Intestinal Parasitic Infections, Treatment and Associated Factors among Pregnant Women in Sao Tome and Principe: A Cross-Sectional Study

Background Intestinal parasitic infections (IPIs) are a public health problem in developing countries such as Sao Tome and Principe (STP) although the pregnancy burden of IPIs is unknown in this endemic country. Thus, the aim of this study was to determine the prevalence of IPIs, prescribed anthelmintics, and associated factors among pregnant women admitted to Hospital Dr. Ayres de Menezes (HAM). Methods A hospital-based cross-sectional study was conducted among pregnant women admitted to the HAM who had undergone antenatal copro-parasitological screening. Data were abstracted from antenatal care (ANC) cards regarding parasitological results and anthelmintic prescriptions. A structured questionnaire face-to-face interview was also applied. Pregnant women with an IPI (210) were compared to noninfected women (151). Data analysis was performed using SPSS version 25.0. Odds ratios (ORs) with 95% confidence intervals (CIs) for factors associated with IPIs were estimated using multiple logistic regression models. A p value <0.05 was considered statistically significant. Results A total of 361 participants (210 IPI and 151 no-IPI) with a mean age of 26.96 (SD: 7.00) were included. The overall prevalence of IPI was 58.2% (95% CI 52.9 to 63.3), mainly due to helminthiasis, with a 55.9% (95% CI 50.7–61.2%) rate. Ascaris lumbricoides (90.9%) was the most predominant parasite species identified followed by Trichuris trichiura (13.8%). Polyparasitism was observed in 25 cases (11.9%). Anthelmintics were prescribed to 23% of pregnant women. S intercalatum (11) and E histolytica (7) infections were not adequately treated. IPI was significantly associated with primary education (AOR 1.73 (95% CI: 1.10–2.71)), unemployment (AOR 1.94 (95% CI: 1.20–3.13)), and parity of five or above (AOR 3.82 (95% CI: 1.32–11.08)). Conclusion This study highlights the IPI burden, associated factors, and missing treatment opportunities among pregnant women with STP. This study is a useful tool for policymakers in STP to enhance the health of women and their unborn babies.

Less frequent but also prone to cause adverse birth outcomes and complications during pregnancy are invasive enteric parasites such as Schistosoma spp. and the protozoan Entamoeba histolytica [1]. Schistosomiasis afects approximately 200 million people and is "second only to malaria in public health importance," with pregnant women being one of the important at-risk groups [5,6]. Entamoeba histolytica, responsible for amebiasis, remains one of the top three parasitic causes of mortality, with an estimated 50 million people afected worldwide [7,8].
Te major concerns regarding IPI in pregnant women are the associated higher risks for adverse maternal outcomes, such as anemia as well as poor pregnancy weight gain [9][10][11]. Additionally, IPI implications in newborn health are the risk of low birth weight (LBW), intrauterine growth retardation, prematurity, stillbirths, and neonatal mortality, a considerable burden in countries that already have a high rate of neonatal mortality and morbidity [12][13][14].
Regarding the anthelmintic treatment of IPI during pregnancy, there is still some controversy [15,16], although it is endorsed by the World Health Organization (WHO) [17][18][19]. Te most recommended drugs for treating IPI in pregnancy are albendazole for helminths, praziquantel for Schistosoma, and metronidazole for protozoan parasites [20,21]. Nonetheless, available evidence shows that most pregnant women in IPI-endemic settings do not receive any deworming medication, with the WHO setting a target of reaching a 75% rate of deworming pregnant women by 2030 [3,22].
Knowing each country's reality and specifc associated factors promoting IPI among pregnant women will allow us to tackle it more accurately. In general, factors associated with IPIs are intimately connected with poverty and are the same for both the general population and pregnant women [23]. For instance, a well-known independent risk factor is WASH, an acronym for inadequate water supply, sanitation, and hygiene, which is also the major factor responsible for mortality and burden of disease due to diarrhea-causing infections in LMICs [24,25].
Sao Tome and Principe (STP) is the second smallest African country, a low-income setting, endemic for helminthiasis and schistosomiasis (S intercalatum) [26,27]. Te current country strategy is preventive IPI chemotherapy to be given only to preschool and school-aged children [28]. Infected pregnant women receive anthelmintic treatment in the second or third trimester on a case-by-case basis although the country's prevalence of anemia among pregnant women is around 61% [29].
To our knowledge, intestinal parasitic infections among pregnant women in Sao Tome and Principe have never been studied. Tis study was undertaken within the context of a broader project on the causes and risk factors contributing to neonatal mortality and adverse birth outcomes in STP [30,31].
We sought to determine the prevalence of IPIs, prescribed anthelmintics, and associated factors among pregnant women admitted to Hospital Dr. Ayres de Menezes.
Tus, these research fndings will help to design targeted interventions for these NTDs among pregnant women in STP and better allocate resources to where they are needed.

