Quantification and determinants of the amount of respiratory syncytial virus (RSV) shed using real time PCR data from a longitudinal household study.

Background A better understanding of respiratory syncytial virus (RSV) epidemiology requires realistic estimates of RSV shedding patterns, quantities shed, and identification of the related underlying factors. Methods RSV infection data arise from a cohort study of 47 households with 493 occupants, in coastal Kenya, during the 2009/2010 RSV season. Nasopharyngeal swabs were taken every 3 to 4 days and screened for RSV using a real time polymerase chain reaction (PCR) assay. The amount of virus shed was quantified by calculating the 'area under the curve' using the trapezoidal rule applied to rescaled PCR cycle threshold output. Multivariable linear regression was used to identify correlates of amount of virus shed. Results The median quantity of virus shed per infection episode was 29.4 (95% CI: 15.2, 54.2) log10 ribonucleic acid (RNA) copies. Young age (<1 year), presence of upper respiratory symptoms, intra-household acquisition of infection, an individual's first infection episode in the RSV season, and having a co-infection of RSV group A and B were associated with increased amount of virus shed. Conclusions The findings provide insight into which groups of individuals have higher potential for transmission, information which may be useful in designing RSV prevention strategies

Transmission of Respiratory Syncytial Virus in Households: Who Acquires Infection From Whom?

Households represent a setting of frequent and intense contacts and hence are conducive to the spread of respiratory viruses, such as respiratory syncytial virus (RSV). Infants are most vulnerable to severe RSV disease but a vaccine is not yet available hence the need to explore alternate strategies of protecting them. Such strategies would require better understanding of who infects the infants. During the RSV season of 2009/2010, we undertook a prospective study in rural Kenya involving 493 members of 47 households each with a child bom after the preceding RSV epidemic and at least one elder sibling. Throughout the epidemic a nasopharyngeal swab (NPS) was collected every 3-4 days irrespective of symptoms, from all household members, and tested for a range of respiratory viruses including RSV using a molecular diagnostic assay. Partial sequencing of the attachment protein (G) gene from positive swabs was used to compare RSV strains within the household. In addition, once-a-week a specimen of oral fluid (OF) from around the gums was collected for RSV-specific antibodies screening and for assessment of sensitivity of the OF in detection of RSV using molecular diagnostics. This is the first prospective study to investigate introduction and transmission of RSV in families using molecular techniques over a complete RSV season. Analysis of RSV infection data is reported in this thesis with particular interest to identifying from where infants derive their infection, estimating the duration of RSV shedding and identity factors influencing the recovery rates, and estimating parameters of RSV susceptibility and transmission probability. In addition, data on diagnostic performance of OF in detection of RSV by molecular methods is presented. A total of 16,924 NPS were collected, representing 86% of planned. RSV was detected in 40 (85%) households and 179 (36%) of the participants. In 28 of the 44 households with complete data, there was transmission of RSV to the infants experiencing their first epidemic. The probable source of RSV infection of the naive infants was a household member in at least 54% of the cases. Co-primary infection between a household member and the RSV-naive infant was ascribed in 4 of the cases. Older children were assigned the primary case for 11 (39%) of the infant cases and 10 (91%) of these were attending school. The infants appeared to play a role transmitting the introduced infections to the other members of the household including to the mothers. These findings support vaccination strategies that target school age children and pregnant women. Both of these vaccination strategies can have profound benefits to RSV naive infants directly by augmenting neutralizing antibodies against RSV (immunization of the pregnant women) and indirectly by reducing transmission from siblings to RSV-naive infants. Results from this study provide increased confidence in the rationale for RSV vaccination of individuals who are not the key targets for protection

Molecular epidemiology of human rhinovirus infections in Kilifi, coastal Kenya

This study reports pediatric surveillance over 3 years for human rhinovirus (HRV) at the District Hospital of Kilifi, coastal Kenya. Nasopharyngeal samples were collected from children presenting at outpatient clinic with no signs of acute respiratory infection, or with signs of upper respiratory tract infection, and from children admitted to the hospital with lower respiratory tract infection. Samples were screened by real-time reverse transcriptase polymerase chain reaction (real-time RT-PCR) and classified further to species by nucleotide sequencing of the VP4/VP2 junction. Of 441 HRV positives by real-time RT-PCR, 332 were classified to species, with 47% (155) being HRV-A, 5% (18) HRV-B, and 48% (159) HRV-C. There was no clear seasonal pattern of occurrence for any species. The species were present in similar proportions in the inpatient and outpatient sample sets, and no significant association between species distribution and the severity of lower respiratory tract infection in the inpatients could be determined. HRV sequence analysis revealed multiple but separate clusters in circulation particularly for HRV-A and HRV-C. Most HRV-C clusters were distinct from reference sequences downloaded from GenBank. In contrast, most HRV-A and HRV-B sequences clustered with either known serotypes or strains from elsewhere within Africa and other regions of the world. This first molecular epidemiological study of HRV in the region defines species distribution in accord with reports from elsewhere in the world, shows considerable strain diversity and does not identify an association between any species and disease severity

Continuous Invasion by Respiratory Viruses Observed in Rural Households During a Respiratory Syncytial Virus Seasonal Outbreak in Coastal Kenya.

