Skip to main content
PLOS ONE logoLink to PLOS ONE
. 2020 Oct 8;15(10):e0239578. doi: 10.1371/journal.pone.0239578

Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya

Gibson Waweru Nyamu 1,2,*, Jimmy Hussein Kihara 3, Elvis Omondi Oyugi 4, Victor Omballa 5, Hajara El-Busaidy 2, Victor Tunje Jeza 1
Editor: Adrian JF Luty6
PMCID: PMC7544053  PMID: 33031456

Abstract

Background

Prevalence of Prevalence of malaria in pregnancy (MiP) in Kenya ranges from 9% to 18%. We estimated the prevalence and factors associated with MiP and anemia in pregnancy (AiP) among asymptomatic women attending antenatal care (ANC) visits.

Methods

We performed a cross-sectional study among pregnant women attending ANC at Msambweni Hospital, between September 2018 and February 2019. Data was collected and analyzed in Epi Info 7. Descriptive statistics were calculated and we compared MiP and AiP in asymptomatic cases to those without either condition. Adjusted prevalence Odds odds ratios (aPOR) and 95% confidence intervals (CI) were calculated to identify factors associated with asymptomatic MiP and AiP.

Results

We interviewed 308 study participants; their mean age was 26.6 years (± 5.8 years), mean gestational age was 21.8 weeks (± 6.0 weeks), 173 (56.2%) were in the second trimester of pregnancy, 12.9% (40/308) had MiP and 62.7% had AiP. Women who were aged ≤ 20 years had three times likelihood of developing MiP (aPOR = 3.1 Cl: 1.3–7.35) compared to those aged >20 years old. The likelihood of AiP was higher among women with gestational age ≥ 16 weeks (aPOR = 3.9, CI: 1.96–7.75), those with parasitemia (aPOR = 3.3, 95% CI: 1.31–8.18), those in third trimester of pregnancy (aPOR = 2.6, 95% CI:1.40–4.96) and those who reported eating soil as a craving during pregnancy (aPOR = 1.9, 95%CI:1.15–3.29).

Conclusions

Majority of the women had asymptomatic MiP and AiP. MiP was observed in one tenth of all study participants. Asymptomatic MiP was associated with younger age while AiP was associated with gestational age parasitemia, and soil consumption as a craving during pregnancy.

Introduction

The commonest plasmodium species that is known to cause malaria in pregnancy (MiP) in Africa is Plasmodium falciparum which can lead to anemia in pregnancy (AiP) [1]. The World Health Organization (WHO) in 2019, reported 11 million pregnant women were infected with plasmodium infection in sub-Saharan Africa, resulting in 872 000 low birth weights [2]. In 2019, the Ministry of Health (MOH), Kenya, estimated MiP to be 6.3% among women attending their first antenatal care (ANC) visit [3]. In Kwale County, Kenya, MiP remains a public health concern with a total of 2316 in 2019 [3]. Anemia in Pregnancy is a well-known risk factor for maternal death, stillbirths, low birth weights and infant prematurity [46]. Previous studies have reported associations between malaria with AiP [7,8] while consumption of soil (geophagy) has been associated with AiP among African women [9]. Besides plasmodium infections, other known causes of AiP include nutritional deficiencies, infectious diseases like HIV, parasitic infections like hookworm infestation, and hemoglobinopathies [4,9].

MiP constitutes a major risk to the mother, fetus, and neonates including stillbirths, spontaneous abortion, premature delivery, maternal anemia, and low birth weight [2,10]. Due to physiological and immunological changes, pregnant women are more at risk of malaria compared to non-pregnant women living in areas of similar malaria endemicity [11]. Factors associated with MiP are low parity, young age, low maternal education level, early gestational age, young maternal age, fewer previous pregnancies, non-ownership or infrequent use of bed nets and maternal unemployment [7,10,1214].

ANC is a package given to pregnant women which entails giving prophylaxis and treatment for anemia and malaria among other services, where the AiP is prevented by providing nutritional counseling including iron supplements, and treating cases of AiP [15]. Interventions aimed at prevention and control of MiP adopted by the MOH–Kenya are Intermittent Preventive Treatment (IPTp) of MiP, with sulfadoxine pyrimethamine (SP) given after 12 weeks gestational period done four weeks apart until the pregnant woman delivers. In areas with high malaria transmission such as Western, Nyanza, and Coast regions, Long-Lasting Insecticidal Nets (LLINs) are provided at the ANC during the first contact [16].

Kenya Malaria Indicator Survey (KMIS) 2015 reported two folds increase in the prevalence of malaria in the coastal region compared to KMIS 2010 [17]. In high-transmission regions like coastal regions in Kenya, where levels of acquired immunity tend to be high P. falciparum infection is usually asymptomatic in pregnancy. However, parasites may be present in the placenta and contribute to AiP even in the absence of documented peripheral parasitaemia [18]. In the study area, women have been reported delaying in starting pre-natal care during pregnancy which may also contribute to delay interventions measures uptakes like IPTp and LLINs usage hence remain a reservoir of parasites contributing to the spread of the disease from one malaria season to the next [19]. Little information is currently available on the epidemiology of malaria and anemia during pregnancy, in Kwale County except the data collected in passive surveillance according to records in the County.

It is from this background that we carried out this study to estimate the prevalence of asymptomatic MiP and AiP and identify the associated factors among women attending their first ANC visit at the largest referral health facility in Kwale County, Kenya.

Methods

Study location

Kwale County is one of six counties in the coastal region of Kenya covering an area of 8,270.3 km2 with a population of 866,820 people [20]. The inhabitants are predominantly Muslim, from the Mijikenda tribe, and practice subsistence farming and smallholder animal husbandry. The weather is hot and humid with two rainy seasons: long rains from April to June and short rains from October through November. The incidence of malaria increases during the long rainy seasons [21].

Msambweni County Referral Hospital (MCRH) is the main referral health facility in Kwale County (Fig 1). The hospital has 155 inpatient beds and189 healthcare workers. Four nurses work at the Mother-Child Health department, who attend on average 125 mothers each month [3].

Fig 1. Map of the study area (Msambweni County Referral Hospital-MCRH).

Fig 1

Study design and population

We conducted a cross-sectional study to determine the prevalence of asymptomatic MiP and AiP.

Pregnant women attending their first ANC visit at MCRH between September 2018 and February 2019. Pregnant women seeking their first ANC were included in the study, especially those with no symptoms of malaria as per clinical assessment (i.e. no fever (temperature >37.5°C), chills, rigor, nausea, vomiting, headache, anorexia, or joint/muscle pains).

We excluded pregnant women who had taken anti-malarial drugs within the past two weeks, antipyretics in case they had fever and those receiving micronutrient.

Definitions of terms

  • Asymptomatic malaria was defined as the presence in the peripheral blood of asexual blood-stage of Plasmodium species, but has no symptoms of malaria per clinical assessment and the pregnant woman reported not taken antipyretics within 48 hours and antimalarials within 14 days.

  • A young age was defined as age ≤ 20 years.

  • Anemia was defined as a hemoglobin < 11 g/dl, mild anemia (10–10.9g/dl), moderate anemia (7–9.9g/dl) and severe anemia (<7g/dl).

  • First trimester was defined as from week 1 to the end of week 12 while the second trimester as from week 13 to the end of week 26 and the third trimester as from week 27 to the end of the pregnancy, as it was classified in a study in Ghana [22].

Sample size determination

Cochran’s formula [23] was used to calculate the sample size required to estimate the prevalence of asymptomatic malaria in pregnant women attending their first ANC visit.The study assumed a 95% confidence interval, 80% power, the prevalence of asymptomatic MiP to be 24% [7] and we adjusted by 10% to cater for those who refused to be enrolled. We calculated the desired sample size as 308 participants.

Sampling procedures

Systematic random sampling method was used to select study participants. Our sampling interval was based on the daily entries in the mother-child health (MCH) register from September 2018 to February 2019. The sampling started by selecting a participant from the daily entries list at random using a table of random numbers and then every kth participant in the frame was selected. A selection interval (k) was determined by dividing the total daily entry listed in order to get the number of the participants required per day. If a randomly-selected participant was not eligible for an interview or refused to be part of the study, the next eligible participant on the list was selected. We sampled our study participants until we arrived at our desired sample size of 308 (Fig 2).

Fig 2. Flow chart of the recruited participants included in this analysis.

Fig 2

Following signing of informed consent, the participants found with malaria parasitemia were treated with Artemether-lumefantrine for those who were in their 2nd and 3rd trimesters while those who were in the 1st trimester were treated with quinine. Those had anemia were given iron supplements and health education on risks and management of anemia during pregnancy at no extra cost.

Data collection

In-person interviews were conducted using a pre-tested structured questionnaire. The questionnaire was developed in English, and translated to Swahili for non-English speaking respondents.

Variables collected were;

  • Socio-demographic characteristics were mother's age, education level, marital status and occupation.

  • Obstetrics variables were gravidity, parity, trimesters, and gestational age in weeks.

  • Clinical history variables were history of fever in the last 48 hours and taken anti pyretic drugs, whether the client has taken antimalarial drugs within the last 2 weeks, the tendency of geophagy, and whether or not on iron supplements.

Laboratory methods

Hemoglobin testing

The index finger was cleaned using 70% isopropyl alcohol and pricked using a sterile lancet by well-trained laboratory technicians. The first drop was wiped away using sterile cotton wool, and then the finger was gently squeezed to obtain approximately 30μl drop of blood onto a micro-cuvette and subsequently into a portable heme-analyzer (Hemo Cue Hb 301, Hemo Cue AB 16, Sweden). To determine anemia status, Hb measurement was obtained within 45 seconds and reported in grams per deciliter (gm/dl).

Malaria testing

Thick and thin blood films were prepared. Absolute methanol was used to fix thin films and Giemsa stain (3%) staining for 30 minutes. The slides were then rinsed with distilled water and air-dried at room temperature. Slides were then viewed under the microscope using 100x objectives on immersion oil. No Parasite Found was reported after 100 fields were examined and no malaria parasites observed.

