Acute respiratory infections caused by bacterial or viral pathogens are among the most common reasons for seeking medical care. Despite improvements in pathogen-based diagnostics, most patients receive inappropriate antibiotics. Host response biomarkers offer an alternative diagnostic approach to direct antimicrobial use. This observational, cohort study determined whether host gene expression patterns discriminate non-infectious from infectious illness, and bacterial from viral causes of acute respiratory infection in the acute care setting. Peripheral whole blood gene expression from 273 subjects with community-onset acute respiratory infection (ARI) or non-infectious illness as well as 44 healthy controls was measured using microarrays. Sparse logistic regression was used to develop classifiers for bacterial ARI (71 probes), viral ARI (33 probes), or a non-infectious cause of illness (26 probes). Overall accuracy was 87% (238/273 concordant with clinical adjudication), which was more accurate than procalcitonin (78%, p<0.03) and three published classifiers of bacterial vs. viral infection (78-83%). The classifiers developed here externally validated in five publicly available datasets (AUC 0.90-0.99). A sixth publically available dataset included twenty-five patients with co-identification of bacterial and viral pathogens. Applying the ARI classifiers defined four distinct groups: a host response to bacterial ARI; viral ARI; co-infection; and neither a bacterial nor viral response. These findings create an opportunity to develop and utilize host gene expression classifiers as diagnostic platforms to combat inappropriate antibiotic use and emerging antibiotic resistance.

The cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4; CD 152) is a negative regulator of T-lymphocyte activation. Particular genotypes of the locus encoding the CTLA-4 glycoprotein have been associated with susceptibility to various autoimmune diseases. To determine their role in susceptibility to systemic lupus erythematosus (SLE), we genotyped 130 patients and 200 ethnically matched controls for allelic determinants at four polymorphic sites, viz., three in the promoter region at positions -1722 (T/C), -1661 (A/G), and -318 (C/T), and one within the first exon at position +49 (A/G), by restriction fragment length polymorphism methods. All genotype frequencies were in Hardy-Weinberg equilibrium. The genotypes at position -1722 were significantly associated with SLE. The frequency of T/T homozygotes was higher in patients than in controls (56% vs 33%, P=0.00003). Conversely, the frequencies of C/C homozygotes and C/T heterozygotes were higher in controls than in patients (15.5% vs 7%, P=0.019; 51.5% vs 37%, P=0.009). Genotypes at positions +49, -318, or -1661 were not significantly associated with SLE. These results show that allelic variation at the -1722 site influences susceptibility to SLE. This is the first report to our knowledge implicating CTLA-4 genotypes at the -1722 locus in susceptibility to any disease.

Objective. Epstein-Barr virus (EBV) is hypothesized to play a role in the development of systemic lupus erythematosus (SLE). Cytotoxic T lymphocyteassociated antigen 4 (CTLA-4) is important in regulating T cell-mediated immunity, encompassing the first line of response to viral infections, and genetic variation in CTLA-4 has been associated with SLE. This study examined the seroprevalence of EBV in a populationbased study of SLE patients from the southeastern United States, and potential interactions with CTLA-4 polymorphisms were assessed.Methods. Cases comprised 230 subjects recently diagnosed as having SLE (144 African American and 86 white) from university and community-based clinics, and controls comprised 276 age-, sex-, and statematched subjects (72 African American and 204 white) recruited from driver's license registries. Antibodies to EBV capsid antigen were determined by enzyme-linked immunosorbent assay, with results expressed as positive or negative using the international standardized ratio (ISR) (a ratio of the sample absorbance to a known standard). CTLA-4 genotypes were identified by polymerase chain reaction-based methods.Results. In African Americans, EBV-IgA seroprevalence was strongly associated with SLE (odds ratio [OR] 5.6, 95% confidence interval [95% CI] 3.0-10.6). In whites, the modest association of SLE with EBV-IgA (OR 1.6) was modified by age, in that the strongest association was observed in those older than age 50 years (OR 4.1, 95% CI 1.6-10.4). The seroprevalence of EBV-IgM and that of EBV-IgG were not associated with SLE. Higher EBV-IgG absorbance ratios were observed in SLE patients, with a significant dose response across units of the ISR in African Americans (P < 0.0001). Allelic variation in the CTLA-4 gene promoter (؊1661A/G) significantly modified the association between SLE and EBV-IgA (P ‫؍‬ 0.03), with a stronger association among those with the ؊1661AA genotype.Conclusion. These findings suggest that repeated or reactivated EBV infection, which results in increased EBV-IgA seroprevalence and higher IgG antibody titers, may be associated with SLE, and that the CTLA-4 genotype influences immune responsiveness to EBV in SLE patients. The observed patterns of effect modification by race, age, and CTLA-4 genotype should be examined in other studies and may help frame new hypotheses regarding the role of EBV in SLE etiology.

