Surveillance of endemic human coronaviruses (HCoV-NL63, OC43 and 229E) associated with pneumonia in Kilifi, Kenya

Introduction: Human coronaviruses (HCoVs) circulate endemically in human populations, often with seasonal variation. We describe the long-term patterns of paediatric disease associated with three of these viruses, HCoV-NL63, OC43 and 229E, in coastal Kenya. Methods: Continuous surveillance of pneumonia admissions was conducted at the Kilifi county hospital (KCH) located in the northern coastal region of Kenya. Children aged &lt;5 years admitted to KCH with clinically defined syndromic severe or very severe pneumonia were recruited. Respiratory samples were taken and tested for 15 virus targets, using real-time polymerase chain reaction. Unadjusted odds ratios were used to estimate the association between demographic and clinical characteristics and HCoV positivity. Results: From 2007 to 2019, we observed 11,445 pneumonia admissions, of which 314 (3.9%) tested positive for at least one HCoV type. There were 129 (41.1%) OC43, 99 (31.5%) 229E, 74 (23.6%) NL63 positive cases and 12 (3.8%) cases of HCoV to HCoV coinfection.  Among HCoV positive cases, 47% (n=147) were coinfected with other respiratory virus pathogens. The majority of HCoV cases were among children aged &lt;1 year (66%, n=208), though there was no age-dependence in the proportion testing positive. HCoV-OC43 was predominant of the three HCoV types throughout the surveillance period. Evidence for seasonality was not identified. Conclusions: Overall, 4% of paediatric pneumonia admissions were associated with three endemic HCoVs, with a high proportion of cases co-occurring with another respiratory virus, with no clear seasonal pattern, and with the age-distribution of cases following that of pneumonia admissions (i.e. highest in infants). These observations suggest, at most, a small severe disease contribution of endemic HCoVs in this tropical setting and offer insight into the potential future burden and epidemiological characteristics of SARS-CoV-2.</ns4:p

The Etiology of Pneumonia in HIV-uninfected Children in Kilifi, Kenya: Findings From the Pneumonia Etiology Research for Child Health (PERCH) Study

Background: In the 1980s, Streptococcus pneumoniae and Haemophilus influenzae were identified as the principal causes of severe pneumonia in children. We investigated the etiology of severe childhood pneumonia in Kenya after introduction of conjugate vaccines against H. influenzae type b, in 2001, and S. pneumoniae, in 2011. Methods: We conducted a case–control study between August 2011 and November 2013 among residents of the Kilifi Health and Demographic Surveillance System 28 days to 59 months of age. Cases were hospitalized at Kilifi County Hospital with severe or very severe pneumonia according to the 2005 World Health Organization definition. Controls were randomly selected from the community and frequency matched to cases on age and season. We tested nasal and oropharyngeal samples, sputum, pleural fluid, and blood specimens and used the Pneumonia Etiology Research for Child Health Integrated Analysis, combining latent class analysis and Bayesian methods, to attribute etiology. Results: We enrolled 630 and 863 HIV-uninfected cases and controls, respectively. Among the cases, 282 (44%) had abnormal chest radiographs (CXR positive), 33 (5%) died in hospital, and 177 (28%) had diagnoses other than pneumonia at discharge. Among CXR-positive pneumonia cases, viruses and bacteria accounted for 77% (95% CrI: 67%–85%) and 16% (95% CrI: 10%–26%) of pneumonia attribution, respectively. Respiratory syncytial virus, S. pneumoniae and H. influenza, accounted for 37% (95% CrI: 31%–44%), 5% (95% CrI: 3%–9%), and 6% (95% CrI: 2%–11%), respectively. Conclusions: Respiratory syncytial virus was the main cause of CXR-positive pneumonia. The small contribution of H. influenzae type b and pneumococcus to pneumonia may reflect the impact of vaccine introductions in this population

Surveillance of endemic human coronaviruses (HCoV-NL63, OC43 and 229E) associated with childhood pneumonia in Kilifi, Kenya.

