of ± 1year and 10 mice strain Balb/c with the age ± 2 month were used in this research. The reaction ofhypersensitivity on the skin post protein membrane bradyzoite injection was indicated by the process ofskin thickening. The diameter skin thickening was measured using cutimeter, in which diameter e” 10 mmindicated positive diagnose. The result showed that optimal dosage of membran protein bradyzoite thatcould be applied to detect toxoplasmosis in livestock and animal using intradermal test were 0,6 ml and0,2 ml for sheep and mice respectively. The sensitivity and specificity level of antigen use (protein membrane)of T. gondii bradizoite stage from local isolate to diagnose toxoplasmosis in mice using intradermal testwere: 85.0 %  and 66.6 % respectively, while in sheep the sensitivity and specificity level were 85.0 % and66.6 % respectively. It can be concluded that  intradermal  test was appropriate to be implemented fordetecting toxoplasmosis in sheep and mice induced with tachyzoite T. gondii.

Pengembangan Antibodi Poliklonal dari Stadium Oosista, Sporosista, dan Sporozoit Eimeria tenella (THE DEVELOPMENT OF POLYCLONAL ANTOBODY FROM EIMERIA TENELLA OOCYST, SPOROCYST, AND SPOROZOITE STADIUM)

Jurnal Veteriner Vol 14, No 2 (2013)
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The research on developing diagnostic method, vaccine, and drugs for coccidiosis has been focused onthe finding of the immunogenic molecule in Eimeria. The identification of this agent will need the antibodywhich can recognize the biomolecule in the antigen. Antibody that has been developed for this purposeshould be analyzed first, and one of the simple methods for analyzing this antibody is through dot blotanalysis. The objective of this research was to analyze the polyclonal antibody which developed from theoocyst, sporocyst, and sporozoite of  E. tenella using dot blot analysis. The antigen for this polyclonalantibody was made from each of the E. tenella stadium by sonication. Fifteen mice, divided into 3 groups,were then injected subcutaneously with each antigen. The sera from these mice were then collected, analyzedby using ELISA and then it will be used for the dot blot analysis. The research result showed that thepolyclonal antibody which has been developed in mice from each antigen can react with the antigen itself.From this result it can be concluded that the developing of this antibody is successful and it can be used forfurther research in immunoproteomic.

Strongyloides spp Distribution on Orangutans in Tanjung Putting National Park, Care Center in Pangkalanbun, and Sebangau National Park

Jurnal Veteriner Vol 14, No 2 (2013)
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Strongyloides spp is a parasitic nematode in livestock, primate and human which is  considered asa danger zoonotic disease. Therefore, study about parasite distribution is very important in order to find outgenetic diversity among orangutan in quarantine, zoo and nature, as an effort to explore infection patternand life cycle of Strongyloides spp on orangutan. Amount of 326 orangutan feces were taken from threedifferent habitat of orangutan in Central Borneo, Tanjung Puting National Park, Orangutan Care Centerand Sebangau National Park. Samples which were collected from Tanjung Puting, Care Center and Sebangauwere 75, 80 and 171 respectively. Those samples were transported to the Parasitology laboratory in Facultyof Veterinary Medicine, Gadjah Mada University, Yogyakarta for examination and detection.  Prevalence ofstrongyloides in Tanjung Putting, Sebangau and Orangutan Care Center were 24%, 14,6% and 13,3%respectively. Among positive samples of Strongyloides, 62,5% were from male orangutans, while 37,5% werefrom female orangutans. Strongyloides in pre adult and baby orangutan were 91,6% and 4,2% respectively.Meanwhile, Strongyloides in adult orangutan were very rare. Orangutan habitat in Sebangau National Parkis an ideal habitat for orangutan, supported by the watery condition of peat land, so that Strongyloides re-infection become difficult. Some factors may have important role in Strongyloidoses, such as behavior,physical condition, nutrition, age, body weight, sex, immunity and social status of orangutan.

Antigen Ekskretori-Sekretori Cacing Jantung (Dirofilaria immitis) Jantan dan Betina yang Berpotensi Sebagai Marka Diagnosis (EXCRETORY-SECRETORY ANTIGENS OF MALE AND FEMALE HEART WORMS (DIROFILARIA IMMITIS) WHICH POTENTIALLY AS A DIAGNOSTIC MARKER)

Jurnal Veteriner Vol 16, No 4 (2015)
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Heart worm (Dirofilaria immitis) is the causative agent of a serious parasitic disease in dogs.Dirofilariasis is generally diagnosed by microfilariae examination and specific antigen testing. Microfilariaeexamination has low sensitivity due to occult infections. The available antigen test at this time is able todetect circulating antigens secreted by adult female worms only. The aim of the present study was toidentify male (MES) and female (FES) heart worms excretory-secretory antigens which have the potentialas a diagnostic targets. Identification of antigen was done by sodium dodecyl sulphate polyacrylamide gelelectrophoresis (SDS-PAGE) and Western Blotting analysis. The results of this study indicated that therewere differences between the MES and the FES profiles. The results showed 12 bands in MES (14–118kDa) and 18 bands in FES (10–205 kDa). Protein with a molecular weight of 59 kDa has the potential asdiagnostic markers of dirofilariasis.

Kekerabatan Genetik Caplak Rhiphicephalus (Boophilus) microplus Asal IndonesiaBerdasarkan Sekuen Internal Transcribed Spacer-2 (GENETIC RELATIONSHIP INDONESIAN RHIPHICEPHALUS (BOOPHILUS) MICROPLUS TICK BASED ON INTERNAL TRANSCRIBED SPACER-2 SEQUENSE )

Jurnal Veteriner Vol 16, No 3 (2015)
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Rhiphicephalus (Boophilus) microplus is important obligatory blood feeding ectoparasites transmittingmany different viral, bacterial and protozoan and plays a role as a vector of Babesia sp., The leria sp. andAnaplasma sp. in cattle. The accuracy in identifying and distinguishing interspecies and intraspeciesdiversity among parasites is needed to understand the epidemiology, biology and capacity as a vector.Variations in the DNA base sequence of the internal transcribed spacer region2 (ITS 2) has been used asa molecular marker for identification in an effort to determine phylogenetic relationships. The aim of thisstudy was to determine the ITS 2 gene nucleotide sequence of R. microplus, which was expected to beuseful for accurate identification the parasite diversity and phylogenetic relationship among many differentspecies. DNA amplification was conducted using BOO2 forward dan BOO2 reverse primers. The DNAsamples containing ITS2 region fragment of 1099 nt were derived from the nucleotide sequence multiplealignments of R.microplus and other ticks genes obtained from Gene bank using Clustal W software, andthen analyzed using the MEGA program version 6. Genetic distances based on nucleotide sequence weredetermined with Kimura 2-parameter method producing the smallest genetic distance of 0 % and 1.2 %.Construction of phylogenetic trees using the Neighbor joining method showed that ticks from variousregions in Indonesia was species complex which have a closer with R.microplus isolates from India, Laos,South Africa, China and Australia R.australis origin.