Institutional Based Quantitative Cross-Sectional Study on Toxoplasmosis Sero-Prevalence, Awareness and Risk Behavior Among Pregnant Women Following Antenatal Care in Asella Teaching and Referral Hospital, Asella, Ethiopia 2018/19.

Background: Toxoplasmosis is caused by infection with the protozoan Toxoplasma gondii (T. gondii), an obligate intracellular parasite. The infections produced a wide range of clinical syndromes in humans, land and sea mammals, and various bird species. Toxoplasmosis is especially important among Immune-compromised patients and pregnant women. Infection of mothers during pregnancy by Toxoplasma gondii may have serious consequences for fetus ranging from miscarriage, central nervous system involvement, retinochoroditis, or at birth subclinical infection. Objective: To determine Toxoplasmosis sero-prevalence, awareness and risk behavior among pregnant women following antenatal care in Asella Teaching and Referral Hospital, Asella, Ethiopia. Methods: An institutional based quantitative cross sectional study design was conducted on sample size of 384 from December 1, 2018 to June 30, 2019. Structured, a pre-tested questionnaire was used to collect the demographic and risk factor related data. Serum sample, collected was tested for Toxoplasma Immunoglobulin G (IgG) and Toxoplasma Immunoglobulin M (IgM) antibodies using anti- IgG and IgM antibodies by Toxoplasma immune-chromatographic test (ICT) IgG-IgM test. A bivariable and multivariable logistic regression model was applied to show association between the dependent and independent variables considering P < 0.05 and the 95% condence interval. Result: The overall sero prevalence of T. gondii in the study area was 81.8 %. Three hundred and fourteen (81.8 %) of the pregnant women were IgG seropositive, fty women (13 %) were IgM seropositive. fty women were positive for both IgG and IgM. None of the pregnant women were positive exclusively for IgG and IgM ani-bodies. eating raw meat (COR=3.480, 95%CI: 1.450-8.352; P= 0.005, AOR=3.798, 95%CI: 1.249-11.550, P=0.19), using unpasteurized milk(COR=3.860,


Background
Toxoplasmosis is caused by the Toxoplasma gondii (T. gondii), an obligate, intracellular protozoan, a widespread zoonotic parasite which can infect all warm-blooded animals (1), and is one of the most common zoonosis in the world (2). Because of the current high rates of infection among the human population (estimated as 30-50%) it is considered a global health hazard (3). In Ethiopia the highest prevalence (95.1%) was reported from Butajira from patients found in 15-49 age groups (4), 94% prevalence among HIV/AIDS patients from Tikur Anbessa Specialized Hospital (5), and 85.4% among pregnant women in Addis Ababa (6). Toxoplasmosis is characterized by a wide range of clinical syndromes. The majority of infections is typically asymptomatic or causes a mild self-limiting ' u' like illness. Infection at primary gestation can be transmitted to the fetus and causes congenital toxoplasmosis and nally resulted in miscarriages, in utero death, retardation growth in neonate, ophthalmic and neurological diseases (7)(8)(9)(10).
The estimated global burden of congenital toxoplasmosis was 19,000 new cases each year resulting in the loss of approximately 1.2 million Disability-Adjusted Life Years (DALYs) (11).
Cats and other felines are the de nitive hosts of T. gondii and thus the only source of environmentally resistant oocysts in nature. From the major routes of infections for humans some of them are: close contact with cats or cat excreta ingestion of oocysts, eating of foods or soil contaminated with oocysts and drinking of contaminated water with oocysts; ingestions of tissue cysts due handling/consumption raw or undercooked meat of infected animals hosts, transplantation of infected organs, blood transfusion and congenital infection. Drinking of unpasteurized milk from infected animals and eating of raw egg can also transmit this parasite (9,(12)(13).
Antenatal screening for T. gondii infection is based detection of anti-toxoplasma speci c IgM and IgG is the backbone in controlling the risk of inherited toxoplasmosis. Maternal-fetal intervention can be achieved through drugs such as spiramycine which prevent congenital tox-oplasmosis by more than 60% (14).
Exposure rates to toxoplasmosis vary greatly to the topographical locality, socioeconomic prominence, traditional and spiritual opinions of a population (15). Therefore, there is a need to determine the prevalence in different settings.
Con rmation of zoonotic infection among cats and livestock animals of Ethiopia shows the presence of the parasite and hence the potential for human infection (16)(17)(18). Current trends in foods and lifestyle with fast foods substituting traditional cooking and popularity of cohort animals, (especially cats, as they require minimal care) and the plenty of stray cats, could increase the probability of experience to toxoplasmosis.
Prevention of congenital toxoplasmosis depends mainly on avoidance of risk factors during pregnancy. But, uncertainty about how most women acquire infection results in advice to avoid numerous risk factors which makes compliance di cult. Therefore, alertness is required with regard to toxoplasmosis and identi cation of signi cant associations between known risk factors and seropositivity to T. gondii among susceptible groups. This will be surely helpful in adopting appropriate prevention and control methods as accommodated for each group. In addition since, only few seroprevalence among females and neonates have been studied and there is no antenatal screening program for pregnant women for toxoplamosis in Ethiopia and also there is no studies have investigated the sero-prevalence, awareness of toxoplasmosis and related preventive behavior patterns among pregnant women in Aris zone, it necessities to have evidence concerning T. gondii in Arsi zone.
Therefore, the aim of this study is to determine Toxoplasmosis sero-prevalence, awareness and risk behavior among pregnant women following antenatal care in Asella Teaching and Referral Hospital, Asella, Ethiopia.

