Pneumococcal carriage and antibiotic susceptibility patterns in mother-baby pairs in a rural community in Eastern Uganda: a cross-sectional study

Ethical statement

We obtained ethical approval for this study from Mbale Regional Referral Hospital Research and Ethics Committee (MRRHREC060418). A letter of introduction to the study was obtained following approval from the Busitema University Faculty of Health Sciences community education program office and used by the researchers to seek permission from the Medical Superintendent of Ngora District Health Centre IV. Written informed consent for participation was obtained from participants preceding the actual data collection exercise. Access to collected data was restricted to persons directly involved in the study only. Participants were free to withdraw their consent to participate in the study at any stage of the study. Such a decision did not affect the medical care they received or their possible participation in future research studies in any way. The procedures were verbally explained to the research participants by the researchers and those who agreed to continue with the study could consent and participate.

Research design and setting

A cross-sectional study was carried out in Ngora district Health Centre IV and Ngora Health Center IV catchment area from 7^th April to 5^th May 2018. It serves a population of approximately 142,487. Ngora district is one of the districts in the Teso sub-region and was part of Kumi district until 2010, when it was established as a separate district by a Uganda parliamentary act. Ngora district covers an area of approximately 715.9 square kilometers and is predominantly inhabited by the Iteso and Kumam ethnicities. According to the Ugandan healthcare hierarchy of organizations, a Health Center IV is expected to serve a population of up to 100, 000 people, meaning at its current capacity, this health facility operates above the level of a Health Center IV¹⁰.

Study population and sampling

Children under five years old and their mothers were selected in a ‘mother-baby pair’ to determine the relationship between carriage of pneumococci in the baby, the prevalence of pneumococci in mothers, and the child’s immunization status. The inclusion criteria were mother-baby pairs from selected villages in the community, and those who visited Ngora Health Center IV for routine immunization with babies under the age of five years. For this study, we defined a baby as any person under the age of five years and a mother was considered as either biological or any other female indirect care of the baby for at least four weeks. This was aimed at comparing samples from the baby, and the person in closest contact with the baby for the longest duration of time in the most recent past to allow presumed cross-transmission/cross-infection. Exclusion criteria were babies above five years, and those presented by male caretakers as those were presumed to spend less time with the babies. Mothers who did not consent or opted out at any stage of the research were excluded as well. The purpose of the research was explained to the mothers and they were asked to voluntarily participate in the research.

Consecutive sampling was used. Every participant meeting the criteria of inclusion was selected at the young child clinic and in randomly selected villages in the community until the required sample size was achieved. This method is relatively easy to employ and reduces the chances for intentional and unintentional manipulation by staff, or errors due to confusion. A total of 152 mother-baby pairs fulfilling the inclusion criteria were sampled. The Sample size was estimated using the Kish and Leslie formula developed in 1965.

Data collection and variables

Demographic data was captured using a pre-tested questionnaire. The questionnaire was pre-tested with 10 mother-baby pairs at Mbale regional referral hospital Young Child Clinic (YCC) because the mother-baby pair in Mbale hospital had homogenous characteristics with our target population in Ngora district. After pre-testing the questionnaires, some questions were rephrased for clarity. The Uganda national immunization guidelines require all children to receive their first PCV dose at six weeks (1.5 months), the second dose at 10 weeks (2.5 months), and the third dose at 14 weeks (3.5 months). Therefore, in the questionnaire, full immunization was defined as any child 14 weeks (3.5 months) of age and above, who had received all the three PCV doses as stipulated in the national immunized schedule. Partial immunization included children above six weeks (1.5 months) of age who had received less than three doses of PCV, whether off schedule, or still on schedule as per the national immunization guidelines. All children below five years of age who had not received any single dose of PCV including those below six weeks (still on schedule) were categorized as not immunized.

Specimen collection and transport

Nasopharyngeal specimens were collected from the posterior nasopharynx of the mother and the baby using sterile cotton swab sticks moistened with 0.9% physiological saline. Sample collection was carried out at the young child clinic in Ngora Health Centre IV and the selected villages in the community by qualified hospital laboratory technicians. Separate swabs were used to collect samples from a mother and a baby in a mother-baby pair. To prevent sample contamination, the swab was placed in a casing containing Amies transport medium and immediately placed into a cool box containing ice packs for transportation to Busitema University Microbiology laboratory for culture and susceptibility testing within 12 hours.

