{"paper_id":"63926443-82f0-4fa0-b124-786bc9f87c47","body_text":"Seroprevalence of teratogenic viruses among women of childbearing age in Botswana. \nI. Gobe1*, K. Baipoledi1, G. Skwenje1, M. Ntamo1, M. Motswaledi1. \n1. School of Allied Health Professions, Faculty of Health Sciences, University of \nBotswana, Gaborone, Botswana. \n*Correspondence: Gobei@ub.ac.bw \nAbstract \nBackground: Teratogenic viruses are viruses than can cross the placenta and infect a growing \nfoetus, resulting in malformations and birth defects. Some of the commonly known teratogenic \nviruses include cytomegalovirus (CMV), rubella, herpes simplex and varicella zoster  (VZV) \nviruses. Most birth defects associated with these infections affect the central nervous system \nand sensory organs leading to symptoms that include mental retardation , hearing loss and \nblindness. The economic burden caused by congenital birth defects is high, as many affected \nchildren require special care, therapeutic and educational services. Despite the risk posed by \nteratogenic viruses during pregnancy, there is no national screening for active CMV, Rubella \nor VZV infection during pregnancy in Botswana and most African countries. Furthermore, data \non the seroprevalence of these viruses among women of childbearing age is limited.  \nMethods and setting: This cross-section study used eighty-nine (89) residual plasma samples \nfrom Scottish Livingstone Hospital Laboratory  in Molepolole-Botswana. Samples were from \nwomen between the ages 15-49 years.  Samples were tested for antibodies against rubella, VZV \nand CMV using enzyme linked immunosorbent assay.  \nResults: Our results show a high seroprevalence of rubella IgG antibodies (97%), even though \na small proportion (3%) of women are still susceptible. There was also a high seroprevalence \nof CMV  IgG (100%) which was accompanied by an equally high CMV IgM of 98%. \nSeroprevalence of VZV IgG was low  (63%) and 3% of the samples showed active VZV \ninfection.  \nConclusions: Teratogenic viruses are a concern in the population. This calls for preventative \nmeasures which include prompt screening and vaccination of susceptible eligible women to \nprevent congenital abnormalities in children.  \nKey words: Teratogenic viruses, Seroprevalence, CMV, Rubella, VZV \n \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \nNOTE: This preprint reports new research that has not been certified by peer review and should not be used to guide clinical practice.\n\nIntroduction \nTeratogenic agents can interfere with normal fetal development. These agents include  but not \nlimited to drugs, viral infections, ma ternal conditions ( e.g. diabetes, epilepsy) and radiation  \n(Kaleelullah and Garugula, 2021). Viruses such as rubella virus, Cytomegalo virus (CMV) and \nvaricella zoster virus (VZV) can cross the placenta and result in fetal tissue damage  \n(Kaleelullah and Garugula, 2021). \nCMV and VZV are double-stranded DNA viruses from the Herpesviridae family. Upon initial \ninfection with one of these viruses, the virus persists indefinitely as a latent infection which \ncould reactivate under immunosuppression  (Richman et al., 2002 ). CMV  infections are \ngenerally asymptomatic in both children and adults (Johnson et al., 2012). Transmission occurs \nthrough contact with body f luids such as u rine and saliva  (Pesch et al., 2021) . However, \ncongenital CMV infection is a major cause of mental retardation, hearing loss and cerebral \npalsy (Johnson et al., 2012) . Congenital CMV infection s may occur following a primary \ninfection or as a re -infection of an expectant mother.  