Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India

1348 The Biobrio An International Quarterly Journal of Life Sciences Website: www.thebiobrio.in SJIF 2024:8.259 ISSN : 2393-9508 e-ISSN: 2582-4902 12(1&2): 1348-1363, 2025 Abhishek Mitra1, Hasib Ansari2, Birendra Kumar Gupta1, Ajay Kumar Sharma1 & Mohammad Sohail2 1University Department of Zoology, Vinoba Bhave University, Hazaribag, Jharkhand, India 2Centre for Tribal Health and Biotechnology Foundation, New Delhi, India. Received : 25th May, 2025 ; Accepted : 25th June, 2025 DOI:- https://doi.org/10.5281/zenodo.17426239 Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India ABSTRACT The escalating burden of malaria in pregnancy (MiP) is of public health concern across the globe, in view of the pathogenesis, epidemiology, clinical sequelae, prevention and treatment of malaria; all have unique features during pregnancy. The combinatorial impact of malaria in pregnancy includes effect of parasitic infection, anaemia, abortions, low birth weight, still births, and maternal mortality further perplexed the situation of prompt diagnosis, treatment and preventive strategies. This prompted us to estimate the population at risk of MIP in Hazaribag, Jharkhand, a malaria-endemic state in central-east India. Cross-sectional evaluation between September 2021 and December 2022 at antenatal clinics (ANC) and delivery units (DU) were performed at Sadar hospital, Hazaribag. Malaria was screened by Giemsa-stained blood smear and/or rapid diagnostic test using peripheral blood. Anaemia was defined as haemoglobin concentration. Pre-tested questionnaires were used to gather socio-demographic, clinical and obstetrical data. 1865 pregnant women were enrolled at the antenatal clinics and 934 at the delivery units. The prevalence of malaria during pregnancy was 4.8% and 3.7% at ANC and DU, and there were 11.4% malaria in women without pregnancy. Interestingly, majority (73%) were infected with P.vivax, and over 67% of the pregnant women were asymptomatic malaria. Anaemia was prevalent in 78% of ANC attendee as compared to 67% in DU, whereas, severe anaemia was 11.8% as compared to 6.3%. Anaemia was significantly associated with malaria; however, severe anaemia was more common among women with parasitaemia. In multivariate analyses, asymptomatic malaria increased over four times the likelihood of having anaemia. In view of sizable prevalence of malaria and in pregnancy with significantly associated anaemia and parasitemia suggests early diagnosis regardless of symptoms and comprehensive drug regime should be offered to pregnant women in association with existing measures in clinical spectrum of infection, delivery and its outcome. Key Words - MIP, Asymptomatic, P.vivax, Jharkhand, Anaemia *Corresponding author : [email protected] INTRODUCTION Tropical malaria, which is caused by the protozoan parasites Plasmodium falciparum and Plasmodium vivax, is responsible for 478 million clinical cases (Snow et al., 2005) and 0.5 to 2 million deaths 1349 annually (Sachs, 2002). Plasmodium vivax, the most widespread parasite causing human malaria, is responsible for an estimated 123-367 million infections annually and is the major cause of malaria in most of Asia and Latin America (Baird, 2007; Hay et al., 2004). Although P. vivax infection is commonly considered to be much more benign than Plasmodium falciparum infection, historical evidence suggests significant mortality associated with P. vivax malaria in the pre-antimalarial era (Dobson, 1994), and death caused by P. vivax malaria has been increasingly recognized over the past few years (Baird, 2007; Barcus et al., 2007; Anstey and Price, 2007). The emergence and spread of drug resistance to commonly used chemotherapeutics are major factors contributing to this increasing burden and most of the mortality and morbidity is borne by children and pregnant women. Pregnant women and their infants are susceptible to common and preventable infectious diseases including malaria but are woefully left unscreened and untreated. According to an estimate, approximately 125 million pregnant women worldwide are exposed to the risks of malaria in pregnancy (MiP) each year, resulting in 200,000 infant deaths (Dellicour et al., 2010; Steketee et al., 2001). Every year, in India, 28 million pregnancies take place with 67,000 maternal deaths (Registrar General of India. Sample Registration System. Special Bulletin on Maternal Mortality in India. 