Predictors of Respiratory Syncytial Virus, Influenza Virus, and Human Metapneumovirus Carriage in Children Under 5 Years With WHO-Defined Fast-Breathing Pneumonia in Pakistan.

BACKGROUND: Pneumonia is a leading cause of morbidity and mortality in children < 5 years. We describe nasopharyngeal carriage of respiratory syncytial virus (RSV), human metapneumovirus (hMPV), and influenza virus among children with fast-breathing pneumonia in Karachi, Pakistan. METHODS: We performed a cross-sectional analysis of nasopharyngeal swabs from children aged 2-59 months with fast-breathing pneumonia, enrolled in the randomized trial of amoxicillin versus placebo for fast-breathing pneumonia (RETAPP) (NCT02372461) from 2014 to 2016. Swabs were collected using WHO standardized methods, processed at the Aga Khan University, Pakistan. Viral detection was performed using LUMINEX xTAG respiratory viral panel assay and logistic regression identified clinical and sociodemographic predictors. FINDINGS: Of the 1000 children tested, 92.2% (n = 922) were positive for viral carriage. RSV, hMPV, and influenza virus were detected in 59 (6.4%), 56 (6.1%), and 58 (6.3%) children and co-infections in three samples (two RSV-hMPV and one influenza-hMPV). RSV carriage was common in infants (56%), we observed a higher occurrence of fever in children with hMPV and influenza virus (80% and 88%, respectively) and fast breathing in RSV (80%) carriage. RSV carriage was positively associated with a history of fast/difficulty breathing (aOR: 1.96, 95% CI 1.02-3.76) and low oxygen saturation (aOR: 2.52, 95% CI 1.32-4.82), hMPV carriage was positively associated with a complete vaccination status (aOR: 2.22, 95% CI 1.23-4.00) and body temperature ≥ 37.5°C (aOR: 2.34, 95% CI 1.35-4.04) whereas influenza viral carriage was associated with body temperature ≥ 37.5°C (aOR: 4.48, 95% CI 2.53-7.93). CONCLUSION: We observed a high nasopharyngeal viral carriage among children with WHO-defined fast-breathing pneumonia in Pakistan. Fever, difficulty in breathing, hypoxia and vaccination status are important clinical predictors for viral nonsevere community-acquired pneumonia.

Secondary attack rates and determinants of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) household transmission in Pakistan: A case-ascertained prospective, longitudinal study.

BACKGROUND: Households are considered ideal settings for studying the transmission dynamics of an infectious disease. METHODS: A prospective study was conducted, based on the World Health Organization FFX protocol from October 2020 to January,2021. Household contacts of laboratory-confirmed index cases were followed up for their symptomatic history, nasal swabs for RT-PCR,and blood samples for anti-SARS CoV-2 antibodies were collected at enrollment and days 7, 14 and 28. We estimated secondary attack rate (SAR), effective household case cluster size and determinants of secondary infection among susceptible household contacts using multivariable logistic regression. RESULTS: We enrolled 77 index cases and their 543 contacts. Out of these, 252 contacts were susceptible at the time of enrollment. There were 77 household clusters, out of which, transmission took place in 20 (25.9%) giving rise to 34 cases. The acquired secondary attack rate (SAR) was 14.0% (95% CI 9.0-18.0). The effective household case cluster size was 0.46 (95%CI 0.33,0.56). Reported symptoms of nausea and vomiting (aOR, 7.9; 95% CI, 1.4-45.5) and fatigue (aOR, 9.3; 95% CI, 3.8-22.7) were associated with SARS-CoV-2 transmission. CONCLUSIONS: We observed a low SARS-CoV-2 secondary attack rate in the backdrop of high seroprevalence and asymptomatic transmission among households in Karachi, Pakistan.

Association of maternal nutritional status and small for gestational age neonates in peri-urban communities of Karachi, Pakistan: findings from the PRISMA study.