Thick films were examined to determine the presence of asexual malaria parasites, quantification of malaria parasites was done by enumerating asexual malaria parasites against 200 white blood cells (WBC). Then, Parasite densities per microliter of blood were determined after multiplying with an assumed WBC count of 8.0*109/l, with the product of numbers of malaria parasites divided by 200 WBC [24]. Also, the speciation of the Plasmodium parasites was done.

Quality control

Quality of Giemsa stain was maintained by testing known positives slides. An independent qualified parasitologist examined 10% of both positive and negative slides which were randomly selected and in the case of any disparity they were read by a third parasitologist and his results were deemed final.

Quality control of our hematology analyzer (HemocueHb 301, Hemo Cue AB 16, Sweden) was performed per as the manufacturer instructions, by analyzing dried samples with known Hb levels before testing participant samples.

Data management and analysis

Data were entered, cleaned, and rechecked using MS Excel 2013. Data were analyzed using Epi Info 7. The following descriptive statistics were calculated: frequencies and proportions for categorical variables, and measures of central tendency (mean, median, and mode) and dispersion (range, interquartile range, and standard deviation) for continuous variables.

We tested the relationship between a variety of predictor variables, including socio-demographic factors and clinical history, and malaria status as the outcome variable, comparing participants who tested malaria positive with those who tested negative. We also compared participants who had anemia to those who did not. Both crude prevalence ratio (cPOR) and adjusted prevalence odds ratio (aPOR), and their 95% confidence intervals were calculated. Variables with p < 0.05 were considered statistically significant. Variables with p-values ≤ 0.20 were included in a logistic regression model using a backward stepwise elimination method to identify independently associated factors.

Ethics approval and consent to participate

We sought written, informed consent from each participant before interviewing and finger pricking for malaria blood slides and hemoglobin level analysis. The authors used oral consent to accommodate the low literacy rates in the populations served by MCRH and append thumb print where the study participants cannot sign or write on consent form. Permission was granted to conduct the study by Kwale department of health and MCRH director. Ethical clearance was obtained from the Pwani University–Ethical Review Committee (ERC/MSc/021/2018).

Results

Socio-demographic characteristics of respondents

A total of 308 respondents were interviewed. Their mean age was 26.6 years (± 5.8 years), 267 (86.9%) were married, 83 (26.9%) had > 8 years of formal schooling, and among them, 29 (9.4%) were formally employed. The mean gestational age was 21.8 weeks (± 6.0 weeks), 29 (9.4%) were in first trimester, 173 (56.2%) were in second trimester and 106 (34.4%) were in third trimester of pregnancy. Those who were primigravidae were 66 (21.4%), secondgravidae were 79 (25.7%) and multigravidae were 163 (52.9%)(Table 1).

Table 1. Socio demographic characteristics of women attending first antenatal care at Msambweni hospital, Kwale County, Kenya.

Characteristics n (%)
Maternal age (n = 308)
≤20 50 (16.2)
>20 258 (83.8)
Education (n = 301)
Had ≤8 years of formal schooling 218 (72.4)
Had > 8 years Eof formal schooling 83 (27.6)
Residence (n = 308)
Rural 275 (89.3)
Urban 33 (10.7)
Marital status (n = 307)
Married 267 (86.9)
Single 32 (10.4)
Divorced 6 (1.9)
Widowed 2 (0.7)
Trimester (n = 308)
First 29 (9.4)
Second 173 (56.2)
Third 106 (34.4)
Gravidity (n = 303)
Primigravidae 66 (21.8)
Secundigravidae 76 (25.1)
Multigravidae 161 (53.1)
Gestational age in weeks (n = 308)
<16 49 (15.9)
≥16 259 (84.1)
Net ownership (n = 308)
Yes 248 (80.5)
No 60 (19.5)
Slept under bed net previous night (n = 248)
Yes 231 (93.1)
No 17 (6.9)
Frequency of sleeping under bed net (n = 248)
Always 205 (82.7)
Sometimes 43 (17.3)
Age of a bed net (in months) (n = 246)
< 6 months 109 (44.3)
  6–12 months 34 (13.8)
> 12 months3e 103 (41.9)

Among the participants, 248 (80.5%) owned bed nets (treated or untreated). Of these, 109 (44.3%) had used a bed net for less than 6 months, 34 (13.8%) had used a bed net for 6–12 months, and 103 (41.8%) had used a bed net for > 12 months. In terms of bed net usage in the current pregnancy, 231 (93.2%) reported having slept under a bed net the previous night while 205 (82.7%) reported always sleeping under a bed net, and 43 (17.3%) reported sometimes sleeping under a bed net (Table 1).

Prevalence and factors associated with asymptomatic malaria

Malaria positivity among the 308 study participants was 12.9% (40/308) and the geometric mean parasite count was 3738 parasites per microliter of blood; 35 (87.5%) tested positive for Plasmodium falciparum, 3 (7.5%) Plasmodium malariae and 2 (5.0%) Plasmodium ovale. In regard to gestational trimesters with plasmodium infections, those in the first trimester were 2/29 (6.9%), second trimester 24/173 (13.9%) and those in the third trimester were 14/106 (13.2%).

The odds of asymptomatic MiP was higher in women who were aged ≤ 20 years (cPOR = 3.5, 95% CI = 1.65–7.23), women who did not own bed nets (cPOR = 2.3, 95% CI 1.08–4.69) and women who owned bed nets but did not sleep under a bed net the night before the interview (cPOR = 2.4, 95% CI 1.14–5.03). After logistic regression analysis, asymptomatic MiP was independently associated with being age ≤ 20 years (aPOR = 4.5 (1.71–12.01) compared with those aged >20 years (Table 2).

Table 2. Factors associated asymptomatic malaria parasitaemia in pregnant women, Kwale County, Kenya.

Potential factors N With Malaria Crude POR Adjusted POR
n (%) (95% CI) (95% CI)
Maternal age (n = 308)
≤ 20 50 14 (28) 3.5 (1.66–7.26) 4.5 (1.71–12.01)
>20
Education (n = 301)
258 26 (10.1) 1 1
Had > 8 years of formal schooling 83 12 (14.5) 1 **
Had ≤ 8 years of formal schooling 218 27 (12.4) 0.8 (0.40–1.74) **
Residence (308)
Urban 33 4 (12.1) 1
Rural 275 36 (13.1) 1.1 (0.36–3.29) **
Trimester (n = 308)
Third 106 14 (13.2) 1 **
First/second 202 26 (12.9) 0.97 (0.48–1.95) **
Gravidity (n = 303)
Primigravidae/Secundigravidae 145 23 (15.7) 1.6 (0.83–3.17) **
Multigravidae 163 17 (10.4) 1 **
Gestational age in weeks (n = 308)
<16 weeks 49 7 (14.3) 1.14 (0.47–2.75) **
≥16 weeks 259 33 (12.7) 1 **
Net ownership (n = 308)
No 60 13 (21.7) 2.3 (1.09–4.71) **
Yes 248 27 (10.9) 1 **
Slept under bed net previous night (n = 248)
No 56 13(23.2) 2.3(1.14–5.01) **
Yes 192 22 (11.5) 1 **
Frequency of sleeping under bed net (n = 248)
Sometimes 43 4 (9.3) 0.8 (0.28–2.62) **
Always 205 22 (10.8) 1 **
Age of a bed net (in months) (n = 246)
≤12 months 143 12 (8.4) 1 **
>12months 103 15 (14.7) 1.9 (0.83–4.16) **

**CI, confidence interval, N, numbers, POR, prevalence odds ratio, aPOR = adjusted prevalence odds ratio.

Prevalence and factors associated with anemia

Anemia was reported in 193 (62.7%) participants, and the mean Hb was 9.6 mg/dl (± 1.3mg/dl); 96 (49.7%) had moderate anemia, 90 (46.6%) had mild anemia and seven (3.6%) had severe anemia. Among those with severe anemia four had malaria, moderate anemia 15/95 (15.8%) and mild anemia were 14/90 (15.6%). Geophagy was reported by 117 (38.6%) participants. Those with a gestational age of ≥ 16 weeks had greater odds of AiP, cPOR = 3.2 (1.72–6.07) compared to those with gestational age <16 weeks. Those who reported eating soil had greater odds of AiP, cPOR = 2.1 (1.27–3.45) compared with those who did not report eating soil. Following logistic regression analysis, AiP was independently associated with gestational age ≥ 16 weeks (aPOR; 3.3, 95% CI: 1.72–6.41), and those who reported eating soil (aPOR 2.0, 95% CI: 1.21–3.41) Table 3.

Table 3. Factors associated with anemia in pregnant women, Kwale County, Kenya.

Potential factors N With anemia Crude POR Adjusted POR
(%) (95% CI) (95% CI)
Maternal age (n = 308)
≤ 20 50 34 (68.0) 1.3 (0.69–2.52) **
>20 258 159 (61.6) 1 **
Education level n = (301)
Had ≤ 8 years of formal schooling 218 142 (65.1) 1.4 (0.85–2.39) **
Had > 8 years of formal schooling 83 47 (56.6) 1 **
Residence (n = 308)
Urban 33 18 (54.5) 1 **
Rural 275 175 (63.6) 1.5 (0.69–3.03) **
Trimester (n = 308)
First 29 11 (37.9) 1
Second 173 102 (58.9) 2.3 (1.04–5.42) **
Third 106 80 (75.5) 5.0 (2.08–12.23) **
Gravidity (n = 308)
Primigravidae 66 42 (63.4) 1.1 (0.58–1.91) **
Secundigravidae 79 49 (62.0) 0.98 (0.56–1.71) **
Multigravidae 163 102 (62.6) 1
Gestational age in weeks (n = 308)
< 16 weeks 49 19 (38.8) 1 1
≥ 16 weeks 259 174 (67.2) 3.2 (1.72–6.14) 3.3 (1.72–6.41)
Net ownership n = 308
No 60 42 (70.0) 1.5 (0.82–2.80) **
Yes 248 151 (60.9) 1 **
Eating soil n = 303
Yes 117 85 (72.7) 2.1 (1.27–3.45) 2.0 (1.21–3.41)
No 186 104 (55.9) 1 1

**CI, confidence interval, POR, prevalence odds ratio, aPOR = adjusted prevalence odds ratio.

Pregnant women with malaria parasitemia were three times more likely to have anemia compared to those without malaria parasitemia (χ2 = 2.79, P-value = 0.005, (aPOR; 3.5, 95% CI: 1.21–8.60) (Table 4).

