Nastiti Wijayanti

Analysis of viral protein-2 encoding gene of avian encephalomyelitis virus from field specimens in Central Java region, Indonesia.

Aim: Avian encephalomyelitis (AE) is a viral disease which can infect various types of poultry, especially chicken. In Indonesia, the incidence of AE infection in chicken has been reported since 2009, the AE incidence tends to increase from year to year. The objective of this study was to analyze viral protein 2 (VP-2) encoding gene of AE virus (AEV) from various species of birds in field specimen by reverse transcription polymerase chain reaction (RT-PCR) amplification using specific nucleotides primer for confirmation of AE diagnosis. Materials and Methods: A total of 13 AEV samples are isolated from various species of poultry which are serologically diagnosed infected by AEV from some areas in central Java, Indonesia. Research stage consists of virus samples collection from field specimens, extraction of AEV RNA, amplification of VP-2 protein encoding gene by RT-PCR, separation of RT-PCR product by agarose gel electrophoresis, DNA sequencing and data analysis. Results: Amplification products of the VP-2 encoding gene of AEV by RT-PCR methods of various types of poultry from field specimens showed a positive results on sample code 499/4/12 which generated DNA fragment in the size of 619 bp. Sensitivity test of RT-PCR amplification showed that the minimum concentration of RNA template is 127.75 ng/µl. The multiple alignments of DNA sequencing product indicated that positive sample with code 499/4/12 has 92% nucleotide homology compared with AEV with accession number AV1775/07 and 85% nucleotide homology with accession number ZCHP2/0912695 from Genbank database. Analysis of VP-2 gene sequence showed that it found 46 nucleotides difference between isolate 499/4/12 compared with accession number AV1775/07 and 93 nucleotides different with accession number ZCHP2/0912695. Conclusions: Analyses of the VP-2 encoding gene of AEV with RT-PCR method from 13 samples from field specimen generated the DNA fragment in the size of 619 bp from one sample with sample code 499/4/12. The sensitivity rate of RT-PCR is to amplify the VP-2 gene of AEV until 127.75 ng/µl of RNA template. Compared to Genbank databases, isolate 499/4/12 has 85% and 92% nucleotide homology.

DC-SIGN (CD209) carbohydrate recognition domain is not polymorphic in dengue virus-infected Indonesian patients.

Dengue virus (DENV) infection is a significant burden in Indonesia and other tropical countries. DENV infection has a wide spectrum of clinical manifestations, i.e. asymptomatic, dengue fever, dengue hemorrhagic fever and dengue shock syndrome. The variety of clinical manifestations may be due to the diversity of genetic constitution of the host. The C-type lectin DC-SIGN (CD209) has been identified as the major dengue receptor on human dendritic cells. There are at least five polymorphisms in exon 5 and 6 of the DC-SIGN encoded gene which have been identified and recorded in dbSNP. The aim of this work is to measure the frequency of these polymorphisms among asymptomatic and hospitalized DENV-infected patients. We enrolled 23 hospitalized and 73 asymptomatic DENV-infected patients. Among the subjects, we performed PCR amplification and DNA direct seqencing for 23 hospitalized DENV-infected patients and 24 asymptomatic DENV-infected patients. The result showed that there were no polymorphic nucleotides in the CD209 encoded gene among the patients.

Assessment of vertical dengue virus transmission in Aedes aegypti and serotype prevalence in Bantul, Indonesia

Abstract Objective To determine the reality of dengue virus (DENV) serotype circulation in Bantul and the potential impact of a vertical transmission in the maintenance of dengue. Methods Mosquitoes were captured using ovitraps in the vicinity of households of patients diagnosed with severe forms of dengue. DENV was detected in mosquitoes by immunochemistry and typed by reverse transcription-polymerase chain reaction. Results Aedes aegypti mosquitoes infected with DENV were found in 14 out of 17 districts in the Bantul Regency. Vertical transmission was demonstrated and serotype prevalence was coherent with the current clinical situation and the previous reports. DENV-3 was the most common serotype (12 districts), followed by DENV-2 (3 districts) and DENV-4 (1 district). No DENV-1 was found. Conclusions Unlike continental countries such as Cambodia or Thailand, where the replacement of serotypes is the rule, maintenance of DENV-3 is a major feature in Java. Vertical transmission is likely to play a major role along with the archipelago structure of Indonesia, which might help maintaining local mosquito populations. Regular survey of circulating DENV and prevalence will help predicting and controlling outbreaks.

