Abstract
Fascioliasis and toxoplasmosis are the two important zoonotic diseases that are endemic in Iran and share some common transmission routes. The present study is aimed at determining the seroprevalence of human fascioliasis and toxoplasmosis in rural and urban areas of Jolfa County, Northwest Iran. In a cross-sectional study, 600 human sera were collected randomly from humans living in Jolfa County including three cities and 13 villages from 2017 to 2018. Anti-Toxoplasma IgG and anti-Fasciola sp. IgG tests have been performed using the enzyme-linked immunosorbent assay. Four (0.7%) out of 600 human sera showed positive levels of anti-Fasciola IgG. Three out of four seropositive humans were from an urban area, and one (25%) was from rural inhabitants. Considering T. gondii infection, 45% of studied human sera were seropositive for anti-T. gondii IgG. In conclusion, this is the first study reporting Fasciola seropositivity in the area. Based on the findings, human fascioliasis is present in the studied area, Northwest Iran, granted in low prevalence. Considering T. gondii seropositivity, the prevalence is high, yet close to the reports from other regions in the province.
1. Introduction
Toxoplasma gondii and Fasciola spp. are completely different parasites; the first one is an obligate apicomplexan intracellular parasite, and the latter is a genus of trematodes or flukes. They are entirely unlike in many aspects; however, both are important zoonotic parasites that are food- and/or waterborne and share some common transmission routes [1]. Toxoplasmosis and fascioliasis have worldwide distribution among animals and humans.
T. gondii has a wide spectrum of intermediate hosts and infects almost all vertebrates including humans [2]. T. gondii is known to be transmitted by meat-containing tissue cysts (meatborne) and vegetables and fruits (plantborne) contaminated by oocysts [3], consistent contact with soil [4], and cats. Additionally, transmission can also occur congenitally [5] and by organ transplantation [6].
It is estimated that approximately one-third of the human population on the planet is infected by T. gondii [2]. Nevertheless, the infection with T. gondii is benign in most cases, being asymptomatic or in some patients with signs such as cervical lymphadenopathy or ocular disease, but immunocompromised individuals and fetuses may be severely diseased [2]. Congenital toxoplasmosis can result in devastating consequences for the fetus such as miscarriage, stillbirth, and neonate with ophthalmic and neurological disorders and also long-term effects after birth [5]. In adults, it is believed to be benign, but behavioral changes are thought to occur in chronically infected individuals [7]. In immunocompromised patients such as patients with AIDS, it can be life-threatening causing HIV-related toxoplasmic encephalitis [8]. The routine diagnosis is based on detecting antibodies against T. gondii; however, molecular diagnosis is available [9].
Fascioliasis on the other hand is a well-known plant- and waterborne zoonotic parasitic disease of herbivores such as ruminants, equids, and camelids but also omnivore mammals such as swine with economic importance and humans with high medical importance [10]. It is caused by two species, F. hepatica and F. gigantica, that parasitize the bile ducts in the liver and the gallbladder of ruminants and humans as the definitive hosts. Fasciola spp. have a life cycle involving snail (genus Radix and Galba) as an intermediate host. A resistance phase named metacercaria occurs on the aquatic plants or in the water is a stage that causes human and animal infections commonly by eating freshwater wild or cultivated plants and water containing metacercaria [11].
F. hepatica is endemic in Europe, Asia, Africa, the Americas, and Oceania, and F. gigantica is found in Africa and Asia [10]. Fascioliasis is an emerging neglected zoonotic infection affecting the health and quality of life in humans. The human infection ranges from asymptomatic to acute or chronic fascioliasis. Nonetheless of the clinical manifestation, the infection can be linked to long-term complications such as liver damage, anemia, and malnutrition [12]. During the prepatent period (early infection), antibody detection by immunological techniques such as ELISA is the only tool available for diagnosis. In the chronic phase, antibody detection, antigen detection, and stool microscopy for detecting eggs are helpful [13].