Primary infections occur in individuals \nwith low antibody titres against  the virus (Johnson et al., 2012). Primary CMV infection poses \na 30-40% risk of fetal transmission while non -primary or reactivation poses over 4%  risk \n(Coppola et al., 2019). Generally, poorer fetal outcomes are common if the pregnant woman is \ninfected during early gestation  (Pesch et al., 2021) . In areas with high CMV seroprevalence, \nseronegative mothers have  high rates of CMV acquisi tion with high placental transmission \n(Coppola et al., 2019) . Global CMV seroprevalence is 83% (Zuhair et al., 2019) . High \nprevalence of >90% is seen in WHO eastern Mediterranean regions (Zuhair et al., 2019)  and \nthe African region (Mhandire et al., 2019). According to the Centres for Disease Control and \nPrevention [CDC], approximately 1  in 200 babies are born with congenital CMV; 1 in 5 of \nthese will develop birth defects , the most common of which is hearing loss which can be \ndetected after birth or later in childhood (CDC,2024).  \nVaricella zoster infections are transmitted through the respiratory route. Primary varicella \nzoster infection causes chicken pox while secondary/reactivation causes shingles (Bhavsar and \nMangat, 2021) . Due to circulating mate rnal antibodies, reactivation of varicella zoster \n(Shingles) does not pose a risk to developing fetus. Primary infection with chicken pox poses \na risk to the fetus and can cause congenital varicella syndrome (Bhavsar and Mangat, 2021) . \nCongenital varicella zoster viral infection is common if primary infection occurs in the first \nhalf of pregnancy with greatest risk between 13 -20 weeks of gestation (Bhavsar and Mangat, \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\n2021). Furthermore, infants who get exposed to the virus 5 days before and 2 days after delivery \nare at an increased risk of experiencing severe disease with increased mortality (Singh et al., \n2022). Congenital varicella syndrome consists of skin lesions, neurologic defects, eye disease \nand skeletal anomalies (Sauerbrei and Wutzler, 2000). Unlike CMV, varicella zoster primary \ninfections can be prevented by vaccination. In  fact, W orld Health Organisation (WHO)  \nrecommends vaccination for all children . However , many developing countries and some \ndeveloped countries like the UK have not included this vaccine in their routine childhood \nimmunization schedules  (Singh et al., 2022) . The seroprevalence of VZV in developed \nvaccinating countries is high  with USA recording a seroprevalence of  >99% in persons 30yrs \nand older (Kilgore et al., 2003) , while Canada recorded 95% in people with HIV (Zou et al., \n2022). In Africa where the vaccine is not routinely administered the seroprevalence is low ; in \nSudan the VZV seroprevalence lies at 50% (Adam et al., 2023) , while Nigeria recorded 66% \nprevalence among children (Oripelaye et al., 2024). \nRubella is an RNA virus from the Rubi virus genus (Das and Kielian, 2021). Rubella infections \nspread through respiratory droplets. These infections are generally mild and are characterised \nby a fever and rash . Nonetheless, in pregnancy, rubella infection can cause miscarriages or \ncongenital rubella syndrome (CRS), (Das and Kielian, 2021). According to the WHO, infection \nof a pregnant woman in the f irst trimester carries a 90% risk of fetal transmission and \nconsequently CRS. CRS symptoms include hearing impairment, eye, and heart defects. Other \nCRS associated disabilities include autism, di abetes mellitus and thyroid dysfunction (WHO, \n2024). CRS is c ommon in populations where women of childbearing age are seronegative. \nAccording to a survey carried out on reports from 1963-2009, the seroprevalence of Rubella in \nAfrica ranged from 68-98% (Goodson et al., 2011). No data from Botswana was available in \nthis report. However, neighbouring South Africa recorded a seroprevalence of 60% (Goodson \net al., 2011). The WHO report of 2020 estimated that about 100 ,000 children are affected by \nCRS annually worldwide (WHO, 2020 rubella vaccine po sition paper). The best preventative \nmeasure against CRS is vaccination. A single dose of rubella vaccine gives 95%  immunity \nwhich is similar to that achieved by natural infection (WHO, 2024 ). Botswana introduced \nrubella vaccine in its childhood immunisation schedule in 2016 (MoH-Botswana). This should \nincrease the seroprevalence of rubella in the long run . However, there is still limited data on \ncurrent seroprevalence of rubella in women of childbearing age.  \nThe economic burden caused by congenital birth defects is high, as many affected children \nrequire special care, therapeutic and educational services. However, despite the risk posed by \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nteratogenic viruses during pregnancy, there is no national screening fo r active CMV, Rubella \nor VZV infection during pregnancy in Botswana.  Furthermore, data on the seroprevalence of \nthese viruses among women of childbearing age is limited.  Therefore, the aim of this study is \nto determine the seroprevalence of these teratogenic viruses among women of childbearing age \nin Botswana. \n \nMaterials and methods \nStudy design and settings \nThe cross-sectional study was conducted between October and December 2023.   Residual \nplasma samples were collected from Scottish Livingstone hospital laboratory in Molepolole, \nBotswana.  Samples were stored at -20oC until further testing at the School of Allied Health \nScience Laboratory, University of Botswana. \nStudy population and sampling strategy \nEighty-nine, residual (EDTA -anticoagulated, separated) blood samples were collected from \nwomen of childbearing age (15 -49 Years) from Scottish Livingstone hospital laboratory in \nMolepolole, Botswana. Non-probability convenience sampling was carried out to include all \nfemales who are in the childbearing age group. Only samples that were stored in the refrigerator \nfor not more than a week were selected.  \nThe sample size was calculated using a single population proportion sample size formula  by \nDaniel (1999). A seroprevalence of 95% was used for sample size calculation for CMV and \nrubella. These proportions were deduced from a n epidemiology review article on CMV in \nAfrica (Mhandire et al. , 2019)  and a rubella seroprevalence study that was ca rried out in \nWestern Cape, South Africa (Corcoran and Hardie, 2005). For VSV, a seroprevalence of 50.4% \nwas used for the calculations as it has been observed in data from Sudan  (Adam et al., 2023).  \nCalculated sample size was 73 for both rubella and CMV and 385 for VSV.  \nEthical considerations \nEthical clearance was obtained from University of Botswana Office of Research and \nDevelopment and the Health Research and Development  committee (HDRC) at the Ministry \nof Health, Botswana. Permission was sought from Scottish Livingstone Hospital  for sample \ncollection. The residual samples from Scottish Livingstone Hospital Laboratory were de -\n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nidentified and given new random identities (“001-089”).  Demographic data collected included \nage and sex only.  \nData collection \nCommercial enzyme -linked immunosorbent assay (ELISA) kits  were supplied by My \nBioSource (San Diego, USA ), which included : Rubella virus IgG ELISA kit - MBS495665, \nHuman VZV IgG ELISA kit -MBS7612746, VZV  IgM ELISA kit -MBS7612747, Human \nCMV IgG ELISA kit -MBS57269157 and human CMV IgM ELISA kit -MBS7269158. The \nELISA kits were used in accordance with the manufacturer’s instructions to measure the \nconcentrations of IgG/IgM against the VZV, CMV and Rubella. \nRubella  \nPlasma samples were diluted 1:100 and then tested according to manufacturer’s instructions. \nA standard curve was prepared using a set of standards containing 0, 10, 50, 100 IU/mL. These \nstandards were supposed to yield OD 620 of <0.2, >0.2, >0.7 and > 1.1, respectively. Using the \nstandard curve and the linear equation calculated, the concentration of each sample was \ncalculated and interpreted as either positive or negative accordingly: >15 IU/mL: positive, 10-\n15 IU/mL: Equivocal and <10 IU/mL: negative.  \nVZV \nSamples and controls were run according to the manufacturer’s instructions. Interpretation of \nresults depended on the results of the controls. For the controls to pass, mean OD 450 of the \nnegative control should be ≤0.1 and the positive control was ≥0.8. A cut-off value was \ncalculated as the mean OD of negative control +0.10. Samples with absorbance values less than \nthe cut-off Value were NON-REACTIVE and were considered NEGATIVE for VZV -\nIgG/IgM. Sample s with absorbance values greater than the  cut-off value were considered \nPOSITIVE for VZV-IgG/IgM.  \nCMV \nBoth CMV IgG -Ab and IgM  ELISA kits applied the competitive enzyme immunoassay \ntechnique, and the intensity of colour was inversely proportional to the amount of antibody \nmeasured. Interpretation of results was aided by the standards that were provided. Standar ds \nA-F with concentrations ranging from 0, 50, 100, 250, 500 and 1000 mg/mL respectively were \nrun with samples. A standard curve was plotted relating the intensity of the colour (O.D450) to \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nthe concentration of standards. The CMV IgG/IgM-Ab concentration in each sample was then \ncalculated from this standard curve. Any sample with an OD 450 of more or equal to that of \nStandard A (0 mg/mL) was deemed negative while samples with OD 450 of less than that of \nStandard A were deemed positive.  \nData analysis  \nThe statistical significance level was set at p<0.05. All statistical analyses were performed \nusing Microsoft Excel 365. Both categorical and quantitative data were used in the study.  To \ndetermine the prevalence/frequencies , calculations were total number of samples that tested \npositive for IgG or IgM antibodies divided by the total number of samples tested multiplied by \n100. Bar charts and box plots were generated using GraphPad Prism 10.3.1. \n \nResults \nA total of 89 samples from women of childbearing age was analysed. Their age range d from \n15- 44 years, with a mean age of 30.4 years.  \nOut of the 89 samples tested 2% (n=2/89) were seronegative for CMV IgM.  100% (n=89/89) \nwere positive for CMV-IgG while 98% (87/89) were positive for CMV- IgM (Figure 1.0).  \nCMV IgM CMV IgG\n0\n20\n40\n60\n80\n100\nNumber of patients\nPositive\nNegative\n \nFigure 1.0: CMV IgM and IgG seroprevalence among 89 samples tested. Antibody testing was \ndone using ELISA method. \n \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nFigure 2.0 below shows the c oncentration distribution of CMV -specific antibodies (IgG and \nIgM). The concentration of CMV IgG ranged from 32-476 mg/mL with a mean of 95.5 mg/mL \nwith lower quartile and upper quartile of 65 -95 mg/ mL respectively. The concentration of \nCMV IgM ranged from 0-481 mg/mL with a mean of 209 mg/mL and lower quartile and upper \nquartile of 261 and 136 respectively. The mean concentration of IgM is higher than that of IgG \nwith a statistically significant p= 0.005.  \nCMV IgG\nCMV IgM\n0\n200\n400\n600\nConcentration (mg/ml)\n \nFigure 2.0. Concentration of CMV IgG and IgM in the sample population \n \n \n \n \n \n \n \n \n \n \n \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nOf the 89-plasma collected and  tested for Rubella virus IgG antibodies, an overall \nseropositivity rate of 96.6 % (n=86/89) was observed.  However, 3.4% (n=3/89) of the samples \ntested negative for Rubella virus IgG antibodies (Figure 3.0).  \nRub IgG positive\nRub IgG negative\n0\n20\n40\n60\n80\n100\nNumber of patients\n \nFigure 3.0: Rubella IgG seroprevalence among 89 samples tested. Antibody testing was done \nusing ELISA method. \n \nThe concentration of Rubella IgG ranged from 1.1 -110 IU/mL with a mean of 62 IU/mL with \na lower quartile and upper quartile of 50 and 75 IU/ML respectively (Figure 4.0).  \n0\n50\n100\n150\nRubella IgG concentration ( IU/ml)\n \n \nFigure 4.0. Concentration of Rubella IgG in the sample population \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nFigure 5.0 below shows the number of people who were seropositive for VZV-IgG and IgM. \nOut of the 89 samples, 53(63%) were positive for VZV IgG while 33 (37%) were negative for \nVZV IgG. 3% (n=3/89) were positive for VZV IgM and 86 (97%) were seropositive for VZV \nIgM. Among the three samples which were positive for IgM, 2 had both IgG and IgM while \none was positive for IgM only.  \n \nVZV IgM VZV IgG\n0\n20\n40\n60\n80\n100\nNumber of patients\nPositive\nNegative\n \nFigure 5.0: VZV IgG seroprevalence among 89 samples tested. Antibody testing was done \nusing ELISA method. \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nDiscussion \n  \nCMV: \nThis study shows a high seroprevalence of CMV IgG which is 100%. The results are consistent \nwith data from other African studies which estimate CMV seroprevalence among women to be \n>90% (Mhandire et al., 2019). Furthermore, the results are consistent with the global narrative \nthat states that CMV seroprevalence correlates with socio -economic status and education \nlevels, with high prevalence seen in under-developed and developing countries (Fowler et al., \n2022). Our study also showed a high seroprevalence of CMV IgM in women of childbearing \nage, in comparison to the  global seroprevalence of CMV IgM, which is estimated to be 13.7% \n(Adane and Getawa, 2021) . These results were puzzling and alarming as they suggest a high \nrisk for congenital malformations in Botswana. In Europe, North America and Latin America, \nthe seroprevalence of CMV IgM is 1- 4.6%,  2.3-4.5%  and 0-0.7% respectively, among women \n(Fowler et al., 2022). In the African countries of Egypt, Kenya, Sudan, Ethiopia and Nigeria, \nthe seroprevalence of CMV I gM was estimated to be 7.3%,  8.1%, 2.5%, 5.5% and 11.1% \nrespectively (Hailemariam et al., 2021). Therefore, the CMV IgM seroprevalence found in this \nstudy is alarmingly high compared to other studies both in Europe and in Africa. CMV IgM is \ngenerally considered as a transient marker of primary infection. However, only 50% of \nindividuals who are CMV IgM positive have actually been shown to have a primary infection \n(Prince and Lapé -Nixon, 2014). Studies have also shown that CMV IgM specificity i s poor \nindicator of recent infection because it is produced during CMV viral reactivation, re-infection \nwith a different CMV strains and it can also persist for over a year after primary CMV infection \n(Prince and Lapé-Nixon, 2014). The high seroprevalence of CMV IgM in our study could mean \nthat active CMV infections are rife in our population  or that IgM is not cleared after activ e \ninfection. The latter might be true because in a study where CMV DNA was detected in HIV \npositive pregnant women in Botswana , the prevalence was found to be 14.6%(Moraka et al., \n2019). Nonetheless, the high CMV IgM suggest something inherent in Botswana’s population \nthat needs to be investigated further. \nSince CMV IgM is inconclusive for primary infection  and PCR testing is expensive , studies \nhave shown that CMV IgG avidity testing is a more specific marker to identify pregnant women \nwith recent CMV infection (Prince and Lapé-Nixon, 2014). Primary infections are worrisome \nbecause the risk of congenital CMV infection has been shown to be approximately 40%  in \ninfants born to mothers who acquire a primary  CMV infection after conception (Prince and \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nLapé-Nixon, 2014) . In spite of this, there is  contradicting evidence that shows that in \npopulations with very  high CMV IgG seropositivity , such as  Brazil (maternal CMV IgG \nseropositivity of >97%), there are more cases of congenital CMV infections (Mussi-Pinhata et \nal., 2018). This indicates that CMV IgG seropositivity is not necessarily protective. Despite \nhigh CMV seroprevalence in Africa, comparative data is lacking. \nRubella: \nOur study shows that the seroprevalence of rubella is comparable to that found in other African \ncountries which range from 68 -98% (Goodson et al., 2011).  