Reduced Transplacental Transfer of Group B Streptococcus Surface Protein Antibodies in HIV-infected Mother-Newborn Dyads.

We evaluated the effect of maternal HIV infection on transplacental antibody transfer specific to 8 group B Streptococcus (GBS) surface proteins among 81 HIV-uninfected and 83 HIV-infected mother-newborn pairs using a multiplex immunoassay. Significantly lower antibody titers were detected in HIV-infected mothers and HIV-exposed uninfected newborns compared to HIV-uninfected mother-newborn dyads. Maternal HIV infection was also associated with reduced transplacental transfer of antibodies for Sip (25.8%), Foldase (30.4%), gba0392 (36.5%), gbs0393 (32.9%), gbs1539 (39.2%), gbs2106 (35.7%), and BibA (19.4%); P < .003. This reduced transplacental antibody might contribute to increased susceptibility for invasive GBS disease in HIV-exposed uninfected infants.

The Incremental Value of Repeated Induced Sputum and Gastric Aspirate Samples for the Diagnosis of Pulmonary Tuberculosis in Young Children With Acute Community-Acquired Pneumonia.

BACKGROUND.: Mycobacterium tuberculosis (Mtb) contributes to the pathogenesis of childhood acute community-acquired pneumonia in settings with a high tuberculosis burden. The incremental value of a repeated induced sputum (IS) sample, compared with a single IS or gastric aspirate (GA) sample, is not well known. METHODS.: Two IS samples were obtained for Mtb culture from children enrolled as cases in the Pneumonia Etiology Research for Child Health (PERCH) study in South Africa. Nonstudy attending physicians requested GA if pulmonary tuberculosis was clinically suspected. We compared the Mtb yield of 2 IS samples to that of 1 IS sample and GA samples. RESULTS: . Twenty-seven (3.0%) culture-confirmed pulmonary tuberculosis cases were identified among 906 children investigated with IS and GA samples for Mtb. Results from 2 IS samples were available for 719 children (79.4%). Of 12 culture-confirmed pulmonary tuberculosis cases identified among children with ≥2 IS samples, 4 (33.3%) were negative at the first IS sample. In head-to-head comparisons among children with both GA and IS samples collected, the yield of 1 GA sample (8 of 427; 1.9%) was similar to that of 1 IS sample (5 of 427, 1.2%), and the yield of 2 GA samples (10 of 300; 3.3%) was similar to that of 2 IS samples (5 of 300; 1.7%). IS samples identified 8 (42.1%) of the 19 culture-confirmed pulmonary tuberculosis cases that were identified through submission of IS and GA samples. CONCLUSIONS.: A single IS sample underestimated the presence of Mtb in children hospitalized with severe or very severe pneumonia. Detection of Mtb is enhanced by combining 2 IS with GA sample collections in young children with acute severe pneumonia.

Standardization of Laboratory Methods for the PERCH Study.

The Pneumonia Etiology Research for Child Health study was conducted across 7 diverse research sites and relied on standardized clinical and laboratory methods for the accurate and meaningful interpretation of pneumonia etiology data. Blood, respiratory specimens, and urine were collected from children aged 1-59 months hospitalized with severe or very severe pneumonia and community controls of the same age without severe pneumonia and were tested with an extensive array of laboratory diagnostic tests. A standardized testing algorithm and standard operating procedures were applied across all study sites. Site laboratories received uniform training, equipment, and reagents for core testing methods. Standardization was further assured by routine teleconferences, in-person meetings, site monitoring visits, and internal and external quality assurance testing. Targeted confirmatory testing and testing by specialized assays were done at a central reference laboratory.

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.

The Effect of Antibiotic Exposure and Specimen Volume on the Detection of Bacterial Pathogens in Children With Pneumonia.

