Molecular Identification of Cutaneous Leishmaniasis Vectors in Alborz Province, North of Iran.

Cutaneous leishmaniasis (CL) is a vector-borne disease widely distributed in tropical and subtropical areas of North and South America, Europe, Asia, and Africa. Considering the increasing number of CL cases in recent years and the fact that no study has been conducted to identify CL fauna and vectors in Alborz province, this study was carried out to identify sand flies and CL vectors in this region. Sand flies were collected from August to October 2021 from plain and mountainous indoor and outdoor areas of the region using sticky paper traps and were detected morphologically. DNA was extracted from the midguts of female sand flies. In this study, 1157 sand flies were collected and identified. The number of sand flies caught from indoor and outdoor places was 367 (31.72%) and 790 (68.28%), respectively. Overall, six species of flies were of the genus Phlebotomus (Raynal, 1937), including Phlebotomus papatasi (P. papatasi, 695 [60.07%]; Scopoli, 1786), P. kandelakii (13 [1.12%]; Shchurenkova, 1926), P. sergenti (232 [20.05%]; Parrot, 1917), P. major (14 [1.21%]; Annandale, 1910), P. caucasicus (4 [0.35%]; Marzinowsky, 1917), P. alexandri (18 [1.56%]; Alexandri Sinton, 1920), and four were of the genus Sergentomyia (Artemiev, 1978), including Sergentomyia tiberiadis (109 [9.42%]; Adler, Theodor & Lourie, 1930), Sergentomyia baghdadis (53 [4.58%]), Sergentomyia sintoni (14 [1.21%]; Sintoni Pringle, 1933), Sergentomyia clydei (5 [0.43%]). P. papatasi spp. were dominant in indoor and outdoor places, with a prevalence of 695 (60.07%). The Leishmania major (L. major) gene was identified in five samples of P. papatasi spp. This suggests that P. papatasi is the potential vector spp. in the study area. Moreover, L. major was confirmed as the aetiological agent of CL cases in Alborz province. The identification of vectors and parasite spp. is very important for the treatment and operational planning of disease vectors.

A Comparative Morphometric Study on Odontobuthus bidentatus and Odontobuthus doriae (Scorpionida: Buthidae) in Iran.

Scorpions are one of the most venomous animals which cause serious public health problems. The sting of scorpions can sometimes be fatal depending on the scorpion species involved. So far, sixty-six (66) scorpion species have been identified in Iran. Annually, about 40-50000 cases of scorpionism are reported in Iran. Odontobuthus doriae and O. bidentatus are among the most medically important scorpion species in Iran, and they are very similar to each other in coloration, carination, and trichobotrial patterns. This morphometric study aimed to compare some of the important morphological characteristics in order to identify the key differences between these two species. A total of 45 morphological characters were measured using calipers and stereomicroscope, and 55 morphological characters and ratios (relative of length to width ratio of morphological characters of scorpions) were analyzed. The independent sample t-test in SPSS software (version 24) was used for the statistical analyses in this study. The mean total length, carapace width, length of fixed and moveable fingers, and chelicerae length of O. doriae were greater than those of O. bidentatus in our study area. The morphological measurements displayed a clear distinction between O. doriae and O. bidentatus in our study area; therefore, they can be used as morphological identification keys for distinguishing between these two species.

New Data on Two Subspecies of Mesobuthus eupeus, the Most Medically Important Scorpion Species in Northwestern Iran.

Scorpions are among the most medically important arthropods in Iran, particularly northwestern areas. To date, five scorpion species, i.e. Mesobuthus eupeus, Mesobuthus caucasicus, Androctonus crassicauda, Hottentotta saulcyi, and scorpio maurus, have been identified. The family Buthidae is responsible for most cases of scorpionism in Iran. The Mesobuthus eupeus species belong to this family and is commonly distributed from Turkey to China, including Iran. Among these species, Mesobuthus eupeus is regarded as the most medically important species and responsible for most cases of envenomation in this area. Morphological differences between some species collected in the study area have been reported. The present study, thus, aimed to identify the subspecies of Mesobuthus eupeus in northwestern Iran. Scorpions were captured in the summer months from 37 localities in three northwestern provinces in Iran: West Azerbaijan, East Azerbaijan, and Ardabil. Scorpion collection was carried out using rock rolling and ultraviolet methods. A total of 376 specimens of Mesobutus eupeus (177males and 199 females) were collected and identified as Mesobuthus eupeus (98.4%) and Mesobuthus eupeus philippovitschi (1.6%). Owing to the findings of our study, M.e.philippovitschi has been added to the scorpion fauna of northwestern parts of Iran for the first time. Unlike M.e. eupeus which is widely distributed from plains to mountainous regions, M.e.philippovitschi has limited distribution and is found mostly along the borders with neighboring countries. This subspecies is the most medically important and most prevalent one in the region. The findings of the present study also provide the basis for future consideration of regional antivenom production for this medically important species.

Prevalence and Early Detection of Hypodermosis in Goats using a Competitive ELISA System in Lorestan, Iran.

