Didik Setyo Heriyanto

Clinical and virological characteristics of hepatitis B or C virus co-infection with HIV in Indonesian patients†

Hepatitis virus‐related liver disease increases substantially the mortality rate of patients with HIV on highly active antiretroviral therapy (HAART). Therefore, early diagnosis of hepatitis B virus (HBV) and hepatitis C virus (HCV) is important. However, the prevalence of HBV and HCV infection in Indonesian patients infected with HIV is unknown. Therefore, this study examined the molecular and clinical characteristics of HBV and HCV in 126 patients infected with HIV, mostly on HAART, at Dr. Sardjito Hospital, Yogyakarta, Indonesia. The rates of triple infection, HIV/HCV co‐infection, HIV/HBV co‐infection, and mono‐infection were 4.8%, 34.1%, 3.2%, and 57.9%, respectively. Seven HCV genotypes were detected, with genotypes 1a, 1b, 1c, 3a, 3k, 4a, and 6n found in 23 (52%), 1 (2%), 4 (9%), 5 (11%), 7 (16%), 3 (6%), and 1 (2%) patients, respectively, indicating multiple modes of transmission. HBV‐DNA was detected in 2/10 patients with hepatitis B surface antigen; both patients were HAART naive. Univariate analysis revealed that male sex, higher education level, injection drug use, sexual contact, alanine aminotransferase ≥40 IU/L, and aspartate aminotransferase‐to‐platelet ratio index > 0.5 were associated with HCV co‐infection. In multivariate analysis, injection drug use (OR: 26.52; 95% CI: 3.52–199.54) and alanine aminotransferase ≥40 IU/L (OR: 6.36; 95% CI: 1.23–32.89) were independently associated with HCV co‐infection. HCV co‐infection was common among Indonesian patients infected with HIV, particularly among injecting drug users, and was a risk factor for disease progression of HIV. J. Med. Virol. 84:857–865, 2012.

Hepatitis B and C virus infection among hemodialysis patients in yogyakarta, Indonesia: Prevalence and molecular evidence for nosocomial transmission

Hemodialysis patients are at an increased risk of acquiring hepatitis B virus (HBV) and hepatitis C virus (HCV) infection. However, the prevalence of hepatitis viral infection and its genotype distribution among hemodialysis patients in Indonesia are unclear. In order to investigate these issues and the possibility of nosocomial transmission, 161 hemodialysis patients and 35 staff members at one of the hemodialysis unit in Yogyakarta, Indonesia, were tested for serological and virological markers of both viruses. HBV surface antigen (HBsAg) was detected in 18 patients (11.2%) and in two staff members (5.7%). Anti‐HCV was detected in 130 patients (80.7%) but not in any staff members. Occult HBV and HCV infection were detected in 21 (14.7%) and 4 (12.9%) patients, respectively. The overall prevalence rates of HBV and HCV infection among patients were 24.2% and 83.2%, respectively. HCV infection was independently associated with hemodialysis duration and the number of blood transfusions. Phylogenetic analysis revealed that 23 of 39 tested HBV strains (59%) were genotype B, 11 (28.2%) were genotype C, and 5 (12.8%) were genotype A. HCV genotype 1a was dominant (95%) among 100 tested HCV strains. Nosocomial transmission was suspected because the genotype distribution differed from that of the general population in Indonesia, and because the viral genomes of several strains were identical. These findings suggest that HBV and HCV infection is common among hemodialysis patients in Yogyakarta, and probably occurs through nosocomial infection. Implementation of strict infection‐control programs is necessary in hemodialysis units in Indonesia. J. Med. Virol. 85:1348–1361, 2013.

Mutations within enhancer II and BCP regions of hepatitis B virus in relation to advanced liver diseases in patients infected with subgenotype B3 in Indonesia

Studies on the characteristics of mutations within the hepatitis B virus (HBV) genome, their roles in the pathogenesis of advanced liver diseases, and the involvement of host properties of HBV‐infected individuals have not been conducted in subgenotype B3‐infected populations. For addressing this issue, 40 cases with HBV surface antigen (HBsAg)—positive advanced liver diseases, including advanced liver cancer and cirrhosis (male 31, female 9, age 54.4 ± 11.6‐year‐old), were collected and compared with 109 cases with chronic hepatitis B (male 71, female 38, age 38.0 ± 13.4‐year‐old). Mutations in enhancer II (Enh II) and basal core promoter (BCP)/precore regions were analyzed by PCR‐direct sequencing method. HBV viral load was examined by real‐time PCR. For all examined regions, the prevalence of mutation was significantly higher in cases with advanced liver diseases. Multivariate analysis showed that, in patients older than 45 years, C1638T and T1753V mutations constituted independent risk factors for the advancement of liver diseases. The presence of C1638T and T1753V mutations may serve as predictive markers for the progression of liver diseases in Indonesia and other countries, where subgenotype B3 infection is prevalent. J. Med. Virol. 84:44–51, 2011.

