RADx-UP Testing Core: Access to COVID-19 Diagnostics in Community-Engaged Research with Underserved Populations.

Research on the COVID-19 pandemic revealed a disproportionate burden of COVID-19 infection and death among underserved populations and exposed low rates of SARS-CoV-2 testing in these communities. A landmark National Institutes of Health (NIH) funding initiative, the Rapid Acceleration of Diagnostics-Underserved Populations (RADx-UP) program, was developed to address the research gap in understanding the adoption of COVID-19 testing in underserved populations. This program is the single largest investment in health disparities and community-engaged research in the history of the NIH. The RADx-UP Testing Core (TC) provides community-based investigators with essential scientific expertise and guidance on COVID-19 diagnostics. This commentary describes the first 2 years of the TC's experience, highlighting the challenges faced and insights gained to safely and effectively deploy large-scale diagnostics for community-initiated research in underserved populations during a pandemic. The success of RADx-UP shows that community-based research to increase access and uptake of testing among underserved populations can be accomplished during a pandemic with tools, resources, and multidisciplinary expertise provided by a centralized testing-specific coordinating center. We developed adaptive tools to support individual testing strategies and frameworks for these diverse studies and ensured continuous monitoring of testing strategies and use of study data. In a rapidly evolving setting of tremendous uncertainty, the TC provided essential and real-time technical expertise to support safe, effective, and adaptive testing. The lessons learned go beyond this pandemic and can serve as a framework for rapid deployment of testing in response to future crises, especially when populations are affected inequitably.

Rickettsioses as Major Etiologies of Unrecognized Acute Febrile Illness, Sabah, East Malaysia.

Orientia tsutsugamushi, spotted fever group rickettsioses, and typhus group rickettsioses (TGR) are reemerging causes of acute febrile illness (AFI) in Southeast Asia. To further delineate extent, we enrolled patients >4 weeks of age with nonmalarial AFI in Sabah, Malaysia, during 2013-2015. We confirmed rickettsioses (past or acute, IgG titer >160) in 126/354 (36%) patients. We confirmed acute rickettsioses (paired 4-fold IgG titer rise to >160) in 38/145 (26%) patients: 23 O. tsutsugamushi, 9 spotted fever group, 4 TGR, 1 O. tsutsugamushi/spotted fever group, and 1 O. tsutsugamushi/TGR. PCR results were positive in 11/319 (3%) patients. Confirmed rickettsioses were more common in male adults; agricultural/plantation work and recent forest exposure were risk factors. Dizziness and acute hearing loss but not eschars were reported more often with acute rickettsioses. Only 2 patients were treated with doxycycline. Acute rickettsioses are common (>26%), underrecognized, and untreated etiologies of AFI in East Malaysia; empirical doxycycline treatment should be considered.

Optimization and Evaluation of a Multiplex Quantitative PCR Assay for Detection of Nucleic Acids in Human Blood Samples from Patients with Spotted Fever Rickettsiosis, Typhus Rickettsiosis, Scrub Typhus, Monocytic Ehrlichiosis, and Granulocytic Anaplasmosis.

Spotted fever group rickettsioses (SFGR), typhus group rickettsioses (TGR), scrub typhus (caused by Orientia tsutsugamushi), ehrlichiosis, and anaplasmosis often present as undifferentiated fever but are not treated by agents (penicillins and cephalosporins) typically used for acute febrile illness. Inability to diagnose these infections when the patient is acutely ill leads to excess morbidity and mortality. Failure to confirm these infections retrospectively if a convalescent blood sample is not obtained also impairs epidemiologic and clinical research. We designed a multiplex real-time quantitative PCR (qPCR) assay to detect SFGR, TGR, O. tsutsugamushi, and infections caused by Anaplasma phagocytophilum and Ehrlichia chaffeensis with the ompA, 17-kDa surface antigen gene, tsa56, msp2 (p44), and vlpt gene targets, respectively. Analytical sensitivity was ≥2 copies/µl (linear range, 2 to 2 × 105) and specificity was 100%. Clinical sensitivities for SFGR, TGR, and O. tsutsugamushi were 25%, 20%, and 27%, respectively, and specificities were 98%, 99%, and 100%, respectively. Clinical sensitivities for A. phagocytophilum and E. chaffeensis were 93% and 84%, respectively, and specificities were 99% and 98%, respectively. This multiplex qPCR assay could support early clinical diagnosis and treatment, confirm acute infections in the absence of a convalescent-phase serum sample, and provide the high-throughput testing required to support large clinical and epidemiologic studies. Because replication of SFGR and TGR in endothelial cells results in very low bacteremia, optimal sensitivity of qPCR for these rickettsioses will require use of larger volumes of input DNA, which could be achieved by improved extraction of DNA from blood and/or extraction of DNA from a larger initial volume of blood.