BACKGROUND: Households are high-intensity close-contact environments favorable for transmission of respiratory viruses, yet little is known for low-income settings. METHODS: Active surveillance was completed on 47 households in rural coastal Kenya over 6 months during a respiratory syncytial virus (RSV) season. Nasopharyngeal swabs (NPSs) were taken from 483 household members twice weekly irrespective of symptoms. Using molecular diagnostics, NPSs from 6 households were screened for 15 respiratory viruses and the remainder of households only for the most frequent viruses observed: rhinovirus (RV), human coronavirus (HCoV; comprising strains 229E, OC43, and NL63), adenovirus (AdV), and RSV (A and B). RESULTS: Of 16928 NPSs tested for the common viruses, 4259 (25.2%) were positive for ≥1 target; 596 (13.8%) had coinfections. Detection frequencies were 10.5% RV (1780), 7.5% HCoV (1274), 7.3% AdV (1232), and 3.2% RSV (537). On average, each household and individual had 6 and 3 different viruses detected over the study period, respectively. Rhinovirus and HCoV were detected in all the 47 households while AdV and RSV were detected in 45 (95.7%) and 40 (85.1%) households, respectively. The individual risk of infection over the 6-month period was 93.4%, 80.1%, 71.6%, 61.5%, and 37.1% for any virus, RV, HCoV, AdV, and RSV, respectively. NPSs collected during symptomatic days and from younger age groups had higher prevalence of virus detection relative to respective counterparts. RSV was underrepresented in households relative to hospital admission data. CONCLUSIONS: In this household setting, respiratory virus infections and associated illness are ubiquitous. Future studies should address the health and economic implications of these observations

Agreement between ELISA and plaque reduction neutralisation assay in detection of respiratory syncytial virus specific antibodies in a birth Cohort from Kilifi, coastal Kenya

Background Severe disease associated with respiratory syncytial virus (RSV) infection occurs predominantly among infants under 6 months of age. Vaccines for prevention are in clinical development. Assessment of the vaccine effectiveness in large epidemiological studies requires serological assays which are rapid, economical and standardised between laboratories. The objective of this study was to assess the agreement between two enzyme linked immunosorbent assays (ELISA) and the plaque reduction neutralisation test (PRNT) in quantifying RSV specific antibodies. Methods Archived sera from 99 participants of the Kilifi Birth Cohort (KBC) study (conducted 2002-2007) were screened for RSV antibodies using 3 methods: ELISA using crude RSV lysate as antigen, a commercial RSV immunoglobulin G (IgG) ELISA kit from IBL International GmbH, and PRNT. Pearson correlation, Bland-Altman plots and regression methods were used in analysis. Results There was high positive correlation between the IBL RSV IgG ELISA and PRNT antibodies (Pearson r=0.75), and moderate positive correlation between the crude RSV lysate IgG ELISA and PRNT antibodies (r= 0.61). Crude RSV lysate IgG ELISA showed a wider 95% limit of agreement (-1.866, 6.157) with PRNT compared to the IBL RSV IgG ELISA (1.392, 7.595). Mean PRNT titres were estimated within a width of 4.8 log2PRNT and 5.6 log2PRNT at 95% prediction interval by IBL RSV IgG and crude RSV lysate IgG ELISA, respectively. Conclusion Although, the IBL RSV IgG ELISA is observed to provide a reasonable correlate for PRNT assay in detecting RSV specific antibodies, it does not provide an accurate prediction for neutralizing antibody levels. An RSV neutralising antibody level is likely to fall within 2.4 fold higher and 2.4 fold lower than the true value if IBL RSV IgG ELISA is used to replace PRNT assay. The utility of an ELISA assay in vaccine studies should be assessed independent of the PRNT method

Human rhinovirus spatial-temporal epidemiology in rural coastal Kenya, 2015-2016, observed through outpatient surveillance

