Bioclimatic analysis and spatial distribution of fascioliasis causative agents by assessment of Lymnaeidae snails in northwestern provinces of Iran.

BACKGROUND: Snails of the Lymnaeidae family are the intermediate hosts of Fasciola species, the causative agents of fascioliasis. The purpose of this study was to determine the prevalence of Fasciola species in lymnaeid snails and to investigate the association of geoclimatic factors and Fasciola species distribution in northwestern provinces of Iran using geographical information system (GIS) data. METHODS: A total of 2000 lymnaeid snails were collected from 33 permanent and seasonal habitats in northwestern Iran during the period from June to November 2021. After identification by standard morphological keys, they were subjected to shedding and crushing methods. Different stages of Fasciola obtained from these snails were subjected to the ITS1 polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for species identification. The associations of weather temperature, rainfall, humidity, evaporation, air pressure, wind speed, elevation, and land cover with the distribution of Fasciola species were investigated. Geographical and statistical analysis was performed using ArcMap and SPSS software, respectively, to determine factors related to Fasciola species distribution. RESULTS: Of the 2000 snails collected, 19 were infected with Fasciola hepatica (0.09%), six with F. gigantica (0.03%), and 13 with other trematodes. Among geoclimatic and environmental factors, mean humidity, maximum humidity, and wind speed were significantly higher in areas where F. hepatica was more common than F. gigantica. The altitude of F. hepatica-prevalent areas was generally lower than F. gigantica areas. No significant relationship was observed between other investigated geoclimatic factors and the distribution of infected snails. CONCLUSIONS: The present study showed the relationship of humidity and wind speed with the distribution of snails infected with F. hepatica or F. gigantica in the northwestern regions of Iran. In contrast to F. gigantica, F. hepatica was more prevalent in low-altitude areas. Further research is recommended to elucidate the relationship between geoclimatic factors and the presence of intermediate hosts of the two Fasciola species.

Comments on "Genetic characterization and phylogenetic analysis of Fasciola species based on ITS2 gene sequence, with first molecular evidence of intermediate Fasciola from water buffaloes in Aswan, Egypt".

Dear Editor-in-Chief, In Annals of Parasitology 2021, 67(1), 55-65, a paper entitled "Genetic characterization and phylogenetic analysis of Fasciola species based on ITS2 gene sequence, with first molecular evidence of intermediate Fasciola from water buffaloes in Aswan, Egypt" was published with great interest [1]. After reading the article carefully and critically, we think some points should be noted. Fasciola species are meiotically functional diploid, can produce sperm and temporarily and store in the seminal vesicles. This type is named spermic fluke [2]. On the other hand, intermediate Fasciola with morphological characteristics intermediates between F. hepatica and F. gigantica with no sperm or aspermic and no sperm in seminal vesicles. However, this is also seen in older flukes [3-5]. It seems that morphological studies based on spermatogenesis ability were necessary for this study. Also, this parasite's anthelmintic resistance is due to aspects of biology, and population structure depends on genetic diversity [6]. We question whether there are any documents about and sequences of mitochondrial markers as COX (Cytochrome Oxidase) and NAD (Nicotinamide Adenine Dinucleotide) to analyze intraspecific phylogenetic relationship in addition to nuclear gene? In Table 3, the pairwise distances between three groups of Fasciola spp. from different livestock animals were low, ranging from 0.004 to 0.01 with an overall mean of 0.008. Genetic diversity is described as a tendency of genetic characteristics to vary and serves as a way for the population to adapt to changing hosts and environments [7]. The nature of the nuclear gene (ITS) is instability. It is better to use mitochondrial sequence data to compare diversity. Also, genetic discrimination grade from infra population to meta population is annotated by Fst value ranging; 0 to 1. Fst values between 0-0.05 indicated a low genetic differentiation population [8]. It seems that by calculating Fst and showing the gene migration based on mitochondrial sequences data of specimens, this study's species population will be obtained. Also, Tajima's D and Fu's F in all loci populations based on GenBank data may show the Fasciola haplotypes' population proximity. Here we recommend, that Omar et al. [1] studies that molecular phylogeny with mitochondrial DNA efectively used for appropriate diferentiation of haplotypes and spermatogenic ability by carmen allium staining helps them find the physiological aspects. Of course, more prominent populations are needed to find intermediate types. [1] Omar M.A, Elmajdoub L.O., Ali A.O., Ibrahim D.A., Sorour S.S., Al-Wabel M.A., Suresh M., Metwally A.M. 2021. Genetic characterization and phylogenetic analysis of Fasciola species based on ITS2 gene sequence, with first molecular evidence of intermediate Fasciola from water buffaloes in Aswan, Egypt. Annals of Parasitology 67: 55-65. doi:10.17420/ap6701.312 [2] Sanderson A. 1953. Maturation and probable gynogenesis in the liver fluke, Fasciola hepatica L. Nature 172: 110-112. doi:10.1038/172110a0 [3] Hayashi K., Ichikawa-Seki M., Mohanta U.K., Singh T.S., Shoriki T., Sugiyama H., Itagaki T. 2015. Molecular phylogenetic analysis of Fasciola flukes from eastern India. Parasitology International 64: 334-338. https://doi.org/10.1016/j.parint.2015.04.004 [4] Ichikawa-Seki M., Tokashiki M., Opara M.N., Iroh G., Hayashi K., Kumar U.M., Itagaki T. 2017. Molecular characterization and phylogenetic analysis of Fasciola gigantica from Nigeria. Parasitology International 66: 893-897. doi:10.1016/j.parint.2016.10.010 [5] Rouhani S., Raeghi S., Mirahmadi H., Fasihi Harandi M., Haghighi A., Spotin A. 2017. Identification of Fasciola spp. in the east of Iran, based on the spermatogenesis and nuclear ribosomal DNA (ITS1) and mitochondrial (ND1) genes. Archives of Clinical Infectious Diseases 12:e57283. doi:10.5812/archcid.57283 [6] Hodgkinson J., Cwiklinski K., Beesley N., Paterson S., Williams D., Devaney E. 2013. Identification of putative markers of triclabendazole resistance by a genome-wide analysis of genetically recombinant Fasciola hepatica. Parasitology 140: 1523. doi:10.1017/S0031182013000528 [7] Bozorgomid A., Rouhani S., Harandi M.F., Ichikawa- Seki M., Raeghi S. 2020. Genetic diversity and distribution of Fasciola hepatica haplotypes in Iran: molecular and phylogenetic studies. Veterinary Parasitology: Regional Studies and Reports 19: 00359. [8] Rouhani S., Raeghi S., Spotin A. 2017. Spermatogenic and phylo-molecular characterizations of isolated Fasciola spp. from cattle, North West Iran. Pakistan Journal of Biological Sciences 20: 204-209.