BACKGROUND.: Antibiotic exposure and specimen volume are known to affect pathogen detection by culture. Here we assess their effects on bacterial pathogen detection by both culture and polymerase chain reaction (PCR) in children. METHODS.: PERCH (Pneumonia Etiology Research for Child Health) is a case-control study of pneumonia in children aged 1-59 months investigating pathogens in blood, nasopharyngeal/oropharyngeal (NP/OP) swabs, and induced sputum by culture and PCR. Antibiotic exposure was ascertained by serum bioassay, and for cases, by a record of antibiotic treatment prior to specimen collection. Inoculated blood culture bottles were weighed to estimate volume. RESULTS.: Antibiotic exposure ranged by specimen type from 43.5% to 81.7% in 4223 cases and was detected in 2.3% of 4863 controls. Antibiotics were associated with a 45% reduction in blood culture yield and approximately 20% reduction in yield from induced sputum culture. Reduction in yield of Streptococcus pneumoniae from NP culture was approximately 30% in cases and approximately 32% in controls. Several bacteria had significant but marginal reductions (by 5%-7%) in detection by PCR in NP/OP swabs from both cases and controls, with the exception of S. pneumoniae in exposed controls, which was detected 25% less frequently compared to nonexposed controls. Bacterial detection in induced sputum by PCR decreased 7% for exposed compared to nonexposed cases. For every additional 1 mL of blood culture specimen collected, microbial yield increased 0.51% (95% confidence interval, 0.47%-0.54%), from 2% when volume was ≤1 mL to approximately 6% for ≥3 mL. CONCLUSIONS.: Antibiotic exposure and blood culture volume affect detection of bacterial pathogens in children with pneumonia and should be accounted for in studies of etiology and in clinical management.

Detection of Pneumococcal DNA in Blood by Polymerase Chain Reaction for Diagnosing Pneumococcal Pneumonia in Young Children From Low- and Middle-Income Countries.

BACKGROUND.: We investigated the performance of polymerase chain reaction (PCR) on blood in the diagnosis of pneumococcal pneumonia among children from 7 low- and middle-income countries. METHODS.: We tested blood by PCR for the pneumococcal autolysin gene in children aged 1-59 months in the Pneumonia Etiology Research for Child Health (PERCH) study. Children had World Health Organization-defined severe or very severe pneumonia or were age-frequency-matched community controls. Additionally, we tested blood from general pediatric admissions in Kilifi, Kenya, a PERCH site. The proportion PCR-positive was compared among cases with microbiologically confirmed pneumococcal pneumonia (MCPP), cases without a confirmed bacterial infection (nonconfirmed), cases confirmed for nonpneumococcal bacteria, and controls. RESULTS.: In PERCH, 7.3% (n = 291/3995) of cases and 5.5% (n = 273/4987) of controls were blood pneumococcal PCR-positive (P < .001), compared with 64.3% (n = 36/56) of MCPP cases and 6.3% (n = 243/3832) of nonconfirmed cases (P < .001). Blood pneumococcal PCR positivity was higher in children from the 5 African countries (5.5%-11.5% among cases and 5.3%-10.2% among controls) than from the 2 Asian countries (1.3% and 1.0% among cases and 0.8% and 0.8% among controls). Among Kilifi general pediatric admissions, 3.9% (n = 274/6968) were PCR-positive, including 61.7% (n = 37/60) of those with positive blood cultures for pneumococcus. DISCUSSION.: The utility of pneumococcal PCR on blood for diagnosing childhood pneumococcal pneumonia in the 7 low- and middle-income countries studied is limited by poor specificity and by poor sensitivity among MCPP cases.

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.

Influenza vaccination of pregnant women and protection of their infants.

BACKGROUND: There are limited data on the efficacy of vaccination against confirmed influenza in pregnant women with and those without human immunodeficiency virus (HIV) infection and protection of their infants. METHODS: We conducted two double-blind, randomized, placebo-controlled trials of trivalent inactivated influenza vaccine (IIV3) in South Africa during 2011 in pregnant women infected with HIV and during 2011 and 2012 in pregnant women who were not infected. The immunogenicity, safety, and efficacy of IIV3 in pregnant women and their infants were evaluated until 24 weeks after birth. Immune responses were measured with a hemagglutination inhibition (HAI) assay, and influenza was diagnosed by means of reverse-transcriptase-polymerase-chain-reaction (RT-PCR) assays of respiratory samples. RESULTS: The study cohorts included 2116 pregnant women who were not infected with HIV and 194 pregnant women who were infected with HIV. At 1 month after vaccination, seroconversion rates and the proportion of participants with HAI titers of 1:40 or more were higher among IIV3 recipients than among placebo recipients in both cohorts. Newborns of IIV3 recipients also had higher HAI titers than newborns of placebo recipients. The attack rate for RT-PCR-confirmed influenza among both HIV-uninfected placebo recipients and their infants was 3.6%. The attack rates among HIV-uninfected IIV3 recipients and their infants were 1.8% and 1.9%, respectively, and the respective vaccine-efficacy rates were 50.4% (95% confidence interval [CI], 14.5 to 71.2) and 48.8% (95% CI, 11.6 to 70.4). Among HIV-infected women, the attack rate for placebo recipients was 17.0% and the rate for IIV3 recipients was 7.0%; the vaccine-efficacy rate for these IIV3 recipients was 57.7% (95% CI, 0.2 to 82.1). CONCLUSIONS: Influenza vaccine was immunogenic in HIV-uninfected and HIV-infected pregnant women and provided partial protection against confirmed influenza in both groups of women and in infants who were not exposed to HIV. (Funded by the Bill and Melinda Gates Foundation and others; ClinicalTrials.gov numbers, NCT01306669 and NCT01306682.).