The neuroprotective effect of Ocimum sanctum Linn. ethanolic extract on human embryonic kidney-293 cells as in vitro model of neurodegenerative disease

Aim: This study aimed to analyze the neuroprotective effect of Ocimum sanctum Linn. ethanolic extract (OSE) on human embryonic kidney-293 (HEK-293) cells as the in vitro model of neurodegenerative diseases. Materials and Methods: In this research, HEK-293 cells divided into five groups consisting of normal and healthy cells (NT), cells treated with Camptothecin 500 µM as the negative control, cells treated with trimethyltin 10 µM (TMT), cells treated with OSE 75 µg/ml, and cells pre-treated with OSE 75 µg/ml then induced by TMT 10 µM (OSE+TMT). MTT assay and phase contrast microscopy were applied to observe the cell viability quantitatively and morphological after Ocimum sanctum Linn extract treatment. Finally, the reverse transcription polymerase chain reaction was employed to study the expression of choline acetyltransferase (ChAT). Results: The MTT assay and phase contrast microscopy showed that OSE pre-treatment significantly increased the viability of TMT-induced apoptotic cells and maintained cell viability of the normal HEK-293 cells. Expression of ChAT markedly reduced on TMT treatment group, but OSE administration stabilized ChAT expression in TMT-induced HEK-293 cells. Conclusion: This present study proved that OSE administration has neuroprotective effect by increased HEK-293 cells viability and maintain ChAT expression.

Phylogenetic Analysis of Newcastle Disease Virus from Indonesian Isolates Based on DNA-Sequence of Fusion Protein-Encoding Gene

Background and Objective: Newcastle disease (ND) is an infectious disease caused by ND virus (NDV), which is the major problem in poultry industries. Although vaccination program has been executed in Indonesia, Newcastle disease is still infecting chicken. The lack of vaccine protection against the disease, presumably due to genetic differences between vaccine strain and virus strain in the field. This study was conducted to analyze phylogenetic of Indonesian NDV isolates based on fusion (F) protein-encoding gene, with aim to determine which genotype group of Indonesian NDV isolates, compared to vaccine strain that circulating in Indonesia. Materials and Methods: Samples were RNA genome of NDV, which were isolated from chicken in Kartosuro, Karanganyar; Galur, Kulon Progo and Sukomoro, Magetan areas. The F gene was amplified by one step RT-PCR method and then was sequenced. The genetic variation among samples, other Indonesian isolates, LaSota and B1 vaccine strain were analyzed by pairwise distance method. Phylogenetic tree based on F gene sequence was generated by Neighbor-Joining method (1000 bootstrap repetition) and Kimura-2 parameter model. Result: The genetic variation of F gene nucleotide and F protein amino acid between samples and vaccine strains were higher compared to the variation between samples and other Indonesian isolates. Phylogenetic analysis showed that the samples were included in genotype VII class II, while LaSota and B1 vaccine strain included in genotype II class II. Conclusion: There were differences of fusion protein antigen and genotype group between Indonesian strains and vaccine strains. This finding encouraged to develop ND vaccine based on Indonesian isolates.

PEMERIKSAAN VIRUS DENGUE-3 PADA NYAMUK Aedes aegypti YANG DIINFEKSI SECARA INTRATHORAKAL DENGAN TEKNIK IMUNOSITOKIMIA MENGGUNAKAN ANTIBODI DSSE10