A deep-sequencing method detects drug-resistant mutations in the hepatitis B virus in Indonesians.

Objective: The long-term administration of a nucleos(t)ide analogue (NA) for the treatment of chronic hepatitis B may encourage the emergence of viral mutations associated with drug resistance. Minor populations of viruses may exist before treatment, but are difficult to detect because of technological limitations. Identifying minor viral quasispecies should be useful in the clinical management of hepatitis B virus (HBV) infection. Methods: Six treatment-naïve Indonesian patients with chronic HBV infection participated in this study. The polymerase region of the HBV genome, including regions with known drug-resistant mutations, was subjected to capillary sequencing and MiSeq sequencing (Illumina). Mutations were analyzed with Genomics Workbench software version 6.0.1 (CLC bio). Results: The mean mapping reads for the six samples was 745,654, and the mean number of amplified fragments ranged from 17,926 to 25,336 DNA reads. Several known drug-resistant mutations in the reverse transcriptase region were identified in all patients, although the frequencies were low (0.12-1.06%). The proportions of the total number of reads containing mutations I169L/M, S202R, M204I/L or N236S were >1.0%. Conclusion: Several known NA-resistant mutations were detected in treatment-naïve patients in Indonesia using deep sequencing. Careful management of such patients is essential to prevent drug-resistant mutations from spreading to other patients.

Protective effects of HLA-DPA1/DPB1 variants against Hepatitis B virus infection in an Indonesian population.

Human leukocyte antigen (HLA) DPA1/DPB1 variants have been reported to influence Hepatitis B virus (HBV) infection. HLA-DPA1/DPB1 plays a pivotal role in antigen presentation to CD4(+) helper T cells and influences the outcome of HBV infection. To investigate the influence of HLA-DP variants on the outcome of HBV infection in an Indonesian population where it has the third-highest prevalence of HBV infection worldwide, we performed a case-control study of 686 participants, including patients with HBV-related advanced or nonadvanced liver disease, patients with spontaneously resolved HBV, and healthy controls. Single-nucleotide polymorphisms in HLA-DPA1 (rs3077) and HLA-DPB1 (rs3135021, rs9277535, and rs228388) were genotyped using real-time TaqMan® genotyping assays. Because rs2281388 deviated from Hardy-Weinberg equilibrium, it was excluded from subsequent analyses. The results of logistic regression analyses showed that the HLA-DPB1 rs9277535 variants were associated with a reduced risk of persistent HBV infection (odds ratio [OR] 0.70, 95% confidence interval [95% CI] 0.52-0.96, P=0.026, additive genetic model; OR 0.60, 95% CI 0.38-0.96, P=0.033, dominant genetic model). The HLA-DPA1 rs3077 variant was associated with a protective effect increasing the spontaneously resolved HBV infection (OR 0.64, 95% CI 0.41-0.98, P=0.039, dominant genetic model). By contrast, the HLA-DPB1 rs3135021 variant was not associated with the outcome of HBV infection, including susceptibility, spontaneously resolved, or disease progression. Combinations of haplotype markers were also associated with HBV susceptibility (CA for rs3077-rs9277535, OR 0.57, 95% CI 0.36-0.92, P=0.021; GA for rs3135021-rs9277535, OR 0.56, 95% CI 0.36-0.86, P=0.0087). In conclusion, these findings confirm that HLA-DPA1/DPB1 variants were associated with the outcomes of HBV infection in an Indonesian population.

GB virus C infection in Indonesian HIV‐positive patients

GB virus C (GBV‐C), a human virus of the Flaviviridae family that is structurally and epidemiologically closest to hepatitis C virus (HCV), has been reported to confer beneficial outcomes in HIV‐positive patients. However, the prevalence of GBV‐C in HIV‐positive individuals in Indonesia is unknown. Since GBV‐C is more prevalent in anti‐HCV positive patients than in anti‐HCV negative subjects, transmission of GBV‐C and HCV could be by the same method. This study examined the prevalence and molecular characteristics of GBV‐C infection in HIV patients in Yogyakarta, Indonesia. The prevalence of GBV‐C among HIV patients (n = 125, median age 31 years) based on the 5′UTR region was 111/125 (88.8%), including 39/48 (81.3%) and 72/77 (93.5%) HIV‐infected patients with and without HCV infection, respectively. GBV‐C isolates were of genotype 2a, 3 and 6 in 58.3%, 12.6% and 28.4% of patients, respectively. Patients with genotype 3 were significantly younger than those with genotypes 2a or 6 (P = 0.001 and P = 0.012, respectively). Genotypes 3 and 6 were significantly associated with injection drug use (P = 0.004 and P = 0.002, respectively) and HCV co‐infection (P < 0.001 for both genotypes), indicating a shared transmission route with HCV. In conclusion, the prevalence of GBV‐C among HIV‐positive patients in Indonesia is high, and three genotypes were detected, namely genotype 2a, 3 and 6.

