Seroprevalence of TORCH Agents in Women with Spontaneous Miscarriage in Jizan, Saudi Arabia

Cytomegalovirus, and Herpes Simplex Virus (TORCH). This study aimed to investigate the seroprevalence of TORCH agents in women with spontaneous miscarriage in Jizan, Kingdom of Saudi Arabia. Methods: A retrospective, cross-sectional study was conducted at a maternity hospital in the Jizan region. The study included 201 pregnant women who visited the hospital between 2018 and 2022. Blood samples were collected


INTRODUCTION
Abortion, defined as a pregnancy loss occurring before 20 weeks of gestation or when the fetus weighs less than 500 g, encompasses unintended, incomplete, and missed abortions. The term "abortion" now specifically refers to surgical or medical procedures performed for therapeutic or voluntary reasons (Kortsmit et al., 2020). Among the various complications affecting the fetus, infections with pathogens from the TORCH group are the most common causes (de Jong et al., 2013). Primary infections with certain TORCH pathogens during pregnancy, particularly in the first trimester, are associated with an increased risk of miscarriage, stillbirth, premature birth, congenital malformations, and fetal or neonatal transient or chronic diseases (de Jong et al., 2013).
TORCH infections, also known as perinatal or congenital infections, are caused by a group of organisms represented by the acronym TORCH: Toxoplasma gondii (TOX), Other (comprising parvovirus, hepatitis viruses, Epstein-Barr virus, human immunodeficiency virus, syphilis), Rubella virus (RV), Cytomegalovirus (CMV), and Herpes Simplex Virus (HSV) (Madrid et al., 2016, Neu et al., 2015. While the diagnosis and management of certain TORCH infections like hepatitis viruses, syphilis, and human immunodeficiency virus are clear and highly effective, others such as TOX, RV, CMV, and HSV often go unnoticed and are difficult to diagnose due to their asymptomatic nature and relatively low virulence, potentially leading to serious fetal consequences (de Jong et al., 2013).
The consequences of TORCH infections during pregnancy can range from intrauterine growth restriction, and congenital malformations, to fetal death, depending on the gestational age at the time of transplacental infection (Wang et al., 2019). While these infections generally cause mild illness in immunocompetent adults, they can have severe complications when acquired during pregnancy, posing risks to the fetus and newborn. Therefore, many countries include screening tests for these infectious organisms as part of routine prenatal care to determine women's immunological status at the beginning of pregnancy (Picone et al., 2020, Bobić et al., 2019. TORCH infections are vertically transmitted and contribute to significant congenital and neonatal morbidity and mortality worldwide (Liang et al., 2019). The traditional TORCH infections include TOX, RV, CMV, and HSV, while the "other" category encompasses various communicable pathogens like hepatitis B and C, HIV, syphilis, Chagas disease, Zika virus, varicella, and parvovirus B19 (Warnecke et al., 2020). In fact, up to 30% of stillbirths are attributed to infectious causes, although further research is needed to determine the global burden of these infections (Megli and Coyne, 2022).
The clinical outcomes of TORCH infections commonly include low birth weight, preterm birth, stillbirth, hearing and vision loss, and long-term neurological and developmental sequelae that can significantly impact affected children throughout their lives (Megli andCoyne, 2022, Warnecke et al., 2020). The exact mechanisms of placental infection and vertical transmission pathways are not yet fully understood, but studies have focused on pathogen invasion of the trophoblast, decidua, maternal capillaries, or other vasculature. Therefore, this study aims to investigate the seroprevalence of TORCH agents in women with spontaneous miscarriage in Jizan, Kingdom of Saudi Arabia.

MATERIALS AND METHODS Study Design and Setting:
This study was designed to be a retrospective and cross-sectional study. It was conducted at the maternity hospital in the Jizan region of Saudi Arabia. The study aimed to assess the frequency of TORCH infections among pregnant women in the region.

Study Population:
The study population consisted of pregnant women who visited a tertiary hospital in the Jizan region during the study period. The inclusion criteria included multigravida prenatal patients who were willing to participate. Pregnant women with a family history of congenital disorders and those with systemic ailments were excluded from the study.

Sample Size and Collection Technique:
A sample size of 201 pregnant women was collected using a convenience sampling technique. The selected participants represented a diverse group of pregnant women from the Jizan region.

Data Collection:
A retrospective analysis of medical records was performed for pregnant patients who visited a tertiary hospital in Jazan region from 2018 to 2022. Data on variables such as age, parity, Bishop Index, doses of misoprostol, and labor induction time were collected. The characteristics of TORCH screening in reproductive-age women were analyzed using logistic regression models.

