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Comparison of sequences of hypervariable region (HVR) subunit S-1 gene of field isolate I-37 infectious bronchitis virus with Connecticut serotype N.L.P Indi Dharmayanti; Risa Indriani; Darminto .
Jurnal Ilmu Ternak dan Veteriner Vol 8, No 2 (2003): JUNE 2003
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Infectious Bronchitis is a contagious and acute respiratory disease in chickens caused by infectious bronchitis virus (IBV).Antigenic differences in IBV are associated with changes in the sequence of the spike glycoprotein (S). The subunit S1 which demonstrates more sequence variability than S-2 have been identified as hypervariable region (HVR-1 and 2). There were several IB virus field isolates included I-37 have been identified in Indonesia by serum neutralization method. However, gene sequence variation in HVR subunit S-1 had not yet been identified. Isolate I-37 was close to the serotype Connecticut 46 (Conn 46). The aim of this study is to identify sequence variation of HVR subunit S-1 gene of isolate I-37 produced by Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) and sequencing. Several procedures were carried out in the study including virus titration, propagation and was concentrated from the allantoic fluid infected with IBV. Then, RNA was extracted for RTPCR. urther the product was sequnced and its homology with IBV references from GenBank was compared by GenMac version 8.0. Result showed that isolate I-37 produced 515 bp of amplification product. Isolate I-37 and Conn 46 are same serotype, yet their HVR subunit S-1 nucleotides and amino acids (protein) differ by 6.9% and 15.6% respectively. It might be concluded that isolate I-37 was variant of Conn 46. Key words: Sequences variation, IBV, I-37 field isolate, HVR subunit S-1 gene

Circulating H5N1 virus among native chicken living around commercial layer farms Simson Tarigan; Risa Indriani; J. Ignjatovic
Jurnal Ilmu Ternak dan Veteriner Vol 20, No 3 (2015): SEPTEMBER 2015
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Soon after the application of vaccination programme against high pathogenic avian influenza H5N1 outbreak of the disease in breeder and commercial layer farms has diminished remarkably in West Java. This study aimed to investigate whether the H5N1 decline is related to the disappearance of source of infection around the farms. Serum samples were collected from 421 native chicken living around commercial layer farms in the Districs of Cianajur and Sukabumi, West Java in March-April 2014. Antibodies to avian influenza virus (AIV) H5N1 were measured using haemaglutination inhibition (HI), ELISAs and immunoblotting that measured presence of antibodies to the haemagglutin of H5N1 strain, as well as the M2e and nucleoprotein (NP) of all avian influenza viruses. Based on the combined results, 8.6% of the native chickens were seropositive to AI virus based on one or more of serological tests. This study provided serological evidence that H5N1 virus was still circulating among native chicken living around commercial layer farms. Many positive sera were however positive for antibodies in one test only: 2.4%, 3.3% and 3.8% by HI test, M2e and NP ELISA, respectively. It could be speculated that the incongruity of the results is due to the fact that HI, MM2e ELISA and NP ELISA all measure different type of antibodies and the duration of these antibodies in serum following infection with H5N1 differ. The fact that H5N1 virus is still circulating around commercial layer farms infers that the commercial farms are still under threat and therefore vaccination and strict biosecurity are still needed.

Pathogenicity and immunogenicity local isolat infectious laryngo tracheitis virus Risa Indriani; Helmy Hamid; R.M Abdul Adjid; Muharam Saepulloh
Jurnal Ilmu Ternak dan Veteriner Vol 9, No 2 (2004): JUNE 2004
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Infectious laryngotracheitis (ILT) is an acute and contagious respiratory diseases of chicken. The virus is Gallid herpes and belong to family herpesviridae. Two local strains of ILT virus those were BGR-6 and BKS-3 were isolated and their pathogenicity and immunogenicity were further observed after five time pareses on coris allantoic of specific pathogenic free embryonated eggs. The pathogenicity of both isolates to be possible for use as seed vaccine were detected based on pathogenicity indices and antibody response. Experimental specific pathogenic free chicken in isolator cages were infected by the isolates using103EID50. ILT virus per dose. Clinical syndromes, pathological anatomic lesions, and immunological response were observed in the infected chickens and another group at uninfected chickens as a control. Results showed that either BGR-6 or BKS-3 caused clinical signs with ITPI scores of 0,05 and 0,03 respectively and there were no mortality of infected chickens. The top antibody responces of BGR-6 and BKS-3 were observed at OD 0.90 and 0.44 respectively. It can be concluded that BGR-6 and BBS-3 had low ITPI scores, but BGR-6 gave higher antibody response and can be used as a candidate for seed vaccine. Key words: Infectious laryngotracheitis, ILT, BGR-6, BKS-3, pathogenicity, immunogenicity