Introduction: Human coronaviruses (HCoVs) circulate endemically in human populations, often with seasonal variation. We describe the long-term patterns of paediatric disease associated with three of these viruses, HCoV-NL63, OC43 and 229E, in coastal Kenya. Methods: Continuous surveillance of pneumonia admissions was conducted at the Kilifi county hospital (KCH) located in the northern coastal region of Kenya. Children aged <5 years admitted to KCH with clinically defined syndromic severe or very severe pneumonia were recruited. Respiratory samples were taken and tested for 15 virus targets, using real-time polymerase chain reaction. Unadjusted odds ratios were used to estimate the association between demographic and clinical characteristics and HCoV positivity. Results: From 2007 to 2019, we observed 11,445 pneumonia admissions, of which 314 (3.9%) tested positive for at least one of the HCoV types surveyed in the study. There were 129 (41.1%) OC43, 99 (31.5%) 229E, 74 (23.6%) NL63 positive cases and 12 (3.8%) cases of HCoV to HCoV coinfection.  Among HCoV positive cases, 47% (n=147) were coinfected with other respiratory virus pathogens. The majority of HCoV cases were among children aged <1 year (66%, n=208), though there was was no change in the proportion infected by age. HCoV-OC43 was predominant of the three HCoV types throughout the surveillance period. Evidence for seasonality was not identified. Conclusions: Overall, 4% of paediatric pneumonia admissions were associated with three endemic HCoVs, with a high proportion of cases co-occurring with another respiratory virus, no clear seasonal pattern, and with the age-distribution of cases following that of pneumonia admissions (i.e. highest in infants). These observations suggest, at most, a small severe disease contribution of endemic HCoVs in this tropical setting and offer insight into their potential future burden and epidemiological characteristics

Is Higher Viral Load in the Upper Respiratory Tract Associated With Severe Pneumonia? Findings From the PERCH Study.

BACKGROUND.: The etiologic inference of identifying a pathogen in the upper respiratory tract (URT) of children with pneumonia is unclear. To determine if viral load could provide evidence of causality of pneumonia, we compared viral load in the URT of children with World Health Organization-defined severe and very severe pneumonia and age-matched community controls. METHODS.: In the 9 developing country sites, nasopharyngeal/oropharyngeal swabs from children with and without pneumonia were tested using quantitative real-time polymerase chain reaction for 17 viruses. The association of viral load with case status was evaluated using logistic regression. Receiver operating characteristic (ROC) curves were constructed to determine optimal discriminatory viral load cutoffs. Viral load density distributions were plotted. RESULTS.: The mean viral load was higher in cases than controls for 7 viruses. However, there was substantial overlap in viral load distribution of cases and controls for all viruses. ROC curves to determine the optimal viral load cutoff produced an area under the curve of <0.80 for all viruses, suggesting poor to fair discrimination between cases and controls. Fatal and very severe pneumonia cases did not have higher viral load than less severe cases for most viruses. CONCLUSIONS.: Although we found higher viral loads among pneumonia cases than controls for some viruses, the utility in using viral load of URT specimens to define viral pneumonia was equivocal. Our analysis was limited by lack of a gold standard for viral pneumonia

Evaluation of Pneumococcal Load in Blood by Polymerase Chain Reaction for the Diagnosis of Pneumococcal Pneumonia in Young Children in the PERCH Study.

BACKGROUND.: Detection of pneumococcus by lytA polymerase chain reaction (PCR) in blood had poor diagnostic accuracy for diagnosing pneumococcal pneumonia in children in 9 African and Asian sites. We assessed the value of blood lytA quantification in diagnosing pneumococcal pneumonia. METHODS.: The Pneumonia Etiology Research for Child Health (PERCH) case-control study tested whole blood by PCR for pneumococcus in children aged 1-59 months hospitalized with signs of pneumonia and in age-frequency matched community controls. The distribution of load among PCR-positive participants was compared between microbiologically confirmed pneumococcal pneumonia (MCPP) cases, cases confirmed for nonpneumococcal pathogens, nonconfirmed cases, and controls. Receiver operating characteristic analyses determined the "optimal threshold" that distinguished MCPP cases from controls. RESULTS.: Load was available for 290 of 291 cases with pneumococcal PCR detected in blood and 273 of 273 controls. Load was higher in MCPP cases than controls (median, 4.0 × 103 vs 0.19 × 103 copies/mL), but overlapped substantially (range, 0.16-989.9 × 103 copies/mL and 0.01-551.9 × 103 copies/mL, respectively). The proportion with high load (≥2.2 log10 copies/mL) was 62.5% among MCPP cases, 4.3% among nonconfirmed cases, 9.3% among cases confirmed for a nonpneumococcal pathogen, and 3.1% among controls. Pneumococcal load in blood was not associated with respiratory tract illness in controls (P = .32). High blood pneumococcal load was associated with alveolar consolidation on chest radiograph in nonconfirmed cases, and with high (>6.9 log10 copies/mL) nasopharyngeal/oropharyngeal load and C-reactive protein ≥40 mg/L (both P < .01) in nonconfirmed cases but not controls. CONCLUSIONS.: Quantitative pneumococcal PCR in blood has limited diagnostic utility for identifying pneumococcal pneumonia in individual children, but may be informative in epidemiological studies

Safety of Induced Sputum Collection in Children Hospitalized With Severe or Very Severe Pneumonia.

