Ashrafi et al. BMC Infectious Diseases (2020) 20:28 https://doi.org/10.1186/s12879-020-4762-0 RESEARCH ARTICLE Open Access Zoonotic transmission of Teladorsagia circumcincta and Trichostrongylus species in Guilan province, northern Iran: molecular and morphological characterizations Keyhan Ashrafi1, Meysam Sharifdini1* , Zahra Heidari2, Behnaz Rahmati1 and Eshrat Beigom Kia3 Abstract Background: Parasitic trichostrongyloid nematodes have a worldwide distribution in ruminants and frequently have been reported from humans in Middle and Far East, particularly in rural communities with poor personal hygiene and close cohabitation with herbivorous animals. Different species of the genus Trichostrongylus are the most common trichostrongyloids in humans in endemic areas. Also, Ostertagia species are gastrointestinal nematodes that mainly infect cattle, sheep and goats and in rare occasion humans. The aim of the present study was to identify the trichostrongyloid nematodes obtained from a familial infection in Guilan province, northern Iran, using morphological and molecular criteria. Methods: After anthelmintic treatment, all fecal materials of the patients were collected up to 48 h and male adult worms were isolated. Morphological identification of the adult worms was performed using valid nematode keys. Genomic DNA was extracted from one male worm of each species. PCR amplification of ITS2-rDNA region was carried out, and products were sequenced. Phylogenetic analysis of the nucleotide sequence data was performed using MEGA 6.0 software. Results: Adult worms expelled from the patients were identified as T. colubriformis, T. vitrinus and Teladorsagia circumcincta based on morphological characteristics of the males. Phylogenetic analysis illustrated that each species obtained in current study was placed together with reference sequences submitted to GenBank database. Conclusions: The finding of current study confirms the zoonotic aspect of Trichostrongylus species and T. circumcincta in inhabitants of Guilan province. The occurrence of natural human infection by T. circumcincta is reported for the first time in Iran and the second time in the world. Keywords: Trichostrongylus colubriformis, Trichostrongylus vitrinus , Teladorsagia circumcincta, Human, Iran Background Trichostrongyloidae, Trichostrongylus is the most com- Trichostrongyloid nematodes (Nematoda: Trichostron- mon genus infecting humans in endemic regions [1]. gyloidae) are significant parasites of the digestive tract of Other genera of trichostrongyloid nematodes such as herbivorous animals. Various genera of these nematodes Haemonchus contortus, Marshallagia marshalli, Nema- are prevalent in ruminants in different countries todirus abnormalis, Ostertagia ostertagi and O. circum- throughout the world [1]. They have an important veter- cincta (Teladorsagia circumcincta) have been reported inary role due to their impact on animal health and se- in humans, particularly in Iran and Azerbaidjan, of vere economic losses in herbivores [1]. Among the former Soviet Union [2–5]. Trichostrongylus species are primarily parasites of live- stock with a worldwide distribution but they have also * Correspondence: [email protected]; [email protected] 1Department of Medical Parasitology and Mycology, School of Medicine, been reported from humans in Middle and Far East and Guilan University of Medical Sciences, Rasht, Iran Africa, particularly in rural communities with poor Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Ashrafi et al. BMC Infectious Diseases (2020) 20:28 Page 2 of 9 personal hygiene and close cohabitation with herbivor- trichostrongyloid infections [22]. However, classification ous animals [2, 4, 6–8]. Epidemiological studies have of Trichostrongylus species and other genera of trichos- demonstrated trichostrongylosis as a prevalent parasitic trongyloid nematodes are based on differences in the infection in domestic ruminants and humans in different morphology of male worms [19]. regions of Iran [2, 9]. Prevalences of 53.6, 46.8, 42.4, Recently, PCR-based tools are applied for reliable iden- 38.7, 37 and 35.8% have been reported in ruminants tification and phylogenetic analysis of nematodes. Based from Isfahan, Khuzestan, Mazandaran, Kermanshah, on the recent studies, the internal transcribed spacer 2 Hormozgan and West Azerbaijan provinces and 71, 67, (ITS2) region of ribosomal DNA is a useful tool for ana- 42, 37, 19, 8, 2 and 1% in humans