Use of Peptide-Based Enzyme-Linked Immunosorbent Assay followed by Immunofluorescence Assay To Document Ehrlichia chaffeensis as a Cause of Febrile Illness in Nicaragua.

Ehrlichia chaffeensis, the etiologic agent of human monocytic ehrlichiosis (HME), has been extensively studied as a cause of acute febrile illness and an emerging tick-borne zoonosis in the United States. Limited data suggest its presence in other regions, including Central and South America but not Nicaragua to date. Diagnosis of E. chaffeensis infection by indirect immunofluorescence assay (IFA) is the reference standard due to its presumed high sensitivity and specificity, but IFA is impractical, variably reproducible, and cumbersome for large epidemiologic studies and for clinical diagnosis in resource-poor regions. We evaluated a high-throughput, objective peptide-based enzyme-linked immunosorbent assay (ELISA) for use alone or in combination with IFA. We found that it performed best as a screening test (sensitivity, 100%; specificity, 84%) to reduce the proportion of serum samples that were required by the more cumbersome and subjective IFA testing to <20%. Using a two-step diagnostic approach (IFA is performed if the ELISA is positive), we identified E. chaffeensis or a serologically and antigenically similar organism as a heretofore unrecognized cause of acute febrile illness in humans in Nicaragua and demonstrated the utility of the peptide ELISA as a screening tool for large-scale clinical studies.

Diagnostic methods to determine microbiology of postpartum endometritis in South Asia: laboratory methods protocol used in the Postpartum Sepsis Study: a prospective cohort study.

BACKGROUND: The South Asian region has the second highest risk of maternal death in the world. To prevent maternal deaths due to sepsis and to decrease the maternal mortality ratio as per the World Health Organization Millenium Development Goals, a better understanding of the etiology of endometritis and related sepsis is required. We describe microbiological laboratory methods used in the maternal Postpartum Sepsis Study, which was conducted in Bangladesh and Pakistan, two populous countries in South Asia. METHODS/DESIGN: Postpartum maternal fever in the community was evaluated by a physician and blood and urine were collected for routine analysis and culture. If endometritis was suspected, an endometrial brush sample was collected in the hospital for aerobic and anaerobic culture and molecular detection of bacterial etiologic agents (previously identified and/or plausible). DISCUSSION: The results emanating from this study will provide microbiologic evidence of the etiology and susceptibility pattern of agents recovered from patients with postpartum fever in South Asia, data critical for the development of evidence-based algorithms for management of postpartum fever in the region.

Respiratory syncytial virus and seasonal influenza cause similar illnesses in children with sickle cell disease.

BACKGROUND: Respiratory syncytial virus (RSV) is a cause of acute chest syndrome (ACS) in sickle cell disease (SCD), but its clinical course and acute complications have not been well characterized. We compared RSV to seasonal influenza infections in children with SCD. PROCEDURE: We defined cases as laboratory-confirmed RSV or seasonal influenza infection in inpatients and outpatients <18 years of age with SCD from 1 September 1993 to 30 June 2011. We used Fisher's exact test to compare proportions, Student's t-test or Wilcoxon rank-sum test to compare continuous variables, and logistic regression to evaluate associations. RESULTS: We identified 64 children with RSV and 91 with seasonal influenza. Clinical symptoms, including fever, cough, and rhinorrhea were similar for RSV and influenza, as were complications, including ACS and treatments for SCD. In a multivariable logistic regression model, older age (OR 1.2 per year, 95% CI [1.02-1.5], P = 0.04), increased white blood cell count at presentation (OR 1.1 per 1,000/µl increase, 95% CI [1.03-1.3], P = 0.008), and a history of asthma (OR 7, 95% [CI 1.3-37], P = 0.03) were independently associated with increased risk of ACS in children with RSV. The hospitalization rate for children with SCD and RSV (40 per 1,000 <5 years and 63 per 1,000 <2 years) greatly exceeds the general population (3 in 1,000 <5 years). CONCLUSIONS: We conclude that RSV infection is often associated with ACS and similar in severity to influenza infection in febrile children with SCD.