This study aimed to determine the prevalence and early detection of hypodermosis in goats by the investigation of Przhevalskiana larvae and sera collected from the infested animals. This study was conducted in Lorestan province, located in the South-West of Iran, from April 2017 up to April 2018. A total of 3350 goats slaughtered in Lorestan abattoirs were investigated by clinical-parasitological examinations in different periods. The larvae were collected from the back and flank regions of the slaughtered goats. The number of infested animals, gender and age, number of maggots present on the body of each animal, location, and larval stage of warble flies were recorded in this study. To detect an infestation in the early period, a total of 150 blood samples were randomly collected from the field animals in Lorestan, Iran. The morphological findings showed that out of 3350 goats examined, 706 (21.07%) goats were infested. Furthermore, three species of Przhevalskiana, including P. Silenus (n=726, 50.07%), P.crossii (n=440, 30.43%), and P. aegagri (n=284, 19.59%) were recognized as the causative agents of goat hypodermosis in this province. No significant difference was observed between genders and/or among the age groups (P>0.05). The anti-Przhevalskiana antibodies in the serum samples were detected using ELISA from August up to mid-September (summer). Clinical diagnosis of infestation was usually performed from late October until mid-March (winter) by visual observations and direct palpation of warbles in the back and flank regions of the animals. It could be concluded that the use of ELISA can help to detect hypodermosis among goats in the early stages.

Species Composition and Spatial Distribution of Scorpions Based on Eco-Environmental Variables in Provinces Along with the Oman Sea and the Persian Gulf in Iran: A GIS-Based Approach.

Scorpions are venomous arachnids with major medical health importance in Iran, specifically in the Southwest. In total, three families of scorpions, including Scorpionidae, Hemiscorpiidae, and Buthidae were reported in Iran. This study was conducted on scorpion ecology to determine the species composition and the dispersion of scorpions based on the ecological and environmental variables in combination with the Geographic Information System (GIS) in Khuzestan, Hormozgan, and Bushehr Provinces along with the Oman Sea and the Persian Gulf in Iran. Scorpions were collected from Hormozgan, Khuzestan, and Bushehr Provinces, Iran using the Ultra Violet light. The specimens were then identified according to their morphological characters utilizing reliable keys. To determine the relationship between the eco-environmental variables and the spatial distribution of species, the GPS points of the collected scorpions were recorded, and the scorpion shapefile was overlaid on digital elevation model, slope, land use, temperature, rainfall, soil texture, and bioclimatic maps. Totally, 25 specimens were reported in three families of Scorpionidae, Hemiscorpiidae, and Buthidae. Furthermore, Razianus zarudnyi, Androctonus crassicauda, Buthacus macrocentrus, Mesobuthus eupeus phillipsii, Odontobuthus bidentatus, and Hemiscorpius lepturus were the common species collected from Hormozgan, Khuzestan, and Bushehr Provinces, Iran. The results of the current study showed that a large number of species preferred the sand texture due to ecomorphological adaptation. Moreover, the poor rangeland vegetation cover was preferred by the majority of the scorpion species, including S. maurus townsendi. According to the results, the combination of the ecological factors related to the suitable habitat of different species of scorpion and GIS will provide the dispersal areas of each species. Furthermore, such databases can be comprehensive and valuable guides for health authorities to reduce and manage scorpion envenomation.

Phylogenetic and Morphological Analyses of Androctonus crassicuda from Khuzestan Province, Iran (Scorpiones: Buthidae).

The Androctonus crassicuda is the most diverse scorpion species in the family of Buthidae, which is endemic to Khuzestan province, Iran. Investigation of the relationship of species by means of a molecular study of specimens is one of the new approaches due to the limitations of the morphological approaches. In the current study, the analysis was based on 32 morphological characteristics of A. crassicuda native to southwest Iran. Moreover, the DNA sequencing of two mitochondrial markers, namely cytochrome oxidase subunit I and 12sRNA loci was performed, and the phylogenetic tree was constructed using maximum likelihood method with 1000 replications using MEGA software (version 7). Based on the results of the phylogenetic tree, A. crassicuda was classified into a monophyletic group. However, the genetic diversity of this species populations was not significant (0.001). The highest and lowest genetic distance of A. crassicuda was compared with the reports obtained in Urmia and west Azerbaijan, Iran. There was a clear divergence between the A. crassicuda isolated from northern and southern areas of Iran. This study showed the importance of geographical and climate features of the region and genetic distance among the populations. The phylogenetic analysis of Androctonus species from other regions showed the highest and lowest genetic distance with A. gonneti (Morocco) and A. amoreuxi (Portugal), respectively. The comparison of the morphological characteristics and morphometric results revealed that metasoma characteristics are important in the identification of A. crassicuda. The results of the analysis of the morphometric values of A. crassicuda were mainly compatible with the phylogenetic trees and supported the traditional morphological classification, thereby presenting a clearly definition of the genera of Androctonus species.

Serodiagnosis of Przhevalskiana spp. Infestation in Goats Using a Competitive ELISA.