in Khuzestan, Isfahan, lyzing genetic variations and phylogenetic relationships Tehran, Hormozgan, Kermanshah, Mazandaran, West in nematodes belonging to the Trichostrongylidae family Azerbaijan and Sistan & Baluchestan provinces in the [6, 7, 20, 21]. The aim of the present study was identifi- past decades respectively [2]. Recent studies indicate a cation, molecular characterization and phylogenetic ana- decreasing trend in the prevalences of trichostrongylosis lysis of human specimens of Trichostrongylus and in both animals and humans in Iran. However, human Teladorsagia obtained from the patients during a famil- trichostrongylosis is still a health problem in some re- ial infection in Langroud district of Guilan province gions of the country. The prevalences of 18.1% [9], based on ITS2 region of ribosomal DNA. 3.05% [6] and 2.1% [10] have been reported from Khou- zestan, Guilan, and Mazandaran provinces in recent Methods years respectively. Study area Until now, ten species of the genus Trichostrongylus Guilan province is one of the 31 provinces of Iran located have been reported from humans in various parts of Iran, at the southern littoral of Caspian Sea in northern Iran which is unique in the world when the number of col- and is comprised of sixteen districts. Langroud district lected species is taken into consideration. Among these with an area of 480 km2 (37° 11′ 49″ N, 50° 9′ 13″ E) is species, T. colubriformis and T. orientalis were more com- located at the eastern part of the province (Fig. 1). Overall, mon in past decades in the country [2, 6, 11–13], but re- 140,686 (70,675 males and 70,011females) were recorded cent studies in the north of Iran have shown T. to live in 49,351 families in the study region in 2016. colubriformis as the predominant species in infected indi- viduals [6, 7, 14, 15]. Humans mostly acquire the infection Sample collection and morphological identification through consumption of fresh vegetables contaminated Seven members of a family living in Langroud district, with infective filariform larvae [6, 7]. Trichostrongylosis is located at the eastern part of Guilan province, were diag- usually asymptomatic in low intensity infections, but ab- nosed to have trichostrongylosis following comprehen- dominal pain, diarrhea, anorexia, nausea, weakness, mild sive clinical and laboratory examinations. Abdominal, anemia, low-grade peripheral eosinophilia, pulmonary and epigastric and right upper quadrant pain, diarrhea, low cutaneous symptoms are the most common manifesta- grade fever, rigors especially at nights, allergic manifesta- tions in heavy infections [8, 16, 17]. tions including urticaria associated with itching, flatu- Ostertagia species, commonly known as the brown lence and weight loss were the most important clinical stomach worm, are gastrointestinal nematodes that infect manifestations declared by the patients. Hypereosinophi- cattle, sheep and goats and other wild ruminants world- lia ranging from 25 to 60% were seen. No parasite ova wide, especially in temperate and cool climates. O. oster- were obtained following examination of three stool sam- tagi mainly infects cattle, but also sheep, goats and other ples on every other day using formalin-ether and Kato- domestic and wild ruminants [1]. It has been reported Katz techniques. Since the patients were living in a high- from two persons in central part of Iran and one person risk area of human fasciolosis they were treated for acute in Azerbaijan of former Soviet Union [3]. T. circumcincta, disease using Egaten (10 mg/kg). No evidence of treat- which was originally placed in the genus Ostertagia, is a ment success was observed about four weeks after drug parasitic nematode of gastric glands of the stomach that therapy. Surprisingly, all patients shed Trichostrongylus infects sheep and goats all around the world and leads to ova following a new set of three stool examination. The weight loss, decreased wool production and death [18]. infected individuals were treated with a single dose of The prevalence rates of 17.3 to 47.2% for T. circumcincta albendazole (400 mg) followed by 200 mg of mebenda- have been reported in sheep and goats in different prov- zole per day for 3 days [14]. All fecal materials of the pa- inces of Iran [19–21]. A single human case of T. circum- tients was collected up to 48 h following cincta has been reported from Azerbaijan of former Soviet chemotherapy in large screw capped plastic