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Background: Accurate diagnosis of enteric fever is challenging, particularly in low- and middle-income countries, due to the overlap of clinical and laboratory features with other pathogens. To better understand the difficulties in enteric fever diagnosis, we evaluated the characteristics of patients clinically diagnosed with enteric fever and the real-world performance of TUBEX TF, one of the most used tests in Indonesia., Methodology/principal Findings: Patients were recruited through the AFIRE (Etiology of Acute Febrile Illness Requiring Hospitalization) study at eight Indonesian hospitals. Blood culture was performed for all patients, and TUBEX TF was performed for suspected enteric cases. Salmonella PCR and ELISA tests were performed at a reference lab. Sensitivity and specificity of TUBEX TF and IgM and IgG anti-S. Typhi ELISA were determined. Of 301 patients clinically diagnosed with enteric fever, 50 (16.6%) were confirmed by blood culture and/or PCR. Confirmed cases were mostly school-aged children presenting with fever, anorexia, dizziness and/or abdominal pain with normal leukocyte count or leukopenia. TUBEX TF demonstrated a sensitivity of 97.6% to 70.7% and specificity of 38.3% to 67.2% at cutoffs of 4 and 6, respectively. Acute IgG demonstrated the best sensitivity and specificity, at 90.7% and 82.7%, respectively, and the best ROC characteristics., Conclusions/significance: A substantial proportion of enteric fever was misdiagnosed at all study hospitals, likely due to the overlap of clinical characteristics and lab parameters with those of other common pathogens. The TUBEX TF rapid serological assay demonstrated suboptimal performance in our setting and tended to over-diagnose enteric fever. The role of IgG from acute specimens for identification of enteric fever cases merits additional consideration., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Host genetic factors, such as the genes for various cytokines and adhesion molecules, play a significant role in determining susceptibility to malaria infection. Polymorphisms in host genes have been correlated with malaria infection in both African and Asian regions. The purpose of this study was to investigate the association between both cytokine and adhesion molecule genotypes with susceptibility to malaria infection in humans. Ten cytokine polymorphism loci (IL4 + 33, IL4-590, IL6-174, IL10-1082, IL10-1035, IL12p40, TNF-238, TNF-308, TNF-1031, and TNF-β) and three adhesion molecule polymorphism loci (CD36 exon 10, ICAM-1 Kilifi, and ICAM-1 exon 6) were genotyped using PCR-RFLP analysis. We conducted this study on 178 asymptomatic malaria subjects and 122 uninfected subjects. Results showed that certain CD36 exon 10 and IL10-3575 polymorphisms were associated with asymptomatic infection. The heterozygous (GT) and homozygous (GG) genotypes for CD36 exon 10 are associated with an increased risk of malaria infection. On the other hand, the homozygous genotype (AA) for IL10-3575 reduced the risk of asymptomatic malaria infection. No significant differences were found for the other polymorphisms studied. We also found that a polymorphism in CD36 exon 10 was strongly associated with asymptomatic malaria caused specifically by Plasmodium vivax. These findings suggest that the G allele of CD36 exon 10 is associated with an increased risk of asymptomatic malaria infection. On the other hand, the genotype AA for IL10-3575 was associated with a reduced risk of malaria infection., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Blood culturing remains the "gold standard" for bloodstream infection (BSI) diagnosis, but the method is inaccessible to many developing countries due to high costs and insufficient resources. To better understand the utility of blood cultures among patients in Indonesia, a country where blood cultures are not routinely performed, we evaluated data from a previous cohort study that included blood cultures for all participants. An acute febrile illness study was conducted from July 2013 to June 2016 at eight major hospitals in seven provincial capitals in Indonesia. All participants presented with a fever, and two-sided aerobic blood cultures were performed within 48 hours of hospital admission. Positive cultures were further assessed for antimicrobial resistance (AMR) patterns. Specimens from participants with negative culture results were screened by advanced molecular and serological methods for evidence of causal pathogens. Blood cultures were performed for 1,459 of 1,464 participants, and the 70.6% (1,030) participants that were negative by dengue NS1 antigen test were included in further analysis. Bacteremia was observed in 8.9% (92) participants, with the most frequent pathogens being Salmonella enterica serovar Typhi (41) and Paratyphi A (10), Escherichia coli (14), and Staphylococcus aureus (10). Two S. Paratyphi A cases had evidence of AMR, and several E. coli cases were multidrug resistant (42.9%, 6/14) or monoresistant (14.3%, 2/14). Culture contamination was observed in 3.6% (37) cases. Molecular and serological assays identified etiological agents in participants having negative cultures, with 23.1% to 90% of cases being missed by blood cultures. Blood cultures are a valuable diagnostic tool for hospitalized patients presenting with fever. In Indonesia, pre-screening patients for the most common viral infections, such as dengue, influenza, and chikungunya viruses, would maximize the benefit to the patient while also conserving resources. Blood cultures should also be supplemented with advanced laboratory tests when available., Competing Interests: The authors have declared that no competing interests exist.