Multiplex Urinary Antigen Detection for 13 Streptococcus pneumoniae Serotypes Improves Diagnosis of Pneumococcal Pneumonia in South African HIV-Infected Adults.

A serotype-specific urinary antigen detection (UAD) assay for 13 serotypes included in the pneumococcal conjugate vaccine (PCV13) was recently reported as a useful diagnostic tool for pneumococcal pneumonia. We aimed to assess the diagnostic accuracy of the UAD in HIV-infected South African adults. Urine specimens from a well-defined cohort of HIV-infected South African adults with pneumonia were evaluated retrospectively in the UAD assay. Pneumonia was considered pneumococcal if either sputum Gram stain, sputum culture, blood culture, or the immunochromatographic (ICT) BinaxNow S. pneumoniae test (composite diagnostic) was positive. Among 235 enrolled pneumonia patients, the UAD assay was more frequently positive (104 [44.3%]) than the composite diagnostic (71 [30.2%]; P < 0.001) and increased the pneumococcal etiology from 30.2% by an additional 22.6% to 52.8%. The UAD assay detected more pneumococcal etiologies (45.0%) than the serotype-independent ICT (23.4%, P < 0.001). UAD identified 6/7 patients with PCV13 serotype bacteremia without misclassification of bacteremia episodes due to non-PCV13 serotypes. UAD was positive for 5.1% of asymptomatic HIV-infected persons, with higher rates among those with nasopharyngeal carriage. Concordance between serotypes identified by UAD and by Quellung reaction and PCR serotyping was 70/86 (81.4%). UAD identified the dominant serotype in multiple serotype carriage. This study confirms the utility of the UAD assay for HIV-infected adults comparing favorably with other diagnostic tests. A highly valent UAD may become a new standard for detection of pneumococcal pneumonia in adults. Prior to PCV introduction, at least 53% of pneumonia cases were due to pneumococci in HIV-infected South African adults.

Serotype-Specific Cell-Mediated Immunity Associated With Clearance of Homotypic Group B Streptococcus Rectovaginal Colonization in Pregnant Women.

We investigated the association between group B Streptococcus (GBS) serotype-specific capsular polysaccharide cellular immunity, measured with enzyme-linked immunospot (ELISPOT) interferon γ release assay at 20 weeks gestation in pregnant women, and its effect on rectovaginal serotype-specific GBS colonization up to 37 weeks gestation. Among women colonized by serotype III at enrollment, interferon γ ELISPOT positivity was more common in those in whom colonization was cleared (44.4%) than in those in whom colonization persisted (7.4%; P = .008), with a similar trend observed for serotype Ia. Presence of serotype-specific capsular polysaccharide cell-mediated immunity contributes to the clearance of GBS rectovaginal colonization.

Increased risk for group B Streptococcus sepsis in young infants exposed to HIV, Soweto, South Africa, 2004-2008(1).