Seroprevalence of highly pathogenic avian influenza H5 subtype clade 2.3.2 on ducks and muscouvy ducks in small holders farm Eny Martindah; Risa Indriani; S Wahyuwardani
Jurnal Ilmu Ternak dan Veteriner Vol 19, No 4 (2014): DECEMBER 2014
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Seroprevalence studies of HPAI H5 subtype in ducks and muscouvy duck in smallholders farm was carried out in Serang and Tangerang District, Banten Province. The study comprised a serological survey to define the distribution and prevalence of HPAI H5 subtype infection on ducks and muscouvy ducks as well as attempted isolation of the virus from these species. Unit of sample in each stage was randomly choosen by multy stage random sample. Blood samples were taken from ducks and muscouvy ducks that had never been vaccinated, purposively. Sera were tested using Haemaglutination Inhibition Test, antigen H5N1 (clade 2.3.2), while the cloaca and trachea swab samples was injected into specific pathogens free (SPF) embryonated 9-11 days old, to isolate the virus. Results showed that H5 subtype virus could be isolated from tracheal swabs of ducks in the various age groups. The seroprevalence of H5 subtype virus in Banten Province was 25.5%, in which, 24.3% occured in ducks and in muscouvy duck in the rate of 1.2%, with titer HI positive was > 3log2. Based on species, seroprevalence level HPAI H5 subtype in ducks was 3-4 times higher than the level of seroprevalence of HPAI H5 subtype in muscouvy duck, which indicated that the H5 subtype virus more likely to circulate in the ducks flock than in muscouvy duck. This study noted that both muscouvy duck and ducks appeared to play a significant role in the epidemiology of the disease.

The development of an Enzyme Linked Immunosorbent Assay for detecting Injectious laryngotrachitis viral antibodies in chicken serum Risa Indriani; R.M Abdul Adjid; Darminto .; Helmy Hamid
Jurnal Ilmu Ternak dan Veteriner Vol 7, No 2 (2002): JUNE 2002
Publisher : Indonesian Center for Animal Research and Development (ICARD)

The aim of this study was to develop an Enzyme-linked immunosorbent assay (ELISA) for detection of antibody against gallid herpes virus, the causal agent of infectious laryngotracheitis (ILT) in chicken. Its application in experimental chicken under laboratory condition was also evaluated. Results showed that ELISA for ILT was developed successfully with sensitivity and specificity was 98% and 97,14% respectively. The ELISA was able to determine the dynamic of antibodies respond in experimental chickens following vaccination and artificial infection with ILT virus. It was concluded that this ELISA offers a simple, sensitive and specific antibody assay for detection of antibodies against ILT virus in chickens arising from vaccination or infection. Key words: ELISA, antibody, chicken, Infectious laryngotrachitis

Detection of antibody responses by using haemagglutination inhibiton test and the protection titer of avian influenza virus H5N1 subtype Risa Indriani; N.L.P.I Dharmayanti; A Wiyono; Darminto .; L Parede
Jurnal Ilmu Ternak dan Veteriner Vol 9, No 3 (2004): SEPTEMBER 2004
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Study on the detection of antibody responses using haemagglutination inhibition (HI) test and the protection titer to Avian influenza (AI) virus H5N1 subtype local isolate has been conducted at the Research Institute for Veterinary Science (RIVS). A total number of 50 village chicken (10 chicken served as un-injected controls) and 30 quail were injected intramuscularly with inactivated virus of AI H5N1 subtype local isolate. Serum samples were collected 3 weeks after injection and were tested using haemagglutination inhibition tests. The correlation between antibody titer and its protection to AI virus H5N1 local isolate were measured by challenging the birds with AI virus H5N1 local isolate The HI test was then used to determine field serum samples. A total number of 48 village chicken from three (3) Districts (Bekasi, Tangerang and Bogor) and 96 quails from two (2) farms in District of Sukabumi which were all vaccinated with commercial AI adjuvant vaccine were sampled. The study revealed that village chicken and quails showed antibody responses after 3 weeks vaccination and that titer of ≥ 3 log 2 was able to protect chicken and quails when they were challenged with local isolate virus. Based on this result, village chicken field samples from Districts of Tangerang, Bekasi and Bogor showed antibody titer which will protect 50, 100 and 85% of the flocks respectively. While quail field samples from Farm I and Farm II in District of Sukabumi showed antibody titer which will protect 60-100% and 0-80% of the flocks respectively. It is concluded that the study has successfully measured antibody titer to AI virus H5N1 subtype which protect village chicken and quails from local isolate virus challenge so that the results will be used to analyze field serum samples after vaccination program to eradicate AI from Indonesia. Key words: Antibody responses, haemagglutination inhibition test, protection titer, AI virus H5N1subtype