2004-06), 1 million women left with chronic ill health, and 1 million neonatal deaths (UNICEF, 2009). Pregnancy is an event of immunologic tolerance, whereby a woman accepts the implantation of the fetal allograft in her uterus and a unique period for vulnerability to malaria infection. Pregnant women with low levels of pre-existing immunity are especially vulnerable to the most severe complications of malaria during pregnancy. These complications include cerebral malaria, severe anaemia, miscarriage, intrauterine fetal death, premature birth, stillbirth, and both maternal and infant mortality (Steketee et al., 2001; Cot and Deloron, 2003). In regions where malaria is endemic, pregnant women are more susceptible to Plasmodium infections than non-pregnant women. While the most serious outcomes are typically seen in first-time mothers (primigravidae) (Rogerson et al., 2007; Menendez, 2006), in areas with low or unstable transmission, women of all pregnancy orders are at similar risk (Menendez, 2006). Compared to non-pregnant women, pregnant women are three times more likely to develop severe malaria, with mortality rates from such cases reaching up to 50% (Monif, 2004; WHO, 2006). Despite the severe and potentially fatal consequences of malaria during pregnancy for the mother, fetus, and newborn, these harmful effects are largely preventable through available interventions (Desai et al., 2007) or timely and appropriate treatment following early and accurate diagnosis (Menendez et al., 2007; Gamble et al., 2007; Ter Kuile and Steketee, 2007). Since malaria infection during pregnancy is often asymptomatic, the most widely used control strategy is Intermittent Preventive Treatment in pregnancy (IPTp). This approach aims to clear existing infections at the time of administration and provide short-term prophylaxis to prevent new infections. However, growing concerns about widespread resistance to commonly used antimalarial drugs (Bardaji et al., 2012) have highlighted the urgent need for alternative and more effective interventions. Diagnosing malaria during pregnancy is particularly challenging due to various factors, including the immunological changes across different stages of pregnancy, increased vulnerability to severe disease, obstetric complications, and the sequestration of parasites in the spleen and placenta. These challenges are often compounded by varying clinical presentations and different forms of anaemia. Consequently, the development of prompt and reliable diagnostic tools remains a critical focus of malaria in pregnancy (MiP) research. Plasmodium falciparum malaria during pregnancy is a well-established cause of maternal and fetal morbidity and mortality. Although P. vivax infection Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India 1350 has received less attention, it is increasingly recognized as a significant contributor to maternal anaemia and low birth weight (Duffy et al., 2001; Singh et al., 1999; Nosten et al., 1999), particularly in regions where both species frequently coexist. Of the approximately 50 million pregnancies occurring each year in malaria-endemic countries, nearly half take place in areas where P. vivax is prevalent. While P. vivax infection during pregnancy has been acknowledged for many years (Duffy et al., 2001), its specific impact has only been systematically assessed in recent studies. Research from Thailand and India has shown that pregnant women infected with P. vivax are more likely to be anaemic and to deliver infants with lower birth weights compared to uninfected women. However, these effects tend to be less severe than those linked to P. falciparum infection. In both studies (Nosten et al., 1999; Singh et al., 