HIV prevalence in Indonesia is increasing, and only 64% of infected individuals know their status. In a prospective cohort of 1,453 hospitalized patients with unexplained fever, 46 (3.2%) had HIV, including 15 (1.1%) patients without a prior HIV diagnosis. Among 31 subjects previously known to have HIV, 21 (68%) had been receiving combination antiretroviral therapy (cART) at the time of enrollment. Of 39 HIV cases with HIV RNA levels ≥ 100 copies/mL, sequencing for genotype analysis and resistance testing was successful in 30 (77%) subjects. The most common HIV subtypes were AE (90%) and B (10%). Five (16.7%) subjects had resistance mutations to nucleoside and non-nucleoside reverse transcriptase inhibitors, and all of them were on cART. No evidence of transmitted drug resistance was found in newly diagnosed individuals. Hospital-based screening may be an efficient method to expand HIV testing and identify a significant number of new cases. Access to care, close monitoring, expansion of anti-retroviral options, and ensuring availability of CD4 determinations, viral load testing, and genotyping are crucial to control of the epidemic in Indonesia.

Primaquine is an effective anti-hypnozoite drug for Plasmodium vivax and Plasmodium ovale. However, it can trigger erythrocyte hemolysis in people with glucose 6-phosphate dehydrogenase (G6PD) deficiency. In a previous report from South Central Timor (SCT), Indonesia, we described the prevalence of Vanua Lava, Chatham, and Viangchan variants; in this study, other G6PD variants (Kaiping, Coimbra, Gaohe, Canton, and Mahidol) were subsequently analyzed. For clarity, all of these results are described together. The 381 DNA samples from the previous study during 2013-2014 were analyzed for G6PD variants by using PCR-restriction fragment length polymorphism (RFLP). The prevalence of G6PD deficiency in SCT was 6.3% (24/381 cases), including 4.2% (16/381 cases), 0.5% (2/381 cases), and 1.6% (6/381 cases) for Coimbra, Kaiping, and Vanua Lava variants, respectively. No other variants were found in this population. A significant association was found between ethnicity and the distribution of G6PD Kaiping in female subjects. A positive association was shown between G6PD activity and heterozygous females carrying Coimbra genotype, hemizygous males carrying Vanua Lava, Plasmodium falciparum infection in female subjects, and P. vivax infection in male subjects. Further molecular analysis of heterozygous females, particularly in malaria-endemic areas, is needed for mapping distribution of G6PD deficiency status in Indonesia.