BACKGROUND.: Induced sputum (IS) may provide diagnostic information about the etiology of pneumonia. The safety of this procedure across a heterogeneous population with severe pneumonia in low- and middle-income countries has not been described. METHODS.: IS specimens were obtained as part a 7-country study of the etiology of severe and very severe pneumonia in hospitalized children <5 years of age. Rigorous clinical monitoring was done before, during, and after the procedure to record oxygen requirement, oxygen saturation, respiratory rate, consciousness level, and other evidence of clinical deterioration. Criteria for IS contraindications were predefined and serious adverse events (SAEs) were reported to ethics committees and a central safety monitor. RESULTS.: A total of 4653 IS procedures were done among 3802 children. Thirteen SAEs were reported in relation to collection of IS, or 0.34% of children with at least 1 IS specimen collected (95% confidence interval, 0.15%-0.53%). A drop in oxygen saturation that required supplemental oxygen was the most common SAE. One child died after feeding was reinitiated 2 hours after undergoing sputum induction; this death was categorized as "possibly related" to the procedure. CONCLUSIONS.: The overall frequency of SAEs was very low, and the nature of most SAEs was manageable, demonstrating a low-risk safety profile for IS collection even among severely ill children in low-income-country settings. Healthcare providers should monitor oxygen saturation and requirements during and after IS collection, and assess patients prior to reinitiating feeding after the IS procedure, to ensure patient safety

Pertussis-Associated Pneumonia in Infants and Children From Low- and Middle-Income Countries Participating in the PERCH Study.

BACKGROUND:  Few data exist describing pertussis epidemiology among infants and children in low- and middle-income countries to guide preventive strategies. METHODS:  Children 1-59 months of age hospitalized with World Health Organization-defined severe or very severe pneumonia in 7 African and Asian countries and similarly aged community controls were enrolled in the Pneumonia Etiology Research for Child Health study. They underwent a standardized clinical evaluation and provided nasopharyngeal and oropharyngeal swabs and induced sputum (cases only) for Bordetella pertussis polymerase chain reaction. Risk factors and pertussis-associated clinical findings were identified. RESULTS:  Bordetella pertussis was detected in 53 of 4200 (1.3%) cases and 11 of 5196 (0.2%) controls. In the age stratum 1-5 months, 40 (2.3% of 1721) cases were positive, all from African sites, as were 8 (0.5% of 1617) controls. Pertussis-positive African cases 1-5 months old, compared to controls, were more often human immunodeficiency virus (HIV) uninfected-exposed (adjusted odds ratio [aOR], 2.2), unvaccinated (aOR, 3.7), underweight (aOR, 6.3), and too young to be immunized (aOR, 16.1) (all P ≤ .05). Compared with pertussis-negative African cases in this age group, pertussis-positive cases were younger, more likely to vomit (aOR, 2.6), to cough ≥14 days (aOR, 6.3), to have leukocyte counts >20 000 cells/µL (aOR, 4.6), and to have lymphocyte counts >10 000 cells/µL (aOR, 7.2) (all P ≤ .05). The case fatality ratio of pertussis-infected pneumonia cases 1-5 months of age was 12.5% (95% confidence interval, 4.2%-26.8%; 5/40); pertussis was identified in 3.7% of 137 in-hospital deaths among African cases in this age group. CONCLUSIONS:  In the postneonatal period, pertussis causes a small fraction of hospitalized pneumonia cases and deaths; however, case fatality is substantial. The propensity to infect unvaccinated infants and those at risk for insufficient immunity (too young to be vaccinated, premature, HIV-infected/exposed) suggests that the role for maternal vaccination should be considered along with efforts to reduce exposure to risk factors and to optimize childhood pertussis vaccination coverage

Standardization of Clinical Assessment and Sample Collection Across All PERCH Study Sites.

