Introduction
Infection with trematodes is important for the hygiene and health of humans and animals, and can negatively affect the agriculture and livestock economy [1]. Snails from the Lymnaeidae and Physidae families, as groups of the order Basommatophora, have an important role in the evolution and transmission of parasitic trematodes. About 18,000 species of trematodes use snails as their first intermediate host [2]. Snails of the Lymnaeidae family live in stagnant water with slow water flow, more oxygen, lower temperatures, and suitable vegetation. Forty species of snails from the Lymnaeidae family have been found, of which 7 have been identified in Iran, where the dominant species was Lymnaea auricularia [3]. Due to the average temperature, high annual rainfall, and high relative humidity, the area around the Caspian Sea is a very favorable habitat for the growth and reproduction of snails. The present study aims to determine the diversity and cercariae infection in the snails from the Lymnaeidae and Physidae families in Guilan province, northern Iran, and assess the effects of physicochemical parameters of water on their abundance and distribution.

Methods
Locating and sampling the freshwater snails from the Lymnaeidae and Physidae families was done using a cluster random sampling method from 117 regions in the aquatic habitats of Guilan province (reservoirs, canals, rivers and agricultural lands) from June 2021 to June 2022. The snails were identified based on morphometric identification keys [4]. Water temperature was measured and recorded with a mercury thermometer at the place and time of the sample. Water pH was measured using a pH meter; electrical conductivity (EC) was measured using an EC meter, and salinity was measured using a salinity meter. In the laboratory, the found snails were poured into a 100-mesh sieve and washed with a pestle to remove mud, foreign objects, and plants. To identify and recognize the morphology of snails, the right-handed and left-handed snails were first placed separately in a 24-well cell culture plate, and 1 mL of deionized water was added to it. A hole was created on the lid at the top of each well for air exchange. To remove the cercariae of the snails, the cell culture plates containing the snails were transferred to a refrigerated incubator at a temperature of 20 °C, and the removal of cercariae was stimulated with artificial light (10 watt LED lamp) [5]. To check the morphological characteristics of the cercariae, 10 μL of the liquid inside the wells was placed on the slide, and after adding 0.5% vital dye, it was covered with the slide. These samples were studied under an optical microscope and identified using the cercariae identification key.

Results
Out of 39486 snails, 19726 were lymnaea auricularia (49.96%), 4911 were lymnaea palustris (12.44%), and 14849 were Physa acuta (37.6%) (Table 1).


The prevalence of trematode cercariae infection in snails was 2.36%, (3.65% in lymnaea auricularia, 4.29% in lymnaea palustris, and none in Physa acuta). The identified cercariae were from the groups of Xiphidiocercariae (0.94%), Gymnocephalous (0.02%), Echinostome (0.8%), Lophocercous (0.16%), and Furcocercous (0.44%). The abundance of snails was 36.36% in the autumn, 31.39% in the summer, 22.25% in the winter, and 10% in the spring. The prevalence of infection in the snail lymnaea auricularia with Xiphidiocercariae was 1.58%; with Gymnocephalous, 0.04%; with Echinostome, 1.28%; with Lophocercous, 0.32%; and with Furcocercous, 0.43%. The prevalence of infection in the snail Lymnaea palustris with Xiphidiocercariae was 1.11%; with Echinostome, 1.39%; and with Furcocercous, 1.79%. There was no cercaria contamination in the snail Physa acuta. The one-year prevalence of infection in snails was 32.97% in spring, 28.28% in summer, 15.95% in autumn, and 22.8% in winter. The highest prevalence of infection with Xiphidiocercariae, Gymnocephalous, and Echinostome was in the summer (1.32, 0.04%, and 1.07%, respectively), while the highest prevalence of infection with Lophocercous and Furcocercous was in the autumn (0.19%) and the spring (1.78%), respectively. The variables of water temperature and waster pH had a significant negative relationship with the prevalence of cercariae infection in snails (P<0.05) (Table 2).


The abundance of snail had a significant positive relationship with the variables of water salinity and water EC (P<0.05) (Table 2).

Conclusion
Snails from the Lymnaeidae family are of great medical and veterinary importance because they play an important role in the life cycle of trematodes. In this study, the abundance of snails and the prevalence of infection in them were higher in the summer and autumn seasons than in winter and spring. Xiphidiocercariae, Gymnocephalous, Echinostome, Lophocercous and Furcocercous cercariae were isolated from the Lymnaeidae snails. In this study, two species of lymnaea auricularia and lymnaea palustris were identified from the Lymnaeidae family; from the Physidae family, Physa acuta was identified. In this study, water salinity had a significant direct relationship with the abundance of snail, while in another study, the increase in water salinity caused a decrease in the population of snails [6]. In our study, there was an negative significant relationship between the prevalence of cercariae infection in snails and water temperature, but  water temperature had no relationship with the abundance of snails. There was a significant relationship between water pH and the prevalence of infection, but water pH was not related to the abundance of snails. This findings are consistent with Soldanova’s report in 2010 [7]. In Mazandaran province of Iran, 3.9% of lymnaea auricularia snails were infected with cercaria trematodes from the Plagiorchiida, Diplostomidae, Clinostomidae and Echinostomatidae families. In the study by Imani et al [8] in 2013, dor Lymnaea gedrosiana snails, the prevalence of infection was reported 8.03% (81.98% with Xiphidiocercariae, 32.26% with Furcocercous, 5.19% with Echinostoma, and 1.24% with Monostomes) in the northwest of Iran [9]. In a recent study conducted in Guilan province, species of Lymnaea auricularia, Lymnaea gedrosiana, Lymnaea palustris, Lymnaea trancatula, Lymnaea stagnalis, Physa acuta and Planorbis species were identified, where infection with Gymnocephalus was observed in Lymnaea auricularia (0.66%) and Lymnaea gedrosiana (0.45%) [10]. This study in Guilan province with many water resources, environmental conditions and geographical areas suitable for the breeding of snails, and the occurrence of large epidemics of parasitic diseases can be important from a medical and veterinary point of view. More studies are recommended in this field.

Ethical Considerations
Compliance with ethical guidelines
This study was approved by the Ethics Committee of Urmia University (Ethics Code: IR.URMIA.REC.1400.002).

Funding
This study was funded by the Urmia University.

Authors' contributions
Study concept and design, acquisition, analysis, or interpretation of data, critical revision, administrative, technical, or material support, and study supervision: All authors; Drafting of the manuscript, funding acquisition: Armin Aligolzadeh Kenarsari and Mohammad Yakhchali; Statistical analysis: Armin Aligolzadeh Kenarsari, Mohammad Yakhchali, Keyhan Ashrafi Fashi.

Conflicts of interest
The authors declare that there is no conflict of interest.

Acknowledgments
The authors would like to express their gratitude to the Faculty of Veterinary Medicine of Urmia University and Guilan University of Medical Sciences for their cooperation in this study.


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