Background: Chikungunya virus (CHIKV) is often overlooked as an etiology of fever in tropical and sub-tropical regions. Lack of diagnostic testing capacity in these areas combined with co-circulation of clinically similar pathogens such as dengue virus (DENV), hinders CHIKV diagnosis. To better address CHIKV in Indonesia, an improved understanding of epidemiology, clinical presentation, and diagnostic approaches is needed., Methodology/principal Findings: Acutely hospitalized febrile patients ≥1-year-old were enrolled in a multi-site observational cohort study conducted in Indonesia from 2013 to 2016. Demographic and clinical data were collected at enrollment; blood specimens were collected at enrollment, once during days 14 to 28, and three months after enrollment. Plasma samples negative for DENV by serology and/or molecular assays were screened for evidence of acute CHIKV infection (ACI) by serology and molecular assays. To address the co-infection of DENV and CHIKV, DENV cases were selected randomly to be screened for evidence of ACI. ACI was confirmed in 40/1,089 (3.7%) screened subjects, all of whom were DENV negative. All 40 cases initially received other diagnoses, most commonly dengue fever, typhoid fever, and leptospirosis. ACI was found at five of the seven study cities, though evidence of prior CHIKV exposure was observed in 25.2% to 45.9% of subjects across sites. All subjects were assessed during hospitalization as mildly or moderately ill, consistent with the Asian genotype of CHIKV. Subjects with ACI had clinical presentations that overlapped with other common syndromes, atypical manifestations of disease, or persistent or false-positive IgM against Salmonella Typhi. Two of the 40 cases were possibly secondary ACI., Conclusions/significance: CHIKV remains an underdiagnosed acute febrile illness in Indonesia. Public health measures should support development of CHIKV diagnostic capacity. Improved access to point-of-care diagnostic tests and clinical training on presentations of ACI will facilitate appropriate case management such as avoiding unneccessary treatments or antibiotics, early response to control mosquito population and eventually reducing disease transmission., Competing Interests: The authors have declared that no competing interests exist.

Background: The burden of leptospirosis in Indonesia is poorly understood. Data from an observational study conducted from 2013 to 2016 in seven cities across Indonesia was used to estimate the incidence of leptospirosis and document its clinical manifestations in patients requiring hospitalization., Methods: Specimens from patients hospitalized with acute fever were collected at enrollment, 14-28 days, and 3 months. Demographic and clinical information were collected during study visits and/or retrieved from medical records and double-entered into clinical report forms. After initially screening for dengue virus and other pathogens, specimens were tested at a central Reference Laboratory for anti-Leptospira IgM using commercial ELISA kits and for Leptospira DNA using an in-house quantitative real-time PCR assay., Results: Of 1464 patients enrolled, 45 (3.1%) confirmed cases (by PCR and/or sero-coversion or four-fold increase of IgM) and 6 (0.4%) probable cases (by high titer IgM) of leptospirosis were identified by the Reference Laboratory. Disease incidence at sites ranged from 0 (0%) cases in Denpasar to 17 (8.9%) cases in Semarang. The median age of patients was 41.2 years (range of 5.3 to 85.0 years), and 67% of patients were male. Twenty-two patients (43.1%) were accurately diagnosed at sites, and 29 patients (56.9%) were clinically misdiagnosed as having another infection, most commonly dengue fever (11, 37.9%). Clinically, 20 patients (39.2%) did not present with hyperbilirubinemia or increased creatinine levels. Two patients (3.9%) died, both from respiratory failure. Fifteen patients (29.4%) clinically diagnosed with leptospirosis at sites were negative based on IgM ELISA and/or PCR at the Reference Laboratory., Conclusions: Leptospirosis remains an important cause of hospitalization in Indonesia. It can have diverse clinical presentations, making it difficult to differentiate from other common tropical infections. PCR combined with ELISA is a powerful alternative to the cumbersome gold-standard microscopic agglutination test, particularly in resource-limited settings.