Emerging Tick-borne Infections in the Upper Midwest and Northeast United States Among Patients With Suspected Anaplasmosis.

Background: Emerging tick-transmitted illnesses are increasingly recognized in the United States (US). To identify multiple potential tick-borne pathogens in patients from the Upper Midwest and Northeast US with suspected anaplasmosis, we used state-of-the-art methods (polymerase chain reaction [PCR] and paired serology) to test samples from patients in whom anaplasmosis had been excluded. Methods: Five hundred sixty-eight patients without anaplasmosis had optimal samples available for confirmation of alternative tick-borne pathogens, including PCR and/or paired serology (acute-convalescent interval ≤42 days). Results: Among 266 paired serology evaluations, for which the median acute-convalescent sampling interval was 28 (interquartile range, 21-33) days, we identified 35 acute/recent infections (24 [9%] Borrelia burgdorferi; 6 [2%] Ehrlichia chaffeensis/Ehrlichia muris subsp eauclairensis [EC/EME]; 3 [1%] spotted fever group rickettsioses [SFGR], and 2 [<1%] Babesia microti) in 33 (12%) patients. Two had concurrent or closely sequential infections (1 B burgdorferi and EC/EME, and 1 B burgdorferi and SFGR). Using multiplex PCR and reverse-transcription PCR, we identified 7 acute infections (5/334 [1%] Borrelia miyamotoi and 2/334 [1%] B microti) in 5 (1%) patients, including 2 with B microti-B miyamotoi coinfection, but no Borrelia mayonii, SFGR, Candidatus Anaplasma capra, Heartland virus, or Powassan virus infections. Thus, among 568 patients with ruled-out anaplasmosis, 38 (6.7%) had ≥1 agent of tick-borne illness identified, with 33 patients (35 infections) diagnosed by paired serology and 5 additional patients (7 infections) by PCR. Conclusions: By identifying other tick-borne agents in patients in whom anaplasmosis had been excluded, we demonstrate that emerging tick-borne infections will be identified if specifically sought.

Multiplex 5' nuclease quantitative real-time PCR for clinical diagnosis of malaria and species-level identification and epidemiologic evaluation of malaria-causing parasites, including Plasmodium knowlesi.

Molecular diagnosis of malaria offers many potential advantages over microscopy, including identification of malaria to the species level in an era with few experienced microscopists. We developed high-throughput multiplex 5' nuclease quantitative PCR (qPCR) assays, with the potential to support large studies, to specifically identify Plasmodium falciparum, P. vivax, P. ovale, P. malariae, and P. knowlesi. We compared qPCR to microscopy and confirmed discordant results with an alternative target PCR assay. The assays specifically detected 1 to 6 parasites/µl of blood. The clinical sensitivities (95% confidence intervals [CIs]) of the 4-plex assay to detect microscopically confirmed malaria were 95.8% (88.3 to 99.1%) for P. falciparum, 89.5% (75.2 to 97.1%) for P. vivax, 94.1% (71.3 to 99.9%) for P. ovale, and 100% (66.4 to 100%) for P. malariae. The specificities (95% CIs) were 98.6% (92.4 to 100%) for P. falciparum, 99% (84.8 to 100%) for P. vivax, 98.4% (94.4 to 99.8%) for P. ovale, and 99.3% (95.9 to 100%) for P. malariae. The clinical specificity for samples without malaria was 100%. The clinical sensitivity of the 5-plex assay for confirmed P. knowlesi malaria was 100% (95% CI, 69.2 to 100%), and the clinical specificity was 100% (95% CI, 87.2 to 100%). Coded retesting and testing with an alternative target PCR assay showed improved sensitivity and specificity of multiplex qPCR versus microscopy. Additionally, 91.7% (11/12) of the samples with uncertain species by microscopy were identified to the species level identically by both our multiplex qPCR assay and the alternative target PCR assay, including 9 P. falciparum infections. Multiplex qPCR can rapidly and simultaneously identify all 5 Plasmodium species known to cause malaria in humans, and it offers an alternative or adjunct to microscopy for clinical diagnosis as well as a needed high-throughput tool for research.

