Additional Diagnostic Yield of Adding Serology to PCR in Diagnosing Viral Acute Respiratory Infections in Kenyan Patients 5 Years of Age and Older

Abstract

The role of serology in the setting of PCR-based diagnosis of acute respiratory infections (ARIs) is unclear. We found that acuteand convalescent-phase paired-sample serologic testing increased the diagnostic yield of naso/oropharyngeal swabs for influenza virus, respiratory syncytial virus (RSV), human metapneumovirus, adenovirus, and parainfluenza viruses beyond PCR by 0.4% to 10.7%. Although still limited for clinical use, serology, along with PCR, can maximize etiologic diagnosis in epidemiologic studies. Serology was until recently the principal method for diagnosing viral pneumonia (1-6). Serology, however, can yield false-negative results if the collection of acute-or convalescent-phase blood samples is delayed or too early (4). Moreover, serology is logistically cumbersome and clinically uninformative, because people must return several weeks after the acute illness has resolved (if they survive) for convalescent-phase blood collection. The advent of nucleic acid-based tests (e.g., PCR) of upper respiratory tract specimens offers sensitive, timely diagnostic alternatives (6). The role of serologic testing in the setting of PCR-based diagnosis is unclear.We enrolled persons that were Ն5 years old (median, 11.0 years; range, 5 to 81 years) presenting with acute respiratory illness, defined as cough, difficulty breathing, or chest pain and a documented temperature of Ն38.0°C or oxygen saturation of Ͻ90%, in rural western Kenya from 1 January 2009 and 28 February 2010 (7). At the time of presentation to the clinic, we collected acute-phase blood and nasopharyngeal and oropharyngeal swab specimens in viral transport media. Patients were asked to return 4 to 6 weeks later for a convalescent-phase serum blood draw. Quantitative real-time reverse transcription-PCR (qRT-PCR) of specimens was carried out for adenovirus, respiratory syncytial virus (RSV), human metapneumovirus (hMPV), influenza A and B viruses, and parainfluenza virus (PIV) types 1 to 3 using singleplex qRT-PCR assay (7,8). The qRT-PCR results were considered positive if the threshold cycle (C T ) was Ͻ40.0. Paired sera were tested for antibodies to influenza viruses using hemagglutination inhibition assays and for antibodies to RSV, hMPV, adenovirus, and PIV using standard methods for indirect IgG enzyme-linked immunoassay, in which all assays were optimized for reagent/ diluent composition and concentration and incubation time/ temperatures to obtain the highest limit of detection for anti- a The study was performed on samples collected from individuals in western Kenya. The samples were collected 1 January 2009 to 28 February 2010. The median number of days between acute-and convalescent-phase sera was 42, and the range was 32 to 72 days. b Although the acute and convalescent-phase sera for 232 patients were tested for all viruses, the serum samples for fewer of these patients had qRT-PCR results for some pathogens, resulting in lower denominators (i.e., 203 or 204). c One patient was positive for both parainfluenza virus 1 and parainfluenza vir...

Serology Enhances Molecular Diagnosis of Respiratory Virus Infections Other than Influenza in Children and Adults Hospitalized with Community-Acquired Pneumonia

“…The proportion of diagnostic seroresponses was higher for all patients with high virus loads (lower C T values), which would be predicted for primary virus infections and is consistent with case-control studies that have shown higher virus loads among RT-PCR-positive symptomatic patients than among matched asymptomatic controls (9,16,21). The relative prevalences of virus detection and patterns of age distribution found in this study were similar for RT-PCR and serology and were consistent with those reported in other CAP studies (9,10,19). Finally, for most viruses, positive test results obtained by both methods were temporally concordant with each other and with the seasonal circulation patterns of the respective respiratory viruses described by regional NREVSS data over the study period.…”

“…Molecular studies of respiratory virus infections that have included serology have generally increased the number of acute infections identified (12,19,20). However, the added diagnostic value of serology for the etiologic assessment of CAP in U.S. populations has not been fully evaluated.…”

Use of Multiple Imputation to Estimate the Proportion of Respiratory Virus Detections Among Patients Hospitalized With Community-Acquired Pneumonia

“…Serologic testing for select viruses (AdV, HMPV, influenza A and B, PIV, and RSV) was performed on available paired acute and convalescent serum specimens. Influenza serology used both hemagglutination inhibition and microneutralization assays to define positive results [14, 15], and serology for other viruses was based on CDC-developed indirect immunoglobulin (Ig)G enzyme immunoassays [11, 12]. Bacterial culture was performed on specimens from blood, sputum, pleural fluid, endotracheal aspirates, and bronchoalveolar lavage.…”

“…Real-time polymerase chain reaction (PCR) assays are most commonly performed on respiratory specimens in etiology studies, because they have improved sensitivity and turnaround time for respiratory virus detection relative to culture [6–10]. Although not timely, serology may capture infections even if a virus is not sufficiently present to be detected by PCR in respiratory specimens; thus, acute and convalescent (paired sera) serology results can increase the diagnostic yield for respiratory viruses in CAP studies [3, 4, 11–13]. However, even in the research setting, paired sera are not always available from each patient, particularly because convalescent serology requires specimen collection posthospital discharge or death.…”

Respiratory Syncytial Virus: Infection, Detection, and New Options for Prevention and Treatment

“…Acute-and convalescent-phase serology paired with PCR increases the diagnostic yield of RSV, in patients 5 years of age and older, from 8.3% with PCR alone to 11.3% with acute-and convalescent-phase serology (210). Enzyme immunoassays (EIAs) against envelope glycoproteins have a sensitivity greater than HEp-2 tube culture.…”