Although group B Streptococcus (GBS) is a leading cause of severe invasive disease in young infants worldwide, epidemiologic data and knowledge about risk factors for the disease are lacking from low- to middle-income countries. To determine the epidemiology of invasive GBS disease among young infants in a setting with high maternal HIV infection, we conducted hospital-based surveillance during 2004-2008 in Soweto, South Africa. Overall GBS incidence was 2.72 cases/1,000 live births (1.50 and 1.22, respectively, among infants with early-onset disease [EOD] and late-onset [LOD] disease). Risk for EOD and LOD was higher for HIV-exposed than HIV-unexposed infants. GBS serotypes Ia and III accounted for 84.0% of cases, and 16.9% of infected infants died. We estimate that use of trivalent GBS vaccine (serotypes Ia, Ib, and III) could prevent 2,105 invasive GBS cases and 278 deaths annually among infants in South Africa; therefore, vaccination of all pregnant women in this country should be explored.

Incidence and serotype distribution of invasive group B streptococcal disease in young infants: a multi-country observational study.

BACKGROUND: Group B Streptococcus (GBS) is a leading cause of serious infection in very young infants. Robust incidence data from many geographic regions, including Latin America and Asia, are however lacking. METHODS: A multicenter, hospital-based observational study was performed in Panama, Dominican Republic, Hong Kong and Bangladesh. All represented urban, tertiary referral hospitals, except Bangladesh. GBS cases (microbiological isolation from normally sterile sites in infants aged 0-89 days) were collected over 12 months. RESULTS: At 2.35 (95% CI: 1.74-3.18) cases per 1000 live births, the incidence of early onset GBS disease (EOD) was highest in the Dominican Republic, compared with 0.76 (95% CI: 0.41-1.39) in Hong Kong and 0.77 (95% CI: 0.44-1.35) in Panama, while no cases were identified in Bangladesh. Over 90% of EOD cases occurred on the first day of life, with case fatality ratios ranging from 6.7% to 40%, varying by center, age of onset and clinical presentation. Overall, 90% of GBS (EOD and late onset disease) was due to serotypes Ia, Ib and III. CONCLUSIONS: The incidence rate of early onset GBS infection reported in Dominican Republic was not dissimilar from that described in the United States prior to screening and intrapartum antibiotic prophylaxis, while the incidence in Hong Kong was higher than previously reported in the Asian region. The failure to identify GBS cases in Bangladesh highlights a need to better understand the contribution of population, healthcare and surveillance practice to variation in reported incidence. Overall, the identified disease burden and serotype distribution support the need for effective prevention methods in these populations, and the need for community based surveillance studies in rural areas where access to healthcare may be challenging.

Genomic load from sputum samples and nasopharyngeal swabs for diagnosis of pneumococcal pneumonia in HIV-infected adults.

Quantitative lytA real-time PCR (rtPCR) results from nasopharyngeal (NP) swabs distinguish community-acquired pneumococcal pneumonia (CAP) from asymptomatic colonization. The use of an optimized cutoff value improved pneumococcal etiology determination compared to that of traditional diagnostic methods. Here, we compare the utility of lytA rtPCR from induced sputum and from NP swabs. Pneumococcus was considered the cause of CAP in HIV-infected South African adults if blood culture, induced-sputum culture or Gram stain, urine antigen test, or whole-blood lytA rtPCR revealed pneumococcus or if lytA rtPCR from NP swabs gave a result of >8,000 copies/ml. lytA rtPCR was also performed on induced sputum. Pneumococcus was detected by lytA rtPCR from sputum in 149 (67.1%) of 222 patients with available induced sputum, whereas the results of either Gram stain or culture of sputum were positive in 105 of 229 patients (45.9%; P < 0.001). The mean copy numbers from sputum were higher when the sputum cultures were positive than when the sputum cultures were negative (7.9 versus 5.6 log10 copies/ml; P < 0.001). Against the composite diagnostic standard, a cutoff value of 10,000 copies/ml for good-quality sputum lytA rtPCR had a sensitivity of 78.1% and a specificity of 80.0%. This cutoff value performed similarly to the previously identified cutoff value of 8,000 copies/ml for NP swab lytA rtPCR (area under the curve receiver operating characteristic [AUC-ROC], 80.4% for sputum of any quality versus 79.6% for NP swabs). The AUC-ROC for good-quality sputum was 83.2%. Overall, lytA rtPCR performs similarly well on induced sputum as on NP swabs for most patients but performs slightly better if good-quality sputum can be obtained. Due to the ease of specimen collection, NP swabs may be preferable for the diagnosis of pneumococcal pneumonia.