Use of Structured Electronic Health Records Data Elements for the Development of Computable Phenotypes to Identify Potential Adverse Events Associated with Intravenous Immunoglobulin Infusion.

INTRODUCTION: Detection of adverse reactions to drugs and biologic agents is an important component of regulatory approval and post-market safety evaluation. Real-world data, including insurance claims and electronic health records data, are increasingly used for the evaluation of potential safety outcomes; however, there are different types of data elements available within these data resources, impacting the development and performance of computable phenotypes for the identification of adverse events (AEs) associated with a given therapy. OBJECTIVE: To evaluate the utility of different types of data elements to the performance of computable phenotypes for AEs. METHODS: We used intravenous immunoglobulin (IVIG) as a model therapeutic agent and conducted a single-center, retrospective study of 3897 individuals who had at least one IVIG administration between 1 January 2014 and 31 December 2019. We identified the potential occurrence of four different AEs, including two proximal AEs (anaphylaxis and heart rate alterations) and two distal AEs (thrombosis and hemolysis). We considered three different computable phenotypes: (1) an International Classification of Disease (ICD)-based phenotype; (2) a phenotype-based on EHR-derived contextual information based on structured data elements, including laboratory values, medication administrations, or vital signs; and (3) a compound phenotype that required both an ICD code for the AE in combination with additional EHR-derived structured data elements. We evaluated the performance of each of these computable phenotypes compared with chart review-based identification of AEs, assessing the positive predictive value (PPV), specificity, and estimated sensitivity of each computable phenotype method. RESULTS: Compound computable phenotypes had a high positive predictive value for acute AEs such as anaphylaxis and bradycardia or tachycardia; however, few patients had both ICD codes and the relevant contextual data, which decreased the sensitivity of these computable phenotypes. In contrast, computable phenotypes for distal AEs (i.e., thrombotic events or hemolysis) frequently had ICD codes for these conditions in the absence of an AE due to a prior history of such events, suggesting that patient medical history of AEs negatively impacted the PPV of computable phenotypes based on ICD codes. CONCLUSIONS: These data provide evidence for the utility of different structured data elements in computable phenotypes for AEs. Such computable phenotypes can be used across different data sources for the detection of infusion-related adverse events.

Host-response transcriptional biomarkers accurately discriminate bacterial and viral infections of global relevance.

Diagnostic limitations challenge management of clinically indistinguishable acute infectious illness globally. Gene expression classification models show great promise distinguishing causes of fever. We generated transcriptional data for a 294-participant (USA, Sri Lanka) discovery cohort with adjudicated viral or bacterial infections of diverse etiology or non-infectious disease mimics. We then derived and cross-validated gene expression classifiers including: 1) a single model to distinguish bacterial vs. viral (Global Fever-Bacterial/Viral [GF-B/V]) and 2) a two-model system to discriminate bacterial and viral in the context of noninfection (Global Fever-Bacterial/Viral/Non-infectious [GF-B/V/N]). We then translated to a multiplex RT-PCR assay and independent validation involved 101 participants (USA, Sri Lanka, Australia, Cambodia, Tanzania). The GF-B/V model discriminated bacterial from viral infection in the discovery cohort an area under the receiver operator curve (AUROC) of 0.93. Validation in an independent cohort demonstrated the GF-B/V model had an AUROC of 0.84 (95% CI 0.76-0.90) with overall accuracy of 81.6% (95% CI 72.7-88.5). Performance did not vary with age, demographics, or site. Host transcriptional response diagnostics distinguish bacterial and viral illness across global sites with diverse endemic pathogens.

Unsuspected dengue and acute febrile illness in rural and semi-urban southern Sri Lanka.