BACKGROUND.: Variable adherence to standardized case definitions, clinical procedures, specimen collection techniques, and laboratory methods has complicated the interpretation of previous multicenter pneumonia etiology studies. To circumvent these problems, a program of clinical standardization was embedded in the Pneumonia Etiology Research for Child Health (PERCH) study. METHODS.: Between March 2011 and August 2013, standardized training on the PERCH case definition, clinical procedures, and collection of laboratory specimens was delivered to 331 clinical staff at 9 study sites in 7 countries (The Gambia, Kenya, Mali, South Africa, Zambia, Thailand, and Bangladesh), through 32 on-site courses and a training website. Staff competency was assessed throughout 24 months of enrollment with multiple-choice question (MCQ) examinations, a video quiz, and checklist evaluations of practical skills. RESULTS.: MCQ evaluation was confined to 158 clinical staff members who enrolled PERCH cases and controls, with scores obtained for >86% of eligible staff at each time-point. Median scores after baseline training were ≥80%, and improved by 10 percentage points with refresher training, with no significant intersite differences. Percentage agreement with the clinical trainer on the presence or absence of clinical signs on video clips was high (≥89%), with interobserver concordance being substantial to high (AC1 statistic, 0.62-0.82) for 5 of 6 signs assessed. Staff attained median scores of >90% in checklist evaluations of practical skills. CONCLUSIONS.: Satisfactory clinical standardization was achieved within and across all PERCH sites, providing reassurance that any etiological or clinical differences observed across the study sites are true differences, and not attributable to differences in application of the clinical case definition, interpretation of clinical signs, or in techniques used for clinical measurements or specimen collection

Microscopic Analysis and Quality Assessment of Induced Sputum From Children With Pneumonia in the PERCH Study.

BACKGROUND.: It is standard practice for laboratories to assess the cellular quality of expectorated sputum specimens to check that they originated from the lower respiratory tract. The presence of low numbers of squamous epithelial cells (SECs) and high numbers of polymorphonuclear (PMN) cells are regarded as indicative of a lower respiratory tract specimen. However, these quality ratings have never been evaluated for induced sputum specimens from children with suspected pneumonia. METHODS.: We evaluated induced sputum Gram stain smears and cultures from hospitalized children aged 1-59 months enrolled in a large study of community-acquired pneumonia. We hypothesized that a specimen representative of the lower respiratory tract will contain smaller quantities of oropharyngeal flora and be more likely to have a predominance of potential pathogens compared to a specimen containing mainly saliva. The prevalence of potential pathogens cultured from induced sputum specimens and quantity of oropharyngeal flora were compared for different quantities of SECs and PMNs. RESULTS.: Of 3772 induced sputum specimens, 2608 (69%) had 25 PMNs per LPF, measures traditionally associated with specimens from the lower respiratory tract in adults. Using isolation of low quantities of oropharyngeal flora and higher prevalence of potential pathogens as markers of higher quality, 25 PMNs per LPF) was the microscopic variable most associated with high quality of induced sputum. CONCLUSIONS.: Quantity of SECs may be a useful quality measure of induced sputum from young children with pneumonia

The Diagnostic Utility of Induced Sputum Microscopy and Culture in Childhood Pneumonia.

BACKGROUND.: Sputum microscopy and culture are commonly used for diagnosing the cause of pneumonia in adults but are rarely performed in children due to difficulties in obtaining specimens. Induced sputum is occasionally used to investigate lower respiratory infections in children but has not been widely used in pneumonia etiology studies. METHODS.: We evaluated the diagnostic utility of induced sputum microscopy and culture in patients enrolled in the Pneumonia Etiology Research for Child Health (PERCH) study, a large study of community-acquired pneumonia in children aged 1-59 months. Comparisons were made between induced sputum samples from hospitalized children with radiographically confirmed pneumonia and children categorized as nonpneumonia (due to the absence of prespecified clinical and laboratory signs and absence of infiltrate on chest radiograph). RESULTS.: One induced sputum sample was available for analysis from 3772 (89.1%) of 4232 suspected pneumonia cases enrolled in PERCH. Of these, sputum from 2608 (69.1%) met the quality criterion of <10 squamous epithelial cells per low-power field, and 1162 (44.6%) had radiographic pneumonia. Induced sputum microscopy and culture results were not associated with radiographic pneumonia, regardless of prior antibiotic use, stratification by specific bacteria, or interpretative criteria used. CONCLUSIONS.: The findings of this study do not support the culture of induced sputum specimens as a diagnostic tool for pneumonia in young children as part of routine clinical practice