Study Design.
A hospital-based cross-sectional study was carried out on pregnant women admitted to the HAM maternity unit for delivery.

2.2.
Setting. Te archipelago of Sao Tome and Principe is an African island nation, with 219,161 inhabitants [29]. Te level of poverty is high, with 47% of the population practicing open defecation and only 69.8% having access to clean and safe drinking water [29].
Tis study was conducted at the HAM maternity unit, which has an annual average delivery rate of 4500 births, approximately 82.4% of all deliveries in the country, as it is the only hospital in the country. Antenatal care (ANC) coverage in STP is high, and women are asked to perform a routine copro-parasitological exam during the pregnancy period. Te exam consists of stool sampling using the direct wet mount method and formol-ether concentration method and then observation by trained parasitologists who declare it positive when various stages of the parasites, such as trophozoites, cysts, ova, and larvae, are identifed. Te results from copro-parasitological exams, as well as prescribed anthelminthics, are registered by the nurses in the ANC pregnancy card that pregnant women carry along all ANC visits and bring to the HAM at the time of labor.

Participants.
All pregnant women admitted to the HAM maternity unit for delivery constituted the source population, whereas the study populations were selected pregnant women admitted to the HAM maternity unit during the study period. Te recruitment occurred from July 2016 to November 2018.

Eligibility Criteria.
All women admitted to the HAM maternity unit for delivery with a gestational age of 28 weeks or more were eligible to be enrolled in the study. Tose who gave birth in the rural basic maternity units but were immediately transferred to HAM for postpartum medical evaluation or newborn follow-up were also eligible to be included in the study.
Te exclusion criteria included the following: (1) pregnant women with gestational age less than 28 weeks, (2) women who had no ANC pregnancy card, (3) women with cognitive impairment, (4) women who were unstable due to postpartum complications, and (5) adolescent or illiterate mothers who had not obtained permission from their parents or legal guardians to participate in the study. Pregnant women without ANC copro-parasitological screening (147) and pregnant women with HIV, sickle cell disorder, and malaria (10) were also excluded from this study.
A total of 361 pregnant women with a copro-parasitological exam performed during the ANC period and with the result registered in the pregnant ANC card were included. Pregnant women were divided into two groups, pregnant women with intestinal parasite infection (at least one) versus those without any IPI for associated factor assessment. A fowchart of participation in the study is shown in Figure 1.

Sampling Method.
Te software used for sample calculation was Raosoft (http://www.raosoft.com/samplesize. html), and a minimum sample size of S � 355 was recommended. For the original study, participants were enrolled based on the following assumptions: a two-sided 95% confdence level and a power of 80% to detect an odds ratio of at least two for neonatal adverse birth outcomes. Since the sample size was not calculated for the present outcomes, a power analysis was performed, varying from 79% to 89% for this study.
Pregnant women were selected randomly until the required sample size was achieved. Participants were invited to participate in the study after admission to the maternity unit of HAM. Each day, from the pile of admission folders, every second interval folder was selected and then carried on asking for consent for enrollment. Women's consent to participate in the study was obtained at the time of admission at HAM, but the interview was held after a woman was stabilized and ready to be discharged.

Data Source.
Te data source for this study consisted of data abstracted from the ANC pregnancy card plus a face-toface interview. Te ANC pregnancy card of each study participant was used to collect information such as the copro-parasitological result and anthelmintic prescription. Data on the sociodemographic characteristics of the participants were gathered using a structured questionnaire through a face-to-face interview with the main investigator before the women were discharged. Issues covered included the mother's and father's socioeconomic characteristics and reproductive history. All data were recorded in the app survey tool.
Polyparasitism was defned as the presence of two or more parasite species in the same host.
Te sociodemographic variables studied were as follows: (i) maternal age, (ii) maternal education, (iii) maternal occupation, (iv) marital status, (v) number of antenatal care visits, and (vi) area of residence. Mother occupation variable was categorized into two groups: employed (professional and service) and unemployed (housewife, farmer, business, and student). Education was included as a categorical variable: no education, primary education, secondary education, and higher education. Te residence was defned as urban or rural, the frst when women were living in the capital city (Água Grande) and rural in all other districts. We also analyzed hygiene and sanitation variables such as having access to improved water and type of toilet (toilets with fush and pit latrines) or open defecation. Parity was categorized as nullipara (0), multipara (1-4), and grand multipara (5 or above).