1999), P. vivax infection was most frequently observed during first pregnancies, with prevalence peaking early in the second trimester. Limited and earlier studies on malaria in pregnancy (MiP) in India have highlighted the significant role malaria plays in contributing to maternal and neonatal morbidity and mortality (Singh et al., 1999; Singh et al., 1998; Brooks et al., 2008; Singh et al., 1995). Preliminary findings, primarily from central India, suggest that both Plasmodium falciparum and P. vivax infections are associated with adverse pregnancy outcomes. However, these studies largely focused on symptomatic pregnant women infected with P. vivax (Singh et al., 1999; Singh et al., 1995), leaving significant gaps in understanding the broader impact of vivax malaria during pregnancy. Notably, there is a scarcity of data from Jharkhand-an understudied, tribal-dominated region with perennial malaria transmission-where malaria remains widespread and is responsible for a substantial number of annual deaths, second only to Odisha in India, according to recent reports (Dhingra et al., 2010; Hussain et al., 2011). This underscores the critical need for comprehensive investigations in the region, particularly concerning the pathology of malaria during pregnancy. As noted in previous research (Singh et al., 1999), malaria infection during pregnancy is linked to an increased risk of neonatal and infant mortality, further emphasizing the urgency of focused research in high-burden areas like Jharkhand. Given the limited data on asymptomatic and Plasmodium vivax infections during pregnancy in India, this study was undertaken with the objective of better estimating the burden of malaria in pregnancy (MiP), determining the prevalence of asymptomatic malaria, and assessing the relative contribution of P. falciparum and P. vivax infections during pregnancy and at delivery. To the best of our knowledge, such a comprehensive epidemiological and clinical profile has not previously been investigated in pregnant women from Hazaribag, Jharkhand-an underrepresented, malaria-endemic region of India. Notably, this study represents the first attempt to evaluate the interaction between anaemia, pregnancy, and asymptomatic malaria, stratified by clinical presentation, in an adult population residing in a perennial transmission zone characterized by codominance of P. vivax and P. falciparum. Conducted in Hazaribag, located in the eastern Indian state of Jharkhand, the study ultimately aims to inform and support the development of evidence-based policies to reduce the burden of MiP in this highrisk region. METHODS Study Sites/design and Population This study was based on cross-sectional surveys conducted at three healthcare units: antenatal care (ANC) clinics, delivery units (DU), and the inpatient antepartum ward of Sadar Hospital in Hazaribag district, Jharkhand, India (Figure 1). Jharkhand, located in eastern India, is a malaria-endemic state with a significant tribal population. Over the past three years, the state has recorded an average Slide Positivity Rate (SPR) of 6.8% among symptomatic individuals, with Plasmodium falciparum accounting for 44% of cases (National Vector Borne Disease Control Programme, 2007). Malaria remains a major public health challenge in this region, particularly in tribal-dominated areas. Abhishek Mitra, Hasib Ansari, Birendra Kumar Gupta, Ajay Kumar Sharma & Mohammad Sohail 1351 According to the Government of India's 2005 Draft National Policy on Tribals, the central-eastern beltwhich includes Jharkhandcontributes approximately 15-20% of all malaria cases in the country. In 2009 alone, Jharkhand reported 230,686 malaria cases, with P. falciparum responsible for 39.53% (Jharkhand, 2009). Hazaribag district, where the present study was conducted, is recognized as a malaria-endemic area within this high-burden state and was selected for its representative nature of stable malaria transmission in tribal settings. 