Dynamics of pneumococcal transmission in vaccine-naive children and their HIV-infected or HIV-uninfected mothers during the first 2 years of life.

Pneumococcal vaccine-naïve mother-child dyads in South Africa had nasopharyngeal swabs taken 9 times within the first 2 years of the children's lives between January 2007 and May 2009. To quantify the strength of the association of serotype-specific carriage in mother-child dyads, a stochastic transmission model was fitted to the data. Children were more susceptible to individual serotypes included in the 7-valent pneumococcal conjugate vaccine (PCV7) transmitted by their mothers than vice versa; however, children infected their mothers with these serotypes more frequently than mothers infected children. The child-to-mother steady-state forces of pneumococcal acquisition were between 0.36 and 3.29 (per 1,000 days) compared with 0.06-0.51 for mother-to-child transmission. Although children of mothers infected with human immunodeficiency virus were more often exposed to PCV7 serotypes by their mothers, their risk of acquisition remained low compared with the risk of child-to-mother transmission. Mothers acquired pneumococci at lower rates (per 1,000 days) from unmeasured exposure within families and in the wider community (range, 0.12-1.69 per 1,000 days) than did children (range, 1.10-5.21 per 1,000 days). Pneumococcal immunization of young children is expected to have an indirect effect of reducing PCV7 serotype maternal colonization and possibly disease even in settings such as ours, in which there is a high prevalence of human immunodeficiency virus-infected mothers.

Evaluation of Trans-Vag broth, colistin-nalidixic agar, and CHROMagar StrepB for detection of group B Streptococcus in vaginal and rectal swabs from pregnant women in South Africa.

Maternal vaginal colonization with group B streptococcus (GBS) is a major risk factor for invasive GBS infection in newborns. The CDC-recommended method for detecting GBS colonization is to culture vaginal and rectal swabs in a selective broth followed by subculture on blood agar or a selective medium. A high incidence of antimicrobial resistance in the fecal microflora can compromise the recovery of GBS from the selective broth. Here, we compared CHROMagar StrepB (CA), Columbia colistin-nalidixic agar (CNA), and Trans-Vag selective broth enrichment for the isolation of GBS from 130 vaginal and 130 rectal swabs from pregnant women. The swabs were randomized for plating first on either CA or CNA, and they then were inoculated in Trans-Vag broth. GBS was cultured from 37.7% of the vaginal swabs and 33.1% of the rectal swabs. There were no differences in the detection rates for the vaginal swabs between CA (31.5%), CNA (26.2%), and the selective broth (30.0%). The sensitivities in relation to a composite score were 83.7%, 69.4%, and 79.6%, respectively. However, recovery of GBS from the rectal swabs was significantly higher from CA (29.2%; P<0.0001) and CNA (23.8%; P=0.002) than from the selective broth (9.2%). The sensitivities were 88.4%, 72.1%, and 27.9%, respectively. The order of plating on the solid medium was significant (P=0.003), with GBS detection rates of 30.8% and 24.6% when swabs were plated first and second, respectively. These findings show that a selective broth is not suitable for the recovery of GBS from rectal swabs in settings such as ours, due to masking of the GBS colonies by persistent microflora.

Interrelationship of Streptococcus pneumoniae, Haemophilus influenzae and Staphylococcus aureus colonization within and between pneumococcal-vaccine naïve mother-child dyads.

BACKGROUND: A high prevalence of bacterial nasopharyngeal co-infections has been reported in children, however, such data is limited in adults. We examined the interaction of Haemophilus influenzae, Staphylococcus aureus and Streptococcus pneumoniae pharyngeal colonization in mother-child dyads. METHODS: Pneumococcal-vaccine naïve children and their mothers had pharyngeal swabs undertaken at 1.6, 2.5, 3.5, 4.5, 7.4, 9.5, 12.5, 16.2 and 24.2 months of child's age. Swabs were cultured for S. pneumoniae, H. influenzae and S. aureus using standard microbiologic methods. Multivariate generalized estimating equation-models were used to explore the associations of the three bacteria within and between children and their mothers. RESULTS: In children, the observed probability of co-colonization was higher than expected. Well-defined associations in colonization between the bacteria were observed in children but not among mothers. In children, a synergistic association was observed between S. pneumoniae and H. influenzae (Adjusted odds ratio (AOR): 1.75, 95% CI: 1.32-2.32) and a negative association between S. pneumoniae and S. aureus (AOR: 0.51, 95% CI: 0.39-0.67) or H. influenzae and S. aureus (AOR: 0.24, 95% CI: 0.16-0.34) colonization. Additionally, all three bacteria had a higher likelihood of concurrent colonization. There was a strong association in colonization by the bacteria in children and their mothers, including increased likelihood of maternal colonization if the child was colonized by S. pneumoniae (AOR: 1.84, 95% CI: 1.28-2.63) and H. influenzae (AOR: 6.34, 95% CI: 2.24-18.0). CONCLUSIONS: The effects of immunization of children with pneumococcal-conjugate-vaccine in settings such as ours needs monitoring with regard to potential changes of pharyngeal bacterial ecology which could occur in vaccinated and -unvaccinated age-groups.

