Emergent spotted fever group Rickettsiae infections among hard ticks in Islamic Republic of Iran.

Background: Tick-borne rickettsioses have become a health concern worldwide following the increasing incidence in recent decades. However, there is limited information about these diseases in Islamic Republic of Iran. Aim: This cross-sectional study was conducted to estimate the Rickettsia infection among ixodid ticks collected from cattle, sheep and goats in Islamic Republic of Iran. Methods: The DNA of ixodid ticks collected from cattle, sheep and goats in 54 villages of Zanjan Province, Islamic Republic of Iran, were collected and analysed using a spectrophotometer. Rickettsial-positive samples were screened by targeting the htrA gene and fragments of gltA gene were analysed. The variables were analysed using descriptive statistics and the χ2 test was used to compare the variables. Results: A total of 528 ticks were tested. Overall, Rickettsia infection rate was 6.44%. Nine of the 12 tick species were infected. Rickettsial positive rates in Hyalomma marginatum and Dermacentor marginatus were 21.33% and 12.77%, respectively. R. aeschlimannii, the predominant rickettsia, was detected only in Hy. marginatum. R. raoultii, R. sibirica and R. slovaca comprised about half of the positive ticks and were recovered from more than one tick species. Conclusion: Considering the discovery of infected ticks in the Islamic Republic of Iran, there is a need to establish a tick control programme in the country, paying attention to populations at high-risk.

Morphometric Indices and Venom Protein Profile in Different Populations of Androctonus crassicauda.

Background: Androctonus crassicauda is the most medically relevant animal and understanding its morphological characteristics is essential in the production of antiscorpion sera. Methods: Adults of A. crassicauda were collected from different areas of Zanjan Province and the morphometric parameters and the cuticular fluorescence patterns of samples were studied. The crude venom of samples was extracted by electric stimulation, and their biochemical properties were analyzed by the SDS-PAGE method. Results: Values of the morphometric parameters depended on sex and altitude of the area. Except for values of the pectinal organ, these parameters in females were higher than in males. No significant difference was in the number, shape, and intensity of cuticular fluorescence patterns. The body length of males in high and lowlands was 72.53±1.53 and 77.33±2.70mm, respectively. Females' body lengths in that area were 81.66±2.19 and 86.55±2.33mm, respectively. Analysis of toxin proteins showed two isotypes that the 12, 13, 15, 16, 18, and 19kDa proteins were in all areas. However, the 41 and 74kDa proteins, and 46 and 63kDa proteins were detected in low and highlands, respectively. Conclusion: Black fat-tailed scorpion has a considerable dominancy and developing preventive programs and providing treatment facilities in studied areas are necessary. Values of the morphological parameters and venom electrophoresis patterns depended on the geographical location. Therefore, pool crude toxin is suggested for the production of effective antivenoms. Moreover, additional field complementary works in the geographic information system based niche modeling and mass fingerprinting of scorpion venoms are suggested for screening effective isotypes.

Molecular Survey of Mitochondrial Genes in Different Populations of the Black Fat-Tailed Scorpion, Androctonus crassicauda.

Background: Androctonus crassicauda is the most medically relevant scorpion and understanding its genetic forms is essential for improvement of anti-venom sera, and risk management of scorpionism. Present study was designed to identify the variations of mitochondrial genes in different populations of A. crassicauda. Methods: Adults of A. crassicauda were collected from Zanjan Province during 2016-2017. Genomic DNA of samples was extracted and fragments of mitochondrial 16S, COI and ND1 genes were amplified and some of the amplicons were sequenced. Haplotype of samples were identified by multiple alignment of sequences, then phylogenetic trees of haplotypes were constructed. Results: Fragments of 352bp, 618bp and 680bp were amplified from 16S, COI and ND1 genes respectively. Nucleotide sequence in COI fragments was conserved, however, five haplotypes with some specific polymorphic sites were detected in 16S and ND1 fragments. Haplotype I was dominant and found in all areas. Other haplotypes were rare and limited to specific regions. Analysis of the phylogenetic trees inferred from 16S and COI genes, confirmed a strong positive correlation between geographic and genetic distance. Conclusion: Mitochondrial COI, 16S and ND1 genes were detected suitable for identifying the population structure. Five genotypes were found using 16S and ND1 genes. To prepare and improve the anti-venoms quality, additional studies are necessary to identify the toxin electrophoretic profile and geographical/ecological niche models of these genotypes in future.

First report of target site insensitivity to pyrethroids in human flea, Pulex irritans (Siphonaptera: Pulicidae).

The human flea, Pulex irritans, is the most important ectoparasite of humans. Intensive use of pyrethroids for its control has led to insecticide resistance. Monitoring pyrethroid resistance and its underlying mechanisms is essential for flea control. The aims of this study were to identify the susceptibility status of human flea to permethrin and to detect the presence of knockdown resistance (kdr) mutation and its frequency in populations of P. irritans. Adults of P. irritans were collected from Zanjan Province, northwest of Iran, during 2013-2017. Different populations of this flea were exposed to permethrin 0.75% for one and 8 h and then the mortality rate, as well as KD50 and KD95 times were calculated. Total RNA and gDNA of samples were extracted, and the fragments of cDNA encoding the partial voltage-gated sodium channel (VGSC) peptides were amplified using degenerated primers. Specific PCR and TaqMan real-time assays were conducted to characterize the vgsc gene and to detect the presence of mutation and genotyping of the populations. Mortality rates were in the range from 32% to 67% for one-hour and 73% to 90% for eight-hour exposure to permethrin 0.75%. KD50 and KD95 times varied in a range from 46 to 241 and 177 to 899 min, respectively. Sequencing of 70 amplified fragments of gDNA resulted in a 578-bp product. These fragments contained two introns (92 and 63 bp) and three exons (141, 189, and 92 bp) encoding 138 amino acids that encompassed IIS4-IIS6 and the partial linker between domains II and III of VGSC. All the studied populations showed L1014F mutation, substitution of CTT for TTT at the 1014 allele. The result of TaqMan assay for 624 samples showed 96.6% homogenous and 3.36% heterozygous mutant. The development of permethrin resistance and the presence of the L1014F mutation at high frequency in flea populations indicate that pyrethroids are likely ineffective in controlling human flea. Therefore, novel alternative control methods are needed to combat this human ectoparasite.