Table 4. Chi-square analysis of proportions with and without malaria parasitemia and anemia.

Anemia
Malaria parasitemia Yes No Total Crude POR χ2 p value Adjusted POR
Yes 33 (17.1%) 7 (6.1%) 40 3.2 (1.36–7.46) 6.8 0.009 3.5 (1.46–8.60)
No 160(82.9%) 108 (93.9%) 268 ref ref
Total 193 115 308

POR, prevalence odds Ratio, Ref, reference.

Discussion

Numerous studies have shown that anemia and malaria contribute to morbidity and mortality among pregnant women. In this study, we found that one in eight women had asymptomatic MiP and more than half of the women had AiP. Asymptomatic MiP was associated with young age (≤ 20 years). Anemia prevalence was also associated with pregnant women who reported eating soil, were in their first and third trimesters of pregnancy, and had P. falciparum infections.

The prevalence of asymptomatic MiP in our study population was 12.9%, which is similar to the prevalence among pregnant women in Ethiopia (9.1%) [25] but lower than the prevalence found in Burkina Faso (24%) among pregnant women [7]. In this study, we included only pregnant women seeking ANC services for the first time in their current pregnancy and the majority of study participants (82.7%) reported always using bednet, whereas the study in Burkina Faso included pregnant women seeking ANC services at any point in their pregnancies. These may be factors that lead in the difference between this two reported prevalence. In this study, we found that the odds of MiP among young women (≤ 20 years of age) were greater than the odds of MiP among older women. This may reflect continuing development of malarial immunity [13,26,27]. Contrast to our findings, a study in Gabon showed that there is no difference found between younger and older pregnant women [28].

The current study reports higher proportion of P. falciparum infections in pregnant women who were in second and third trimesters and less proportion in the first trimester. Studies in Nigeria have also reported high malaria prevalence in pregnant women who were in their second trimester [29,30]. Another study in Mali reported pregnant women in their first trimester were two times more likely to get malaria compared to the third trimester [31]. The probable explanation may be our study had a few numbers in the first trimester compared with 2nd and 3rd trimesters hence the higher prevalence. With an increase in the number of pregnant women in their first trimester, there is the possibility that there could be changes from the present results.

In this study, we reported the highest proportion of Plasmodium infections among the primigravidae (19.7%) followed by secundigravidae (12.7%) and multigravidae (10.4%) with parasitemia declining with increasing gravidity.

These results are consistent with previous studies which found Plasmodium infections are more common in primigravidae compared to multigravidae [7,30,32]. The reason for the present result of gravidae-associated predisposition to P. falciparum infections may be due to the fact that adults who live in malaria-endemic regions generally have some acquired immunity to malaria infection. This acquired immunity diminishes significantly in pregnancy particularly in primigravidae. It has also been suggested by various authors that the early onset of antibody response in multigravidae and the delayed antibody production in primigravidae may be responsible for the gravidity-dependent and differential prevalence of falciparum malaria among pregnant women [18,33].

Intervention measures for first visit pregnant women at antenatal clinic for malaria in pregnancy are IPTp and provision of LLINs among others [16]. Although the role of IPTp is known to reduce maternal malaria episodes and improve pregnancy outcomes [34], the current study did not include pregnant women who had taken IPTp hence we could not ascertain the role played by IPTp. Our study evaluated through interviewing study participants on bed net ownership and usage. Majority of our respondents owned, slept under a bed net and almost all participants reported sleeping under a bed net the previous night. The KMIS, 2015, reported similar high rates of bed net ownership and use in pregnant women living in malaria-endemic zones, with 83.7% of pregnant women reporting sleeping under a bed net the night before they were interviewed [17]. Bed net ownership and usage have been reported in several studies to be protective against malaria infections [27,35]. Our observed high rate of bed net ownership may be the result of Kenya’s Malaria Control Program conducted a mass net distribution campaign and all households were issued bed nets in Kwale County, in 2017 [36]. Bed nets ownership and usage have been reported to have protective effects for Plasmodium infections [37]. In this study, pregnant women with no bed nets had 90% higher odds of asymptomatic MiP compared to those who owned bed nets, though was not statistically significant.

The study reported more than half of the respondents (60%) had AiP. The etiology of anemia is variable and potentially multi-factorial, and thus several underlying morbid and co-morbid conditions may contribute to the prevalence of anemia. Similar to our findings, one study conducted in the Pumwani maternity hospital in Nairobi, Kenya reported an AiP prevalence of 57% [38], and another study in southern Ethiopia reported an AiP prevalence of 60% [34]. Lower prevalence of AiP have also been reported in southwest Ethiopia (23.5%) and northwest Ethiopia (16.5%) [39,40].

The present study found that a significant number of women with asymptomatic MiP had AiP. Malaria in pregnancy is known to cause AiP, this association has also been reported in other studies in sub-Saharan Africa [7,8].

We also found high reported rates of eating soil (geophagy), consistent with other studies that have found high rates of geophagy among pregnant African women as well as associations between geophagy and anemia [9,41].

Our results indicate that, anemia is more common among women in their third trimester than women in their first trimester, similar to findings reported in other studies [42,43]. Hemoglobin decreases through to the end of the third trimester. Anemia is a function of plasma volume and red cell mass; both of which increase during pregnancy; but the increase in plasma volume is proportionately greater than the increase in red cell mass [44].

Our study had several limitations

We collected data from September 2018 to February 2019, a period during which there is low malaria transmission. This could have resulted in the under estimation of the overall prevalence of asymptomatic MiP in our study area. A continuous monitoring throughout the year of MiP incidence will account for seasonality burden [21].

In addition, the study was hospital-based, excluding pregnant women who did not seek ANC services. While this may limit the generalizability of findings to the community, few women fail to seek antenatal care in our study area. Determination of factors associated with asymptomatic MiP and AiP in hospital-based studies provides a proxy indicator of predictors in the community of that particular facility when community-based surveys are not feasible.

Lastly, this study did not explore other factors that may contribute to anemia, including nutritional factors, soil-transmitted helminthes infection, and hereditary conditions such as sickle cell disease thus limiting our ability to assess the contribution of other causes of anemia during pregnancy. However, diagnosis of anemia was based on laboratory analysis and did not depend on clinical assessment as reported by other researchers.

Conclusion

Asymptomatic Plasmodium infections and anemia are common in women attending their first ANC visit at Msambweni County Referral Hospital in Kwale County. Most of the Plasmodium infections in this area are caused by P. falciparum. We did not observe a clear gravidity pattern of asymptomatic MiP, however was associated with younger maternal age (≤20 years). Anemia in pregnancy was associated with Plasmodium infections, women who reported to have geophagy tendency and those who were their third trimester. Majority of the study participants in this study registered for antenatal care in their second and third trimester. This practice is detrimental as it does not allow for early detection and correction of pregnancy related complications such as anemia. Therefore, we recommend to the Msambweni County referral hospital in conjunction with Kwale department of health should organize for regular outreach to the community targeting pregnant women for health education should be provided to positively influence the knowledge and attitudes of pregnant women of child-bearing age to malaria and anemia, including early antenatal registration.

Acknowledgments

We would like to thank the Technical University of Mombasa, Kenya, for their support during the study. Special thanks go to study participants and to the Kwale County Department of Health authorities for their collaboration. We also thank the many Vector Borne Disease Control Unit (VBDCU) Laboratory officers, including Said Lipi, Joyce Bandika, Peter Siema, Charles Ng’ang’a, and Elton Mzungu for their dedication and meticulous microscopy. We extend a special thank you to nurse Wendy Losier, for her assistance with the consenting process and data collection. We also thank Dr. Robert Perry, Dr. Elizabeth Wanja, and Dr. Shama Cash-Goldwasser for their valuable insight and comments on the manuscript.

Data Availability

All relevant data are within the paper.

Funding Statement

The author(s) received no specific funding for this work.