BACKGROUND: Early pregnancy nutritional status can be associated with adverse birth outcomes such as small-for-gestational age (SGA) and low birth weight (LBW). BMI (Body Mass Index) and MUAC (Mid-upper arm circumference) are easy to use assessments and are indicative of the pre-pregnancy nutritional status if obtained in the first trimester. This study primarily assesses the association of maternal nutritional status using BMI and MUAC with SGA in a community-based cohort of Pakistani women. It also aims to determine the predictive ability of MUAC and BMI in predicting SGA. Secondarily, we assessed the association between maternal nutrition and large for gestational age (LGA) and LBW. METHODS: This study is a secondary analysis of an ongoing pregnancy cohort "Pregnancy Risk Infant Surveillance and Measurement Alliance (PRISMA)"in Ibrahim Hyderi and Rehri Goth, Karachi. PRISMA participants who were enrolled between January 2021 to August 2022 were included given they had a gestational age < 14 weeks confirmed via ultrasound, MUAC and BMI measurements were available and birth weight was captured within 72 hours. Multivariable logistic regression was used to determine an association between maternal nutritional status and SGA. The PRISMA study was approved by the Aga Khan University Ethics Review Committee (2021-5920-15,518). RESULTS: Of 926 women included in the analysis, 26.6% (n = 247) had a low MUAC (< 23 cm) while 18.4% (n = 171) were underweight (BMI < 18.5 kg/m2). Nearly one third of low MUAC and underweight women delivered SGA infants (34.4 and 35.1% respectively). Underweight women and women with low MUAC had a statistically significant association with SGA (Underweight: OR 1.49, 95% CI 1.1,2.4; Low MUAC-OR 1.64, 95% CI 1.2,2.3) as well as LBW (Underweight: OR-1.63, 95% CI 1.1,2.4; Low MUAC-OR-1.63, 95% CI 1.2,2.3). ROC curves showed that MUAC and BMI had modest predictability for SGA (AUC < 0.7). CONCLUSION: Maternal nutritional status as indicated by BMI and MUAC are strongly associated with adverse pregnancy outcomes including SGA, LGA and LBW. Although MUAC and BMI are widely used to determine maternal nutritional status, they have poor predictive ability for newborn size. Further research is needed to identify other tools or a combination of tools to better predict adverse birth outcomes in resource-limited settings and plan interventions.

Clinical experience of next generation sequencing based expanded carrier screening in high-risk couples from a tertiary healthcare center in Pakistan.

INTRODUCTION: Carrier screening for genetic conditions has long been a part of preconception and prenatal care. While the use of expanded carrier screening (ECS) is widely common in HICs (high income countries), the clinical actionability of ECS in LMICs (low middle income countries) with high consanguineous unions is not well-understood. METHOD: Retrospective chart review of couples who presented to the Prenatal Genetics Clinic at Aga Khan University Hospital, between the period of June 2018 and November 2022. All the statistical analyses were performed using the statistical software STATA version 17.0. RESULTS: Of the 202 individuals tested, 166 (82%) were identified to be carriers of at least one gene associated with a monogenic condition. Out of the 302 genes tested, individuals were found to be carriers of conditions associated with 87 genes. Clinical actionability of ECS was established in a total of 45 (45%) high risk couples undergoing screening using ECS. CONCLUSION: We report the first clinical experience of using next generation sequencing based ECS in 101 high-risk couples seeking consultation in the Genetics Clinic, at a tertiary healthcare center in Pakistan, showing that over 80% high-risk individuals are carriers of at least one condition and 45% of couples receive an actionable result to making autonomous informed reproductive decisions in future pregnancies.

Early to mid-pregnancy HbA1c levels and its association with adverse pregnancy outcomes in three low middle-income countries in Asia and Sub-Saharan Africa.

BACKGROUND: Hyperglycemia during pregnancy leads to adverse maternal and fetal outcomes. Thus, strict monitoring of blood glucose levels is warranted. This study aims to determine the association of early to mid-pregnancy HbA1c levels with the development of pregnancy complications in women from three countries in South Asia and Sub-Saharan Africa. METHODS: We performed a secondary analysis of the AMANHI (Alliance for Maternal and Newborn Health Improvement) cohort, which enrolled 10,001 pregnant women between May 2014 and June 2018 across Sylhet-Bangladesh, Karachi-Pakistan, and Pemba Island-Tanzania. HbA1c assays were performed at enrollment (8 to < 20 gestational weeks), and epidemiological data were collected during 2-3 monthly household visits. The women were followed-up till the postpartum period to determine the pregnancy outcomes. Multivariable logistic regression models assessed the association between elevated HbA1c levels and adverse events while controlling for potential confounders. RESULTS: A total of 9,510 pregnant women were included in the analysis. The mean HbA1c level at enrollment was found to be the highest in Bangladesh (5.31 ± 0.37), followed by Tanzania (5.22 ± 0.49) and then Pakistan (5.07 ± 0.58). We report 339 stillbirths and 9,039 live births. Among the live births were 892 preterm births, 892 deliveries via cesarean section, and 532 LGA babies. In the multivariate pooled analysis, maternal HbA1c levels of ≥ 6.5 were associated with increased risks of stillbirths (aRR = 6.3, 95% CI = 3.4,11.6); preterm births (aRR = 3.5, 95% CI = 1.8-6.7); and Large for Gestational Age (aRR = 5.5, 95% CI = 2.9-10.6). CONCLUSION: Maternal HbA1c level is an independent risk factor for predicting adverse pregnancy outcomes such as stillbirth, preterm birth, and LGA among women in South Asia and Sub-Saharan Africa. These groups may benefit from early interventional strategies.

Knowledge, attitudes, and practices toward seasonal influenza vaccination among healthcare workers and pregnant women in Pakistan: A mixed methods approach.

Vaccine hesitancy is a significant public health issue globally. We aim to document the barriers toward seasonal influenza vaccine uptake among healthcare workers (HCWs) and pregnant women (PW) in Pakistan. We performed a concurrent mixed methods study in four cities (Karachi, Islamabad, Quetta, and Peshawar) across Pakistan from September to December 2021. The quantitative component consisted of independent cross-sectional surveys for PW and HCWs, and the qualitative component comprised of in-depth interviews (IDIs) and focus group discussions (FGDs) among HCWs. Simple linear regression was used to determine the association of sociodemographic variables with knowledge, attitudes, and practices. Overall, 750 PW and 420 HCWs were enrolled. Among the PW, 44% were willing to receive the vaccine if available free of cost. Only 44% of the HCWs were vaccinated; however, 86% intended to get vaccinated and were willing to recommend the vaccine to their patients. HCWs refused vaccine due to side-effects (65%), cost (57%), and allergies (36%). An education level of secondary school and above was predictive of higher attitude and knowledge scores while having received the COVID-19 vaccine was associated with higher practice scores for both PW and HCWs. Several themes emerged from the interviews: 1) HCWs' knowledge of influenza and its prevention, 2) HCWs' perception of motivators and barriers to influenza vaccine uptake and 3) HCWs' attitudes towrd vaccine promotion. We report low influenza vaccine coverage among HCWs and PW in Pakistan. Educational campaigns addressing misconceptions, and improving affordability and accessibility through government interventions, can improve vaccine uptake.

Investigating the impact of prior COVID-19 on IgG antibody and interferon γ responses after BBIBP-CorV vaccination in a disease endemic population: A prospective observational study.

Background and Aims: COVID-19 vaccinations have reduced morbidity and mortality from the disease. Antibodies against severe acute respiratory syndrome coronavirus 2 (SARS­CoV­2) have been associated with immune protection. Seroprevalence studies revealed high immunoglobulin G (IgG) antibody levels to SARS-CoV-2 in the Pakistani population before vaccinations. We investigated the effect of BBIBP-CorV vaccination on circulating IgG antibodies and interferon (IFN)-γ from T cells measured in a cohort of healthy individuals, with respect to age, gender, and history of COVID-19. Methods: The study was conducted between April and October 2021. BBIBP-CorV vaccinated participants were followed up to 24 weeks. Antibodies to SARS-CoV-2 Spike protein and its receptor-binding domain (RBD) were measured. IFNγ secreted by whole blood stimulation of Spike protein and extended genome antigens was determined. Results: Study participants with a history of prior COVID-19 displayed a higher magnitude of IgG antibodies to Spike and RBD. IgG seropositivity was greater in those with prior COVID-19, aged 50 years or younger and in females. At 24 weeks after vaccination, 37.4% of participants showed IFN-γ responses to SARS-CoV-2 antigens. T cell IFN-γ release was higher in those with prior COVID-19 and those aged 50 years or less. Highest IFN-γ release was observed to extended genome antigens in individuals both with and without prior COVID-19. Conclusion: We found that IgG seropositivity to both Spike and RBD was affected by prior COVID-19, age and gender. Importantly, seropositive responses persisted up to 24 weeks after vaccination. Persistence of vaccine induced IgG antibodies may be linked to the high seroprevalence observed earlier in unvaccinated individuals. Increased T cell reactivity to Spike and extended genome antigens reflects cellular activation induced by BBIBP-CorV. COVID-19 vaccination may have longer lasting immune responses in populations with a higher seroprevalence. These data inform on vaccination booster policies for high-risk groups.

Co-carriage of Staphylococcus aureus and Streptococcus pneumoniae among children younger than 2 years of age in a rural population in Pakistan.

Objectives: Staphylococcus aureus and Streptococcus pneumoniae are common colonizers of the human nasopharynx. In this study, we describe S. aureus nasopharyngeal carriage and evaluate its association with S. pneumoniae carriage post-10-valent pneumococcal conjugate vaccine (PCV10) introduction in Pakistan. Methods: A serial cross-sectional study was undertaken from 2014 to 2018, children <2 years were randomly selected, and nasopharyngeal swabs were collected using standard WHO guidelines. S. aureus and S. pneumoniae isolates were identified using standard methods and tested for antimicrobial susceptibility by the standard Kirby-Bauer disk-diffusion method as per Clinical & Laboratory Standards Institute (CLSI) recommendations. Regression analysis was used to determine predictors associated with S. aureus carriage. Results: We enrolled 3140 children. S. aureus carriage prevalence was 5.6% (176/3140), and 50.1% (81/176) of the isolates were methicillin-resistant S. aureus (MRSA). S. aureus carriage was higher in the absence of pneumococcus compared to isolates in which pneumococcus was present (7.5% vs 5.0%). S. aureus carriage was negatively associated with pneumococcal carriage, being in 3rd and 4th year of enrollment, and vaccination with two and three PCV10 doses, in addition, fast breathing, ≥2 outpatients visits, and rainy season were positively associated. The following resistance rates were observed: 98.9% for penicillin, 74.4% for fusidic acid, and 23.3% for gentamicin, 10.2% for erythromycin, and 8.5% for cotrimoxazole. All isolates were susceptible to amikacin. Conclusions: Overall S. aureus carriage prevalence was low, PCV10 vaccine was protective against the carriage. The proportion of MRSA carriage and antimicrobial resistance was high in this community warranting continuous monitoring for invasive infections.

Differential coverage for vaccines in the expanded program on immunization (EPI) among children in rural Pakistan.