2008, 56% of women had at least one antenatal clinic (ANC) visit, while only 18% delivered in health facilities-59% in urban areas and just 13% in rural settings (International Institute for Population Sciences, 2009). Sadar Hospital, the primary district hospital serving Hazaribag's predominantly rural population, features a dedicated obstetric unit with 40 beds. Between 2010 and 2013, it recorded a high volume of annual deliveries, ranging from approximately 4,800 to 5,500. The hospital also managed a substantial number of ANC visits during the same period, averaging between 5,200 and 6,600 per year. Screening and enrollment The study comprised two components, with participant recruitment targeting all pregnant women presenting at antenatal care (ANC) units and delivery units (DU) ward. For the ANC component, pregnant women aged ≥ 17 years who reported for routine care at the study site were screened and enrolled, provided they were willing and gave informed consent. In the DU component, women aged ≥ 18 years presenting for delivery were enrolled upon providing written informed consent. For the inpatient component, pregnant women admitted with a diagnosis of malaria, anaemia, or febrile illnesses of unknown origin were screened. Those with a confirmed malaria-related diagnosis were enrolled after obtaining informed consent. At each point of contact, all women underwent clinical assessment, peripheral blood smear examination for malaria parasites, and measurement of axillary body temperature prior to enrollment. The detailed enrollment strategy, sampling procedures, and classification into broad clinical groups are illustrated in the schematic flow chart shown in Figure 2. ANC procedures Trained study personnel conducted interviews with enrolled participants to collect data on sociodemographic characteristics (e.g., date of birth, socio-economic status, literacy), reproductive Figure 1Map of Jharkhand Showing Study Site Hazaribag district, with a total population of 1,734,005 according to the 2011 census, was selected for this study as it represents a rural-cumsemi-urban region with low but perennial malaria transmission. Over the past three years, Hazaribag recorded an average Slide Positivity Rate (SPR) of 7.3% among symptomatic individuals, with Plasmodium falciparum accounting for 14% of cases (Jharkhand, 2008; State Malaria Control Program Annual Report, Ranchi, Jharkhand, Directorate of Health Services). The population is socio-culturally diverse, comprising tribal communities, Scheduled Castes, Scheduled Tribes, and other caste groups, characterized by unique social stratification and notable gender disparities. Situated in the tropical zone, the district experiences an average annual rainfall of 1234.5 mm and has geo-climatic and ecological conditions favorable for year-round malaria transmission. According to the District Level Household and Facility Survey (DLHS3) conducted between December 2007 and April Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India 1352 history including gravidity, recent history of fever, use of anti-malarial medications, and preventive measures against malaria. A comprehensive physical examination was performed for each participant, including assessment of gestational age-estimated by palpation of the uterine fundus in conjunction with the date of the last menstrual period-along with measurement of axillary temperature using a digital thermometer and evaluation of other vital signs. DU procedures Pregnant women enrolled at the delivery units (DUs) were interviewed to collect information on sociodemographic and anthropometric characteristics, obstetric complications, history of fever and antimalarial drug use during pregnancy, use of antimalarial preventive measures, birth outcomes, and mode of delivery. After delivery, peripheral venous blood samples (3-5 ml) were collected for malaria blood film preparation, rapid diagnostic testing (RDT), haemoglobin measurement, and additional Peripheral venous blood samples (3-5 ml) were collected from all participants for malaria blood film preparation, rapid diagnostic testing (RDT), haemoglobin measurement, and other biochemical and molecular analyses. Women who tested