The utility of the hematoxylin and eosin staining in patients with suspected Hirschsprung disease

BackgroundWhile immunohistochemistry (IHC) methods have been widely conducted for the diagnosis of Hirschsprung disease (HSCR) in developed countries, there are very few studies on their use in developing countries where hematoxylin and eosin (HE) staining is a key element of the diagnosis of HSCR. We aimed to determine the accuracy of HE staining in the diagnosis of HSCR using S100 IHC as the reference standard in Indonesia.MethodsAll histopathology performed for the suspicion of HSCR patients from January 2013 to August 2015 in Dr. Sardjito Hospital, Yogyakarta, Indonesia, were retrospectively reviewed.ResultsOur study included 23 HSCR patients: 9 males and 14 females. The HE staining revealed 14 negative (absence of ganglion cells) cases (61%) and 9 positive (presence of ganglion cells) cases (39%). In S100 IHC, out of the 9 positive cases by HE staining, 6 (67%) were confirmed for having ganglion cells; and out of the 14 negative cases by HE staining, 12 (86%) were reported negative and 2 (14%) were positive by S100 IHC staining. The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy rates of the HE staining were 80% (95% CI: 0.51–0.95), 75% (95% CI: 0.36–0.96), 85.7% (95% CI: 0.56–0.98), 66.7% (95% CI: 0.31–0.91), and 78.3% (95% CI: 0.58–0.90), respectively.ConclusionsOur study showed that HE staining has relatively moderate accuracy for the diagnosis of HSCR. The use of HE staining is still recommended for the diagnosis of HSCR given the limitation of resource allocation for more expensive IHC technologies in developing countries.

Hepatitis E virus infection in two different regions of Indonesia with identification of swine HEV genotype 3

Hepatitis E is an emerging disease with a high incidence globally. Few data are available on hepatitis E virus (HEV) infection in Indonesia. To obtain molecular information on HEV infection in two regions of Indonesia with different customs and swine breeding conditions, serum samples from 137 swine farm workers, 100 blood donors and 100 swine (27 fecal samples also obtained) in Yogyakarta (Central Java) and from 12 and 64 swine farm workers, 42 and 135 local residents and 89 and 119 swine in Tulungagung (East Java) and Mengwi (Bali), respectively, from our previous study, were compared. Serological tests for anti‐HEV antibodies by ELISA, HEV‐RNA detection by RT‐PCR and phylogenetic analysis were performed. The total prevalence of anti‐HEV antibodies in humans was higher in Bali (11.6%) than in Java (5.1%; P = 0.015). No significant differences in anti‐HEV prevalence among swine farm workers and local residents in Java were found. The finding of swine HEV genotype 3 in specimens from Yogyakarta and genotype 4 from Tulungagung and Bali is somewhat different from other reports. We suggest other factors in addition to close contact with swine might play an important role in HEV transmission of non‐endemic/related custom groups. To the best of our knowledge, this is the first report on swine HEV genotype 3 in Indonesia.

Rare Helicobacter pylori Infection May Explain Low Stomach Cancer Incidence: Ecological Observations in Bali, Indonesia

The incidence rate of stomach cancer in Bali, Indonesia, is estimated to be strikingly lower than that in Japan. We conducted an on-site ecological study to investigate the association between the stomach cancer incidence and Helicobacter pylori (H. pylori) infection. Recruiting 291 healthy persons (136 men and 155 women) from the general population in Bali, Indonesia, we conducted a urea breath test (UBT) to examine H. pylori infection, along with a pepsinogen test to detect chronic atrophic gastritis and urine analysis to estimate sodium and potassium excretion. UBT positivities were 9% (2-15, 95% confidence interval) for men and 7% (1-12) for women, and positive cases for H. pylori IgG antibodies were 1% (0-3) for men and 3% (0-5) for women, significantly lower than the respective values in Japan. Positive pepsinogen tests in Bali were 0% (0-0) for men and 1% (0-4) for women, also significantly lower than the Japanese figures. Computed values for daily salt excretion were 13.3±4.1 g (mean ± SD) for men and 11.1±3.1 g for women, as high as corresponding Japanese consumption values. Moreover, the estimated potassium excretion was 3.2±0.7 g for men and 2.8±0.6 g for women in Bali, significantly higher than the figures in Japan. There were no associations across genetic polymorphisms of IL-beta, TNF-alpha, and PTPN11 with UBT positivity. The low incidence of stomach cancer in Bali may thus mainly be due to the rare H. pylori infection. Namely, the bacterium infection seems to be a critical factor for gastric cancer rather than host or other environmental factors.