Protection of avian influenza (AI) vaccines for poultry against infection of field isolates A/Chicken/West Java/Smi-Pat/2006 and A/Chicken/West Java/Smi-Mae/2008 under laboratory condition Risa Indriani; N.l.p.I. Dharmayanti; R.M.A. Adjid
Jurnal Ilmu Ternak dan Veteriner Vol 16, No 2 (2011): JUNE 2011
Publisher : Indonesian Center for Animal Research and Development (ICARD)

The aim of this research was to study level of protection of avian influenza (AI) commercial vaccines available in Indonesia (subtipe H5N1, H5N2 and H5N9) against infection of HPAI field isolates of A/Chicken/West Java/Smi-Pat/2006 and A/Chicken/West Java/Smi-Mae/2008. There were 7 commercial vaccines used in this study, the each vaccines were injected in to 3 weeks old of layer chichickenen intramuscularly. At 3 weeks after vaccination, ten chichickenens from each group were challenged separately with the A/Chicken/West Java/Smi-Pat/2006 and A/Chicken/West Java/Smi-Mae/2008 isolates intranasaly with dose 106 ELD50 per 0,1 ml per chicken. Ten unvaccinated chicken were included in the challenge test as control. The study demonstrate that the AI vaccines with subtipe H5N1 protected chicken (100%) against virus of A/Chicken/West Java/Smi-Pat/2006 and 90-100% against virus A/Chicken/West Java/Smi-Mae/2008. Viral shedding were not seen by 2 days post challenge. The AI vaccines with subtipe H5N2 protected chicken at 20-30% against virus of A/Chicken/West Java/Smi-Pat/2006 and protected chicken at 70-100% against virus of A/Chicken/West Java/Smi-Mae/2008. Viral shedding still detected at 8 days post challenge. The AI vaccines AI with subtipe H5N9 did not protect chicken (0%) against virus A/Chicken/West Java/Smi-Pat/2006 and protected chicken at 50% against virus A/Chicken/West Java/Smi-Mae/2008. Viral shedding still detected by 8 days post challenge. This study concluded that AI vaccines with subtipe H5N1 are better than other AI subtipe vaccines in preventing HPAI virus A/Chicken/West Java/Smi-Pat/2006 dan A/Chicken/West Java/Smi-Mae/2008 infections under laboratory condition. Key Words: Avian Influenza, Vaccine, Poultry, Chicken

Antibody response and protection of inactivated-local isolate vaccine for infectious bronchitis in laying chicken Risa Indriani; Darminto .
Jurnal Ilmu Ternak dan Veteriner Vol 6, No 2 (2001): JUNE 2001
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Infectious bronchitis (IB) is an acute highly contagious viral respiratory disease of poultry caused by Coronavirus. IBV infection consists of many serotypes and can only be controlled by vaccination. An effective IB vaccine should be prepared from local isolates, due to the antigenic variation among serotypes. The aims of this research were to develop inactivated IB vaccine derived from IBV local isolate and to determine the efficacy of that vaccine in layer flocks. Five layer chicken groups were used in this experiments, group I was vaccinated with commercial IBV live vaccine thrice, group II was vaccinated with commercial IBV live vaccine once and repeated with inactivated local IBV isolate twice, group III was vaccinated with commercial IBV live vaccine once and repeated with commercial inactivated twice, group IV was vaccinated with IBV live vaccine once, and group V was not vaccinated. After the chickens reached at a stable egg production they were challenged with IBV local isolates. Antibody responses were examined by means of haemagglutination hibitition (HI) test and HI titres were expressed as log2 of the reciprocal of the highest dilution of serum causing inhibition of a log2 HA titre of 2. The mean titres of antibody responses of chicken in group I, II, III, IV, and V was 4.9 ± 0.87, 6.8 ± 0.97, 7.7 ± 0.46, 2.9 ± 0.94, and 2.0 ± 1.67 respectively. The levels of protection against challenges were determined by viral isolation, this in group I, II, III, IV, and V was 63, 73, 60, 50, and 0% respectively. Clinical symptom of egg quality was slightly reduced in group I, IV, and V and it were unchanged in group II and III. Group II gave better in number of egg production than the other groups. The results indicated that the IBV inactivated localisolate vaccine gave high titres of antibody and higher protection rates than that of commercial IBV inactivated vaccine. Inaddition, IBV local isolate vaccinated group prevented from declining egg production after challenged with IBV local isolate. Key words: Infectious bronchitis, layer, antibody titre, vaccine, challenge virus