positive on the RDT or were found to be anaemic were referred immediately to a hospital physician for appropriate treatment. Additionally, hospital staffs were notified of any parasitaemic individuals identified through microscopy to ensure timely clinical management. Figure 2Schematic Flow Chart Summarizing the Sampling Strategy and Groups biochemical and molecular analyses. Women who tested positive by RDT or blood smear were referred promptly for treatment. In addition to assessing malaria prevalence among women at the DUs, clinical and demographic data and samples were also stratified by mode of delivery-normal, cesarean, and stillbirth-as well as by birth outcomes, including preterm, term, and post-term deliveries. These detailed results are presented in Table 1. Abhishek Mitra, Hasib Ansari, Birendra Kumar Gupta, Ajay Kumar Sharma & Mohammad Sohail 1353 Laboratory procedures Thick and thin blood smears prepared from peripheral blood samples of ANC and DU participants were stained with Giemsa and examined under high-power microscopy. Parasite density was quantified by counting the number of asexual parasite forms per 200 leukocytes, assuming a standard leukocyte count of 8,000 leukocytes/?l of blood (Trape, 1985). The thin smear was specifically used for Plasmodium species identification. All slides were independently crosschecked by trained technical staff following stringent diagnostic criteria to confirm Plasmodium infection. Additionally, the commercial First Response Malaria pLDH/HRP2 combo rapid diagnostic test (RDT) kits (Premier Medical Corporation, Mumbai, India) were employed as per the manufacturer's instructions to screen pregnant women for malaria. Haemoglobin Concentration Hemoglobin (Hb) levels were measured during the initial ANC and DU visits. Hb concentrations were determined from peripheral blood samples using a portable HemoCue hemoglobinometer (HemoCue AB, Ängelholm, Sweden), following the manufacturer's instructions. The Hb values were recorded on the study questionnaire and verified by a laboratory technician. Women were classified as anemic if their Hb was below 11 g/dL. Further classification identified moderate to severe anemia with Hb levels below 8 g/dL and 7 g/dL, respectively, as the primary outcome. Those with Hb levels of 9 g/dL or higher were categorized as mildly anemic or non-anemic, based on the WHO anemia classification (McLean et al., 2012). Study definitions Malaria was defined by the presence of asexual blood stages of Plasmodium parasites in peripheral blood or a positive rapid diagnostic test (RDT), regardless of the species or symptoms. Symptomatic malaria infection was identified when there was a history of fever within the past week or a temperature of ≥ 37.5°C, along with the presence of asexual forms of P. falciparum or P. vivax detected on a thick blood smear or a positive †Numbers may not add to sample size secondary to missing data. * Defined as 3 or more pregnancies ** For ANC enrollees, gestational age assessed by fundal height. For DU enrollees, gestational age was assessed by Ballard score. Table 1-Baseline characteristics of pregnant women attending antenatal and delivery units C haracteristic s Antenatal clinics n=1271 Delivery units n=870 N, (%) N, ( %) Age (Years) <20 166(13.1) 109(12.5) 20 - 34 983(77.4) 708(81.4) ≥35 122(9.5) 53(6.1) Prior pregnancies Primigravid 423(33.3) 338(38.38) Secundigravid 578(45.5) 209( 24.1) Multigravid* 270(21.2) 323(37.1) Gestational age at enrollment (weeks)** <20 weeks 567(44.6) n/a 20 - 36 weeks 641(50.4) 57(6.5) ≥37 weeks 63(5) 813(93.5) Caste Schedule caste 169(13.3) 93(10.7) General caste 428(33.7) 307(35.3) Other backward caste 311(24.5) 219(25.2) Scheduled trib e 363(28.5) 251(28.8) Education No formal schooling 357(28.1) 321(36.9) Attended school any length of time 914(71.9) 549(28.8) Socioeconomic characteristics Owns TV 567(44.6) 387(44.5) Owns bi cycle 1173(92.2) 687(78.9) Owns house 958(75.4) 643(73.9) Owns refrigerator 123(9.6) 83(905) Roof material Mud 622(48.9) 513(58.9) Corrugated iron/asbestos sheet 242(19) 182(20.9) cement/concrete 329(25.8) 107(12.3) Other 78(6.1) 68(7.8) Wall material Mud/sand/dung 673(52.9) 478(54.9) Mud bricks 127(9.9) 93(10.7) Cement bricks 419(32.9) 267(30.7) Other 52(4.1) 32(3.7) Primary cooking fuel wood 619(48.7) 387(44.5) charcoal 437(34.4) 279(32.1) Gas 153(12.1) 136(15.6) Other 62(4. 