A recent study from Morocco \nfurther recorded an IgG seroprevalence of 85% among pregnant women  (Zahir et al., 2020) . \nGenerally, the cut-off point for protective rubella antibodies is 10  IUM/mL (Kempster et al., \n2020). Our results show  a mean protective im munity that is significantly higher than the \nrecommended cut threshold. These results show that most of the women of childbearing age \nare protected from rubella infections.  The Ministry of Health in Botswana introduced rubella \nvaccine in 2016, hence the vaccinated population are not yet at childbearing age. Therefore, \nprotective immunity shown by this study reflects immunity due to natural infection. These \nresults also show that rubella infections are very common which is a concern especially for \nthose that are seronegative and are planning to be pregnant. \n Although our study show that most participants are protected, there is still that 3% of women \nof childbearing age who still carry the risk of contracting rubella and consequently CRS, if they \nget pregnant. This calls for interventions such as screening and vaccinating those that are \nseronegative. Since rubella vaccine as a life attenuated vaccine may potentially be unsafe for \npregnant women (WHO, 2024) it would be best to  avail the vaccine to all women of \nchildbearing age who are planning to be pregnant. \n VZV \nSeroprevalence of VZV IgG among women of childbearing age in Botswana was the lowest \nwith significant number of individuals being unprotected and susceptible. We found a small \npercentage of the study participants who had recent or active VSV  infection as shown by \npositive IgM results in the absence of IgG. Limited data is available on the seroprevalence of \nVZV in Africa, the few studies done show a significantly low seroprevalence when compared \nto developing nations as found in Sudan which recorded a seroprevalence of 50% (Adam et al., \n2023).The seroprevalence of VZV in developed vaccinating countries is high with USA \nrecording a seroprevalence of >99% in persons 30 years and older (Kilgore et al., 2003), while \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nCanada recorded 95% among people living with HIV (Zou et al., 2022) . The low \nseroprevalence of VZV which indicates a high proportion of susceptible women is worrisome. \nCongenital  VZV infections are common if primary infection occurs in the first half of \npregnancy with greatest risk between 13 -20 weeks of gestation  (Bhavsar and Mangat, 2021) . \nFurthermore, infants who get exposed to the virus 5 days before and 2 days after delivery are \nat an increased risk of experiencing severe disease with increased mortality (Singh et al., 2022). \nThis calls for countries to heed WHO recommendation of including VZV vaccine in routine \nvaccination programs and target susceptible women who are planning to get pregnant.   \nLimitations: There were several limitations to this study. First the sample size of the study \nwas small. This was especially true for VZV data where an estimated sample size of 385 was \nsupposed to be used but only 89 samples were analysed . Furthermore, co -morbidities which \ncould affect the data such as HIV were not taken into consideration. Further studies need to be \ndone to take into consideration these limitations.  \nConclusions: Our study shows a  high seroprevalence of rubella  IgG (97%), even though a \nsmall proportion (3%) of women are still susceptible. There is also a high seroprevalence of \nCMV IgG (100%). However, according to literature, this does not necessarily translate to low \ncongenital CMV infections hence prompt screening of pregnant women for active CMV \ninfections is necessary. The low seroprevalence of VZV IgG (63%) indicates that a significant \nproportion of the population is susceptible.  \nRecommendations: The results call for improved strategies to reduce the burden of congenital \ndefects. These could include early identification through maternal or new -born screening, \nvaccination, behavioural interventions and treatment for infected pregnant women and infants.   \nAcknowledgements \nThe authors wish to express gratitude to the staff and management of Scottish Livingstone \nhospital laboratory in Molepolole, where samples were obtained.  \nCompeting interests:  \n The authors declare that they have no financial or personal relationships that may have \ninappropriately influenced them in writing this article. \nAuthor contributions \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nIrene Gobe conceived the idea. The experiments were carried out by K. Baipoledi, G. Sekwenje \nand M.  Ntamo under the supervision of Irene Gobe. I. Gobe also analysed the results and \ndrafted the article. Prof. M. Motswaledi participated in data analysis, critiqued the content, and \ngave final approval of the version to be published. \nData availability \nData sharing regarding this article is available upon written request to the corresponding author. \nDisclaimer \nThe views and opinions expressed in this article are those of the author(s) and do not necessarily \nreflect the official policy or position of any affiliated agency of the authors. \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nReferences \nADAM, O., MUSA, A., KAMER, A., BENHARRAT, S. & HÜBSCHEN, J. M. 2023. Active circulation of \nvaricella zoster virus among different age groups in Sudan. Epidemiology & Infection, 151, \ne10. \nADANE, T. & GETAWA, S. 2021. Cytomegalovirus seroprevalence among blood donors: a systematic \nreview and meta-analysis. Journal of International Medical Research, 49, \n03000605211034656. \nBHAVSAR, S. M. & MANGAT, C. 2021. Congenital varicella syndrome. \nCOPPOLA, T., MANGOLD, J. F., CANTRELL, S. & PERMAR, S. R. 2019. Impact of maternal immunity on \ncongenital cytomegalovirus birth prevalence and infant outcomes: a systematic review. \nVaccines, 7, 129. \nCORCORAN, C. & HARDIE, D. R. 2005. Seroprevalence of rubella antibodies among antenatal patients \nin the Western Cape. South African Medical Journal, 95. \nDAS, P. K. & KIELIAN, M. 2021. Molecular and structural insights into the life cycle of rubella virus. \nJournal of virology, 95, 10.1128/jvi. 02349-20. \nFOWLER, K., MUCHA, J., NEUMANN, M., LEWANDOWSKI, W., KACZANOWSKA, M., GRYS, M., \nSCHMIDT, E., NATENSHON, A., TALARICO, C. & BUCK, P. O. 2022. A systematic literature \nreview of the global seroprevalence of cytomegalovirus: possible implications for treatment, \nscreening, and vaccine development. BMC Public Health, 22, 1659. \nGOODSON, J. L., MASRESHA, B., DOSSEH, A., BYABAMAZIMA, C., NSHIMIRIMANA, D., COCHI, S. & \nREEF, S. 2011. Rubella epidemiology in Africa in the prevaccine era, 2002–2009. The Journal \nof infectious diseases, 204, S215-S225. \nHAILEMARIAM, M., MEKONNEN, Z., CLAEYS, G. & PADALKO, E. 2021. Congenital cytomegalovirus \ninfections:(no) focus on Africa: a review. Gynecology and Obsteatrics, 11, 1-6. \nJOHNSON, J., ANDERSON, B. & PASS, R. F. 2012. Prevention of maternal and congenital \ncytomegalovirus infection. Clinical obstetrics and gynecology, 55, 521-530. \nKALEELULLAH, R. A. & GARUGULA, N. 2021. Teratogenic genesis in