Methods And Materials
Study area and period A institutional based cross-sectional study design was done at Asella Teaching and Referral) Hospitals from December 01, 2018 to February 30, 2019. Asella Teaching and Referral Hospital found in Assela town, Arsi Zone, at 175 kilometers to the east of Addis Ababa, the capital city of Ethiopia.

Study design and period
An institutional based quantitative cross-sectional study design was conducted from December 01, 2018 up to February 30, 2019 to determine the sero-prevalence of toxoplasmosis, awareness and its associated risk factors in Asella Teaching and Referral Hospital,

Source population
All pregnant women in Asella referral and teaching Hospital catchment area.

Study population
All pregnant women following antenatal care in Asella referral and teaching Hospital.

Eligibility Criteria
Inclusion criteria: A pregnant woman who was come to selected Hospitals for the rst time for antenatal care during sample collection period for current pregnancy.
Exclusion criteria: Mothers who can't respond because of serious illness and who was come more than once during sample collection period to selected Hospital for antenatal service.

Sample Size Determination
The sample size was calculated to estimate the prevalence of toxoplasma antibodies in this population. For the calculation, prevalence was assumed to be 50% because there is no previous research done on this parasite among pregnant women in Arsi zone by considering a con dence interval of 95% and the acceptable difference to be 5%, the minimum required sample size was 384. The sample was collected by recruiting all consenting pregnant woman at their rst visit to the antenatal clinic of Asella Teaching and Referral Hospital.

Sampling Techniques
All pregnant women who were come to Asella Teaching and Referral Hospital for the 1 st time for antenatal care and willing to participate in the study were included until required sample size was achieved.