9) 68(7.8) Mode of delivery among pregnant women Normal n/a 586(67.3) Caesarean n/a 179(20.6) Still Birth n/a 105(12.1) Birth Outcome Pre - Term Delivery(≤36 weeks) n/a 129(14.8) Term Delivery (31 - 41 weeks) n/a 623(71.6) Post-Term Delivery (after 41 weeks) n/a 118(13.5) Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India 1354 RDT. Severe malaria was characterized by a malaria episode accompanied by any of the following conditions: cerebral malaria, severe anemia, renal failure, pulmonary edema, hypoglycemia, shock, spontaneous bleeding, or repeated convulsions. Maternal height and weight were measured during the first ANC and DU visits, and body mass index (BMI) was calculated as weight in kilograms divided by height in meters squared. A low BMI was defined as less than 22.0 kg/m². A documented fever was considered an axillary temperature of ≥ 37.5°C. Ethics Statement and Subject Consent All human blood samples used in this study were collected after obtaining consent from the study participants under protocols activities approved by the Institutional Ethics Committee (IEC) of the Vinoba Bhave University, Hazaribag, Jharkhand and human ethical guidelines as reflected in the guidelines of the Medical Ethics Committee, Ministry of Health, Govt. of India. All study participants provided informed consent. The protocol is approved from IEC, VBU having memo no. VBU/R/888/2012 dated 05-06-2012. Data management and analysis All clinical, demographic, and anthropometric data were thoroughly reviewed for accuracy, and any inconsistencies were resolved prior to analysis. The data were entered into MS Excel, and statistical analyses were conducted using SPSS version 16 (SPSS Inc., Chicago, IL, USA) and GraphPad Prism version 5.0 (GraphPad Software, Inc., CA, USA). For comparing means between two groups, the Student's t-test was applied to normally distributed data, while non-parametric tests (Mann-Whitney U) were used when the data did not follow a normal distribution. Categorical variables are reported as frequency counts (percentages) and compared using either the chi-square test or Fisher's exact test, as appropriate. Continuous variables are presented as means (± standard error) and compared using t-tests or analysis of variance (ANOVA), depending on the context. Because most participants were unsure of their exact birthdates, ages were categorized into ranges based on their estimates. Risk factors for P. falciparum or P. vivax parasitemia were first assessed through univariate analysis, followed by multivariate analysis to adjust for significant predictors. Statistical significance was set at a p-value of less than 0.05. RESULTS Recruitment and Enrollment Between September 2021 and December 2022, 1,890 pregnant women were screened during antenatal care (ANC) visits. Of these, 1,746 were willing to hear the study protocol, 1,271 consented and provided peripheral samples, while 475 declined participation. In parallel, 444 non-pregnant women consented to peripheral sampling. The final study population included 1,271 pregnant women and 444 non-pregnant women. Pregnant women were further stratified by trimester: 135 in the first, 492 in the second, and 644 in the third. The nonpregnant cohort comprised 227 malaria-positive and 217 malaria-negative women. In delivery units (DU), an additional 870 pregnant women were screened and enrolled. Antenatal Clinics (ANC) Most ANC participants were aged 18-38 years and had some formal education (Table 1). Nearly all were Hindi-speaking (97.6%) and non-smokers (98.7%). A majority owned their homes (75.4%) and were engaged in household work (76.7%), while 12.3% were involved in farming. The median number of ANC visits was one (range: 0-9), and 33.3% were primigravidae. Slightly more than half attended ANC during the