Dengue virus (DENV), a globally emerging cause of undifferentiated fever, has been documented in the heavily urbanized western coast of Sri Lanka since the 1960s. New areas of Sri Lanka are now being affected, and the reported number and severity of cases have increased. To study emerging DENV in southern Sri Lanka, we obtained epidemiologic and clinical data and acute- and convalescent-phase serum samples from patients >2 years old with febrile illness. We tested paired serum samples for DENV IgG and IgM and serotyped virus by using isolation and reverse transcription PCR. We identified acute DENV infection (serotypes 2, 3, and 4) in 54 (6.3%) of 859 patients. Only 14% of patients had clinically suspected dengue; however, 54% had serologically confirmed acute or past DENV infection. DENV is a major and largely unrecognized cause of fever in southern Sri Lanka, especially in young adults.

Unsuspected rickettsioses among patients with acute febrile illness, Sri Lanka, 2007.

We studied rickettsioses in southern Sri Lanka. Of 883 febrile patients with paired serum samples, 156 (17.7%) had acute rickettsioses; rickettsioses were unsuspected at presentation. Additionally, 342 (38.7%) had exposure to spotted fever and/or typhus group rickettsioses and 121 (13.7%) scrub typhus. Increased awareness of rickettsioses and better tests are needed.

Severe pandemic H1N1 and seasonal influenza in children and young adults with sickle cell disease.

Influenza causes excess morbidity in sickle cell disease (SCD). H1N1 pandemic influenza has been severe in children. To compare H1N1 with seasonal influenza in SCD (patients younger than 22), we reviewed medical records (1993-2009). We identified 123 cases of laboratory-confirmed influenza (94 seasonal, 29 H1N1). Those with seasonal influenza were younger (median 4.4 vs 8.7 years old, P = .006) and had less asthma (24% vs 56%, P = .002). Those with H1N1 influenza more often had acute chest syndrome (ACS; 34% vs 13%, P = .01) and required intensive care (17% vs 3%, P = .02), including mechanical ventilation (10% vs 0%, P = .02). In multivariate analysis, older age (odds ratio [OR] 1.1 per year, P = .04) and H1N1 influenza (OR 3.0, P = .04) were associated with ACS, and older age (OR 1.1 per year, P = .02) and prior ACS (OR 3.3 per episode in last year, P < .006) with intensive care. Influenza, especially H1N1, causes critical illness in SCD and should be prevented.

Leptospirosis as frequent cause of acute febrile illness in southern Sri Lanka.

To determine the proportion of fevers caused by leptospirosis, we obtained serum specimens and epidemiologic and clinical data from patients in Galle, Sri Lanka, March-October 2007. Immunoglobulin M ELISA was performed on paired serum specimens to diagnose acute (seroconversion or 4-fold titer rise) or past (titer without rise) leptospirosis and seroprevalence (acute). We compared (individually) the diagnostic yield of acute-phase specimens and clinical impression with paired specimens for acute leptospirosis. Of 889 patients with paired specimens, 120 had acute leptosoirosis and 241 had past leptospirosis. The sensitivity and specificity of acute-phase serum specimens were 17.5% (95% confidence interval [CI] 11.2%-25.5%) and 69.2% (95% CI 65.5%-72.7%), respectively, and of clinical impression 22.9% (95% CI 15.4%-32.0%) and 91.7% (95% CI 89.2%-93.8%), respectively. For identifying acute leptospirosis, clinical impression is insensitive, and immunoglobulin M results are more insensitive and costly. Rapid, pathogen-based tests for early diagnosis are needed.

Multiplex 5' nuclease-quantitative PCR for diagnosis of relapsing fever in a large Tanzanian cohort.

Relapsing fever (RF) is caused by tick- and louse-borne Borrelia spp., is characterized by recurrent fever, and is often misdiagnosed as malaria. Because of submicroscopic bacteremia, microscopy can be insensitive between febrile bouts. We designed a multiplex quantitative PCR (qPCR) assay to distinguish RF Borrelia from Plasmodium falciparum and P. vivax. The assay specifically (100%) amplified pathogenic RF Borrelia (1 copy/reaction). We then tested blood from participants within a Tanzanian cohort assessed at scheduled intervals and with fever. Among 8,617 blood samples from 2,057 participants surveyed routinely, 7 (0.08%) samples and 7 (0.3%) participants had RF DNA (median, 4.4 × 10(3) copies/ml). Of 382 samples from 310 febrile persons, 15 (3.9%) samples from 13 (4.2%) participants had RF DNA (median, 7.9 × 10(2) copies/ml). Five (1.3%) samples from 4 (1.3%) participants were found to harbor Borrelia by microscopy. We conclude that multiplex qPCR holds promise for improved clinical diagnosis and epidemiologic assessment of RF.