Identification of Novel Voltage-Gated Sodium Channel Mutations in Human Head and Body Lice (Phthiraptera: Pediculidae).

In recent years, the increase of head louse infestation in Iran (7.4%) and especially in West-Azerbaijan Province (248%) has raised the hypothesis of insecticide resistance development. There are different mechanisms of resistance to various groups of insecticides, and knockdown resistance (kdr) is a prominent mechanism of resistance to pyrethroids, an insecticide group which is used conventionally for pediculosis control. For detection of kdr-type well-known amino acid substitutions (M815I-T917I-L920F) and additional sodium channel mutations potentially associated with kdr resistance in head and body lice, louse populations were collected from West-Azerbaijan and Zanjan Provinces of Iran. Six novel mutations were found to be located in the IIS1-2 extracellular loop (H813P) and IIS5 (I927F, L928A, R929V, L930M, and L932M) of the α-subunit. Genotyping results showed that all specimens (100%) have at least one of these or the well-known mutations. Therefore, the presence of kdr-related and novel mutations in the sodium channel is likely to be the reason for the frequent use of pyrethroid insecticides due to treatment failure against lice. Further studies are now required to evaluate the prevalence of the kdr-like mutant allele for monitoring of insecticide resistance and the management of head and body lice in other provinces of the country.

Genetic Characterization of Fasciola Isolates from West Azerbaijan Province Iran Based on ITS1 and ITS2 Sequence of Ribosomal DNA.

BACKGROUND: Fascioliasis, caused by Fasciola hepatica and F. gigantica, has medical and economic importance in the world. Molecular approaches comparing traditional methods using for identification and characterization of Fasciola spp. are precise and reliable. The aims of current study were molecular characterization of Fasciola spp. in West Azerbaijan Province, Iran and then comparative analysis of them using GenBank sequences. METHODS: A total number of 580 isolates were collected from different hosts in five cities of West Azerbaijan Province, in 2014 from 90 slaughtered cattle (n=50) and sheep (n=40). After morphological identification and DNA extraction, designing specific primer were used to amplification of ITS1, 5.8s and ITS2 regions, 50 samples were conducted to sequence, randomly. RESULT: Using morphometric characters 99.14% and 0.86% of isolates identified as F. hepatica and F. gigantica, respectively. PCR amplification of 1081 bp fragment and sequencing result showed 100% similarity with F. hepatica in ITS1 (428 bp), 5.8s (158 bp), and ITS2 (366 bp) regions. Sequence comparison among current study sequences and GenBank data showed 98% identity with 11 nucleotide mismatches. However, in phylogenetic tree F. hepatica sequences of West Azerbaijan Province, Iran, were in a close relationship with Iranian, Asian, and African isolates. CONCLUSIONS: Only F. hepatica species is distributed among sheep and cattle in West Azerbaijan Province Iran. However, 5 and 6 bp variation in ITS1 and ITS2 regions, respectively, is not enough to separate of Fasciola spp. Therefore, more studies are essential for designing new molecular markers to correct species identification.