Data Quality Control.
Te questionnaires were administered in Portuguese, the national language. Continuous follow-up and supervision of data collection were made by the supervisors. Te collected data were checked daily for completeness. Te feld investigator (a pediatrician) executed and was responsible for the main activities as follows: (1) obtaining consent and enrollment of the mothers, (2) face-to-face interviews, (3) abstraction of data from antenatal pregnancy cards, and (4) data entry into the app survey tool.

Statistical Analyses.
Data analysis was carried out in three stages. Te frst stage involved pooling data for descriptive statistics with categorical variables being presented as frequencies and percentages and quantitative variables as the mean and standard deviation. In the second stage, the chi-square test and Fisher's exact test were used to describe the relationship between pregnant women with IPI and the categorical explanatory variables. Te third stage involved a univariable analysis to identify the candidate variables for the multivariable analysis (with a p value <0.25). In all these  Moreover, all methods in our study were performed in accordance with the relevant guidelines and regulations in practice.
Written informed consent was obtained from all participants after the purpose of the research was explained orally by the researcher. Approval by the participants' parents or legal guardians was asked in the case of adolescents under 16 years of age or illiterate women.
Te participants or their legal representatives also consented to have the results of this research work to be published. Participation in the survey was voluntary, as participants could decline to participate at any time during the study.

Characteristics of the Population.
A total of 361 pregnant women with a mean age of 26.96 (SD: 7.00) years old were included in this study.
Most of the participants had primary education, lived in rural areas, and were unemployed as described in Table 1. More than three-quarters of the participants were multiparous, and half had complete ANC attendance. A high proportion had access to improved water sources, and half of the pregnant women had sanitation.

Prevalence of Parasitic Infections.
Of the total number of pregnant women, 210 (58.2%, 95% CI 52.9 to 63.3) were infected with at least one pathogenic parasite. Te types of parasites identifed in the 210 copro-parasitological specimens are presented in Table 2.
Coinfection with diferent parasites (polyparasitism) in the same host was observed in 25 pregnant women (11.9%), as described in Table 3.

IPI Pregnant Women Treatment.
Te anthelminthic prescriptions registered in the ANC pregnancy cards were albendazole, mebendazole, metronidazole, and piperazine. Table 4 further describes the prescription according to the type of parasite. Regarding the helminthiasis group, a total of 23.8% (46) received anthelminthic treatment. Adequate treatment prescription (albendazole, mebendazole, or piperazine) was identifed in 40 pregnant women with helminthic-IPI. Five received metronidazole inadequately.
Anthelminthic prescriptions for women infected with S intercalatum and E histolytica were all identifed as inadequate (Table 4).

Pregnant Women with IPI versus Pregnant Women with
No-IPI. Te comparison of sociodemographic characteristics between pregnant women with IPIs and those without IPIs is shown in Table 1.

Factors Associated with IPIs in Pregnant
Women: Univariable Analysis. Table 5 concerns the univariable regression analysis to identify candidates for the multivariable analysis among factors for IPI during pregnancy. Te results indicate that the odds of having an IPI during pregnancy increased twice for pregnant women with primary education (cOR 2.58; 95% CI 1.14-5.83; p � 0.023). Te odds of having an IPI increased twice for unemployed women (cOR 2.03; 95% CI 1.28-3.19; p � 0.002). Women with a parity of 5 or above had 4.19 times higher odds of having an IPI during pregnancy (95% CI 1.49-11.79; p � 0.006) than nulliparous women. Women with 4 to 7 visits and those with 8 or more ANC visits were 68% (cOR 0.32; 95% CI 0.11-0.89) and 63% (cOR 0.37; 95% CI 1.13-1.04) less likely to have an IPI when compared to those with 1 to 3 ANC visits, with a statistical signifcance of p � 0.030 and p � 0.060, respectively.

Factors Associated with IPIs in Pregnant Women: Multivariable Analysis.
Te results obtained from the multivariable logistic regression model are also depicted in Table 5, showing the adjusted association strength between the factors that remained in the fnal model and intestinal parasite infections in pregnant women. Results indicate that  women with primary education, not working and with a parity of 5 or above, are more likely to have an IPI during pregnancy.