Socio demographic and clinical features
After clear training was given for ve midwifes, ve medical laboratory technologists and ve facilitators from who were selected from Asella teaching and referral hospitals data collection was started. To collect Socio demographic and clinical features a pretested questionnaire was administered by Midwifes and trained research assistants' information from pregnant women at selected hospitals antenatal care unit was gathered. The questions was produced information on four major areas: (a) socio-demographic information of the participants such as age, ethnicity, level of education, employment status, marital status, area of residence; (b) details of prior adverse birth outcomes, from multigravida i.e. miscarriages, still births, children with delayed milestones; (c) disease-related risk factors such as cat ownership, personal hygiene (frequency of hand washing), kitchen hygiene (frequency of washing utensils, cutting board and knife after preparation of meat), consumption of raw meat, cooking of meat, frequency of consuming unwashed raw vegetables and fruits, frequency of having non-homemade meals and exposure to soil; and nally, (d) the awareness of toxoplasmosis and sources of information. After implementation the questionnaire, each subject will be educated on signi cance and prophylactic actions against toxoplasmosis using an information sheet. in univariable logistic regression analysis was included in the nal multivariable logistic regression model to construct the likely model (P < 0 .05). The model will be reduced by backwards elimination of non-signi cant variables (P > 0.05) based on likelihood ratio test to de ne the model that would best t the data.

Dissemination of Research nding
The research ndings will be presented to the Arsi University and other stake holders, shared to Arsi Zonal Health Bureau, Regional Health Bureau, Federal Minister of Health (FMOH) and organizations/partners working on human health in the country. Finally effort will be made to publish the ndings on peer reviewed scienti c journal.
Signi cance and bene ciary of the study The nding will be signi cant both for researcher, communities and government in adding information regarding Toxoplasmosis sero-prevalence, awareness, and risk behavior among pregnant women following antenatal care in Arsi zone, Ethiopia. Nationally it develops the understanding of relationship between having cat, eating raw meat and vegetables, blood transfusion, organ transplantation, drinking contaminated water, eating soil and so on with congenital toxoplasmosis.

Expected Outcome
A wide range of data based on which rigorous generalizations and recommendations can be made based on the data was obtained from the diversity score survey and laboratory analysis of sample.

Quality assurance
Strict measures were taken throughout the analytic process. Five percent of the Questionnaire was pre-tested among pregnant women attending at Asella Referal and teaching Hospital (Asella town), and Questionnaire was revised accordingly. Data collectors were trained for 3 days on how to con duct the interview and the sampling process. Completed questionnaires were reviewed immediately to ensure accuracy and legibility. Quality control samples were tested parallel with the research samples and standard operating procedures were followed during the laboratory investigation.

Socio-demographic characteristics
This study included a total of 384 pregnant women with a response rate of 100%. Majority (65%) of them were urban dwellers and the mean age (± SD) of the study subjects were 26.2 (± 6.55) years. The majority (51%) of the study participants were found within the age group of 15-24.9 years. Almost all (98.2%) of the study participants were not have any information about Toxoplasmosis and its mode of transmission (Table-1).

Sero-prevalence and Risk factors for Toxoplasmosis
In this study, the serum sample was collected from a total of 384 pregnant women for the serological test, out of these, 314 (81.8%) were found to be positive for anti-bodies speci c to T. gondii. Furthermore, 264(68.8%) were found to be positive only for IgG; the rest 50(13 %) were positive both for IgM and IgG (