Evaluation of the impact of HIV-1 infection and density of common nasopharyngeal bacterial colonizers in South African children immunized with 7-valent pneumococcal conjugate vaccine.

BACKGROUND: Due to limitations in standard culture methods, the impact of pneumococcal conjugate vaccine (PCV) immunization on nasopharyngeal bacterial carriage density is unclear, including among HIV-infected children. METHODS: The prevalence and density of serotype/serogroup-specific pneumococcal and other nasopharyngeal colonizing bacteria were investigated in archived swabs of HIV-infected and HIV-uninfected, PCV-7 immunized (at 6, 10 and 14 weeks of age) South African children collected at 9 and 16 months of age. During the course of the study, PCV-immunization of children in Soweto was limited to study-participants, as the vaccine had not been introduced into the public immunization program. RESULTS: At 9 months of age, the prevalence of overall pneumococcal colonization was lower in HIV-infected (58.6%) than HIV-uninfected children (69.9%, p = 0.02), mainly due to lower prevalence of non-vaccine-serotype colonization (27.8% vs. 40%, respectively; p = 0.047). The mean-log10 density of pneumococcal colonization was, however, higher in HIV-infected (4.81 CFU/ml) than HIV-uninfected pneumococcal colonized children (4.44 CFU/ml; p = 0.014); mainly due to higher mean-log10 density of PCV7-serotype colonization (4.21 vs. 3.72 CFU/ml; p = 0.014). No difference in the prevalence or density of overall pneumococci was found at 16 months of age. The prevalence of non-vaccine serotype colonization remained 1.7 fold higher in HIV-uninfected (60.4%) than HIV-infected children (50.9%, p = 0.049). Other differences included a lower prevalence of H. influenzae colonization in HIV-infected (42.3% and 56%) than HIV-uninfected children (64.2% and 73.4%) at both 9 and 16 months of age respectively; however, the density of colonization was similar. CONCLUSION: Increased carriage density of residual PCV7-serotypes might cause HIV-infected children to have a higher risk of pneumococcal disease. The higher carriage density observed in HIV-infected children could be attributed to a combination of factors, including HIV treatment and impaired host immunity. Additional studies are needed.

Effect of HIV-exposure and timing of antiretroviral treatment initiation in children living with HIV on antibody persistence and memory responses to Haemophilus influenzae type b and pneumococcal polysaccharide-protein conjugate vaccines.

BACKGROUND: We investigated the effect of in utero HIV-exposure, timing of antiretroviral treatment (ART) initiation, and ART interruption on memory responses and persistence of immunity induced by pneumococcal (PCV) and Haemophilus influenzae type b (HibCV) polysaccharide-protein conjugate vaccines. METHODS: Children were enrolled (6-12 weeks of age), and vaccinated with a three-dose primary series of 7-valent PCV (PCV7) and HibCV at 6, 10 and 14 weeks of age. Study groups included infants infected with HIV perinatally with CD4+ ≥ 25% initiating ART following immunological or clinical deterioration (ART-Def), or immediately upon enrolment followed by interruption at 40 (ART-Immed/40w) or 96 weeks (ART-Immed/96w); and HIV-uninfected infants with (HEU), and without HIV (HIV-unexpsoed) exposure in utero. Within each group, children were randomized to receive either a booster dose of PCV7 or HibCV at 15 months of age. PCV serotype-specific and polyribosyl ribitol phosphate (PRP) IgG were measured pre-boost, two-weeks post-boost and at two-years of age. Opsonophagocytic activity (OPA) to serotypes 9V, 19F and 23F was measured post-booster dose. RESULTS: Persistence of IgG to PCV vaccine-serotypes and anti-PRP was similar in all groups of children living with HIV (CLWH) compared to HIV-unexposed children. Anamnestic responses to PCV and HibCV were also similar in all three groups of CLWH compared to HIV-unexposed children. CLWH, however, tended to have lower functional antibody (OPA) titers than HIV-unexposed children after the PCV booster dose for some serotypes. Immunity to PCV and HibCV was similar between the ART-Immed/40w and ART-Immed-96w groups. There were no differences in IgG kinetics between HEU and HIV-unexposed children. CONCLUSIONS: A three dose primary series, with or without PCV or HibCV booster doses in CLWH initiated on ART during infancy, would likely be similarly effective in preventing invasive bacterial disease as in HIV-unexposed children.