fetal malformations. Cureus, 13. \nKEMPSTER, S. L., ALMOND, N., DIMECH, W., GRANGEOT-KEROS, L., HUZLY, D., ICENOGLE, J., EL \nMUBARAK, H. S., MULDERS, M. N. & NÜBLING, C. M. 2020. WHO international standard for \nanti-rubella: learning from its application. The Lancet Infectious Diseases, 20, e17-e19. \nKILGORE, P. E., KRUSZON‐MORAN, D., SEWARD, J. F., JUMAAN, A., VAN LOON, F. P., FORGHANI, B., \nMCQUILLAN, G. M., WHARTON, M., FEHRS, L. J. & COSSEN, C. K. 2003. Varicella in Americans \nfrom NHANES III: implications for control through routine immunization. Journal of medical \nvirology, 70, S111-S118. \nMHANDIRE, D., ROWLAND-JONES, S., MHANDIRE, K., KABA, M. & DANDARA, C. 2019. Epidemiology \nof Cytomegalovirus among pregnant women in Africa. Journal of infection in developing \ncountries, 13. \nMORAKA, N. O., MOYO, S., MAYONDI, G., LEIDNER, J., IBRAHIM, M., SMITH, C., WEINBERG, A., LI, S., \nTHAMI, P. K. & KAMMERER, B. 2019. Cytomegalovirus Viremia in HIV-1 Subtype C Positive \nWomen at Delivery in Botswana and Adverse Birth/Infant Health Outcomes. JAIDS Journal of \nAcquired Immune Deficiency Syndromes, 81, 118-124. \nMUSSI-PINHATA, M. M., YAMAMOTO, A. Y., ARAGON, D. C., DUARTE, G., FOWLER, K. B., BOPPANA, \nS. & BRITT, W. J. 2018. Seroconversion for cytomegalovirus infection during pregnancy and \nfetal infection in a highly seropositive population:“The BraCHS Study”. The Journal of \ninfectious diseases, 218, 1200-1204. \nORIPELAYE, M. M., OLANREWAJU, F. O., AJANI, A. A., AKINBORO, A. O., ENITAN, A. O. & ONINLA, O. \nA. 2024. Seroprevalence of varicella-zoster virus among people living with human \nimmunodeficiency virus in Ile-Ife, Nigeria: A cross-sectional study. Journal of Clinical \nSciences, 21, 14-19. \nPESCH, M. H., KUBOUSHEK, K., MCKEE, M. M., THORNE, M. C. & WEINBERG, J. B. 2021. Congenital \ncytomegalovirus infection. BMJ, 373. \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint \n\nPRINCE, H. E. & LAPÉ-NIXON, M. 2014. Role of cytomegalovirus (CMV) IgG avidity testing in \ndiagnosing primary CMV infection during pregnancy. Clinical and Vaccine Immunology, 21, \n1377-1384. \nSAUERBREI, A. & WUTZLER, P. 2000. The congenital varicella syndrome. Journal of Perinatology, 20, \n548-554. \nSINGH, S., SHARMA, A., RAHMAN, M. M., KASNIYA, G., MAHESHWARI, A. & BOPPANA, S. B. 2022. \nCongenital and perinatal varicella infections. Newborn (Clarksville, Md.), 1, 278. \nZAHIR, H., ARSALANE, L., ELGHOUAT, G., MOUHIB, H., ELKAMOUNI, Y. & ZOUHAIR, S. 2020. \nSeroprevalence of rubella in pregnant women in Southern Morocco. The Pan African \nMedical Journal, 35. \nZOU, J., KRENTZ, H. B., LANG, R., BECKTHOLD, B., FONSECA, K. & GILL, M. J. Seropositivity, risks, and \nmorbidity from varicella-zoster virus infections in an adult PWH cohort from 2000–2020.  \nOpen Forum Infectious Diseases, 2022. Oxford University Press, ofac395. \nZUHAIR, M., SMIT, G. S. A., WALLIS, G., JABBAR, F., SMITH, C., DEVLEESSCHAUWER, B. & GRIFFITHS, \nP. 2019. Estimation of the worldwide seroprevalence of cytomegalovirus: a systematic \nreview and meta‐analysis. Reviews in medical virology, 29, e2034. \nDaniel W.W. (1999). Biostatistics: A foundation for Analysis in Health Sciences. 7th edition. \nNew York: John Wiley & sons. \nWHO, 2024. Rubella fact sheet. https://www.who.int/news-room/fact-sheets/detail/rubella. \nRetrieved 9/August/2024.  \n(WHO, 2024: https://www.who.int/news-room/fact-sheets/detail/rubella. Retrieved, 29 July \n2024). \n . CC-BY 4.0 International licenseIt is made available under a \nperpetuity. \n is the author/funder, who has granted medRxiv a license to display the preprint in(which was not certified by peer review)preprint \nThe copyright holder for thisthis version posted September 22, 2024. ; https://doi.org/10.1101/2024.09.20.24314038doi: medRxiv preprint","source_license":"CC-BY-4.0","license_restricted":false}