BACKGROUND: Pakistan has a well-established Expanded Program on Immunization (EPI) however vaccine-preventable diseases still account for high infant and child mortality rates. This study describes the differential vaccine coverage and determinants of vaccine uptake in rural Pakistan. METHODS: From October 2014 to September 2018, we enrolled children younger than 2 years of age from the Matiari Demographic Surveillance System in Sindh, Pakistan. Socio-demographic and vaccination history were collected from all participants. Vaccine coverage rates and timeliness were reported. Socio-demographic variables for missed and untimely vaccination were studied in multivariable logistic regression. RESULTS: Of the 3140 enrolled children, 48.4 % received all EPI recommended vaccines. Only 21.2 % of these were age appropriate. Around 45.4 % of the children were partially vaccinated, and 6.2 % were unvaccinated. Highest coverage was seen for the first dose of pentavalent (72.8 %), 10-valent Pneumococcal Conjugate Vaccine (PCV10) (70.4 %) and Oral Polio Vaccine (OPV) (69.2 %) and the lowest coverage was for measles (29.3 %) and rotavirus (1.8 %) vaccines. Primary caretakers and wage earners with a higher level of education were protective against missed and untimely vaccination. Enrollment in the 2nd, 3rd and 4th study year was negatively associated with being unvaccinated whereas distance from a major road was positively associated with non-adherence to schedule. CONCLUSION: Vaccine coverage was low among children in Matiari, Pakistan, and majority received delayed doses. Parents' education status and year of study enrollment was protective against vaccine dropout and delayed vaccination whereas geographical distance from a major road was a predictor. Vaccine promotion and outreach efforts may have had a beneficial impact on vaccine coverage and timeliness.

Knowledge, attitudes, and practices towards seasonal influenza vaccination among pregnant women and healthcare workers: A cross-sectional survey in Afghanistan.

Background: Despite recommendation by the World Health Organization (WHO), influenza vaccination coverage among high-risk groups remains suboptimal in Afghanistan. This study aims to document the knowledge, attitudes, and practices of seasonal influenza vaccine uptake among two priority groups, pregnant women (PWs) and healthcare workers (HCWs). Methods: This cross-sectional study enrolled PWs and HCWs in Kabul, Afghanistan, from September to December 2021. Data on vaccine intention and uptake, knowledge, and attitudes towards vaccination were collected. Simple linear regression was used to predict the impact of sociodemographic characteristics on the KAP score. Results: A total of 420 PWs were enrolled in Afghanistan. The majority (89%) of these women had never heard of the influenza vaccine but 76% intended to receive the vaccine. Of the 220 HCWs enrolled, 88% were unvaccinated. Accessibility and cost were factors which encouraged vaccination among HCWs. Fear of side effects and affordability were identified as key barriers. HCWs reported high level of vaccine intention (93%). PWs aged under 18 years (ß: 6.5, P = 0.004), between 18 and 24 years (ß: 2.9, P = 0.014), currently employed (ß: 5.8, P = 0.004), and vaccinated against COVID-19 (ß: 2.8, P = 0.01) were likely to have a higher attitude score. Among HCWs, being female was a predictor for poor vaccination practice (ß: -1.33, P < 0.001) whereas being vaccinated against COVID-19 was a predictor for higher practice score (ß: 2.4, P < 0.001). Conclusion: To increase influenza vaccination coverage among priority groups, efforts should be made to address issues such as lack of knowledge, limited availability, and cost barriers.

Pneumococcal Carriage in Infants Post-PCV10 Introduction in Pakistan: Results from Serial Cross-Sectional Surveys.

The 10-valent pneumococcal vaccine was introduced in Pakistan's Expanded Program on Immunization (EPI) in 2013 as a 3 + 0 schedule without catchup. We conducted three annual cross-sectional surveys from 2014−2016 to measure vaccine-type (VT) carriage in infants from a rural part of Pakistan. Nasopharyngeal specimens were collected by random sampling of infants from two union councils of Matiari. Samples were then transported to the Infectious Disease Research Laboratory (IDRL) at the Aga Khan University within 6−8 h of collection. Serotypes were established using sequential multiplex PCR. Of the 665 children enrolled across three surveys, 547 were culture-positive for pneumococcus. VT carriage decreased from 21.8% in 2014 to 12.7% in 2016 (p-value for trend <0.001). Those who were not vaccinated or partially vaccinated were found to be at higher risk of carrying a VT serotype ((aOR 2.53, 95% CI 1.39, 4.63 for non-vaccinated) and (aOR 3.35, 95% CI 1.82, 6.16 for partially vaccinated)). On the other hand, being enrolled in the most recent survey was negatively associated with VT carriage (aOR 0.51, 95% CI 0.28, 0.93). We found that PCV10 was effective in decreasing the carriage of vaccine-type serotypes in Pakistani infants.