ABSTRACT Dengue viruses, globally the most prevalent arboviruses, are transmitted to humans by persistently infected Aedes mosquitoes. The most important vector of Dengue virus is the mosquito Ae.aegypti, which should be the main target of surveillance and control activities. Virologic surveillance for dengue viruses in its vector has been used as an early warning system to predict outbreaks. Detection of Dengue virus antigen in mosquito head squash using immunocytochemical streptavidin biotin peroxidase complex (SBPC) assay is an alternative method for dengue vector surveillance. The study aimed to develope immunocytochemical SBPC assay to detect Dengue virus infection in head squash of Ae.aegypti. The study design was experimental. Artificially-infected adult Ae. aegypti mosquitoes of DENV 3 were used as infectious samples and non-infected adult Ae. aegypti mosquitoes were used as normal ones. The immunocytochemical SBPC assay using monoclonal antibody DSSE10 then was applied in mosquito head squash to detect Dengue virus antigen. The results were analyzed by descriptive analysis. The immunocytochemical SBPC assay can detect Dengue virus antigen in mosquito head squash at day 2 postinfection. There are some false positive results found in immunocytochemical SBPC assay. Key Word: Dengue, immunocytochemistry, DSSE10ABSTRACT Leptospirosis is a zoonotic disease transmitted by Leptospira bacteria by rats as the main reservoir. Cases of leptospirosis occurred in several districts in Sleman. One of them is Berbah Sub-district with one case of leptospirosis in 2011. The purpose of this study to identify the biotic and abiotic environmental conditions, species of rodents captured in the study site, trap success, and Leptospira bacteria in the blood serum and kidney of rats. The study was conducted in the Jogotirto Village, Berbah Sub-district, Sleman District. As many as 150 traps had been used in 2 trapped indoor and 2 trapped outdoor during 3 days. Measurements, observations environmental conditions and interviews about rats and rats disease transmission around leptospirosis cases. Rats had been trapped identified, blood and kidneys was taken to be examined in the presence of Leptospira bacteria by using PCR (Polymerase Chain Reaction). Data were analyzed descriptively in tables and graphs. Biotic and abiotic environmental conditions favor transmission of leptospirosis. Rats spesies had been found were Rattus tanezumi, Suncus murinus, and R. tiomanicus with trap succes indoor as much 10.5% and outdoor as much 5.2%. People had less knowledge about rats and rats disease transmission. Keywords: rats, knowledge, SlemanABSTRACT Introduction: Dengue Haemorrhagic Fever (DHF) is still a public health problem in Singkawang Municipality which was an endemic area. DHF surveillance is expected to inform endemicity of an area, season of transmission and disease progression that can be use to make the system more effective and efficient. Methods: Observational study by using a structured questionnaire. Interview was conducted to all DHF surveillance officers. Evaluated had been done to the variable of input, process, and output of the surveillance system. We conducted an on the job training to all DHF surveillance officers after the evaluation. Results: 66.7% officers never got any trainings of surveillance, 83.3% had double duty, budgeting limited to physical needs, facilities and infrastructures. Process variable, data collection was late; analysis and recommendation had not been directed to the distribution of cases, the relationship between risk factors and the mortality of DHF incidence, and environment changing, feedback; data distribution had not been implemented optimally. Output variable was still weak, no surveillance epidemiology profile. Attribute surveillance such as simplicity, flexibility, and positive predictive value were good, but still weak in acceptability, sensitivity, representativeness, and timeliness. Short-term evaluation resulted that there was an increasing knowledge of surveillance officers (p value <0.05). Mid-term evaluation resulted that there was an increasing of completeness and accuracy of DHF report from 80% to 100%, active case finding, epidemiology investigation conducted to all DHF cases. Discussion and Conclusions : DHF surveillance system in Singkawang needs to be improved, there were many attributes of surveillance system that had not done well. Training of surveillance system is needed to improve capability and capacity of the surveillance officers. Keywords: Evaluation, Surveillance, DHF, SingkawangABSTRACT Malaria is still becoming public health problem in Tabalong District, South Kalimantan Province. Malaria case in Tabalong during 3 last years experiences improvement, and spread in four Sub District. The study aimed to explains the epidemiology of malaria in Santuun village, Muara Uya Sub District. Santuun village is most endemic area in Muara Uya Subdistrict, malaria control activity which have been done are passive case detection in Health Center / Hospital and entomological survey. Malaria distribution in Santuun village based on time related to rainfall, at high rainfall of malaria cases tend to increase. Most of malaria cases found at male group. Anopheles habit in Santuun is rainwater pond, sleazy pool, cracks, rice field. Anopheles which were found : An. aconitus, An. balabacensis, An. nigerimus, An. barbirostris, An. kochi, An. vagus, An.minimus. Keyword : Malaria, Santuun, Muara Uya