Serotype variation among infectious bronchitis viral isolates taken from several areas of Java Risa Indriani; Darminto .
Jurnal Ilmu Ternak dan Veteriner Vol 5, No 4 (2000): DECEMBER 2000
Publisher : Indonesian Center for Animal Research and Development (ICARD)

Infectious bronchitis (IB) is an acute highly contagious viral respiratory disease of poultry caused by virus belongs to the family of Coronaviridae. The virus consist of many serotypes with low level of cross-protectivity among serotypes. Field data showed that the outbreaks of IB were frequently reported in chicken flocks, although vaccinations against the disease have been practiced. Hence, the study on serotype relationship among isolates of the viruses is essentially required. The aim of this study was to isolate and characterize IB viruses from chicken flocks in some areas of Java. Isolation of the virus was carried out in nine-day old embrionated chicken eggs and identified by means of agar gel precipitation (AGP) tests against standard antisera to IB virus. The serotypes of the IB viral isolates were determined by cross-neutralization tests in nine day old embryonated chicken eggs using r value derived from homologous and heterologous serum titres as criteria. This study obtained 12 IB viral isolates which were identified on the basis of the ability to cause lesions in chicken embryos and positive to agar gel presipitation test against standard positive antiserum to the virus. Based on the cross-neutralization tests in embryonated chicken eggs, isolate I.9 was formed to have relationship closed to Mass-41 serotype, while I.2, I. 3, and I.7 isolates were closely to the serotype of Con-46. Virus isolates (I.5, I.14, I.24, and I.25) were decided to have no serotype relationships to either Mass-41 or Con-46 serotype. Since the I.5, I.14, I.24 and I.25 isolates were not neutralized by antisera against the previous identified local infectious bronchitis viral isolates, and that were considered to be distinct serotype to the previously identified local IB viral isolates. Key words: Infectious bronchitis, virus, embryonated egg, cross neutralization test.

Characterisation of enzymatic activities of H5N1 influenza virus Simson Tarigan; Risa Indriani; Darminto .
Jurnal Ilmu Ternak dan Veteriner Vol 12, No 2 (2007): JUNE 2007
Publisher : Indonesian Center for Animal Research and Development (ICARD)

One of the two glycoproteins projected from the surface of the influenza virus is identified as neuraminidase. This enzyme enables the virus to spread in the host, and therefore it plays vital roles in the viral pathogenicity. From the viewpoint of disease control, neuraminidase is used as the target for the development of anti-flu drugs, and for the development of diagnostic test to differentiate infected from vaccinated animals (DIVA). Since the roles of the enzyme are very important, information regarding the characteristics and the procedure to measure its activity, which is the purpose of this study, is essential. The optimum incubation time of the neuraminidase-substrate (fetuin) reaction and the optimum pH of the buffer were determined. The stability of the enzyme against heating, supplementation or chelating of calcium ion, and b-propiolactone treatment were analysed. This study showed that neuraminidase from H5N1-influenza virus was, in regards to the characteristics investigated in this study, was comparable to that from Clostridium perfringens. The optimum incubation time for the viral and Clostridial neuraminidases were 60 and 30 minutes, respectively; whereas, the optimum pH for both neuraminidase was 6-7. At pH 8, both neuraminidase were inactive. Supplementation of calcium ion tended to increase activity but chelating of the cation did not have any observable effects. Treatment with 0.2% b-propiolactone for 6 hours reduced the activity, whereas heating at 60°C for 60 minutes abolished all activity. Since inactivation by b-propiolactone is partially only, neuraminidase assay could be performed safely in ordinary laboratories using b-propiolactone-treated-influenza virus, rather than the life virus. The thermolabile nature of the enzyme will complicate any attempt to purify the enzyme. Key Words: H5N1, Neuraminidase, Stability, Thiobarbituric Assay

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