References

  • 1.Alaku I., Abdullahi A., and Kana H., Epidemiology of Malaria Parasites Infection among pregnant women in some part of Nasarawa State, Nigeria. Developing Country Studies, 2015. 5(2). [Google Scholar]
  • 2.WHO. WORLD MALARIA REPORT 2019 14 th June 2020 [cited 2020 14th June]; Available from: https://www.who.int/malaria/publications/world-malaria-report-2019/en/.
  • 3.DHIS2, District Health Information Systems. 2019.
  • 4.McClure E.M., et al. , The association of parasitic infections in pregnancy and maternal and fetal anemia: a cohort study in coastal Kenya. PLoS neglected tropical diseases, 2014. 8(2). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Helmy M.E., Elkhouly N.I., and Ghalab R.A., Maternal anemia with pregnancy and its adverse effects. Menoufia Medical Journal, 2018. 31(1): p. 7. [Google Scholar]
  • 6.Uneke C., Duhlinska D., and Igbinedion E., Prevalence and public-health significance of HIV infection and anaemia among pregnant women attending antenatal clinics in south-eastern Nigeria. Journal of health, population, and nutrition, 2007. 25(3): p. 328 [PMC free article] [PubMed] [Google Scholar]
  • 7.Douamba Z., et al. , Asymptomatic malaria correlates with anaemia in pregnant women at Ouagadougou, Burkina Faso. BioMed Research International, 2012. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Isah A.Y., Amanabo M.A., and Ekele B.A., Prevalence of malaria parasitemia amongst asymptomatic pregnant women attending a Nigerian teaching hospital. Annals of African Medicine, 2011. 10(2). [DOI] [PubMed] [Google Scholar]
  • 9.Stephen G., et al. , Anaemia in pregnancy: prevalence, risk factors, and adverse perinatal outcomes in Northern Tanzania. Anemia, 2018. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Gajida A., Iliyasu Z., and Zoakah A., Malaria among antenatal clients attending primary health care facilities in Kano state, Nigeria. Annals of African medicine, 2010. 9(3). [DOI] [PubMed] [Google Scholar]
  • 11.Tarning J., et al. , Population pharmacokinetics of dihydroartemisinin and piperaquine in pregnant and nonpregnant women with uncomplicated malaria. Antimicrobial Agents and Chemotherapy, 2012. 56(4): p. 1997–2007. 10.1128/AAC.05756-11 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Olubukola A., Odunayo A., and Adesina A., Anemia in pregnancy at two levels of health care in Ibadan, south west Nigeria. Annals of African Medicine, 2011. 10(4). [DOI] [PubMed] [Google Scholar]
  • 13.Agomo C.O. and Oyibo W.A., Factors associated with risk of malaria infection among pregnant women in Lagos, Nigeria. Infectious Diseases of Poverty, 2013. 2(1): p. 19 10.1186/2049-9957-2-19 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Nega D., et al. , Anemia associated with asymptomatic malaria among pregnant women in the rural surroundings of Arba Minch Town, South Ethiopia. BMC research notes, 2015. 8(1): p. 110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 15.MOH. Focused Antenatal Care,. 2015 11th [cited 2020 June]; Available from: https://www.researchgate.net/publication/241510221.
  • 16.MOH. Malaria in Pregnancy. 2020 [cited 2020 14th, June]; Available from: http://www.nmcp.or.ke/index.php/malaria-in-pregnancy.
  • 17.MOH. Kenya Malaria Indicator Survey. 2015 14 th June 2020; Available from: https://dhsprogram.com/pubs/pdf/MIS22/MIS22.pdf.
  • 18.Desai M., et al. , Epidemiology and burden of malaria in pregnancy. The Lancet infectious diseases, 2007. 7(2): p. 93–104. 10.1016/S1473-3099(07)70021-X [DOI] [PubMed] [Google Scholar]
  • 19.McClure E.M., et al. , The association of parasitic infections in pregnancy and maternal and fetal anemia: a cohort study in coastal Kenya. PLoS Negl Trop Dis, 2014. 8(2): p. e2724 10.1371/journal.pntd.0002724 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.KNBS, Kenya Population and Housing Census Volume I: Population By County and Sub-County.Vol. I. 2019. 38 p. Available from: http://www.knbs.or.ke. 2019.
  • 21.Bisanzio D., et al. , Use of prospective hospital surveillance data to define spatiotemporal heterogeneity of malaria risk in coastal Kenya. Malaria journal, 2015. 14(1): p. 482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Tay S.C., et al. , Malaria and anaemia in pregnant and non-pregnant women of child-bearing age at the University Hospital, Kumasi, Ghana. 2013. [Google Scholar]
  • 23.Cochran W.G., Sampling techniques. 2007: John Wiley & Sons. [Google Scholar]
  • 24.WHO, Basic malaria microscopy. Geneva: World Health Organization; 2010. [Google Scholar]
  • 25.Nega D., et al. , Prevalence and predictors of asymptomatic malaria parasitemia among pregnant women in the rural surroundings of Arbaminch Town, South Ethiopia. PLoS One, 2015. 10(4). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Brabin B.J., An analysis of malaria in pregnancy in Africa. Bulletin of the World Health Organization, 1983. 61(6): p. 1005 [PMC free article] [PubMed] [Google Scholar]
  • 27.Matangila J.R., et al. , Asymptomatic Plasmodium falciparum infection is associated with anaemia in pregnancy and can be more cost-effectively detected by rapid diagnostic test than by microscopy in Kinshasa, Democratic Republic of the Congo. Malaria journal, 2014. 13(1): p. 132. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Bouyou-Akotet M.K., et al. , Prevalence of Plasmodium falciparum infection in pregnant women in Gabon. Malaria journal, 2003. 2(1): p. 18. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Yakubu D., Kamji N., and Dawet A., Prevalence of Malaria Among Pregnant Women Attending Antenatal Care at Faith Alive Foundation, Jos, Plateau State, Nigeria. Noble International Journal of Scientific Research, 2018. 2(4): p. 19–26. [Google Scholar]
  • 30.Ivoke N., et al. , Falciparum Malaria Parasitaemia Among Pregnant Women Attending Antenatal Clinics in a Guinea-Savannah Zone, Southwestern Ebonyi State, Nigeria. International Journal of Scientific and Engineering Research, 2013. 4(9): p. 1876–1883. [Google Scholar]
  • 31.Dicko A., et al. , Risk factors for malaria infection and anemia for pregnant women in the Sahel area of Bandiagara, Mali. Acta Tropica, 2003. 89(1): p. 17–23. 10.1016/j.actatropica.2003.07.001 [DOI] [PubMed] [Google Scholar]
  • 32.Ouma P., et al. , Malaria and anaemia among pregnant women at first antenatal clinic visit in Kisumu, western Kenya. Tropical Medicine & International Health, 2007. 12(12): p. 1515–1523. [DOI] [PubMed] [Google Scholar]
  • 33.Enato E., et al. , Plasmodium falciparum malaria in pregnancy: prevalence of peripheral parasitaemia, anaemia and malaria care-seeking behaviour among pregnant women attending two antenatal clinics in Edo State, Nigeria. Journal of Obstetrics and Gynaecology, 2009. 29(4): p. 301–306. 10.1080/01443610902883320 [DOI] [PubMed] [Google Scholar]
  • 34.WHO. Intermittent preventive treatment of malaria in pregnancy (IPTp). 2017 [cited 2020 14th June]; Available from: http://www.who.int/malaria/areas/preventivetherapies/pregnancy/en/.
  • 35.Agan T., et al. , Prevalence of asymptomatic malaria parasitaemia. Asian pacific journal of tropical medicine, 2010. 3(1): p. 51–54. [Google Scholar]
  • 36.Waweru, M. Mass Distribution Of 15mn Mosquito Nets Kicks Off In 23 Counties. Capital news 2017 [cited 2020 1st, August]; Available from: https://www.capitalfm.co.ke/news/2017/03/mass-distribution-15mn-mosquito-nets-kicks-off-23-counties/.
  • 37.Mwangi M.N., Safety and efficacy of iron supplementation in pregnant Kenyan women. 2014: Wageningen University. [Google Scholar]
  • 38.Okube O.T., et al. , Prevalence and factors associated with anaemia among pregnant women attending antenatal clinic in the second and third trimesters at pumwani maternity hospital, Kenya. Open Journal of Obstetrics and Gynecology, 2016. 6(01): p. 16. [Google Scholar]
  • 39.Melku M., et al. , Prevalence and predictors of maternal anemia during pregnancy in Gondar, Northwest Ethiopia: an institutional based cross-sectional study. Anemia, 2014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40.Zekarias B., et al. , Prevalence of anemia and its associated factors among pregnant women attending antenatal care (ANC) in Mizan Tepi University Teaching Hospital, South West Ethiopia. Health Science Journal, 2017. 11(5): p. 1–8. [Google Scholar]
  • 41.Mathee A., et al. , A cross-sectional analytical study of geophagia practices and blood metal concentrations in pregnant women in Johannesburg, South Africa. South African medical journal, 2014. 104(8): p. 568–573. 10.7196/samj.7466 [DOI] [PubMed] [Google Scholar]
  • 42.Saute F., et al. , Malaria in pregnancy in rural Mozambique: the role of parity, submicroscopic and multiple Plasmodium falciparum infections. Tropical Medicine & International Health, 2002. 7(1): p. 19–28. [DOI] [PubMed] [Google Scholar]
  • 43.Glover-Amengor M., Owusu W., and Akanmori B., Determinants of anaemia in pregnancy in Sekyere West District, Ghana. Ghana medical journal, 2005. 39(3): p. 102 [PMC free article] [PubMed] [Google Scholar]
  • 44.Gedefaw L., et al. , Anaemia and associated factors among pregnant women attending antenatal care clinic in Walayita Sodo town, Southern Ethiopia. Ethiopian journal of health sciences, 2015. 25(2): p. 155–164. 10.4314/ejhs.v25i2.8 [DOI] [PMC free article] [PubMed] [Google Scholar]

Decision Letter 0

Adrian JF Luty

Transfer Alert

This paper was transferred from another journal. As a result, its full editorial history (including decision letters, peer reviews and author responses) may not be present.

14 May 2020

PONE-D-20-11384

Prevalence and factors associated with asymptomatic malaria and anemia among pregnant women in Kwale County, Kenya: A hospital based cross sectional study

PLOS ONE

Dear Mr Nyamu,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Both the expert reviewers have expressed significant concerns in several areas, many of which are overlapping, and with all of which I entirely agree. Your revised manuscript, should you choose to submit such, must therefore take into account all their specific comments and suggestions for improvement. The statistical analyses especially need attention as indicated in the detailed comments of Reviewer #2 in particular. There are also typographical/grammatical errors throughout that require your close attention.

We would appreciate receiving your revised manuscript by Jun 28 2020 11:59PM. When you are ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter.

To enhance the reproducibility of your results, we recommend that if applicable you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). This letter should be uploaded as separate file and labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. This file should be uploaded as separate file and labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. This file should be uploaded as separate file and labeled 'Manuscript'.

Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out.

We look forward to receiving your revised manuscript.

Kind regards,

Adrian J.F. Luty, PhD

Academic Editor

PLOS ONE

Journal requirements:

When submitting your revision, we need you to address these additional requirements:

1.    Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at http://www.plosone.org/attachments/PLOSOne_formatting_sample_main_body.pdf and http://www.plosone.org/attachments/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. Please carefully proofread your manuscript for typographical errors. For example, in the abstract “Prevalence of malaria in pregnant (MIP) in Kenya …” and in the methods section “We conducted across-sectional study …”.