Iran is one of the endemic countries for human fascioliasis [14] and toxoplasmosis [15], especially in the Caspian Sea basin (North of Iran); both infections are in the highest prevalence among the human population [14, 15]. In 1989 in Gilan Province (north of the country), thousands of individuals had signs of fascioliasis. It was the first largest outbreak of human fascioliasis in the world infecting about 10000 humans. Ten years later, the second outbreak affecting 5000 people occurred in the same region [14]. Considering toxoplasmosis, the seroprevalence is reported as high as 64.7% [16] and 74.6% [17] in the Caspian Sea basin (North of Iran).
It is necessary to have updated information about endemic infectious diseases in every region to be armed for future unwilling events. Because there is no comprehensive study on the seroprevalence of human fascioliasis and toxoplasmosis in Jolfa District, Northwest Iran, the present study is aimed at determining the prevalence of the infection in the area.
2. Materials and Methods
The present study was carried out on the serum samples collected for the previously published study on the seroprevalence of cystic echinococcosis in Jolfa County [18], which was reconsidered and approved by the Vice-Chancellor of Research and Technology, Urmia University of Medical Sciences, Urmia, Iran (research project number: 2855, ethical code: IR.UMSU.REC.1400.148).
2.1. Study Area and Sampling
Jolfa County is located in the north of East Azerbaijan Province with an area of 1670 km2, Northwest Iran. It is located on the south border of Aras River (Figure 1). It is restricted to two countries, Azerbaijan and Armenia. The county’s population is estimated at around 61358 humans, 44704 in urban and 16654 individuals in rural districts [18]. The Jolfa County has a semiarid climate.
2.2. Sample Collection
This cross-sectional study was conducted with 600 human blood samples collected randomly from humans living in Jolfa County including three cities and 13 villages from 2017 to 2018. The samples were collected from cities including Hadishahr 238 (Central 82, South 79, and East 77), Jolfa 96, and Siyahrud 32 sera and villages including Komar-e Sofla 6, Daran 30, Kordasht 6, Ushtabin 49, Nowjeh Mehr 20, Marazad 20, Qarah Bolagh 23, Luvarjan 10, Ersi 24, Shoja 15, and Iri-ye Sofla 31 samples. Sera were separated from blood and kept frozen at -20°C until the examination.
2.3. Determining Anti-Toxoplasma and Anti-Fasciola IgG
Anti-Toxoplasma IgG and Anti-Fasciola IgG testing have been performed using the enzyme-linked immunosorbent assay (ELISA kits; Pishtaz Teb, Iran). The tests were carried out based on the manual of the ELISA kits. The sensitivity of 92% and specificity of 93% for anti-Fasciola IgG and sensitivity of 100% and specificity of 99% for anti-Toxoplasma IgG are claimed by the company. The kits contained positive and negative controls.
2.4. Geographic Information System (GIS)
A GIS map of East Azerbaijan Province locating Jolfa County stating sampling areas and T. gondii seropositivity was built using ArcGIS v. 10.6 software.
2.5. Data Analysis
Data were analyzed by SPSS version 23 (IBM SPSS Statistics for Windows, version 27.0, Armonk, NY: IBM Corp) using respective tests depending on the nature of the data including chi-square, Mann–Whitney, and binary logistic regression tests. value < 0.05 was considered significant.
2.6. Limitations of the Study
Analysis of some risk factors related to toxoplasmosis and fascioliasis was a limitation of the present study. These data were not available in the questionnaires, and they could not be retrieved to include in the analysis, such as contact with cats, eating raw meat, and living with livestock.
3. Results
3.1. Studied Population
Among 600 humans, 352 were female (58.7%) and 248 were male (41.3%). The mean age of the participants was 40.43 years (, range 2-90 years, ).
3.2. Anti-Fasciola IgG
Four (0.7%) out of 600 human sera including three females and a male showed positive levels of anti-Fasciola IgG. Furthermore, three (0.8%) out of 366 urban (all from southern Hadishahr) and one (0.4%) out of 234 rural (from Daran) inhabitants were seropositive for Fasciola spp. Because the number of seropositive humans was low, further data analysis could not be performed. The demographics of positive cases are available in Table 1 as descriptive data.