Sample Collection and Laboratory Testing:
After obtaining verbal informed consent, pregnant women visiting the hospital for regular antenatal checkups provided blood samples (5-10 ml) along with clinical data. Previous pregnancy outcomes, including premature labor, low birth weight, microcephaly, intrauterine death (IUD), and congenital defects, were documented. The serum was separated through centrifugation and stored frozen. TORCH agents IgG and IgM antibodies (T. gondii, rubella, CMV, and HSV types 1 and 2) were estimated using commercially available indirect ELISA kits following the manufacturer's instructions.

Statistical Analysis:
The collected data, including TORCH IgG and IgM ELISA results, were coded and analyzed using SPSS Version 22 (IBM Corp., Armonk, NY, USA). Descriptive statistics were generated, and the strength of the relationship between variables was measured using appropriate statistical tests. The correlation between variables of interest was assessed using the chi-square test. A significance level of p < 0.05 was regarded as statistically significant at the 5% level.

RESULTS
The sociodemographic characteristics of the pregnant women (n=201) were revealed in Table 1. In terms of age, the majority of them fell within the 31-40 age range, comprising 49.3% of the sample, followed by the 21-30 age group at 34.8%. A smaller percentage of participants belonged to the 15-20 age group (2.5%), while those above 50 years old accounted for only 0.5%. In relation to educational level, the largest proportion of pregnant women were graduates (54.2%), with the smallest proportion as illiterate (2.5%). Examining occupation, the most common occupation reported by pregnant women was being a housewife (   Cytomegalovirus, there were no significant differences in IgG (p=0.968) and IgM (p=0.499) seropositivity across the years, with IgG ranging from 60.3% to 64.3% and IgM ranging from 0% to 7.9%. In terms of Herpes 1, there were no significant differences in IgG (p=0.738) and IgM (p=0.832) seropositivity across the years, with IgG ranging from 86.3% to 92.3% and IgM ranging from 0% to 1.3%. Lastly, for Herpes 2, no significant differences were found in IgG (p=0.924) and IgM (p=0.826) seropositivity across the years, with IgG ranging from 91.7% to 96.2% and IgM ranging from 0% to 1.4%.  ranging from 9% to 16.7%. Regarding Rubella, no significant differences were observed in both IgG (p=0.120) and IgM (p=0.732) seroprevalence across age groups, with IgG ranging from 71.1% to 100% and IgM ranging from 4% to 10%. For Cytomegalovirus, there was a significant difference in IgM seroprevalence across age groups (p=0.003), ranging from 27.8% to 60%. However, there were no significant differences in IgG seroprevalence (p=0.760), ranging from 38.9% to 98.1%. In terms of Herpes 1, no significant differences were found in IgG (p=0.808) and IgM (p=0.792) seroprevalence across age groups, with IgG ranging from 85.3% to 100% and IgM ranging from 0% to 1.4%. Lastly, for Herpes 2, no significant differences were observed in IgG (p=0.859) and IgM (p=0.778) seroprevalence across age groups, with IgG ranging from 91.8% to 100% and IgM ranging from 0% to 1.5%. Table 4: Seroprevalence of TORCH IgG and IgM antibodies among women with abortion based on women's age: (n=201). Table 5 presents the presence of TORCH IgG antibodies and hematological parameters (Hb, PCV, and RBCs) among women with abortion (n=201). For Toxoplasmosis, there were no significant differences in the distribution of low, normal, and high levels of Hb (p=0.252), PCV (p=0.012), and RBCs (p=0.052) based on IgG positivity. In Rubella, there were no significant differences in the distribution of hematological parameters based on IgG positivity. However, there was a significant association between IgG positivity for Cytomegaly virus and the distribution of low, normal, and high levels of Hb (p=0.004) and PCV (p=0.011). For Herpes 1 and Herpes 2, no significant associations were observed between IgG positivity and hematological parameters. It is important to note that the p-values indicate the level of statistical significance for the associations observed . The presence of TORCH IgM antibodies and hematological parameters (Hb, PCV, and RBCs) among women with abortion (n=201) is indicated in Table 6. For Toxoplasmosis, there were no significant differences in the distribution of low, normal, and high levels of Hb (p=0.912), PCV (p=0.403), and RBCs (p=0.053) based on IgM positivity. In Rubella, there were no significant associations between IgM positivity and hematological parameters. Similarly, for Cytomegaly virus, no significant associations were observed between IgM positivity and hematological parameters. For Herpes 1 and Herpes 2, no significant associations were found between IgM positivity and hematological parameters. It's important to note that the p-values indicate the level of statistical significance for the observed associations. The association between demographic variables and TORCH IgG status among women with abortion (n=201) revealed significant associations with gestational period and multiple gestation for certain TORCH infections (Table 7). Specifically, in the 1st trimester, a higher percentage of women tested positive for Rubella IgG compared to the negative group, while a higher percentage of women tested negative for Herpes 1 IgG compared to the positive group. Moreover, the negative group showed a higher percentage of women with multiple gestations compared to the positive group for Cytomegaly virus IgG and Herpes 1 IgG. These findings suggest that the gestational period and multiple gestation may play a role in the development of TORCH IgG antibodies among women with abortion, emphasizing the importance of considering these factors in understanding TORCH infections. Table 7: association between the demographic variables of participants and the TORCH IgG: (n=201).