3. Please include additional information regarding the survey or questionnaire used in the study and ensure that you have provided sufficient details that others could replicate the analyses. For instance, if you developed a questionnaire as part of this study and it is not under a copyright more restrictive than CC-BY, please include a copy, in both the original language and English, as Supporting Information.

4. Please provide details of the obtained Ethics approval and the informed written participant consent in the methods section of your manuscript. Currently this information is only available in the ethics statement on the online submission form.

5. We note that you have stated that you will provide repository information for your data at acceptance. Should your manuscript be accepted for publication, we will hold it until you provide the relevant accession numbers or DOIs necessary to access your data. If you wish to make changes to your Data Availability statement, please describe these changes in your cover letter and we will update your Data Availability statement to reflect the information you provide.

6. Please amend either the abstract on the online submission form (via Edit Submission) or the abstract in the manuscript so that they are identical.

7. We note that Figure 1 in your submission contain map images which may be copyrighted. All PLOS content is published under the Creative Commons Attribution License (CC BY 4.0), which means that the manuscript, images, and Supporting Information files will be freely available online, and any third party is permitted to access, download, copy, distribute, and use these materials in any way, even commercially, with proper attribution. For these reasons, we cannot publish previously copyrighted maps or satellite images created using proprietary data, such as Google software (Google Maps, Street View, and Earth). For more information, see our copyright guidelines: http://journals.plos.org/plosone/s/licenses-and-copyright.

We require you to either (1) present written permission from the copyright holder to publish these figures specifically under the CC BY 4.0 license, or (2) remove the figures from your submission:

1.    You may seek permission from the original copyright holder of Figure 1 to publish the content specifically under the CC BY 4.0 license. 

We recommend that you contact the original copyright holder with the Content Permission Form (http://journals.plos.org/plosone/s/file?id=7c09/content-permission-form.pdf) and the following text:

“I request permission for the open-access journal PLOS ONE to publish XXX under the Creative Commons Attribution License (CCAL) CC BY 4.0 (http://creativecommons.org/licenses/by/4.0/). Please be aware that this license allows unrestricted use and distribution, even commercially, by third parties. Please reply and provide explicit written permission to publish XXX under a CC BY license and complete the attached form.”

Please upload the completed Content Permission Form or other proof of granted permissions as an "Other" file with your submission.

In the figure caption of the copyrighted figure, please include the following text: “Reprinted from [ref] under a CC BY license, with permission from [name of publisher], original copyright [original copyright year].”

2.    If you are unable to obtain permission from the original copyright holder to publish these figures under the CC BY 4.0 license or if the copyright holder’s requirements are incompatible with the CC BY 4.0 license, please either i) remove the figure or ii) supply a replacement figure that complies with the CC BY 4.0 license. Please check copyright information on all replacement figures and update the figure caption with source information. If applicable, please specify in the figure caption text when a figure is similar but not identical to the original image and is therefore for illustrative purposes only.

The following resources for replacing copyrighted map figures may be helpful:

USGS National Map Viewer (public domain): http://viewer.nationalmap.gov/viewer/

The Gateway to Astronaut Photography of Earth (public domain): http://eol.jsc.nasa.gov/sseop/clickmap/

Maps at the CIA (public domain): https://www.cia.gov/library/publications/the-world-factbook/index.html and https://www.cia.gov/library/publications/cia-maps-publications/index.html

NASA Earth Observatory (public domain): http://earthobservatory.nasa.gov/

Landsat: http://landsat.visibleearth.nasa.gov/

USGS EROS (Earth Resources Observatory and Science (EROS) Center) (public domain): http://eros.usgs.gov/#

Natural Earth (public domain): http://www.naturalearthdata.com/

8. Please include your tables as part of your main manuscript and remove the individual files. Please note that supplementary tables (should remain/ be uploaded) as separate "supporting information" files

9. We note you have included a table to which you do not refer in the text of your manuscript. Please ensure that you refer to Table 3 in your text; if accepted, production will need this reference to link the reader to the Table.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: No

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: No

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: No

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The investigators have surveyed pregnant women in their district of Kenya for malaria and anaemia at first antenatal visit. With a cohort of just over 300 women they have a reasonable power to identify risk factors for these conditions, as well as estimating their prevalence with reasonable precision. The observations are not particularly novel, but the study is of some interest in examining these important pregnancy-associated morbidities.

Major comments

1. Abstract results: make it clear whether the results presented are adjusted odds ratios throughout.

2. Risk factors for AIP: The authors need to better explain how anaemia risk is apparently associated with BOTH first trimester, AND gestation >16 weeks. There are problems with results presentation in Table 3 and it seems they have got the reference and comparator groups switched for the gestation comparison. In addition, part of the results in the table are presented as percentages of women with risk factor who are anaemia (the table column heading) and part are presented as percentage of anaemic women who fall into each risk factor category (not what we want). Please redo this table and recheck your AORs and revise text accordingly.

3. Discussion: differences in malaria prevalence between Kenya and Burkina Faso are more likely related to malaria transmission intensity, prevalence falls with gestation, and falls more in women receiving IPTp. (Related to this, please also discuss role of IPTp in controlling MIP in conclusions).

4. Similarly, end P 11, haemodilution over pregnancy seems more likely to explain declining hb over pregnancy. If postulating dfetal needs please provide reference.

Minor comments

1. Abstract: line 1 “pregnancy”. Results 3rd sentence rewrite “ Women who had MIP were 12.9%”. Last line of results: rewrite for clarity.

2. There were a lot of spacing issues in the pdf, e.g. words joined together, or words and brackets without spaces before them. Examples in abstract methods: odds ratios(OR) and confidence intervals(CI)- there are many others.

3. Introduction 3rd sentence change “malaria with AIP and consumption of soil (geophagy)”, needs different punctuation?

4. Next sentence: is this risk factors for MIP (parasitic infections? High gravidity is NOT associated with MIP) Or AIP?

5. “In sub-Saharan Africa, MIP affects approximately 125 million pregnant women every year”- this is a major misreading of the cited reference.

6. Top of P 5 “a cross sectional”

7. Inclusion/exclusion: “as well as those who”

8. Sample size “desired level” of what was 5%?

9. Data management “ between a variety…”

10. Page 8: “prevalence and risk factors for malaria parasitemia”. Latent malaria is not a recognised phrase. Next line remove “who”.

11. Parasite counts are best expressed as geometric mean not median.

12. P 9 define mild, moderate, severe anaemia.

13. Page 11 first paragraph contains multiple grammatical errors, please rewrite

14. Third paragraph same page please rewrite.

15. A lot of the references are incomplete (lacking volumes/pages), or not enough details are given to retrieve them, or have formatting issues. Refs 2, 5, 6 (what is MOH? Where can this be obtained- similar comments for some others), 7, 17-22, 33.

16. Table groups need editing to ensure e.g. it is clear which group women with 8 y school fall into. Same for gestation. And make table 2 N column wider so numbers are readable.

17.

Reviewer #2: Malaria in pregnancy (MiP) remains a serious public health issue in sub-Saharan African countries despite efficient preventive measures, making difficult for the overall control of malaria. This present study aims to determine the prevalence and risk factors associated with asymptomatic malaria and anaemia among pregnant women in Kwale county, Kenya. The topic of asymptomatic malaria (potential reservoir) is relevant, however, as presented, this article is not original. In addition, the manuscript still requires significant grammatical editing to improve its readability and clarity.

The authors should highlight the fact that asymptomatic malaria is an important concern especially in the first ANC visit where any preventive measures are not yet implemented. Also, they have to underline the importance of the first trimester which is a critical risk period for pregnant women in terms of malaria and anaemia as found in their results.

There are below some comments for the authors that they should take into consideration for improving the manuscript. Moreover, a numbered manuscript would have making easier the review.

1) Suggested title: “Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anaemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya. Use asymptomatic malaria is less specific.

2) Abbreviations: Malaria in pregnancy (MiP and not MIP), Anaemia in pregnancy (AiP and not AIP)

3) Abstract:

a. Background sub-session: Prevalence of malaria in pregnancy, not … in pregnant

b. Results sub-session: please specify what type of odds you used for AiP (crude or adjusted)

4) Key words: important key words do not appear. Please use “asymptomatic malaria” instead of “parasitaemia” which could lead confusion as including symptomatic and asymptomatic cases.

5) Introduction

a. The authors should provide more detail on the current policies against malaria and anaemia during pregnancy used in Kenya, particularly IPTp administration and timeline, iron and folate supplementation? Who in charge of these measures (Government or pregnant women themselves)?

b. Page 3, line 6: “Other factors associated with MiP are……” Use “Other” supposes that you have cited first factors which is not the case. Please rephrase the sentence to make it clear.

c. Page 3, line 7: The authors state that “high parity” is associated with malaria. What do you mean by high parity? Multigravidae women? If yes, I think it’s a wording mistake as it’s well-known that is the primigravidae who are higher risk of MiP, so ‘less parity”.

d. The rational of the study is unclear as presented. Please, give more details explaining what the study brings for the scientific community which are not already known. The only fact that “factors contributing to MiP are not well-described in this part of Kenya” is not sufficient.

e. What is the most common species of malaria parasites in Kenya? I suppose “P. falciparum”. Hence, it would interesting to adjust the title of manuscript accordingly.

6) Methods

a. Ethical statement should be presented in the main text.

i. Did the pregnant women receive IPTp at the 1st ANC if they were eligible?

ii. Did the pregnant women with asymptomatic malaria receive curative treatment? What and how (uncomplicated and severe malaria)? If not, why? Same concerns regarding the anaemia, particularly severe cases?

iii. Any written informed consent? Any ethical committee approval?

b. Study population: give more detail on the strategy of participants selection

c. Did you consider among the symptoms of malaria the history of fever the past 48 h before the visit?

d. Exclusion criteria: Pregnant women who had taken antimalarial drugs within the past two weeks were excluded. What about the women who had taken fever drug? There is a risk to consider women asymptomatic while they just took fever drug the day before the visit.

e. Sample size: The authors have considered a prevalence of MiP from a study in Burkina Faso (24%) while the malaria transmission is different to both countries.

f. Data collection: give more details on socio-demographic characteristics, obstetric and clinical history. How did you assess the LLIN use? Please define the different trimesters of pregnancy (1st, 2nd, 3rd)? Give more details on how the gestational age was assessed?

g. Quality control: Please precise if the 10% of slides chosen was for all sides or positive slides.

h. Statistical analysis: What procedure did you use for variable selection in the final model (multivariate model)?