Discussion
Previously there wasn't known any information about the sero-prevalence of T. gondii in Arsi zone and this is the rst report of T. gondii infection in Asella, Ethiopia. The sero-prevalence of T.gondii among pregnant women in the study area was found to be 81.8 %(  25). The present discovery is also higher than the sero-prevalence of T. gondii in the lower altitude and higher humidly cities of Kenya: Kisumu (52%) and Mombasa (57%) (26). But silt lower than Bench Maji Ethiopia 85.3% (27) and Addis Ababa 85.4% (28).
Amongst the sero-positive females, the majority of them were found to have a chronic or past infection. However, fty women (50/314, 16 %) or 13% of the 384 women was found to have a recent infection (Table-2) which is higher than the studies conducted in Arba Minch Hospital 9(3.9%) and Mizan Aman General Hospital 7(3.0%) in Ethiopia (19,27 ). Bearing in mind the asymptomatic nature of the disease and the opportunity of congenital spread, the serologic nding of this study should not be overlooked. This is because the health -care facilities in the study area lack speci c tests for T. gondii and diagnosis isn't completely made which may result in misdiagnosis or delayed diagnosis.
In this study, there were 21 different risk factors have been also evaluated for the sero-prevalence of T. gondii but only : eating raw meat, drinking unpasteurized milk, eating raw egg and having three or more children (Table-1) were showed signi cant association. This nding is consistent with the studies conducted in Tanzania (22), Mali (29) and have of south of Iran (30).
In these ndings, contact with garden soil, owing to a domestic cat or presence of rat at home, and consumption of raw vegetables, inadequate washing of fruits and vegetables, educational status, contact with cat's feces and soil without gloves weren't showed signi cantly associated with T. gondii infection (Table-1) which contradicted other studies conducted in Ethiopia (19,27,30), China (32) and Brazil (33).This difference might be due to the differences in socioeconomic status, personal hygiene practices, feeding habits, differences in testing methods, variation immune staus of the study participants and sample size of each study.
Cats and dogs are thought to be the important ampli ers of infection of T. gondii (34). In the Ethiopian context, it is common to see domestic cats live and sleep to get her with human beings. Cats can directly contaminate humans, other animals, and their surrounding through their feces (35). Similarly, after contact with cat's feces, dogs can contribute to the spread of the organism through mechanical contamination of garden soil, vegetables, and human beings. Therefore, human beings can possibly acquire the infection or ingest the oocyst of the parasite through their dirtied hands after direct contact with cats or dogs, garden soil, and surfaces or ingesting of contaminated vegetables. Oocysts of T. gondii are hard free living phases of the parasite, and consequently are a major contributor to infections associated with the above-mentioned risk factors (34). Since they on the other hand, cats or dogs may acquire the infection from other warm blooded animals through carnivore contamination, and in sequence infect human beings (36).
This study was showed as the difference in educational status of the study participants have no effect on the seroprevalence, because only 1.8% was heard about Toxoplasmosis even though they were in different educational level and 98.2% doesn't have any information (table-1).This agrees with the study done in Hawassa and Yirgalem Hospitals southern, Ethiopia which indicated as 99.6% of the study participants was no information about Toxoplasmosis (37). But in reality it is extremely important to increase awareness on the transmission of toxoplasmosis at all level.
Even though it weren't showed signi cant association with Toxoplasmosis in this study, contact with: cat's feces, contaminated soil and water, raw or under cocked fruits and vegetables, gestational age, information about toxoplasmosis, abortion history and presence of rat and domestic cat's it is important to prevent these risk factors as it is evidence by different researches as these were some of the risk factors for Toxoplasmosis (9,27,32,(38)(39)(40)(41)(42). But some of current ndings agree with research done in northwest Ethiopia which showed as there are no any signi cant risk factors associated with sero-positivity in relation socio-demographic characters, gestational age, gravidity, consumption of raw vegetable, and blood transfusion (28,43).

Conclusions
The sero-prevalence of T. gondii antibodies was high among the pregnant women. Those who consumed raw: meat, milk and egg were at higher risk of T. gondii infection. Hence, blood screening for Toxoplasmosis, health education and awareness creation among women of reproductive age group in general and pregnant women in particular should be done during antenatal follow up. It is also alarming to add Toxoplasmosis screening during antenatal follow up. Moreover, there is need to control urban stray cat population to reduce the risk of zoonotic transmission of the parasite and further epidemiological studies to ascertain additional risk factors, economic and health impact of toxoplasmosis are called for. Availability of data and materials The data sets used and /or analyzed during the current study are available from the corresponding author or on reasonable request.

Con ict of interest
The authors declare that there is no con ict of interests.

Funding
The source of this research is Arsi University. The fund is used to buy reagents, kits, data collection and diagnosis only.
Authors' contributions SA provided conceptual framework for the project, data collection, guidance for interpretation of the data, perform data analysis, participated in the Parasitological work, performance of statically analysis and guidance for data interpretation. I also read and approved the nal manuscript.