Prevalence and density-related concordance of three diagnostic tests for malaria in a region of Tanzania with hypoendemic malaria.

Accurate malaria diagnosis has dual roles in identification of symptomatic persons for effective malaria treatment and also enumeration of asymptomatic persons who contribute to the epidemiologic determinants of transmission. Three currently used diagnostic tests, microscopy, rapid diagnostic tests (RDTs), and real-time PCR, all have different sensitivities and specificities, which are parasite density dependent. Here, we compare their concordance among 451 febrile episodes in a cohort of 2,058 children and adults followed over 6 months in a region in central Tanzania with hypoendemic malaria. Microscopy, a histidine-rich protein-based RDT, and two different real-time PCR gene probes detected Plasmodium falciparum in 20, 54, 41, and 78 episodes of fever, respectively. They had complete concordance in only 9 episodes. Real-time PCR with an 18S probe was more sensitive than with a mitochondrial probe for cytochrome b despite higher copy numbers of mitochondrial DNA. Both PCR yields were increased 4-fold by glycogen/acetate precipitation with low-speed centrifugation. Duplicate PCR increases low-density malaria detection. RDT had the highest number of unique positives, presumably from persistent antigen despite the absence of parasites, although RDT did not detect 3 parasitemias with over 1,000 parasites/µl. In a latent class analysis, real-time PCR had significantly higher sensitivity than did microscopy or RDT. Agreement between real-time PCR, RDT, and microscopy was highest in March and April, when both the P. falciparum parasite rate and parasite densities are highest. Real-time PCR is more sensitive and specific than RDT and microscopy in low-prevalence, low-parasite-density settings.

The Host Response to Viral Infections Reveals Common and Virus-Specific Signatures in the Peripheral Blood.

Viruses cause a wide spectrum of clinical disease, the majority being acute respiratory infections (ARI). In most cases, ARI symptoms are similar for different viruses although severity can be variable. The objective of this study was to understand the shared and unique elements of the host transcriptional response to different viral pathogens. We identified 162 subjects in the US and Sri Lanka with infections due to influenza, enterovirus/rhinovirus, human metapneumovirus, dengue virus, cytomegalovirus, Epstein Barr Virus, or adenovirus. Our dataset allowed us to identify common pathways at the molecular level as well as virus-specific differences in the host immune response. Conserved elements of the host response to these viral infections highlighted the importance of interferon pathway activation. However, the magnitude of the responses varied between pathogens. We also identified virus-specific responses to influenza, enterovirus/rhinovirus, and dengue infections. Influenza-specific differentially expressed genes (DEG) revealed up-regulation of pathways related to viral defense and down-regulation of pathways related to T cell and neutrophil responses. Functional analysis of entero/rhinovirus-specific DEGs revealed up-regulation of pathways for neutrophil activation, negative regulation of immune response, and p38MAPK cascade and down-regulation of virus defenses and complement activation. Functional analysis of dengue-specific up-regulated DEGs showed enrichment of pathways for DNA replication and cell division whereas down-regulated DEGs were mainly associated with erythrocyte and myeloid cell homeostasis, reactive oxygen and peroxide metabolic processes. In conclusion, our study will contribute to a better understanding of molecular mechanisms to viral infections in humans and the identification of biomarkers to distinguish different types of viral infections.

Previously Derived Host Gene Expression Classifiers Identify Bacterial and Viral Etiologies of Acute Febrile Respiratory Illness in a South Asian Population.