7) Results

a. Table 1: The proportion of pregnant women in the first trimester at the 1st ANC visit was 9.4% while the authors found that the proportion of pregnant women with gestational age < 16 wg was 15.9%. Why this discrepancy when the first trimester finished at 15 wg.

b. Table 2:

i. Why did you keep in the final multivariate model, the variable gestational age even if not significant in bivariate analysis?

ii. In the same way, why did you keep in the multivariable model “slept under bed net previous night” and “frequency of sleeping under bed net”. Both variables seems to be correlate.

iii. However, you drop out the variable “gravidity” which should be forced in the final model even if not significant because it’s a well-know factors strongly associated with MiP. Furthermore, I would like to suggest to the authors to make an sensitivity analysis by pooling primigravidae and secundigravidae in comparison to multigravidae.

c. Prevalence and factors associated with latent malaria:

i. Please define latent malaria?

ii. What is the prevalence of asymptomatic malaria among pregnant women in the 1st, 2nd and 3rd trimesters at the first ANC visit?

iii. What are the proportion of different species of parasites (P.f.; P.o; P.v; P.m)

iv. Please define POR at the first time it use in the text.

d. Prevalence and factors associated with anaemia:

i. Among the 3.6% of severe anaemia, how many are infected by malaria?

8) Discussion

a. Regarding the factors associated to MiP:

i. The authors should also discuss what happens among women in the first trimester of pregnancy. We can observe that women are more at risk of infection than those in 2nd and 3rd trimester (19.7% vs. 12.7% and 10.4%, respectively).

ii. The only factor associated with MiP is young age (< 20 y). The authors should consider to check an interaction between age and gravidity as both are correlated. Hence, this could be explained by that young pregnant women are mostly primigravidae? This deserves a couple of sentence in the discussion.

b. Regarding the factors associated to AiP: First and 3rd trimester are both associated with AiP. This could be also explained by the haemoglobin level variation due to physiopathology of the pregnancy. This should be included in the discussion

c. Study limitations: The authors have stated several limitations for the study. It is a good point. However, they have to explain how they have controlled this bias to ensure the validity of the study.

9) Conclusion: The authors should revise their conclusion in order to highlight the originality of the study.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files to be viewed.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org. Please note that Supporting Information files do not need this step.

PLoS One. 2020 Oct 8;15(10):e0239578. doi: 10.1371/journal.pone.0239578.r002

Author response to Decision Letter 0


1 Jul 2020

Reviwer #1

Questions/Concerns Comments Corrections

Major comments

Concern 1 Abstract results: make it clear whether the results presented are adjusted odds ratios throughout. Corrected done it is adjusted prevalence odds ratios that has been used throughout the abstract

Concern 2 Risk factors for AIP: The authors need to better explain how anaemia risk is apparently associated with BOTH first trimester, AND gestation >16 weeks. There are problems with results presentation in Table 3 and it seems they have got the reference and comparator groups switched for the gestation comparison. In addition, part of the results in the table are presented as percentages of women with risk factor who are anaemia (the table column heading) and part are presented as percentage of anaemic women who fall into each risk factor category (not what we want). Please redo this table and recheck your AORs and revise text accordingly. Yes, I have reanalyzed the anemia data, by use of backward elimination method, there seemed an interaction between trimester’s variable and gestational variable. By removing trimester variable for it had a bigger p value, gestational >16 weeks resulted to AOR; 3.3 (1.72-6.41). I have changed the comparison groups accordingly both in the table and in the text

Concern 3 . Discussion: differences in malaria prevalence between Kenya and Burkina Faso are more likely related to malaria transmission intensity, prevalence falls with gestation, and falls more in women receiving IPTp. (Related to this, please also discuss role of IPTp in controlling MIP in conclusions). Interventions measures for first visit pregnant women at antenatal clinic for malaria in pregnancy are; IPTp and provision of LLINs among others [17]. Although the role of IPTp is known to reduce maternal malaria episodes and improve pregnancy outcomes [33], the current study did not include pregnant women who had taken antimalarials hence we could not ascertain the role played by IPTp and majority of our study participants (82.7%) in the current study reported to always using bednet, whereas the study in Burkina Faso included pregnant women seeking ANC services at any point in their pregnancies. This may be factors in the difference between these two reported prevalence.

Concern 4 Similarly, end P 11, haemodilution over pregnancy seems more likely to explain declining hb over pregnancy. If postulating dfetal needs please provide reference.

Anemia is a function of plasma volume and red cell mass; both of which increase during pregnancy; but the increase in plasma volume is proportionately greater than the increase in red cell mass [40]. Explanation has been given

Minor comments

Concern 1 Abstract: line 1 “pregnancy”. Results 3rd sentence rewrite “ Women who had MIP were 12.9%”. Last line of results: rewrite for clarity.

Corrected; pregnant to pregnancy

Rewritten; Women who had plasmodium infections were 12.9 % (40/308)

Concern 2 There were a lot of spacing issues in the pdf, e.g. words joined together, or words and brackets without spaces before them. Examples in abstract methods: odds ratios(OR) and confidence intervals(CI)- there are many others. Attention to details in spacing has been addressed throughout the manuscript

Concern 3 Introduction 3rd sentence change “malaria with AIP and consumption of soil (geophagy)”, needs different punctuation?

It has been re-written

Concern 4 . Next sentence: is this risk factors for MIP (parasitic infections? High gravidity is NOT associated with MIP) Or AIP?

It has been re-written

Concern 5

“In sub-Saharan Africa, MIP affects approximately 125 million pregnant women every year”- this is a major misreading of the cited reference.

It has been re-written

Concern 6 Top of P 5 “a cross sectional”

It has been re- written

Concern7

Inclusion/exclusion: “as well as those who”

It has been re- written

Concern 8

Concern 8 Sample size “desired level” of what was 5%?

Two-sided significance level defined at 5%

Concern 9 Data management “ between a variety…”

It has been re-written

Concern 10 Page 8: “prevalence and risk factors for malaria parasitemia”. Latent malaria is not a recognised phrase. Next line remove “who”.

Latent replaced by asymptomatic and re-written

Concern 11 Parasite counts are best expressed as geometric mean not median. Geometric mean was 3738

Concern 12 . P 9 defines mild, moderate, severe anaemia.

Mild, moderate and severe anemia has been defined in definition of terms

Concern 13 Page 11 first paragraph contains multiple grammatical errors, please rewrite

It has been re looked and corrected accordingly

Concern 14 Third paragraph same page please rewrite. It has been re looked and corrected accordingly

Concern 15 A lot of the references are incomplete (lacking volumes/pages), or not enough details are given to retrieve them, or have formatting issues. Refs 2, 5, 6 (what is MOH? Where can this be obtained- similar comments for some others), 7, 17-22, 33.

The references has been corrected and the links are provided where necessary

Concern 16 Table groups need editing to ensure e.g. it is clear which group women with 8 y school fall into. Same for gestation. And make table 2 N column wider so numbers are readable.

The table 2; corrected < 8 year in schooling and gestational age in weeks >16

It has been expanded and fonts increased for readability

Reviwer #2

Comments /questions Corrections/ responses

Concern 1 Suggested title: “Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anaemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya. Use asymptomatic malaria is less specific Plasmodium falciparum was the predominant species more than 80% hence we concur with the reviewer

Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya.

Concern 2 Abbreviations: Malaria in pregnancy (MiP and not MIP), Anaemia in pregnancy (AiP and not AIP) Have corrected the whole manuscript where applicable MIP to MiP and AIP to AiP

Concern 3 Abstract:

a) Background sub-session: Prevalence of malaria in pregnancy, not … in pregnant Corrected to read Prevalence of malaria in pregnancy, not … in pregnant

b) Results sub-session: please specify what type of odds you used for AiP (crude or adjusted) We have corrected the omission, the odds Ratio was Adjusted Odd Ratio

c) Key words: important key words do not appear. Please use “asymptomatic malaria” instead of “parasitaemia” which could lead confusion as including symptomatic and asymptomatic cases Have replaced “Parasitaemia”, with “asymptomatic malaria”

Concern 4 Introduction

a. The authors should provide more detail on the current policies against malaria and anaemia during pregnancy used in Kenya, particularly IPTp administration and timeline, iron and folate supplementation? Who in charge of these measures (Government or pregnant women themselves)?

“We have included these paragraphs

The focused antenatal care is a package given to pregnant women which entails giving prophylaxis and treatment for anemia and malaria, among other services, where the AiP is prevented by providing nutritional counseling including iron supplements, and treating cases of AiP [20].

An intervention aimed at prevention and control MiP adopted by the Ministry of Health (MOH) Kenya, are; Intermittent Preventive Treatment (IPTp) of MiP, with Sulfadoxine Pyrimethamine (SP) given after 12 weeks gestational period done four weeks apart until the pregnant woman delivers, in areas with high malaria transmission such as Western, Nyanza and Coast regions and Long-lasting Insecticidal Nets (LLINs) at the ANC in the first contact among others [21]

Page 3, line 6: “Other factors associated with MiP are……” Use “Other” supposes that you have cited first factors which are not the case. Please rephrase the sentence to make it clear. Besides plasmodium infections other factors known to cause AiP include nutritional deficiencies, infectious diseases like HIV, parasitic infections like hookworm infestation, and the hemoglobinopathies [4, 5].

c. Page 3, line 7: The authors state that “high parity” is associated with malaria. What do

you mean by high parity? Multigravidae women? If yes, I think it’s a wording mistake as it’s well-known that is the primigravidae who are higher risk

of MiP, so ‘less parity”.