Discussion
Women living in LMICs are known to have a higher risk of acquiring an IPI during their pregnancy and consequently sufering complications from these NTDs [1]. Tis study aimed to identify the prevalence, treatments, and associated factors among pregnant women with an intestinal parasitic infection. Tis study confrms a high prevalence of intestinal parasitic infection among pregnant women in Sao Tome and Principe, as the overall prevalence was 58.2% (95% CI 52.9 to 63.3). Our prevalence is higher than that in nearby countries, namely, Ghana with 23.0% [33,34] and Nigeria with 20.8% [35], but lower than that in other countries, such as Ethiopia, where it reaches 70.6% [36][37][38]. Soil-transmitted helminths (STHs) or geohelminths were the main IPI group among pregnant women enrolled in our study, in contrast to Ghana, which reports higher rates of intestinal protozoans [34].
Although higher in our study (90.9%), other studies from endemic countries also found a preponderance of Ascaris lumbricoides among pregnant women, such as Venezuela with 57% [13], Ethiopia with 32.7% [36], and Kenya with 6.5% [39]. In STP, previous data concerning IPI among children reported up to an 86.7% rate of infection, mainly with Ascaris lumbricoides (56.3%) and Trichuris trichiura (52.5%), reinforcing a previously existing high burden of helminthiasis in the country [38]. Terefore, these high rates of ascariasis and trichuriasis in STP illustrate that transmission occurs due to soil and domestic water supply with fecal pollution around homes with poor sanitation and improper sewage disposal [39].
In contrast, a very low rate of hookworm (1.4%) and strongyloidiasis (0.5%) in comparison to other studies [37,38] was found, probably due to infrequently walking barefoot among the adult population in the country preventing larvae penetration in the feet skin [1,37].
Regarding schistosomiasis, we found a 3% prevalence of S intercalatum. Overall estimates report that 10 million women in Africa per year have schistosomiasis during pregnancy although we could not fnd any study regarding S intercalatum infection in pregnant women in literature [40,41]. Tis paucity of data can be related to the fact that S intercalatum is restricted to a few central African countries, is transmitted by Bulinus forskalii, and has a mild pathogenicity linked to rectal schistosomiasis and minor liver pathology [32,42]. We observed that most pregnant women from our study with an S intercalatum infection had an urban residence. Tis "urbanization" of S intercalatum was previously reported in another study from central Africa [43]. Tis "urban" transmission of schistosoma occurs while walking through fooded streets (temporary snail breeding sites) instead of the traditional transmission in rural areas when people become infected through contact with parasiteharboring snails in natural water sources during activities such as fshing, farming, or swimming [44].
Concerning Entamoeba histolytica, a lower rate of 1.9% was found in this study compared to other settings, such as Venezuela, with a 12% rate [13]. Other intestinal protozoa, Table 3: Polyparasitic infections (coinfection of two or more parasite species in the same pregnant women).