Background: The epidemiology of acute febrile illness, a common cause of hospitalization in Indonesia, has not been systematically studied., Methodology/principal Findings: This prospective observational study enrolled febrile patients (temperature ≥38°C) aged ≥1 year from July 2013 until June 2016 at eight government referral teaching hospitals in seven provincial capitals in Indonesia. Patients were managed according to the hospital standard-of-care (SOC), and blood samples were drawn for molecular and serological assays. Clinical data, laboratory results, and specimens for additional tests were collected at enrollment, days 14-28, and at three months. Regular follow-up visits were then scheduled for every three months either until symptoms resolved or until one year. In total, this study included 1,486 adult and pediatric patients presenting with multi-organ (768, 51.7%), gastrointestinal (497, 33.0%), respiratory (114, 7.7%), constitutional (62, 4.2%), skin and soft-tissue (24, 1.6%), central nervous system (17, 1.1%), or genitourinary (4, 0.3%) manifestations. Microbiological diagnoses were found in 1,003/1,486 (67.5%) participants, of which 351/1,003 (35.0%) were not diagnosed during hospitalization using SOC diagnostic tests. Missed diagnoses included all cases caused by Rickettsia spp., chikungunya, influenza, and Seoul virus. The most common etiologic agents identified were dengue virus (467, 46.6%), Salmonella spp. (103, 10.3%), and Rickettsia spp. (103, 10.3%). The overall mortality was 89 (5.9%)., Conclusions/significance: Febrile illness in Indonesia has various microbiologic etiologies and substantial overall mortality. Diagnostic limitations and lack of epidemiologic data resulted in potentially treatable, and at times fatal, diseases being missed., Competing Interests: The authors have declared that no competing interests exist.

Background: Malaria remains a significant public health issue in Eastern Indonesia, where multidrug resistant Plasmodium falciparum and Plasmodium vivax are highly prevalent. The objective of this study was to describe treatment-seeking behaviour and household costs prior to a change to a unified treatment policy of dihydroartemisinin-piperaquine in Mimika district, Papua province in 2006., Methods: In 2005 a randomized cross-sectional household survey was conducted to collect data on demographics, socio-economic status (SES), treatment-seeking, case management, and household costs. Information on the cost of illness was also collected from patients exiting health facilities, in order to compare the cost of episodes diagnosed as P. vivax compared with those diagnosed as P. falciparum., Results: 825 households were included in the survey. Of the 764 individuals who sought treatment for fever outside the home in the last month, 46% (349/764) went to a public health facility. Of the 894 reported visits to healthcare providers, 48% (433) resulted in a blood test, of which 78% (337) were reportedly positive. Only 10% (17/177) of individuals who reported testing positive for P. falciparum or mixed infection received the first-line treatment of chloroquine with SP, and 38% (61/159) of those with a diagnosis of P. vivax reportedly received the first-line treatment of chloroquine and primaquine. Overall, public facilities were more likely to prescribe the correct prevailing first-line drug combinations than private providers (OR = 3.77 [95% CI 2.31-6.14], p < 0.001). The mean cost to the household of an episode of P. vivax was similar to the cost of P. falciparum [US$44.50 (SD: 46.23) vs US$48.58 (SD: 64.65)]., Conclusions: Private providers were a popular source of treatment for malaria, but adherence to the national guidelines was low and the economic burden of malaria for both P. falciparum and P. vivax infections was substantial. Engagement with the private sector is needed to ensure that patients have access to affordable good quality, effective diagnostics and anti-malarials for both P. falciparum and P. vivax.

Background: Most malarious countries outside of Africa are co-endemic for Plasmodium falciparum and Plasmodium vivax. The comparative burden of anaemia in the community caused by these two species is incompletely characterized., Methods: A three-stage, cross-sectional, community survey was used to determine the proportion of moderate or severe anaemia (haemoglobin <7 g/dL) attributable to patent P. vivax, P. falciparum and mixed parasitaemia in Papua, Indonesia. Adjusted population-attributable fractions were calculated from multivariable logistic regression models. Eight hundred and twenty-five households were surveyed with a total of 5255 occupants, 3890 (74 %) of whom were present and provided a blood sample. Plasmodium falciparum parasitaemia was present in 8.1 % (n = 315) of participants, P. vivax in 6.4 % (n = 250) and mixed infections in 1.9 % (n = 72). Overall, P. falciparum was associated with a mean reduction in haemoglobin of 1.16 g/dL compared to those without patent parasitaemia [95 % confidence interval (95 % CI) 0.91, 1.41 g/dL]. The corresponding values for P. vivax and mixed infections were 0.66 g/dL (95 % CI 0.35, 0.96) and 1.25 g/dL (0.71, 1.80), respectively. Overall, 16.7 % (95 % CI 8.52, 24.2 %) of haemoglobin concentrations <7 g/dL in the community were estimated to be attributable to patent parasitaemia. The fractions for infants and 1-5 years old were 34.4 % (95 % CI -3.30, 58.3 %) and 23.2 % (95 % CI 3.34, 39.0 %), respectively. Plasmodium vivax was associated with a greater than threefold higher attributable fraction of anaemia in infants compared with P. falciparum [27.6 % (95 % CI -3.20, 49.2 %) versus 7.94 % (-5.87, 20.0 %)]., Conclusion: Despite comparatively low-level endemicity, malaria is associated with a significant proportion of all cases of community anaemia in southern Papua. Contrary to its benign reputation, P. vivax is an important and preventable risk factor for anaemia during infancy-a probable consequence of relapsing disease prior to the development of immunity.