BACKGROUND: Pathogen-based diagnostics for acute respiratory infection (ARI) have limited ability to detect etiology of illness. We previously showed that peripheral blood-based host gene expression classifiers accurately identify bacterial and viral ARI in cohorts of European and African descent. We determined classifier performance in a South Asian cohort. METHODS: Patients ≥15 years with fever and respiratory symptoms were enrolled in Sri Lanka. Comprehensive pathogen-based testing was performed. Peripheral blood ribonucleic acid was sequenced and previously developed signatures were applied: a pan-viral classifier (viral vs nonviral) and an ARI classifier (bacterial vs viral vs noninfectious). RESULTS: Ribonucleic acid sequencing was performed in 79 subjects: 58 viral infections (36 influenza, 22 dengue) and 21 bacterial infections (10 leptospirosis, 11 scrub typhus). The pan-viral classifier had an overall classification accuracy of 95%. The ARI classifier had an overall classification accuracy of 94%, with sensitivity and specificity of 91% and 95%, respectively, for bacterial infection. The sensitivity and specificity of C-reactive protein (>10 mg/L) and procalcitonin (>0.25 ng/mL) for bacterial infection were 100% and 34%, and 100% and 41%, respectively. CONCLUSIONS: Previously derived gene expression classifiers had high predictive accuracy at distinguishing viral and bacterial infection in South Asian patients with ARI caused by typical and atypical pathogens.

Geospatial analysis of dengue emergence in rural areas in the Southern Province of Sri Lanka.

BACKGROUND: Dengue is a major cause of acute febrile illness in Sri Lanka. Dengue has historically been considered an urban disease. In 2012-2013, we documented that acute dengue was surprisingly associated with self-reported rural residence in the Southern Province of Sri Lanka. METHODS: Patients admitted with an acute febrile illness were enrolled from June 2012-May 2013 in a cross-sectional surveillance study at the largest tertiary care hospital in the Southern Province. Acute dengue was diagnosed by serology and virology testing. Site visits were performed to collect residential geographical coordinates. Spatial variation in odds of acute dengue was modeled using a spatial generalized additive model predicted onto a grid of coordinate pairs covering the Southern Province. RESULTS: Of 800 patients, 333 (41.6%) had laboratory-confirmed acute dengue. Dengue was spatially heterogeneous (local probability of acute dengue 0.26 to 0.42). There were higher than average odds of acute dengue in the rural northeast of the Southern Province and lower than average odds in the urbanized southwest of the Southern Province, including the city Galle. CONCLUSIONS: Our study further affirms the emergence of dengue in rural southern Sri Lanka and highlights both the need for real-time geospatial analyses to optimize public health activities as well as the importance of strengthening dengue surveillance in non-urban areas.

Effectiveness of the 10-valent pneumococcal conjugate vaccine against radiographic pneumonia among children in rural Bangladesh: A case-control study.

BACKGROUND: Pneumococcal conjugate vaccine (PCV) effectiveness against radiographic pneumonia in South Asia is unknown. Bangladesh introduced PCV10 in 2015 using a three dose primary series (3 + 0). We sought to measure PCV10 effectiveness for two or more vaccine doses on radiographic pneumonia among vaccine-eligible children in rural Bangladesh. METHODS: We conducted a matched case-control study over two years from 2015 to 2017 using clinic and community controls in three subdistricts of Sylhet, Bangladesh. Cases were vaccine eligible 3-35 month olds at Upazila Health Complex outpatient clinics with World Health Organization-defined radiographic primary endpoint pneumonia (radiographic pneumonia). Clinic controls were matched to cases within a one week time window by age, sex, and clinic and had an illness unlikely to be Streptococcus pneumoniae; community controls were healthy and similarly matched within a one week time window by age and sex, and distance from the clinic. We estimated adjusted vaccine effectiveness (aVE) using conditional logistic regression. RESULTS: We matched 1262 cases with 2707 clinic and 2461 community controls. Overall, aVE using clinic controls was 21.4% (95% confidence interval, -0.2%, 38.4%) for ≥2 PCV10 doses and among 3-11 month olds was 47.3% (10.5%, 69.0%) for three doses. aVE increased with higher numbers of doses in clinic control sets (p = 0.007). In contrast, aVE using community controls was 7.6% (95% confidence interval, -22.2%, 30.0%) for ≥2 doses. We found vaccine introduction in the study area faster and less variable than expected with 75% coverage on average, which reduced power. Information bias may also have affected community controls. CONCLUSIONS: Clinic control analyses show PCV10 prevented radiographic pneumonia in Bangladesh, especially among younger children receiving three doses. While both analyses were underpowered, community control enrollment - compared to clinic controls - was more difficult in a complex, pluralistic healthcare system. Future studies in comparable settings may consider alternative study designs.