3.3. Anti-Toxoplasma IgG
The positive levels of anti-Toxoplasma IgG were observed in 270 (45%) human sera. Considering the sex distribution, the seropositivity for T. gondii infection was found in 161 (45.7%) and 109 (44%) females and males, respectively. There was a significant relationship between T. gondii IgG seropositivity and contact with soil (), and living in rural areas () (Table 2). In addition, two out of four (50%) Fasciola-infected individuals were coinfected with T. gondii.
The lowest T. gondii seropositivity was observed in the southern region of Hadishahr, where three out of four cases of fascioliasis were detected, and the highest seroprevalence was observed in Komar village located in the central region of Jolfa County (Table 3 and Figure 1).
The mean age of T. gondii seropositive individuals (44.66 years) was significantly higher () compared to seronegative ones (36.96 years).
The IgG concentration was significantly different in different occupations, higher in males and humans that washed raw consumed vegetables with water without using respective disinfectants (Table 4).
4. Discussion
In the present study, the seropositivity for anti-Fasciola IgG was 0.7%, the majority of whom were from the same area at the moment of sampling, the southern region of Hadishahr. This may reflect an endemic focus on human fascioliasis in the southern part of the city. Considering T. gondii, 45% of the studied humans were IgG seropositive. Also, half of Fasciola-infected individuals were coinfected with T. gondii.
Jolfa County has a semiarid climate suitable for the development of Fasciola spp. From different regions of East Azerbaijan Province, there are studies reported Fasciola hepatica from livestock [19], but there was no data on the prevalence of Fasciola infection in humans in the province, and the present study is the first report of human fascioliasis in Jolfa County, Northwest Iran. The highest human seroprevalence was believed to be reported from a region in Gilan Province (50%). The highest prevalence rate of fascioliasis in animals is reported in the north, and the lowest prevalence is in Central Iran [20].
Heydarian et al. [21] investigated the seroprevalence of human fascioliasis in Lorestan Province, Western Iran, on 1256 humans. They detected anti-Fasciola antibodies in 16 individuals (1.3%). Also, no significant differences were reported between infection and age groups, sex, level of education, and occupation; yet significant differences were recorded regarding the location of residency, consuming local freshwater plants, and water sources with seropositivity [21]. In another similar study in the same region, Lorestan Province, Eshrati et al. studied seroepidemiology of human fascioliasis on 1053 humans. They reported Fasciola seropositivity in 28 humans (2.66%) among which 18 were females [22]. In the present study, the seropositivity is considerably lower than in the West of Iran. However, in 2016, there was a report from a village in Lorestan Province that resulted in 0.7% of human fascioliasis (6 out of 801) [23], which is exactly similar to the findings of the present study.
There are studies on the seroprevalence of fascioliasis that are reported from different regions of Iran, such as Gorgan, Northeast Iran in 2020, 1.79% (11 out of 612) [24]; Yasuj, Southwest Iran in 2012, 1.8% (18 out of 1000) [25]; Meshkin Shahr (in neighbouring province to ours), Northwest Iran in 2013, 1.96% (9 out of 458) [26]; and Isfahan, Central Iran in 2014, 1.7% (8 out of 471) [27]. Also, human fascioliasis is reported from Mashhad, Northeast Iran [28], and Sistan and Baluchestan Province, Southeast Iran [29]. In Alborz Province, fascioliasis was reported to be significantly lower in patients hospitalized (2.2%) due to COVID-19 compared to healthy controls (4.3%) [30]. In a meta-analysis, the overall prevalence of human fascioliasis in Iran is estimated as 2% (95% CI 1-5) [31].
As it is obvious, human fascioliasis is endemic in most regions of Iran, granted in low prevalence; however, in some areas such as the Caspian Sea basin (North of Iran), there were some outbreaks in the past [14]. Thus, the Jolfa area is not an exception, and based on the results of the present study, human fascioliasis is present in Jolfa County, Northwest Iran, with a close prevalence rate to most of the regions in the country.