second or third trimester, while 44.6% presented before 20 weeks' gestation. Only 46.3% reported iron/folate supplementation and 33.2% took multivitamins. Bed nets were commonly present in households, but insecticidetreated nets (ITNs) were rare (Table 2). Nine women reported taking malaria prophylaxis; however, seven could not identify the drug, while two reported chloroquine use. Malaria prevalence in ANC attendees was 5.4% (68/ 1271; Table 3). Blood smears were positive in 4.3% of women, with an additional 1.1% detected by rapid diagnostic tests (RDTs). The mean parasite density among smear-positive women was 63,236 Abhishek Mitra, Hasib Ansari, Birendra Kumar Gupta, Ajay Kumar Sharma & Mohammad Sohail 1355 asexual forms/μl (range: 600-489,000). Species distribution showed Plasmodium falciparum in 4.4%, P. vivax in 86.8%, and mixed infections in 8.8%. Rural residence was strongly associated with parasitaemia (OR 4.32, 95% CI 1.67-9.46). Primigravidae and secundigravidae were also at higher risk compared to multigravidae (OR 4.75, 95% CI 1.23-11.58). A history of fever within the prior week or fever at enrollment was associated with parasitaemia (4.2% vs. 2.3%, p=0.02). Importantly, 70.6% (48/68) of malaria-positive Prevention measures utilized Antenatal clinics n=1271 N (%) Delivery units n=870 N (%) Bed net in household 937(73.7) 643(73.9) Insecticide-treated bed net in household 43(3.3) 21(2.4) Sleeps under net most nights 873(68.6) 503(57.8) Taken malaria prophylaxis in pregnancy 9(0.7) 3 (0.3) Table 2: Use of malaria prevention measures by pregnant women attending antenatal clinics and delivery units Table 3: Parasitaemia, reported fever, and anaemia among pregnant women attending antenatal clinics and delivery units Antenatal clinics n=1271 N (%) Delivery units n=870 N (%) Peripheral Parasitaemia Overall 68(5.4) 37(4.3) Falciparum 3(0.23) 2(0.22) Vivax 59(4.6) 32(3.67) Mixed 6(0.47) 3(0.34) By gravidity Primigravid 21/423(4.9) 11/338(3.2) Secundigravid 38/ 578 (6.6) 15/209(7.1) Multigravid** 9/27 0 (3.3) 11/323(3.4) Report of fever within 1 week 167(13.1) 93(10.6) Anaemia 1093(86) 626(72) Severe anaemia 148/1093 (13.6) 49/626 ( 7.8 ) Factors at ANC Peripheral Parasitemia % (Positive/Total) Adjusted OR (95%CI) P Adjusted OR (95%CI) P 1 st /2 nd pregnancies 6.3 (64/1001) 4.45 (2.32 - 9.61) 0.0001 4.23 (2.15-8.42) 0.0001 3 rd or greater pregnancies 1.4 (4/270) 1 1 Age < 20 7.2 (12/166) 1.43 (0.34 - 3.76) 0.052 1.31 (0.26 - 2.84) 0.076 Age ≥ 20 5.0 (56/1105) 1 1 Fever within past week 16.1 (27/167) 4.42 (3.64 - 8.21) 0.002 4.62(3.73-9.83) 0.001 No fever within past week 3.7 (41/1104) 1 1 Bednet use* 7.6 (42/563) 1.12 (0.27 - 2.47) 0.072 1.37 (0.48 - 3.24) 0.084 No bednet use 6.7 (26/374) 1 Rural 7.1 (61/857) 4.21 (1.53 - 5.21) 0.003 4.36(2.48-7.32) 0.0001 No Rural 1.7 (7/414) 1 1 Tribal caste 6.3 (23/363) 1.26 (0.64 - 2.96) 0.054 1.42 (0.81 - 3.75) 0.12 No Tribal caste 4.9 (45/908) 1 1 No formal education 6.1 (22/357) 1.22 (0.42 - 2.46) 0.065 1.34 (0.68 - 3.92) 0.084 Formal education 5.0 (46/914) 1 1 Factors at DU 1 st /2 nd pregnancies 5.8 (32/547) 3.9 (0.97 - 11.56) 0.004 3.62 (0.94-7.83) 0.001 3 rd or greater pregnancies 1.5 (5/323) 1 1 Age < 20 8.2 (28/109) 2.32 (1.32 - 9.37) 0.062 2.47 (1.1710.63) 0.14 Age ≥ 20 3.6 (9/761) 1 1 Fever within past week 13.9 (13/93) 4.47 (1.25 - 12.42) 0.0001 4.43 (1.38 - 11.57) 0.0001 No fever within past week 3.1 (24/777) 1 1 Bednet use* 5.3 (27/503) 1.97 (0.83 - 7.62) 0.084 1.62 (0.58 - 6.39) 0.27 No bednet use 2.7 (10/367) 1 1 Rural 7.1 (29/405) 4.22 (0.41 - 4.51) 0.003 3.87 (0.78-13.62) 0.0001 No Rural 1.7 (8/465) 1 1 Tribal caste 5.5 (14/251) 1.51 (0.56 - 3.92) 0.053 1.74 (0.83 - 5.38) 0.59 No Tribal caste 3.7 (23/619) 1 1 No formal education 5.3 (17/321) 1.46 (1.23 - 3.17) 0.57 1.62 (0.87 - 4.63) 0.21 Formal education 3.6 (20/549) 1 1 women were asymptomatic. Most infections occurred between July and January, peaking during the monsoon season (August-October). Multivariate analysis confirmed that primigravidity, recent fever, and rural residence were independent predictors of parasitaemia (Table 4). Table-4 Factors associated with peripheral parasitemia during malaria in pregnancy (MIP) attending antenatal clinic (ANC) and delivery unit (DU) at Sadar Hospital, Hazaribag, Jharkhand, using univariate and multivariate analysis * ITN use was not evaluated in this model since these were very rarely used as well as quite lesser awareness about ITN among women. Incidences of asymptomatic Plasmodium vivax infection and its association with anaemia during pregnancy in malaria endemic population of Hazaribag, Jharkhand, India 1356 Delivery Units (DU) Similar demographic patterns were observed in the DU cohort: most participants were aged 20-36 years, formally educated, non-smokers (100%), and Hindi-speaking (97.2%) (Table 1). Most owned their homes (73.9%) and were primarily engaged in household work (84.3%). The median number of ANC visits was three (range: 0-9). Nearly two-thirds were primigravidae or secundigravidae. Bed nets were common, though ITN ownership remained rare (Table 2). Only three women reported malaria prophylaxis, none of whom could identify the drug. Peripheral parasitaemia was detected in 4.3% of DU participants. Among these, P. falciparum accounted for 5.4% (2/37), P. vivax for 86.5% (32/ 37), and mixed infections for 8.1% (3/37). The mean parasite density was 16,395 asexual forms/μl (range: 870-65,000). Primigravidae had significantly higher parasite densities than multigravidae (36,600 ± 9,743 vs. 7,532 ± 4,623 asexual forms/ μl, p=0.002). Asymptomatic infection predominated (75.7%, 28/37), while 24.3% were symptomatic. Parasitaemia was significantly associated with fever at enrollment (36.4% vs. 9.2%, p=0.005). Rural residence was again a significant risk factor (OR 3.46, 95% CI 1.29-10.4, p=0.03). Gravidity also influenced infection risk, with primigravidae and secundigravidae more likely to be parasitaemic (OR 4.23, 95% CI 1.97-23.2, p=0.04). Most infections occurred between July and September. Anaemia was strongly associated with parasitaemia: 83.7% of infected women were anaemic compared to 47.6% of uninfected (p=0.004), and severe anaemia was more common among infected women (5.7% vs. 2.6%, p=0.02). Group of Subjects Haemoglobin Range (g/dl.) Healthy Women Mean±SE (95%CI) MIP at ANC Mean±SE (95%CI) Women with Malaria Mean±SE (95%CI) MIP at DU Mean±SE (95%CI) P.vivax Infected Subjects at ANC Mean±SE (95%CI) P.vivax Infected Subjects at DU Mean±SE (95%CI) ≥11.5-14 12.3±0.1 (12.1-12.6) 12.1±0.1 (12.2-12.8) 12.2±0.08 (12-12.4) 12.2±0.2 (11.4-13) 12.5±0.1 (12.2-12.8) 12.4±0.1 (12.1-12.7) ≥10.3-≤11.4 11.2±0.06 (11-11.3) 10.4±0.1 (10.1-10.6) 11.2±0.08 (11.1-11.4) 11.3±0.09 (11.1-11.5) 10.8±0.1 (10.5-11) 11.3±0.1 (11-11.5) ≥8.4-≤10.2 10.2±0.07 (10-10.3) 9.6±0.1 (9.4-9.8) 9.8±0.06 (9.7-9.9) 9.6±0.07 (9.3-9.9) 9.6±0.07 (9.4-9.7) 9.4±0.08 (9.2-9.6) ≤8.3 8.2±0.1 (7.8-8.5) 7.9±0.1 (7.6-8.2) 8.2±0.1 (7.7-8.6) 8.4±0.1 (8-8.8) 7±0.3 (6.2-7.8) 7.8.5±0.3 (6.3-9.2) Table 5 - Prevalence range of haemoglobin levels in stratified groups during malaria in pregnancy and control groups at ANC and DU. Group of Subjects Classified Anaemia Healthy Women N (%) [95%CI] N=68 Malaria in Pregnancy At ANC * N (%) [95%CI] N=68 Women with Malaria N (%) [95%CI] N=68 Malaria in Pregnancy At DU * N (%) [95%CI] N=37 * Overall Malaria in Pregnancy N (%) [95%CI] N=105 Non - anaemic 18(26.4) [12.1-12.6] 13(19.1) [12.4-12.8] 13(19.1) [12-12.4] 4(10.8) [11.4-13] 17(16.1) [12.3-12.7] Mild anaemia 15(22) [11-11.3] 14(20.5) [10.1-10.6] 14(20.5) [11.1-11.4] 13(35.1) [11.1-11.5] 27(25.7) [10.6-11.1] Moderate anaemia 31(45.5) [10-10.3] 27(39.7) [9.4-9.8] 37(54.4) [9.7-9.9] 13(35.1) [9.3-9.9] 40(38.1) [9.4-9.7] Severe anaemia 4(5.8) [7.8-8.5] 14(20.5) [7.6-8.2] 4(5.8) [7.7-8.6] 7(18.9) [8-8.8] 21(20) [7.8-8.3] Overall Anaemia (%) 73.6 81 81 89 84 Table 6 - Frequency and status of anaemia in malaria during pregnancy among stratified cases at ANC and DU. Abhishek Mitra, Hasib Ansari, Birendra Kumar Gupta, Ajay Kumar Sharma & Mohammad Sohail 1363 malaria: a major cause of morbidity in early infancy. 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