Nationally representative observational and translational research is needed to address the public health challenges in Indonesia due to the geographic disparity, recently decentralized health system, and diverse infectious disease priorities. To accomplish this, the Indonesian Ministry of Health in collaboration with the US National Institute of Health has established INA-RESPOND (Indonesia Research Partnership on Infectious Disease) - a clinical research network comprising 9 referral hospitals, 7 medical faculties, and 2 research centres across Indonesia. The network provides a forum to conduct research at a national scale and to address scientific questions that would be difficult to address in smaller research settings. Further, it is currently conducting multi-centre research on the etiologies of fever, sepsis, and tuberculosis. There are opportunities to leverage existing network resources for other public health research needs. INA-RESPOND is an Indonesian-led network in a country with diverse population groups and public health needs which is poised to collaborate with researchers, universities, donors, and industry worldwide. This paper describes the network and its goals and values, as well as the management structure, process for collaboration, and future vision.

Background: In Papua, Indonesia, maternal malaria is prevalent, multidrug resistant and associated with adverse outcomes for mother and baby. In March 2006, anti-malarial policy was revised for the second and third trimester of pregnancy to dihydroartemisinin-piperaquine (DHP) for all species of malaria. This study presents the temporal analysis of adverse outcomes in pregnancy and early life following this policy change., Methods: From April 2004 to May 2010, a standardized questionnaire was used to collect information from all pregnant women admitted to the maternity ward. A physical examination was performed on all live birth newborns. The relative risks (RR) and the associated population attributable risks (PAR) of adverse outcomes in women with a history of malaria treatment to the risk in those without a history of malaria during the current pregnancy were examined to evaluate the temporal trends before and after DHP deployment., Results: Of 6,556 women enrolled with known pregnancy outcome, 1,018 (16%) reported prior anti-malarial treatment during their pregnancy. The proportion of women with malaria reporting treatment with DHP rose from 0% in 2004 to 64% (121/189) in 2010. In those with history of malaria during pregnancy, the increasing use of DHP was associated with a 54% fall in the proportion of maternal malaria at delivery and a 98% decrease in congenital malaria (from 7.1% prior to 0.1% after policy change). Overall policy change to more effective treatment was associated with an absolute 2% reduction of maternal severe anaemia and absolute 4.5% decrease in low birth weight babies., Conclusions: Introduction of highly effective treatment in pregnancy was associated with a reduction of maternal malaria at delivery and improved neonatal outcomes. Ensuring universal access to arteminisin combination therapy (ACT) in pregnancy in an area of multidrug resistance has potential to impact significantly on maternal and infant health.