DISCUSSION
The present study aimed to investigate the seroprevalence of TORCH agents (Toxoplasma, Rubella, Cytomegalovirus, and Herpes) in women with spontaneous miscarriage in Jizan, Kingdom of Saudi Arabia. The study provides valuable insights into the seroprevalence patterns and associations of TORCH infections in this specific population. To contextualize these findings, comparisons were made with previous studies to identify similarities and contrasts in seroprevalence rates and risk factors associated with TORCH infections.
Regarding the sociodemographic characteristics of pregnant women, the distribution of age groups, educational levels, occupations, and exposure to cats was consistent with previous studies (Surpam et al., 2006, Prasoona et al., 2015. This suggests that the sample population in this study is representative of pregnant women in terms of these demographic factors. However, it is important to acknowledge that the study's generalizability may be limited to the specific region of Jizan.
Analyzing the seropositivity of TORCH IgG and IgM antibodies among cases of miscarriage, the study found varying rates for different agents. Toxoplasma and

Rubella
showed relatively higher seropositivity rates compared to Cytomegalovirus and Herpes. These findings align with previous studies, indicating consistency in seroprevalence rates across different populations (Prasoona et al., 2015, Acharya et al., 2014. For example, the seroprevalence rate of Toxoplasma IgG antibodies (40.3%) in this study falls within the range reported in other investigations (Paulson et al., 2021). Similarly, the rates for Rubella IgG (24.9%) and Cytomegalovirus IgG (28.9%) antibodies are consistent with previous findings (Mohymen et al., 2009, Adgoy et al., 2020. The lower seroprevalence rates observed for Herpes 1 and Herpes 2 antibodies are also in line with existing literature (de Jong et al., 2013, Wang et al., 2019 . Comparing the seroprevalence rates of our study with previous studies, this study provides further evidence of the global concern posed by TORCH infections during pregnancy. The findings reinforce the importance of screening and prevention strategies to mitigate the risks associated with these infections.
Despite the similarities in seroprevalence rates, some contrasting findings emerged when comparing this study with previous investigations. Notably, this study did not find significant associations between TORCH seropositivity and age groups or gestational periods, which differs from certain previous studies (Surpam et al., 2006, Wang et al., 2019. These contrasting results suggest that the influence of age and gestational period on TORCH seroprevalence may vary across different populations and regions. It is important to acknowledge that this study was limited to a specific geographic area, and further research in diverse populations is necessary to obtain a more comprehensive understanding of TORCH seroprevalence and its associated factors. Another contrasting finding was the significant associations between TORCH infections and multiple gestations observed in this study. Specifically, a higher percentage of women with multiple gestations tested positive for Cytomegalovirus and Herpes 1 IgG antibodies compared to the negative group. This contrasts with some previous studies that did not find a significant association between multiple gestations and TORCH infections (Al-Saeed et al., 2015). These contrasting findings emphasize the complex nature of TORCH infections and the need for further exploration of potential risk factors.
There are several limitations of this study. Firstly, the study sample consisted of women with spontaneous miscarriages from a specific region in Saudi Arabia (Jizan), which may limit the generalizability of the results to other regions of the king §dom. Secondly, the study relied on serological testing for the detection of TORCH antibodies, which may not always accurately reflect the current infection status or the exact timing of the infections. Additionally, the study did not investigate other potential risk factors or confounding variables that could contribute to the development of TORCH infections or miscarriages.

Conclusion:
In conclusion, this study shed light on the seroprevalence of TORCH agents (Toxoplasma, Rubella, Cytomegalovirus, and Herpes) among women with spontaneous miscarriage in Jizan, Kingdom of Saudi Arabia. The results indicate varying rates of seropositivity for different TORCH agents, with Toxoplasma and Rubella showing relatively higher seroprevalence compared to Cytomegalovirus and Herpes. These findings are consistent with previous studies conducted in different populations, highlighting the global concern associated with TORCH infections during pregnancy. Future studies with larger and more diverse populations, considering a wider range of variables and employing more comprehensive diagnostic methods, are recommended to further investigate the complexities of TORCH infections and their implications in cases of spontaneous miscarriage. Ethical Approval: This study was commenced after acquiring approval from Jazan Health Ethics Committee, Ministry of Health (Approval number: 2303). The study was accomplished according to the guidelines of the Declaration of Helsinki. Informed consent of all participants was obtained and the identity of the participants was well protected. Funding: This study was supported by the Deputyship for Research and Innovation, Ministry of Education in Saudi Arabia under the project number MOE-BU-1-2020.