Toxoplasmosis is also prevalent in Iran with a wide range of prevalence, higher in warm and humid climates (Caspian Sea basin) and lower in the desert, arid, and colder areas. In a meta-analysis, the overall prevalence of T. gondii infection is estimated at 39.3% [15]. In the Jolfa area, only one study could be retrieved by searching databases, which was published in 2005 in the Farsi language by Fallah et al. [32]. They studied anti-T. gondii IgG in high school girls in Jolfa using indirect immunofluorescence antibody (IFA) test and reported a prevalence of 21.8%. They also found a significant relationship between having contact with the cat, eating raw liver, and signs (lymphadenopathy, fever, and skin rash) of T. gondii infection. In the present study, anti-Toxoplasma IgG was observed in 45% of the studied population which is considerably higher than the prevalence reported by Fallah et al. [32]. Furthermore, a significant relationship between T. gondii infection and contact with soil and living in rural areas was observed. The lowest T. gondii seropositivity was observed in the southern region of Hadishahr, where three out of 4 cases of fascioliasis were also detected, and the highest seroprevalence was observed in Komar Village located in the central region of Jolfa County. The mean age of T. gondii-seropositive individuals was significantly higher compared to seronegative ones. In Urmia (Northwest Iran), the seroprevalence of feline toxoplasmosis was reported using a modified agglutination test in 35.3% of stray and household cats [33].
The IgG concentration was significantly different in different occupations (highest in farmers and lowest in students), higher in males and humans that washed raw consumed vegetables with water without using respective disinfectants. This may show a probable relationship between a load of oocysts entering the body with a level of IgG. Contact with cats and eating raw livestock liver were not studied in the present study.
In a study carried out by Rajaii et al. on the seroprevalence of T. gondii infection in women of childbearing ages in different regions of East Azerbaijan Province, Northwest Iran, using the IFA test, they studied 1659 women from Tabriz, Maragheh, Ahar, Marand, Sarab, and Mianeh. Of 1659 women, 899 (54.13%) were reported seropositive. They also reported a direct linear relationship between seropositivity and age and higher seropositivity in subjects with lower educational levels and those living in rural regions [34]. In the present study, there was also a similar pattern of some risk factors in Jolfa County, for example, the higher prevalence in rural residents, illiterates, and humans with consistent soil contact and lower prevalence in students, which may show a somehow uniform transmission pattern in East Azerbaijan Province.
5. Conclusion
This is the first study reporting Fasciola spp. seropositivity in Jolfa County. The southern region of Hadishahr is the area with the highest Fasciola seropositivity, which is concerning regarding the health care system to monitor the area for the infection source to be ready and armed with information on any potential future outbreaks in the area. Considering T. gondii seropositivity, the prevalence is high, yet close to the reports from other regions in the province. Furthermore, having contact with soil, living in rural areas, having higher age, and being illiterate increase the risk of being tested seropositive for T. gondii in Jolfa County.
Data Availability
Data in SPSS format is available and presented by contacting the corresponding author in case of a request.
Ethical Approval
All methods in the present study were carried out under relevant guidelines and regulations of the Iran National Committee for Ethics in Biomedical Research. The study was approved by the Ethics Committee of Urmia University of Medical Sciences under the ethical code IR.UMSU.REC.1400.148 and research no. 2855.
Conflicts of Interest
The authors have no competing interests.
Authors’ Contributions
S. Z. conducted laboratory work and prepared the data. S. K., E. Y., G. S., N. A., and S. M. contributed to the previous study on the sera that were studied in the present study (Sakhaei et al. [18]) and contributed to collecting samples and filling out questionnaires. R. J. supervised the study, confirmed the diagnoses, analyzed the data (in cooperation with S. M.), and prepared the manuscript.
Acknowledgments
The authors would like to acknowledge the Vice-Chancellor of Research and Technology, Urmia University of Medical Sciences, Urmia, Iran, for approval and financial support (research no: 2855, ethical code: IR.UMSU.REC.1400.148).