Corrected; Several studies has documented factors associated with MiP are women with less parity,

d. The rational of the study is unclear as presented. Please, give more details explaining what the study brings for the scientific community which are not already known. The only fact that “factors contributing to MiP are not well-described in this part of Kenya” is not sufficient.

Kenya Malaria Indicator Survey (KMIS) 2015 reported 2 folds increase in the prevalence of malaria in the coastal region compared to KMIS 2010 [18]. There are a possibility plasmodium infections in healthy adults, including pregnant women, in moderate to high transmission areas rarely result in fever [19]. Therefore, elimination of malaria is highly unlikely if diagnostic strategies do not include asymptomatic patients, because they will remain a reservoir of parasites contributing to the spread of the disease from one malaria season to the next. Man¬agement and control of MiP and AiP are enhanced by the availability of local prevalence statistics, which is not adequately provided in Kwale County except the data collected in passive surveillance. It is in this background we carried out this study to estimate the prevalence of asymptomatic MiP and AiP and identify the associated factors among women attending their first ANC visit at the largest referral health facility in Kwale County.

e. What is the most common species of malaria parasites in Kenya? I suppose “P. falciparum”. Hence, it would interest to adjust the title of manuscript accordingly.

Has adjusted the title by including the “asymptomatic Plasmodium falciparum infection”

Concern 5 Methods

a) Ethical statement should be presented in the main text.

i) Did the pregnant women receive IPTp at the 1st ANC if they were eligible?

Yes, they did, as per the government of Kenya Policy in prevention and control of Malaria in Pregnancy. .http://www.nmcp.or.ke/index.php/malaria-in-pregnancy

ii. Did the pregnant women with asymptomatic malaria receive curative treatment? What and how (uncomplicated and severe malaria)? If not, why? Same concerns regarding the anaemia, particularly severe cases? Following informed consent, the study participants were explained whose test positive for malaria and will have anemia will benefit by being treated as per the guidelines of malaria and anemia in pregnancy with no extra cost.

iii. Any written informed consent? Any ethical committee approval?

We sought written, informed consent from each participant before interviewing and finger pricking for malaria blood slides and Hemoglobin level analysis. Permission was granted to conduct the study by Kwale Department of Health and Msambweni Hospital Director. Ethical clearance was obtained from the Pwani University -Ethical Review Committee (ERC/MSc/021/2018).

b. Study population: give more detail on the strategy of participant’s selection

Sampling procedures

Systematic random sampling method was used to select study participants. Our sampling interval was based on the daily entries in the mother-child health (MCH) register from September 2018 to February 2019. The sampling started by selecting a participant from the daily entries list at random using a table of random numbers and then every kth participant in the frame was selected. A selection interval (k) was determined by dividing the total daily entry listed in order to get the number of the participants required per day. If a randomly-selected participant was not eligible for an interview or refused to be part of the study, the next eligible participant on the list was selected. We sampled our study participants until we arrived at our desired sample size of 308.

Following informed consent, the study participants were explained whose test positive for malaria and anemia will benefit by being treated as per the guidelines of malaria and anemia in pregnancy with no extra cost.

c. Did you consider among the symptoms of malaria the history of fever the past 48 h before the visit?

Exclusion criteria: Pregnant women who had taken antimalarial drugs within the past two weeks were excluded. What about the women who had taken fever drug? There is a risk to consider women asymptomatic while they just took fever drug the day before the visit. Asymptomatic malaria was defined as the presence in the peripheral blood of asexual blood stage of Plasmodium, irrespective of species but has no symptoms of malaria per clinical assessment (i.e. temperature <37.50C, chills, rigor, nausea, vomiting, headache, anorexia, or joint/muscle pains) and has not taken antipyretics within 48 hours and antimalarials within 14 days.

Concern 6 e. Sample size: The authors have considered a prevalence of MiP from a study in Burkina Faso (24%) while the malaria transmission is different to both countries.

True, we took the prevalence of MiP for Burkina Faso (24%). To our best of our knowledge by the time we were conceptualizing the protocol there was no study for asymptomatic MiP we could get hence we used the African study similar to ours to calculate the prevalence. Also, we thought the differences in malaria prevalence between Kenya and Burkina Faso are more likely related to malaria transmission intensity due to we have similar climate is characterized by ‘long rains’ (April–June) and ‘short rains’ (October–December) rainy seasons.

f. Data collection: give more details on socio-demographic characteristics, obstetric and clinical history.

Variables collected included;

Socio-demographic characteristics; mother's age, education level, marital status and occupation.

Obstetrics variables; gravidity, parity, trimesters, and gestational age in weeks.

Clinical history variables; history of fever in the last 48 hours and taken anti pyretic drugs, whether the client has taken antimalarials drugs, tendency of geophagy, whether patient is on iron supplements

How did you assess the LLIN use?

We depended on what the study participant reported which has been highlighted as a limitation but we used a trained nurse on the protocol to collect the data.

Please define the different trimesters of pregnancy (1st, 2nd, 3rd)?

First trimester was defined as; from week 1 to the end of week 12 while the second trimester is from week 13 to the end of week 26 and the third trimester is from week 27 to the end of the pregnancy

Give more details on how the gestational age was assessed? We used fundal height and last monthly period to estimate the gestational age

Concern 7 g. Quality control: Please precise if the 10% of slides chosen was for all sides or positive slides. An independent qualified parasitologist examined 10% of both positives and negatives slides which were randomly selected

Concern 8 h. Statistical analysis: What procedure did you use for variable selection in the final model (multivariate model)?

Variables with p-value ≤ 0.20 were included in a logistic regression model using a backward stepwise elimination method to identify independently associated factors.

Concern 9

7) Results

a. Table 1: The proportion of pregnant women in the first trimester at the 1st ANC visit was 9.4% while the authors found that the proportion of pregnant women with gestational age < 16 wg was 15.9%. Why this discrepancy when the first trimester finished at 15 wg.

b. Table 2:

i. Why did you keep in the final multivariate model, the variable gestational age even if not significant in bivariate analysis?

ii. In the same way, why did you keep in the multivariable model “slept under bed net previous night” and “frequency of sleeping under bed net”. Both variables seems to be correlate.

iii. However, you drop out the variable “gravidity” which should be forced in the final model even if not significant because it’s a well-know factors strongly associated with MiP. Furthermore, I would like to suggest to the authors to make an sensitivity analysis by pooling primigravidae and secundigravidae in comparison to multigravidae.

c. Prevalence and factors associated with latent malaria:

i. Please define latent malaria?

ii. What is the prevalence of asymptomatic malaria among pregnant women in the 1st, 2nd and 3rd trimesters at the first ANC visit?

iii. What are the proportion of different species of parasites (P.f.; P.o; P.v; P.m)

iv. Please define POR at the first time it use in the text.

d. Prevalence and factors associated with anaemia:

i. Among the 3.6% of severe anaemia, how many

a) The difference is due to the definition of the trimesters that was used in this study. First trimester was defined as; from week 1 to the end of week 12 while the second trimester is from week 13 to the end of week 26 and the third trimester is from week 27 to the end of the pregnancy

b) Have reanalyzed the data (see table 2 and 3)

c) Similar to the answer b, have re- analyzed the data (Table 2) and pooled together the primigravidae and second gravidae in comparison with multigravidae but still was not significant

we have used I have replaced latent malaria with asymptomatic malaria

Gestational trimesters with plasmodium infections, the first trimester were 2/29 (6.9%), second trimester 24/173 (13.9%) and third trimester were 14/106 (13.2%).

Plasmodium falciparum were 35 (87.5%), Plasmodium malarie 3 (7.5%) and Plasmodium ovale 2 (5.0%).

I has been defined as Prevalence Odds Ratio

Those who had severe anemia 4/7 (57.1%) had malaria, moderate anemia 15/95 (15.8%) and mild anemia was 14/90 (15.6%).

Discussion

a. Regarding the factors associated to MiP:

i. The authors should also discuss what happens among women in the first trimester of pregnancy. We can observe that women are more at risk of infection than those in 2nd and 3rd trimester (19.7% vs. 12.7% and 10.4%, respectively).

The current study reported a higher proportion of Plasmodium infections in pregnant women who were both in second and third trimesters and less proportion to the first trimester. Studies in Nigeria have reported high malaria prevalence in pregnant women who were in their second trimesters [27-29]. A study in Mali reported pregnant women in their first trimester were two times more likely to get malaria compared to the third trimester [30]. In contrast, our study reported less proportion of pregnant women in their first trimester and was not associated with malaria. This was probably due to the small number of pregnant women among this category. With an increase in the number of pregnant women in their first trimester, there is the possibility that there could be changes from the present results.

ii. The only factor associated with MiP is young age (< 20 y). The authors should consider to check an interaction between age and gravidity as both are correlated. Hence, this could be explained by that young pregnant women are mostly primigravidae? This deserves a couple of sentence in the discussion.

There was no Interaction

In this study, the highest proportional of Plasmodium infections was observed among the primigravidae (19.7%): followed by secundigravidae (12.7.7%) and multigravidae (10.4 %) with parasitaemia declining with increasing gravidity. These results are consistent with previous reports which found plasmodium infections are more common in primigravidae women compared to multigravidae women [7, 28, 31]. The reason for the present result of gravidae-associated predisposition to P. falciparum infections may be due to the fact that adults who live in malaria-endemic regions generally have some acquired immunity to malaria infection due to immunoglobulin production stimulated by previous malaria infection. This acquired immunity diminishes significantly in pregnancy particularly in primigravidae. It has also been suggested by various authors that the early onset of antibody response in multigravidae and the delayed antibody production in primigravidae may be responsible for the gravidity-dependent and differential prevalence of falciparum malaria among pregnant women [19, 32].

b. Regarding the factors associated to AiP: First and 3rd trimester are both associated with AiP. This could be also explained by the haemoglobin level variation due to physiopathology of the pregnancy. This should be included in the discussion

We found that anemia is more common among women in their third trimester than among women in their first trimester, similar to findings reported in other studies [39, 40]. Hemoglobin decreases until the end of the third-trimester. This might be due the fact that increase in trimester may cause reduction in maternal iron reserves. Anemia is a function of plasma volume and red cell mass; both of which increase during pregnancy; but the increase in plasma volume is proportionately greater than the increase in red cell mass [41]. Also, anemia in the third trimester may be more likely due to higher nutrient demands of the fetus later in pregnancy

c. Study limitations: The authors have stated several limitations for the study. It is a good point. However, they have to explain how they have controlled this bias to ensure the validity of the study.