Performance of the Biomark HD real-time qPCR System (Fluidigm) for the detection of nasopharyngeal bacterial pathogens and Streptococcus pneumoniae typing.

Traditional qPCR assays for pneumococcal detection and serotype characterization require large sample volume, is expensive and labor intensive. We aimed to develop a quantitative nanofluidic Fluidigm assay to overcome some of these shortcomings. A quantitative Fluidigm assay was established to detect 11 bacterial pathogens, 55 pneumococcal serotypes and 6 serotypes of H. influenzae. The Fluidigm assay results were compared to conventional qPCR and culture. All reactions in the Fluidigm assay effectively amplified their respective targets with high sensitivity and specificity compared to qPCR. There was excellent concordance between qPCR and Fluidigm for detection of carriage prevalence (kappa > 0.75) and density (Rho > 0.95). Fluidigm identified an additional 7 (4.2%) serotypes over those detected by qPCR. There was a modest concordance between culture and Fluidigm for the majority of reactions detecting S. pneumoniae serotypes/serogroups (kappa > 0.6), with Fluidigm identifying an additional 113 (39.1%) serotypes. Discordant results between the three methods were associated with a low carriage density. The Fluidigm assay was able to detect common pneumococcal serotypes, H. influenzae serotypes, and other common nasopharyngeal bacterial organisms simultaneously. Deployment of this assay in epidemiological studies could provide better insight into the effect of PCV immunization on the nasopharyngeal microbiota in the community.

Effect of HIV exposure and timing of antiretroviral therapy initiation on immune memory responses to diphtheria, tetanus, whole cell pertussis and hepatitis B vaccines.

OBJECTIVES: We evaluated memory responses and antibody persistence to diphtheria-toxoid, tetanus-toxoid, whole-cell-pertussis (DTwP), and Hepatitis-B vaccines in HIV-unexposed, HIV-exposed-uninfected and HIV-infected children previously randomized to initiate time-limited ART at 6-10 weeks (ART-Immed) or when clinically/immunologically indicated (ART-Def). METHODS: All children received DTwP booster at 15-18 months. Antibodies were measured for pertussis-toxoid, filamentous haemagglutinin (FHA), diphtheria-toxoid, tetanus-toxoid, and hepatitis-B prior to booster, 1-2 weeks post-booster and at 24 months of age. RESULTS: Pre-booster antibody GMC were lower in HIV-infected groups than HIV-unexposed children for all epitopes. Post-booster and at 24 months of age, the ART-Def group had lower GMCs and antibody proportion ≥0.1 IU/ml for tetanus-toxoid and diphtheria-toxoid compared to HIV-unexposed children. At 24 months of age, the ART-Immed group had higher GMCs, and more likely to maintain antibody titres ≥1.0 IU/ml to tetanus-toxoid and diphtheria-toxoid compared to HIV-unexposed children. Compared to HIV-unexposed children, at 15 and 24 months of age, persistence of antibody to HBsAg of ≥10 mIU/ml was similar in the ART-Immed group but lower among the ART-Def group. Antibody kinetics indicated more robust memory responses in HIV-exposed-uninfected than HIV-unexposed children to diphtheria-toxoid and wP. CONCLUSION: HIV-infected children not on ART at primary vaccination had poorer memory responses, whereas HIV-exposed-uninfected children mounted robust memory responses.