Polyparasitism
Frequency (n�) Percentage * (%) Ascaris lumbricoides + Trichuris trichiura 18 Schistosoma intercalatum + Entamoeba histolytica 3 Ascaris lumbricoides + Ancylostoma duodenale 2 Ascaris lumbricoides + Schistosoma intercalatum 2 TOTAL 25 11.9 * From the 210 pregnant women with an IPI, a total of 25 (11.9%) had two diferent parasites identifed in their copro-parasitologial exam. Te helminths group total 224 including polyparasitic infections ( Table 2), but only 202 pregnant women had helminthic monoparasitic infection. IPI: intestinal parasitic infection. * For the anthelmintic prescription analysis, three diferent groups were considered since the recommended drugs are diferent for helminthiasis (albendazole, mebendazole, or piperazine), schistosomiasis (praziquantel) [20,32], and amebiasis (paromomycin, nitroimidazoles as metronidazole or tinidazole) [7,8]. Te sum refers to 220 infections: 202 monoparasitic helminthiasis plus 11 schistosomiasis (S intercalatum), and 7 amebiasis (E histolytica). Schistosomiasis and amebiasis coinfection cases were included. Giardiasis cases (2) were not described due to missing information on anthelmintic prescription. 6 Journal of Tropical Medicine such as Giardia duodenalis, were also very uncommon (0.9%) in our study, perhaps due to methodological limitations in their identifcation since a previous study in the country, using molecular methods (PCR), found a 7.5% prevalence of Giardia duodenalis among children attending HAM [45]. Looking into the associated factors of intestinal parasitic infection, we identifed that education, parity, and employment were signifcantly associated with IPI among pregnant women in this study. Primary education increased the odds of IPI in pregnant women, in accordance with other studies from Ethiopia [37] and Kenya [10], since better education is related to enhanced health-seeking behavior. Te odds of IPI were almost twofold higher in unemployed pregnant women than in those employed. Tis relationship between unemployment and IPIs was already reported in other studies, reinforcing that a low economic standard, typically associated with unemployment, promotes IPIs in endemic settings [46,47].
Te odds of IPI were approximately four times higher among pregnant women with a parity of fve or above, consistent with other studies that reported that age and parity were possible risk factors for parasitosis [1]. For instance, ascariasis in pregnancy was found to be most common in women between 20 and 29 years of age, and the prevalence increased with parity [1], similar to our study. In contrast, other authors state that multiparous women had reduced odds of IPIs compared to nulliparous women with the rationale that they might have experienced more ANC education on how to avoid IPIs in their previous pregnancies [33]. Our fndings suggest a child-driven intrafamily transmission of parasites in STP since grand multiparous women have more children, which is in accordance with previous studies that described a high burden of IPI among children in STP [48].
Open defecation and not having access to improved water were associated with an IPI higher risk although no statistically signifcant diference was found. While the fndings are in line with those reported from studies in Ghana [34], Colombia [49], and Mexico [50], they are contrary to those from Ethiopia [37], where the unavailability of toilet facilities was found to be signifcantly associated with IPI in pregnancy. Beyond this lack of association, we identifed that more than 40% of all participants in this study reported having a daily practice of open defecation. Tus, Sao Tome and Principe will surely beneft if it takes the frst step on the "sanitation ladder" proposed by the World Health Organization Program for Water Supply and Sanitation toward better health for all in the country [25]. Our analysis regarding anthelmintic prescription to infected pregnant women highlights that most were not treated at all, adverting to important "treatment missing opportunities." Tis can be due to issues related to anthelmintic safety, namely, fear of teratogenic efects by health professionals and pregnant women, a signifcant obstacle also reported in other endemic countries [51].
Additionally, this study also reveals the high proportion of inadequate anthelmintic prescriptions. Regarding STH infection, most pregnant women received the recommended drugs (albendazole or mebendazole) although some still received metronidazole, which is specifc to protozoan infection [1]. None of the pregnant women with S intercalatum infection took the recommended praziquantel (40 mg/kg), and two women were even inadequately treated with piperazine [20,32]. Similarly, pregnant women infected with E histolytica were not treated, even though amebiasis treatment should be warranted even in asymptomatic carriers, not only because of the potential of developing the invasive disease but also to diminish the spread of disease [7,52,53].
Te "treatment missing opportunities" identifed by this study should be urgently addressed, frst by training health professionals on proper prescription, and second, in case of pregnant women refusal of treatment during the pregnancy period, they should be referred and followed up for adequate treatment in the postpartum period.
As this is the frst study in Sao Tome and Principe to provide comprehensive data concerning the burden of intestinal parasites among pregnant women, recommendations from the study will assist government health ofcials in policy development. Public health education and awareness campaigns combined with health professionals' education programs would also enhance women's knowledge of IPI prevention and ensure adequate anthelmintic therapeutic practices by health professionals.

Limitations.
Although the study was conducted at a referral center (HAM) which serves most pregnant women in Sao Tome and Principe, its fndings cannot be generalizable to other areas. Rural antenatal care services in STP may have a higher prevalence of IPIs among pregnant women since they are more exposed to potential sources of infection, such as contaminated water, farm animals, and wildlife [45]. In addition, they may also have distinct behavior and poorer hygiene practices and sanitation, increasing their risk of infection in comparison with urban pregnant women [45].

Conclusions
Intestinal parasitic infections are a high burden for pregnant women in Sao Tome and Principe, mainly for those with primary education, unemployed, and grand multiparous. Missing opportunities for IPI treatment, mainly for S intercalatum and E histolytica, should be addressed with health professionals' training and through the follow-up of women who refuse anthelmintic drugs during pregnancy for later postpartum treatment.

Data Availability
Te datasets used and/or analyzed during the current study are all included within the manuscript itself.

Ethical Approval
Te study complies with the Declaration of Helsinki and was approved and consented to by the Ministry of Health of Sao Tome and Principe and by the main board of Hospital Dr. Ayres de Menezes since at the time the study protocol was submitted there was no ethics committee in STP. Only recently has the country's National Ethics Committee been appointed. Previously, study analysis and approval were carried out by the Ministry of Health and the institution where the study was to be performed, which is what the authors have carried out. All methods were performed in accordance with the relevant guidelines and regulations in practice.

Consent
Written informed consent was obtained from all participants (or their parent or legal guardian in the case of an adolescent under 16 or illiterate) after the purpose of the research was explained orally by the investigator. Participation in the survey was voluntary, as participants could decline to participate at any time during the study.