We collected data from September 2018 to February 2019, a period during which there is low malaria transmission. This could have resulted in the underestimation of the overall prevalence of asymptomatic MiP in our study area. A continuous monitoring throughout the year of MiP incidences will account for seasonality burden [21].

In addition, the study was hospital-based, excluding pregnant women who did not seek ANC services. While this may limit the generalizability of findings to the community, few women fail to seek antenatal care in our study area. Determination of factors associated with asymptomatic MiP and AiP in hospital based studies provides a proxy indicator of predictors in the community of that particular facility when community based surveys are not feasible.

Lastly, this study did not explore other factors that may contribute to anemia, including nutritional factors, soil-transmitted helminthes infection, and hereditary conditions such as sickle cell disease thus limiting our ability to assess the contribution of other causes of anemia during pregnancy. However, diagnosis of anemia was based on laboratory analysis and did not depend on clinical assessment as reported by other researchers.

Concern 10 9) Conclusion: The authors should revise their conclusion in order to highlight the originality of the study.

Asymptomatic Plasmodium infections and anemia are common in women attending their first ANC visit at Msambweni County Referral Hospital in Kwale County. Most of the Plasmodium infections in this area are caused by P. falciparum. Asymptomatic MiP was associated with younger maternal age (≤20 years). Anemia in pregnancy was associated with Plasmodium infections, women who reported to have geophagy tendency and those who were their third trimester. In the study area, we recommend pregnant women should not delay their first ANC attendance, for less than 10% attended in their first trimester. All women of childbearing age should be included in measures to control Plasmodium infection and anemia by the National Malaria Control Program, reproductive health department and other non – state actors should. Also, the reproductive health department should carry out health promotion and education on late adolescent and school going pregnancy for delay of sexual debut.

Attachment

Submitted filename: Response to Reviewers#1.docx

Decision Letter 1

Adrian JF Luty

28 Jul 2020

PONE-D-20-11384R1

Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya.

PLOS ONE

Dear Dr. Nyamu,

Thank you for submitting your revised manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a newly revised version of the manuscript that addresses the points raised by the reviewers.

Both reviewers were in agreement that the revised manuscript is an improvement on the original submission, but both have nevertheless still raised some issues that require further attention, in particular concerning grammatical errors. Reviewer #1 has taken the time to provide a copy-edited version, appended to this letter, to assist in improving the English throughout. You should address these concerns in your revision along with the specific remaining issues outlined in the appended comments from the reviewers.

Please submit your revised manuscript by Sep 11 2020 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

  • A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

  • A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

  • An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols

We look forward to receiving your revised manuscript.

Kind regards,

Adrian J.F. Luty, PhD

Academic Editor

PLOS ONE

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: Partly

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: No

Reviewer #2: No

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The authors have addressed many of my comments but I have a few of remaining issues they need to address.

It would have been helpful if the manuscript had page and line numbers.

1. Discussion page 3: The high rates of net ownership cannot really be attributed to the policy of bed net distribution at first ANC visit as women would only receive these nets AFTER enrolment in the study- so no primigravidae would have received nets by this route. The mass distribution campaigns are more likely driving this.

2. Table 3 still needs correction. The column n(%) needs to be the percentage of women who are anaemic as a fraction of the women in that row. From Trimester onwards the figures must be corrected.

3. In the same table the crude POR for secundigravidae is incorrect. They have a higher anaemia prevalence than women in first trimester.

4. Also in Table 3, please switch “gestation >16 weeks” and “gestation ≤ 16 weeks” to be consistent with the rest of the table where lower values come first.

5. In these lines there is an error in n for women ≥16 weeks with anaemia- is it 174?

I have also attached the revised manuscript with highlighting to indicate the many minor formatting, spacing and other editorial issues that need to be corrected. These are mostly around missing spaces, and spaces which should not be there.

Two specific formatting comments:

1. Primigravidae, multigravidae are always written as one word and are nouns. Primi-gravidae is not correct and nor is “primigravidae women.

2. A semi-colon has a different function from the usage here. Colons are appropriate, or the punctuation can be removed.

Reviewer #2: While the revised manuscript has improved, it still requires significant grammatical editing to improve its readability and clarity. Some concepts remain unaddressed for me.

a) In the first review, I have highlighted that the rational of the study is unclear. The authors stated that pregnant women are potential reservoir of P.f parasites; however, the IPTp is one the main strategy protected pregnant women and is administered regardless the malaria status of women. Hence, this should not be a major lack as the women will clean by IPTp. In addition, in their conclusion, the authors recommended to take into account women of reproductive age in malaria control program; but no specific strategy has been suggested. The authors should revise their manuscript to show the scientific add-value of their paper.

b) For positive malaria management, the authors just precised that they used per the government of Kenya

Policy in prevention and control of Malaria in Pregnancy. But they should presented shortly how these cases are managed in the manuscript.

c) Informed consent administration: What is the process for illiterate women who did not write?

d) There is no flow chart diagram? How many pregnant women ineligible were excluded to reach the 308 pregnant women? And what are the reasons? Also compared the baseline characteristics of the pregnant women included and those who did not

e) How many participants have been excluded because of the following criteria: Intake of antipyretic drugs within 48h?

f) I have a concerns with the cut-off used to define 1st, 2nd and 3rd trimester. The authors define first trimester by using a gestational age < 12 wg instead of 14 completed wg, the cut-off usually used. Why ?

g) Two methods were used to measure the gestational age (fundal height and LMP); but there is no precision in what situation one or both are used?

h) QC of positive and negative slides. What is done in case of discrepancy ?

i) The authors showed that the prevalence of malaria were 6.9%; 13.9% and 13.2% in the first, 2nd and 3rd trimester respectively? It is slightly amazing as results as the prevalence should decrease from first trimester to delivery because of preventive measures implemented. How the authors explained this result. this deserve a couple of sentences in the discussion session

j) The authors highlighted the IPTp started from the 12 wg. I'm not sure if this is the current WHO recommendation as the IPTp should started as soon as possible from the 2nd trimester.

k) Number the page of manuscript.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Manfred Accrombessi

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Attachment

Submitted filename: PONE-D-20-11384_R1_reviewer-1.pdf

PLoS One. 2020 Oct 8;15(10):e0239578. doi: 10.1371/journal.pone.0239578.r004

Author response to Decision Letter 1


17 Aug 2020

Thanks for the consideration to review and your reviwers comments have been addressed to our best of knowledge.

Attachment

Submitted filename: Response to Reviewers #2_13_08_2020.docx.pdf

Decision Letter 2

Adrian JF Luty

10 Sep 2020

Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya.

PONE-D-20-11384R2

Dear Dr. Nyamu,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Adrian J.F. Luty, PhD

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

This manuscript is now acceptable for publication; however, the English needs again additional improvement.

For example on the clean version, there are some minor errors identified

- Line 1: Plasmodium should be in italic

- Line 2: Typo error on "department"

- Line 26: "Prevalence of malaria in pregnancy" instead of "both asymptomatic and symptomatic"

- Line 30: delete "asymptomatic". Pregnant women could not be asymptomatic

- Line 33: Odds ==> odds

- Line 44: Asymptomatic MiP, not AiP

- Line 46: Please reconsider "Pregnant"; may be "Pregnancy" or "Pregnant women"

- Line 54: Sub - Saharan Africa ==> sub-Saharan Africa

- Line 55: Delete 89324/142896 and keep only the %

- Line 63: Malaria in pregnancy, not Pregnancy with capital P. And we have already defined an abbreviations for that purpose, so use MiP

- Lines 65-67: Rephrase the sentence. Due to...., pregnant women are more at risk of malaria"

-Lines 70-71: Rephrase the sentence to make it clear. What do you mean by focused antenatal care? Use abbreviation of ANC previously defined

- Lines 105, 108: Combine sub-session of study design and study population ==> study design and population

- Lines 111: Inclusion and exclusion criteria should be included in the "study population" session

- Line 128-136: Rephrase the sub-session "sample size determination" by making an unique block without bullets.

- Line 156: Delete were, there is a repetition of "were" with those of line 155

- Line 174: Why abbreviate No Parasit Found (NPF) as you did not use it after in the mansucript

- Line 217: First bracket missing for 56.2%

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: (No Response)

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: (No Response)

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: (No Response)

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: (No Response)

Reviewer #2: No

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: (No Response)

Reviewer #2: (No Response)

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

Acceptance letter

Adrian JF Luty

25 Sep 2020

PONE-D-20-11384R2

Prevalence and risk factors associated with asymptomatic Plasmodium falciparum infection and anemia among pregnant women at the first antenatal care visit: A hospital based cross-sectional study in Kwale County, Kenya.

Dear Dr. Nyamu:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Adrian J.F. Luty

Academic Editor

PLOS ONE

Associated Data

    This section collects any data citations, data availability statements, or supplementary materials included in this article.

    Supplementary Materials

    Attachment

    Submitted filename: Response to Reviewers#1.docx

    Attachment

    Submitted filename: PONE-D-20-11384_R1_reviewer-1.pdf

    Attachment

    Submitted filename: Response to Reviewers #2_13_08_2020.docx.pdf

    Data Availability Statement

    All relevant data are within the paper.


    Articles from PLoS ONE are provided here courtesy of PLOS

    RESOURCES