Background Early and accurate identification of individuals with viral infections is crucial for clinical management and public health interventions. We aimed to assess the ability of transcriptomic biomarkers to identify naturally acquired respiratory viral infection before typical symptoms are present. Methods In this index-cluster study, we prospectively recruited a cohort of undergraduate students (aged 18–25 years) at Duke University (Durham, NC, USA) over a period of 5 academic years. To identify index cases, we monitored students for the entire academic year, for the presence and severity of eight symptoms of respiratory tract infection using a daily web-based survey, with symptoms rated on a scale of 0–4. Index cases were defined as individuals who reported a 6-point increase in cumulative daily symptom score. Suspected index cases were visited by study staff to confirm the presence of reported symptoms of illness and to collect biospecimen samples. We then identified clusters of close contacts of index cases (ie, individuals who lived in close proximity to index cases, close friends, and partners) who were presumed to be at increased risk of developing symptomatic respiratory tract infection while under observation. We monitored each close contact for 5 days for symptoms and viral shedding and measured transcriptomic responses at each timepoint each day using a blood-based 36-gene RT-PCR assay. Findings Between Sept 1, 2009, and April 10, 2015, we enrolled 1465 participants. Of 264 index cases with respiratory tract infection symptoms, 150 (57%) had a viral cause confirmed by RT-PCR. Of their 555 close contacts, 106 (19%) developed symptomatic respiratory tract infection with a proven viral cause during the observation window, of whom 60 (57%) had the same virus as their associated index case. Nine viruses were detected in total. The transcriptomic assay accurately predicted viral infection at the time of maximum symptom severity (mean area under the receiver operating characteristic curve [AUROC] 0·94 [95% CI 0·92–0·96]), as well as at 1 day (0·87 [95% CI 0·84–0·90]), 2 days (0·85 [0·82–0·88]), and 3 days (0·74 [0·71–0·77]) before peak illness, when symptoms were minimal or absent and 22 (62%) of 35 individuals, 25 (69%) of 36 individuals, and 24 (82%) of 29 individuals, respectively, had no detectable viral shedding. Interpretation Transcriptional biomarkers accurately predict and diagnose infection across diverse viral causes and stages of disease and thus might prove useful for guiding the administration of early effective therapy, quarantine decisions, and other clinical and public health interventions in the setting of endemic and pandemic infectious diseases. Funding US Defense Advanced Research Projects Agency.

SummaryExposure to influenza virus triggers a complex cascade of events in the human host. In order to understand more clearly the evolution of this intricate response over time, human volunteers were inoculated with influenza A/Wisconsin/67/2005 (H3N2), and then had serial peripheral blood samples drawn and tested for the presence of 25 major human cytokines. Nine of 17 (53%) inoculated subjects developed symptomatic influenza infection. Individuals who will go on to become symptomatic demonstrate increased circulating levels of interleukin (IL)-6, IL-8, IL-15, monocyte chemotactic protein (MCP)-1 and interferon (IFN) gammainduced protein (IP)-10 as early as 12-29 h post-inoculation (during the presymptomatic phase), whereas challenged patients who remain asymptomatic do not. Overall, the immunological pathways of leucocyte recruitment, Toll-like receptor (TLR)-signalling, innate anti-viral immunity and fever production are all over-represented in symptomatic individuals very early in disease, but are also dynamic and evolve continuously over time. Comparison with simultaneous peripheral blood genomics demonstrates that some inflammatory mediators (MCP-1, IP-10, IL-15) are being expressed actively in circulating cells, while others (IL-6, IL-8, IFN-a and IFN-g) are probable effectors produced locally at the site of infection. Interestingly, asymptomatic exposed subjects are not quiescent either immunologically or genomically, but instead exhibit early and persistent down-regulation of important inflammatory mediators in the periphery. The host inflammatory response to influenza infection is variable but robust, and evolves over time. These results offer critical insight into pathways driving influenza-related symptomatology and offer the potential to contribute to early detection and differentiation of infected hosts.

Early, presymptomatic intervention with oseltamivir (corresponding to the onset of a published host-based genomic signature of influenza infection) resulted in decreased overall influenza symptoms (aggregate symptom scores of 23.5 vs 46.3), more rapid resolution of clinical disease (20 hours earlier), reduced viral shedding (total median tissue culture infectious dose [TCID50] 7.4 vs 9.7), and significantly reduced expression of several inflammatory cytokines (interferon-γ, tumor necrosis factor-α, interleukin-6, and others). The host genomic response to influenza infection is robust and may provide the means for early detection, more timely therapeutic interventions, a meaningful reduction in clinical disease, and an effective molecular means to track response to therapy.

Cytotoxic lymphocyte antigen-4 (CTLA-4) plays an important role in regulating T cell activation, and may help to limit T cell response under conditions of inflammation. Genetic variability in CTLA-4 has been implicated in the development of several autoimmune diseases. Some studies have described associations between CTLA-4 polymorphisms and systemic lupus erythematosus (SLE), but findings have been inconsistent. We examined polymorphisms in the CTLA-4 gene promoter region (-1722T/C, -1661 A/G, -318C/T) and exon I (+49G/A) with respect to SLE in a population-based case-control study in the southeastern US. Genotypes from 230 recently diagnosed cases and 276 controls were examined separately for African-Americans and whites. We observed no overall associations between SLE and the four CTLA-4 polymorphisms examined. Subgroup analyses revealed effect modification by age for the presence of the -1661G allele, yielding a significant positive association with SLE in younger (<35 years) African-Americans (OR = 3.3). CTLA-4 genotypes also interacted with HLA-DR2 and GM allotype to contribute to risk of SLE. These findings suggest allelic variation in this region of CTLA4 is not a major independent risk factor for SLE, but may contribute to risk of disease in younger African-Americans or in the presence of certain immunogenetic markers.

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