Background Human rhinovirus (HRV) is the predominant cause of upper respiratory tract infections, resulting in a significant public health burden. The virus circulates as many different types (~160), each generating strong homologous, but weak heterotypic, immunity. The influence of these features on transmission patterns of HRV in the community is understudied. Methods Nasopharyngeal swabs were collected from patients with symptoms of acute respiratory infection (ARI) at nine out-patient facilities across a Health and Demographic Surveillance System between December 2015 and November 2016. HRV was diagnosed by real-time RT-PCR, and the VP4/VP2 genomic region of the positive samples sequenced. Phylogenetic analysis was used to determine the HRV types. Classification models and G-test statistic were used to investigate HRV type spatial distribution. Demographic characteristics and clinical features of ARI were also compared. Results Of 5,744 NPS samples collected, HRV was detected in 1057 (18.4%), of which 817 (77.3%) were successfully sequenced. HRV species A, B and C were identified in 360 (44.1%), 67 (8.2%) and 390 (47.7%) samples, respectively. In total, 87 types were determined: 39, 10 and 38 occurred within species A, B and C, respectively. HRV types presented heterogeneous temporal patterns of persistence. Spatially, identical types occurred over a wide distance at similar times, but there was statistically significant evidence for clustering of types between health facilities in close proximity or linked by major road networks. Conclusion This study records a high prevalence of HRV in out-patient presentations exhibiting high type diversity. Patterns of occurrence suggest frequent and independent community invasion of different types. Temporal differences of persistence between types may reflect variation in type-specific population immunity. Spatial patterns suggest either rapid spread or multiple invasions of the same type, but evidence of similar types amongst close health facilities, or along road systems, indicate type partitioning structured by local spread

Impact of viral upper respiratory tract infection on the concentration of nasopharyngeal pneumococcal carriage among Kenyan children

Viral upper respiratory tract infection (URTI) predisposes to bacterial pneumonia possibly by facilitating growth of bacteria such as Streptococcus pneumoniae colonising the nasopharynx. We investigated whether viral URTI is temporally associated with an increase in nasopharyngeal pneumococcal concentration. Episodes of symptomatic RSV or rhinovirus URTI among children <5 years were identified from a longitudinal household study in rural Kenya. lytA and alu PCR were performed on nasopharyngeal samples collected twice-weekly, to measure the pneumococcal concentration adjusted for the concentration of human DNA present. Pneumococcal concentration increased with a fold-change of 3.80 (95%CI 1.95–7.40), with acquisition of RSV or rhinovirus, during 51 URTI episodes among 42 children. In repeated swabs from the baseline period, in the two weeks before URTI developed, within-episode variation was broad; within +/−112-fold range of the geometric mean. We observed only a small increase in nasopharyngeal pneumococcal concentration during RSV or rhinovirus URTI, relative to natural variation. Other factors, such as host response to viral infection, may be more important than nasopharyngeal pneumococcal concentration in determining risk of invasive disease

RSV modelling meeting

A group of investigators gathered to review the landscape of predictive mathematical modelling of RSV intervention programmes, and to identify gaps in knowledge and strategy options being explored. The objective was to set an agenda for future modelling and related research, to explore possible areas for collaborations, and provide an informed status update for various stakeholders

Quantifying social contacts in a household setting of rural Kenya using wearable proximity sensors

International audienc

Surveillance of respiratory viruses among children attending a primary school in rural coastal Kenya

Background: Respiratory viruses are primary agents of respiratory tract diseases. Knowledge on the types and frequency of respiratory viruses affecting school-children is important in determining the role of schools in transmission in the community and identifying targets for interventions. Methods: We conducted a one-year (term-time) surveillance of respiratory viruses in a rural primary school in Kilifi County, coastal Kenya between May 2017 and April 2018. A sample of 60 students with symptoms of ARI were targeted for nasopharyngeal swab (NPS) collection weekly. Swabs were screened for 15 respiratory virus targets using real time PCR diagnostics. Data from respiratory virus surveillance at the local primary healthcare facility was used for comparison. Results: Overall, 469 students aged 2-19 years were followed up for 220 days. A total of 1726 samples were collected from 325 symptomatic students; median age of 7 years (IQR 5-11). At least one virus target was detected in 384 (22%) of the samples with a frequency of 288 (16.7%) for rhinovirus, 47 (2.7%) parainfluenza virus, 35 (2.0%) coronavirus, 15 (0.9%) adenovirus, 11 (0.6%) respiratory syncytial virus (RSV) and 5 (0.3%) influenza virus. The proportion of virus positive samples was higher among lower grades compared to upper grades (25.9% vs 17.5% respectively; χ2 = 17.2, P -value <0.001). Individual virus target frequencies did not differ by age, sex, grade, school term or class size. Rhinovirus was predominant in both the school and outpatient setting. Conclusion: Multiple respiratory viruses circulated in this rural school population. Rhinovirus was dominant in both the school and outpatient setting and RSV was of notably low frequency in the school. The role of school children in transmitting viruses to the household setting is still unclear and further studies linking molecular data to contact patterns between the school children and their households are required