Background: Artesunate-amodiaquine (AS-AQ) is one of the most widely used artemisinin-based combination therapies (ACTs) to treat uncomplicated Plasmodium falciparum malaria in Africa. We investigated the impact of different dosing strategies on the efficacy of this combination for the treatment of falciparum malaria., Methods: Individual patient data from AS-AQ clinical trials were pooled using the WorldWide Antimalarial Resistance Network (WWARN) standardised methodology. Risk factors for treatment failure were identified using a Cox regression model with shared frailty across study sites., Results: Forty-three studies representing 9,106 treatments from 1999-2012 were included in the analysis; 4,138 (45.4%) treatments were with a fixed dose combination with an AQ target dose of 30 mg/kg (FDC), 1,293 (14.2%) with a non-fixed dose combination with an AQ target dose of 25 mg/kg (loose NFDC-25), 2,418 (26.6%) with a non-fixed dose combination with an AQ target dose of 30 mg/kg (loose NFDC-30), and the remaining 1,257 (13.8%) with a co-blistered non-fixed dose combination with an AQ target dose of 30 mg/kg (co-blistered NFDC). The median dose of AQ administered was 32.1 mg/kg [IQR: 25.9-38.2], the highest dose being administered to patients treated with co-blistered NFDC (median = 35.3 mg/kg [IQR: 30.6-43.7]) and the lowest to those treated with loose NFDC-25 (median = 25.0 mg/kg [IQR: 22.7-25.0]). Patients treated with FDC received a median dose of 32.4 mg/kg [IQR: 27-39.0]. After adjusting for reinfections, the corrected antimalarial efficacy on day 28 after treatment was similar for co-blistered NFDC (97.9% [95% confidence interval (CI): 97.0-98.8%]) and FDC (98.1% [95% CI: 97.6%-98.5%]; P = 0.799), but significantly lower for the loose NFDC-25 (93.4% [95% CI: 91.9%-94.9%]), and loose NFDC-30 (95.0% [95% CI: 94.1%-95.9%]) (P < 0.001 for all comparisons). After controlling for age, AQ dose, baseline parasitemia and region; treatment with loose NFDC-25 was associated with a 3.5-fold greater risk of recrudescence by day 28 (adjusted hazard ratio, AHR = 3.51 [95% CI: 2.02-6.12], P < 0.001) compared to FDC, and treatment with loose NFDC-30 was associated with a higher risk of recrudescence at only three sites., Conclusions: There was substantial variation in the total dose of amodiaquine administered in different AS-AQ combination regimens. Fixed dose AS-AQ combinations ensure optimal dosing and provide higher antimalarial treatment efficacy than the loose individual tablets in all age categories.

Tetrahydrobiopterin (BH₄) is a co-factor required for catalytic activity of nitric oxide synthase (NOS) and amino acid-monooxygenases, including phenylalanine hydroxylase. BH4 is unstable: during oxidative stress it is non-enzymatically oxidized to dihydrobiopterin (BH₂), which inhibits NOS. Depending on BH₄ availability, NOS oscillates between NO synthase and NADPH oxidase: as the BH₄/BH₂ ratio decreases, NO production falls and is replaced by superoxide. In African children and Asian adults with severe malaria, NO bioavailability decreases and plasma phenylalanine increases, together suggesting possible BH₄ deficiency. The primary three biopterin metabolites (BH₄, BH₂ and B₀ [biopterin]) and their association with disease severity have not been assessed in falciparum malaria. We measured pterin metabolites in urine of adults with severe falciparum malaria (SM; n=12), moderately-severe malaria (MSM, n=17), severe sepsis (SS; n=5) and healthy subjects (HC; n=20) as controls. In SM, urinary BH₄ was decreased (median 0.16 ¼mol/mmol creatinine) compared to MSM (median 0.27), SS (median 0.54), and HC (median 0.34)]; p<0.001. Conversely, BH₂ was increased in SM (median 0.91 ¼mol/mmol creatinine), compared to MSM (median 0.67), SS (median 0.39), and HC (median 0.52); p<0.001, suggesting increased oxidative stress and insufficient recycling of BH2 back to BH4 in severe malaria. Overall, the median BH₄/BH₂ ratio was lowest in SM [0.18 (IQR: 0.04-0.32)] compared to MSM (0.45, IQR 0.27-61), SS (1.03; IQR 0.54-2.38) and controls (0.66; IQR 0.43-1.07); p<0.001. In malaria, a lower BH₄/BH₂ ratio correlated with decreased microvascular reactivity (r=0.41; p=0.03) and increased ICAM-1 (r=-0.52; p=0.005). Decreased BH4 and increased BH₂ in severe malaria (but not in severe sepsis) uncouples NOS, leading to impaired NO bioavailability and potentially increased oxidative stress. Adjunctive therapy to regenerate BH4 may have a role in improving NO bioavailability and microvascular perfusion in severe falciparum malaria.