Goat warble-fly infestation is one of the parasitic diseases caused by the larvae of przhevalskiana spp. It is known to be a major challenge for health and welfare in infested goats and causes high economic losses in livestock worldwide. The detection of goat grub was previously conducted by direct palpation of second and third stage larvae in the back and flank site of the live animals or visual evaluation of the carcasses in slaughterhouses. However, due to the small size of the first instar larvae of przhevalskiana (less than 1 mm during emerging from the egg), some of the infected cases were ignored and recorded as negative samples. Immunodiagnostic procedures as easy and cost-effective diagnostic methods provide early detection of myiasis in living animals (even when the larvae are still migrating or are undetectable in the animal body).This study was conducted to evaluate the competitive enzyme-linked immunosorbent assay (ELISA) system in order to detect the antibodies of przhevalskiana larvae in the goat sera. In order to prepare the larval antigen, 200 first instar larvae of przhevalskiana were collected from the subdermal region of the back and flanks of the infested goats in Khoramabad slaughterhouse, Khoramabad, Iran, from September 2017 to March 2018. Totally, 37 and 46 sera samples were taken from the infected and uninfected goats. The sensitivity and specificity at cut-off 3SD were obtained at 89.18% and 84.78%. Moreover, the measures of inter-and intra-assay coefficients of variability to express the precision or replicability of ELISA kit results were 5.33% and 2.81%, respectively.

First Case Report of an Unusual Echis genus (Squamata: Ophidia: Viperidae) Body Pattern Design in Iran.

Three families of venomous snakes exist in Iran including Viperidae, Elapidae, and Hydrophidae. Viperidae family is the only family with a widespread distribution. Saw-scaled vipers are important poisonous snakes in Asia and Africa. This name is given to this snake due to the presence of obliquely keeled and serrated lateral body scales. Distribution of this genera is mostly reported in the central and southern regions of Iran. This genus has four main clades: the Echis carinatus, E. coloratus, E. ocellatus, and E. pyramidum. Design pattern in Echis species plays an important role in camouflage and variety of habitat. In the present report, we investigated a specimen from the eastern region of Iran; we examined 25 specimens of Echis that were collected from the eastern region of our country. Among them, only one specimen with a different pattern was found compared with the other 24 specimens by surveying meristic, mensural, and design pattern characters using valid key identifiers. The similarities between the specific Echis with a different pattern and other 24 specimens were also studied and compared. The results of this investigation clearly showed that although the pattern of the lateral white line and block on dorsal body of the specific Echis snake was different, since the meristic and mensural characters were similar to other Echis snakes it can be concluded that this specimen is not a different species; the difference in these patterns may be due to a minor genetic mutation of that specimen. It is the first case report of Echis carinatus sochureki Stemmler, 1969 from Iran with a different pattern.

Sequence analysis of lysozyme C from the scorpion mesobuthus eupeus venom glands using semi-nested rt-PCR.

BACKGROUND: Lysozyme is an antimicrobial protein widely distributed among eukaryotes and prokaryotes and take part in protecting microbial infection. Here, we amplified cDNA of MesoLys-C, a c-type lysozyme from the most common scorpion in Khuzestan Province, Southern Iran. METHODS: Scorpions of Mesobuthus eupeus were collected from the Khuzestan Province. Using RNXTM solution, the total RNA was extracted from the twenty separated venom glands. cDNA was synthesized with extracted total RNA as template and modified oligo(dT) as primer. In order to amplify cDNA encoding a lysozyme C, semi-nested RT-PCR was done with the specific primers. Follow amplification, the fragment was sequenced. RESULTS: Sequence determination of amplified fragment revealed that MesoLys-C cDNA had 438 bp, encoding for 144 aa residues peptide with molecular weight of 16.702 kDa and theoretical pI of 7.54. A putative 22-aminoacids signal peptide was identified. MesoLys-C protein was composed of one domain belonged to c-type lysosyme/ alphalactalbumin. CONCLUSION: Multiple alignment of MesoLys-C protein with the related cDNA sequences from various organisms by ClustalW program revealed that some of the conserved residues of other c-type lysosymes were also seen in MesoLys-C. However, the comparison suggested that Mesobuthus eupeus of Khuzestan and east Mediterranean Mesobuthus eupeus belonged to different subspecies.

Pedipalps and venom vesicle anomalies in two families of scorpions (Scorpiones: Hemiscorpiidae, Buthidae) from Iran.

The developmental anomalies are reported in this study. The first and second abnormally are presented in right pedipalps of Paraorthochirus and Orthochirus (Buthidae) while Paraorthochirus pedipalp just includes coxa, trochanter and without other parts (femur, patella, movable and fixed finger). The right pedipalp of Orthochirus specimen has abnormally too; it has all parts ofpedipalp except complete fixed finger. In both of scorpions, the left pedipalp is normal. Another case is present in venom vesicle of Hemiscourpius (Hemiscorpiidae). Pictures and morphometric measurements for three specimens are given.