Laboratory procedures

Samples were cultured on sheep blood agar and chocolate agar, followed by 24 hours of incubation at 37°C anaerobically. The isolates were identified morphologically by colony appearance and Gram staining. Optochin sensitivity and bile solubility testing were conducted on colonies that were potentially identifiable as S. pneumoniae by an alpha-hemolytic appearance on the culture media and lancet-shaped Gram-positive cocci appearing in pairs.

A 0.5 McFarland standard of S. pneumoniae was made from a 24-hour subculture by suspending colonies in sterile normal saline and inoculated by swabbing onto a plate of Mueller-Hinton agar supplemented with 7% sheep blood for susceptibility testing. Antibiotic susceptibility to penicillin G (1U), chloramphenicol (30µg), tetracycline (10µg), clindamycin (2µg), erythromycin (30µg), and ceftriaxone (30µg) was determined using modified Kirby-Bauer agar disc diffusion methods and the disc zone diameters were interpreted using the Clinical and Laboratory Standards Institute Guidelines¹².

Data management

Collected data were entered in Microsoft Excel 2010, cleaned, coded, and imported to SPSS Version 16.0 statistical package for analysis. Data were analyzed using descriptive statistics, frequencies, and bivariate analyses (cross-tabulations). The primary outcome was a pneumococcal carriage and the secondary outcome was antibiotic resistance. Statistical frequency distribution tables and graphs were used for data presentation in terms of proportions, absolute values, percentages, and confidence intervals for point approximations at a 95% level of confidence, with P<0.05 considered as statistically important.

Data quality control

Laboratory procedures were performed by laboratory scientists under the close supervision of a clinical microbiologist to ensure quality results were obtained. Data were double entered into Microsoft Excel for accuracy and reliability. ATCC 49619 S. pneumoniae was used as a control strain for quality assurance during isolation and drug susceptibility testing.

Demographic characteristics of the study participants

At the young child clinic, a total of ninety-three pairs were sampled, two of whom were excluded because the babies were presented by male caretakers, and one mother opted out due to fear of discomfort associated with the procedure for sample collection. Ninety pairs were recruited. In the community, a total of sixty-nine pairs were sampled, five were excluded because they presented children above five years of age, while two were presented by male caretakers. Sixty-two were recruited, making a total of 152 mother-baby pairs.

The study participants comprised 152 mothers and 152 babies. Of the 152 babies, 74 were male and 78 were female with the age range of 0–59 months. The youngest mother was 16 years, whereas the oldest was 44 years. None of the mothers who participated in the study reported having formal employment¹³.

Prevalence of pneumococci in mother-baby pairs

During the study, 304 samples were collected; 152 from the mothers and 152 from the children, making 152 mother-baby pairs. All samples were cultured, and antibiotic susceptibility was carried out on the isolated pneumococci. Out of 152 samples from the mothers, only five (5/152) isolates of pneumococci were obtained whereas seven (7/152) were isolated from the babies. Only one mother-baby pair (1/152) was found to be colonized with pneumococci in both mother and baby and the rest of S. pneumoniae colonized either the mother or baby.

Immunization coverage

During data collection, the immunization status of the baby was categorized as fully immunized, not immunized, and partially immunized among different age groups (Figure 1). There was high immunization coverage among the children above 12 months old but lower in the 3.5-<12 age group.

Figure 1. PCV-10 immunization coverage among children under five years old.

Antibiogram

The antibiotic susceptibility testing was done on positive isolates for both mother and baby.

Generally, a high trend of anti-microbial resistance was observed among the S. pneumoniae isolated (Table 1). The highest resistance patterns were recorded with chloramphenicol (50%) and tetracycline (50%), whereas the lowest resistance was recorded in clindamycin (17%).

Babies that were fully immunized had a lower likelihood of being colonized by S. pneumoniae than their non-immunized counterparts P<0.05. Other factors examined by this study were not significantly associated with colonization with S. pneumoniae among the babies.

Limitations

We were not able to serotype the pneumococci isolated to determine the circulating serotypes and to link pneumococcal carriage to the development of the disease.

Underlying data

OSF: Pneumococcal carriage and antibiotic susceptibility patterns in mother-baby pairs in a rural community in Eastern Uganda: a cross-sectional study. https://doi.org/10.17605/OSF.IO/H9X7R¹³

Data are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication).