Molecular identification, risk factor assessment, and phylogenetic analysis of tick-borne pathogens in symptomatic and asymptomatic cattle from South-Eastern Iran.

Tick-borne pathogens (TBPs) represent a substantial threat to cattle globally, exerting adverse impacts on production, health, and economic viability. This study delves into the prevalence and implications of TTBPs in cattle sourced from resource-limited smallholder livestock farms situated in southeastern Iran, proximate to Afghanistan and Pakistan. Blood and tick specimens were systematically collected from a cohort of 230 cattle, comprising 150 asymptomatic and 80 symptomatic individuals. Genomic DNA isolated from blood samples underwent rigorous examination for the presence of key TBPs, including Anaplasma marginale, A. phagocytophilum, A. bovis, A. centrale, Babesia bigemina, and Theileria annulata, utilizing multiple genetic markers. Nucleotide sequence analysis facilitated the reconstruction of phylogenetic relationships. The study also evaluated various potential risk factors, such as clinical status, gender, age, breed, tick infestation, and management practices, to elucidate their associations with TTBPs. Among the cattle cohort, a staggering 87.8% (202/230) tested positive for at least one pathogen. Prevalence statistics encompassed A. marginale (72.2%), T. annulata (68.3%), A. phagocytophilum/A. platys-like complex (66.1%), A. centrale (16.7%), B. bigemina (10.0%), and A. bovis (6.1%). Remarkably, mixed infections involving two, three, and four pathogens were detected in 23%, 52.1%, and 2.2% of animals, respectively. Notably, all asymptomatic cattle were positive for at least one TBP. Tick infestation was observed in 62.2% (143/230) of cattle, predominantly caused by Hyalomma anatolicum (82.5%), Rhipicephalus (Boophilus) annulatus (13.1%), and R. sanguineus sensu lato (4.4%). Risk factors linked to TBPs encompassed tick infestation, older age, and crossbred animals. Clinical presentations among symptomatic cattle encompassed fever, anemia, weight loss, anorexia, jaundice, and enlarged superficial lymph nodes. This study underscores the pivotal role of asymptomatic carriers in the propagation of TTBPs within endemic regions. Furthermore, it emphasizes the potential for the implementation of molecular diagnostics to unmask subclinical infections, thereby affording the opportunity for targeted interventions aimed at ameliorating the burden of TTBPs in resource-constrained smallholder dairy farms.

MicroRNAs as Biomarkers for Early Diagnosis, Prognosis, and Therapeutic Targeting of Ovarian Cancer.

Ovarian cancer is the major cause of gynecologic cancer-related mortality. Regardless of outstanding advances, which have been made for improving the prognosis, diagnosis, and treatment of ovarian cancer, the majority of the patients will die of the disease. Late-stage diagnosis and the occurrence of recurrent cancer after treatment are the most important causes of the high mortality rate observed in ovarian cancer patients. Unraveling the molecular mechanisms involved in the pathogenesis of ovarian cancer may help find new biomarkers and therapeutic targets for ovarian cancer. MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression, mostly at the posttranscriptional stage, through binding to mRNA targets and inducing translational repression or degradation of target via the RNA-induced silencing complex. Over the last two decades, the role of miRNAs in the pathogenesis of various human cancers, including ovarian cancer, has been documented in multiple studies. Consequently, these small RNAs could be considered as reliable markers for prognosis and early diagnosis. Furthermore, given the function of miRNAs in various cellular pathways, including cell survival and differentiation, targeting miRNAs could be an interesting approach for the treatment of human cancers. Here, we review our current understanding of the most updated role of the important dysregulation of miRNAs and their roles in the progression and metastasis of ovarian cancer. Furthermore, we meticulously discuss the significance of miRNAs as prognostic and diagnostic markers. Lastly, we mention the opportunities and the efforts made for targeting ovarian cancer through inhibition and/or stimulation of the miRNAs.

First report of Babesia occultans in two symptomatic cows in Iran.

This study reports the identification and first molecular characterization of Babesia occultans from naturally infected cows in Iran. Microscopic examination showed pyriform trophozoites, and ring-shaped merozoites (>2.5 µm) in Giemsa-stained blood smears obtained from two symptomatic cows in West-Azarbaijan province, Iran. PCR amplification of the partial 18S rRNA gene including the V4 hypervariable region were carried out on DNA extracted from blood samples. BLAST analyses of the partial 18S rRNA (approximately 400 bp) obtained from two cows revealed the presence of B. occultans and the detected sequences were identical to each other. Comparisons of the partial 18S rRNA sequence of the current isolate with other B. occultans sequences from Tunisia, South Africa, Turkey, Pakistan, and China confirmed the relation of the Iranian isolate to the species B. occultans. Sequence analysis of the obtained B. occultans showed 99.5-100% identity to the previously reported isolates. The sequences of B. occultans had 100% identity to a sequence obtained from ticks in Tunisia. This report is the beginning of a path to further research about B. occultans in vectors and reservoirs throughout Iran.

Identification of Anaplasma marginale in long-eared hedgehogs (Hemiechinus auritus) and their Rhipicephalus turanicus ticks in Iran.

Hedgehogs play a role in the eco-epidemiology of parasites, viruses and bacteria of veterinary and medical importance. In Europe, hedgehogs have been found infected with tick-transmitted Anaplasma phagocytophilum and A. marginale. In this study blood and ticks collected from 53 long-eared hedgehogs (Hemiechinus auritus) living in southeastern Iran close to Afghanistan and Pakistan borders were examined for Anaplasma spp. infection using microscopical and molecular biology methods. At microscopical examination Anaplasma-like inclusion bodies were found at the margin of erythrocytes in the blood smear of one infected hedgehog (prevalence 1.9 %). Each hedgehog was infested with three to eight adult ticks (average 4.7 ± 1.5). On the body of hedgehogs, the ears were the main site for attachment of ticks. Out of 248 collected adult ticks 81.4 % were Rhipicephalus sanguineus sensu lato and 18.5 % were R. turanicus. PCR and sequencing showed the presence of A. marginale infection in the blood of two hedgehogs (3.8 %) and R. turanicus ticks collected from them. This is the first report of A. marginale infection in hedgehogs in Iran. Epidemiological importance of this finding is discussed and current knowledge on the tick fauna of hedgehogs in the country is reviewed.

Histopathological Survey on Sarcocystis Species Infection in Slaughtered Cattle of Zabol-Iran

Objective: Sarcocystosis is an important zoonotic protozoal disease with worldwide distribution and wide range of hosts. The aim of the present study was to determine the intensity of Sarcocystis spp. infection and to show histopathological features of their cystic lesions in slaughtered cattle of Zabol- Iran. Methods: From April to September 2018, 500 tissue samples from esophagus, heart, diaphragm, tongue and masticatory muscles were prepared from 100 slaughtered cattle. All samples were fixed in 10% neutral buffered formalin and routine tissue processing protocol was performed. Results: The microscopic results showed that 92.2% of specimens had thin-walled cysts of S. cruzi and 14% had thick-walled Sarcocystis (S. hirsuta and S. hominis) but macrocyst was only observed in one cattle. The positivity rate of thin walled cysts was 58.8% for heart, 13.9% for masticatory muscles, 10.2% for tongue, 9.3% for esophagus and 7.8% for diaphragm. The positivity rate of thick walled cysts was 32.8% for esophagus, 28.6% for tongue, 22.9% for heart, 15.7% for masticatory muscles and 0% for diaphragm, which could represent either S. hominis or S. hirsuta. The most infected tissue was heart and the least infected tissue was diaphragm. Thin walled cysts (S. cruzi) were mostly found in heart and were less found in diaphragm. However, thick-walled cysts (S. hirsuta and S. hominis) were mostly detected in esophagus. No thick-walled cysts were found in diaphragm muscle. Conclusion: A high positivity rate of sarcocystosis in slaughtered cattle in Zabol abattoir revealed heavily environmental contamination of Sistan region by this important parasitic disease.

Identification of Theileria Species in Sheep and Vector Ticks Using PCR Method in Zabol, Eastern Iran.

BACKGROUND: Theileria is a protozoal parasite that belongs to the phylum Apicomplexa. Theileriosis is an important tick-borne disease caused by various species of Theileria. Among these species, T. lestoquardi (T. hirci) is highly pathogenic, while other species such as T. ovis make Subclinical and mild infections in small ruminant. Therefore, the precise identification of the species and the vector ticks are very essential for epidemiological studies and the design of control programs. METHODS: This research was conducted with the aim of molecular study to identify Theileria species and vectors in Zabol, eastern Iran in 2015. The presence of Theileria in 80 blood samples and vector ticks was evaluated using PCR method. RESULTS: Of 80 blood samples, PCR analysis showed that 50 samples (62.5%) were infected with Theileria. The evaluation of the first phase PCR with Nested PCR showed that infections with Theileria ovis and Theileria lestoquardi were 67.45% and 32.55% cases respectively. Overall, 110 ticks (78 males and 32 females) were collected and generally two genera and six species including Rhipicephalus bursa (9.1%), Rh. sanguineus (29.1%), Rh. turanicus (10.9%) Hyalomma asiaticum asiaticum (23.63%), Hy. excavatum (10.9%), Hy. anatolicum (16.37%) were detected. After evaluating ticks infection by PCR method, three species of Rh. turanicus, Rh. sanguineus and Hy. asiaticum asiaticum, were infected. CONCLUSION: Theileria ovis has a high prevalence among the sheep of zabol and Hy. asiaticum asiaticum, Rh. sanguineus and Rh. turanicus may be the main vectors of Theileria species in this area.

Epidemiology and Molecular Prevalence of Toxoplasma gondii in Cattle Slaughtered in Zahedan and Zabol Districts, South East of Iran.

BACKGROUND: Toxoplasma gondii is an obligate, intracellular parasite which causes the toxoplasmosis in humans and warm-blooded animals. Red meat is an important source for transmission of the infection to humans. This study aimed to determine the prevalence of Toxoplasma among imported and indigenous cattle in the Sistan region. METHODS: One hundred samples from slaughtered cattle were collected from two abattoirs of Zabol and Zahedan, South East of Iran in 2015. Each sample was a mixture of three muscle, including tongue, cardiac, and triceps. Additional data of each cattle, including sex, breed, age, indigenous or imported, location of slaughter, management practices, and feeding system were obtained through observations and interviews. Infection by T. gondii was determined by PCR method. RESULTS: The prevalence of Toxoplasma in indigenous cattle was 6% and in imported cattle was 26%, and this difference was statistically significant (P=0.006). Moreover, the prevalence of Toxoplasma was statistically associated with management practices (P=0.01) and feeding system (P=0.001). However, relationship between the prevalence of Toxoplasma with age, breed, sex, and location of slaughter was not statistically significant. CONCLUSION: Since the prevalence of Toxoplasma among imported cattle is higher than indigenous cattle, so strict supervision for importing livestock from neighboring countries is necessary.

Oxidative stress and hepatic injury induced in mice fed a Sarcocystis hirsuta cyst extract.

We studied the toxic effects of a Sarcocystis hirsuta cyst extract fed to mice. Degenerative changes were found in mice gavage-fed fresh, frozen, and heat-treated S. hirsuta cyst extract. There were increases in the levels of serum aspartate aminotransferase and alanine aminotransferase as well as hepatic and brain malondialdehyde (MDA) levels along with concomitant decreases in catalase (CAT) and superoxide dismutase (SOD) activities of mice receiving fresh and frozen S. hirsuta extracts. Gavage feeding of heat-treated S. hirsuta cyst extract had no effects on liver enzymes or brain MDA content, but the liver MDA level did increase. Mice in the heat-treated cyst group showed reduced CAT and SOD activities as well as increased hepatic MDA levels compared to those in the control group. These results indicate that an extract of S. hirsuta cyst can induce oxidative stress and hepatic injury, even after heat treatment.

Detection of Theileria annulata carriers in Holstein-Friesian (Bos taurus taurus) and Sistani (Bos taurus indicus) cattle breeds by polymerase chain reaction in Sistan region, Iran.

Theileria annulata is common in tropical and subtropical regions especially in Iran and causes great economic losses in cattle industry. In Iran the epidemiological aspects of bovine theileriosis in different breeds of cattle is poorly understood. The aim of present study is comparison of the number of T. annulata carriers in the two major cattle breeds (Holstein-Friesian and Sistani) in Sistan of Iran by giemsa and polymerase chain reaction (PCR) methods. During winter 2013, 160 native cattle, from the two major breeds in Sistan, with the mean age of more than one year and without typical clinical symptoms of theileriosis were selected. At first, a thin layer smear was held from their ear sublime vein blood for Giemsa staining method. In order to do PCR assay, jugular vein blood sample of each cow was taken. The PCR employs primers specific for the 721-bp gene fragment encoding the 30-kDa major merozoite surface antigen of T. annulata. By PCR method, 38 (47.5 %) Holstein blood samples and 22 (27.5 %) Sistani blood samples had DNA of T. annulata and considered positive (The correlation was significant at values of P < 0.05). By checking 160 blood smears with light microscope and lens × 100, only 10 samples (6.25 %) were positive for T. annulata. Statistical comparison between PCR and smear method showed that the PCR method is more sensitive and accurate in comparison to Giemsa staining method to diagnose the asymptomatic carriers of T. annulata.

A study on prevalence and identification of Ovine Theileria and Babesia infection in Zabol using PCR method.

Infection with Babesia and Theileria Causes high mortality and economical losses in livestock and has a relatively high prevalence in Iran. In Zabol, animals are susceptible to this disease because of presence of vector ticks, weather conditions and smuggle animal across the border and they cause great damages to the economy and production. However, few studies have been done for differentiation of Theileria and Babesia in sheep in this area. The purpose of the present study was to determine the prevalence of Babesiosis and Theileriosis in sheep in Zabol. A number of 80 animals were randomly selected and their blood samples were examined. The presence of Theileria and Babesia parasites in each sample was determined with PCR and microscopic examination. Of 80 blood samples obtained from sheep, 8 cases (10 %) were positive by microscopic examination where 6 samples were infected with Theileria and 2 samples with Babesia. The result of PCR method determined the prevalence of 66.25 % (53 samples) and 3.75 % (3 samples) for Theileria and Babesia, respectively. The correlation between these two methods (PCR and microscopic examination) was determined with Kappa statistical test. Based on the obtained results, it is concluded that Babesiosis has a low prevalence among the sheep of Zabol. This is the first report in which ovine Babesiosis has been studied in this region using molecular identification techniques.

Endoparasites of the Long-Eared Hedgehog (Hemiechinus auritus) in Zabol District, Southeast Iran.

OBJECTIVE: The long-eared hedgehog (Hemiechinus auritus) is a nocturnal animal living in Central and Southeast Iran. However, there are few studies concerning endoparasites, some of which are zoonotic, of the hedgehogs in the north and northwest of Iran. The aim of the present study is to investigate endoparasites in long-eared hedgehogs, living in Zabol district, Southeast Iran. MATERIALS AND METHODS: Stool and blood samples collected from 50 hedgehogs (35 males and 15 females) that were trapped alive were examined with Clayton-Lane flotation and Giemsa staining methods. Furthermore, 10 road-killed hedgehog carcasses were necropsied. The adult parasites were collected and identified under a light microscope. RESULTS: Spirurida eggs in the stool samples and Anaplasma inclusion bodies in red blood cells were determined in 32% and 52% of the samples, respectively. Physaloptera clausa, Mathevotaenia erinacei, Nephridiacanthus major, and Moniliformis moniliformis were identified in the necropsy. CONCLUSION: To the best of our knowledge, ours is the first study concerning endoparasites of long-eared hedgehogs in Iran. Furthermore, M. erinacei was for the first time reported as a parasitic fauna in Iran.

Comparative prevalence of hepato-pulmonary hydatidosis among native and imported cattle in north of Sistan and Baluchestan: Iran.

The objective of present study was comparison of hepato-pulmonary hydatidosis among imported and native cattle. The present study was performed during 2011-2012. 2,657 and 525 imported and native cattle were inspected in slaughter house of Zahedan and Zabol respectively. The present and number of hydatid cysts were evaluated. Also the fertility rate of cysts has been estimated. The prevalence of infection in imported and native cattle were 15.1 and 5.3 % respectively and the statistical correlation was significant (P value <0.001). Also the fertility rate of hydatid cysts in the imported and native cattle were 16 and 18.2 % respectively and the statistical correlation was not significant (P value >0.05). The results of present study shows significant differences in hydatidosis rate between native Iranian and imported cattle in Sistan and Baluchestan provinces of Iran. Slaughtering of imported livestock should be done in industrial centers to prevent potential increasing in canine echinococcosis and consequently human and farm animals hydatidosis.

Comparison of internal transcribed spacers and intergenic spacer regions of five common Iranian sheep bursate nematodes.

BACKGROUND: Accurate identification of sheep nematodes is a critical point in epidemiological studies and monitoring of drug resistance in flocks. However, due to a close morphological similarity between the eggs and larval stages of many of these nematodes, such identification is not a trivial task. There are a number of studies showing that molecular targets in ribosomal DNA (Internal transcribed spacer 1, 2 and Intergenic spacer) are suitable for accurate identification of sheep bursate nematodes. The objective of present study was to compare the ITS1, ITS2 and IGS regions of Iranian common bursate nematodes in order to choose best target for specific identification methods. METHODS: The first and second internal transcribed spacers (ITS1and ITS2) and intergenic spacer (IGS) of the ribosomal DNA (rDNA) of 5 common Iranian bursate nematodes of sheep were sequenced. The sequences of some non-Iranian isolates were used for comparison in order to evaluate the variation in sequence homology between geographically different nematode populations. RESULTS: Comparison of the ITS1 and ITS2 sequences of Iranian nematodes showed greatest similarity among Teladorsagia circumcincta and Marshallagia marshalli of 94% and 88%, respectively. While Trichostrongylus colubriformis and M. marshalli showed the highest homology (99%) in the IGS sequences. Comparison of the spacer sequences of Iranian with non-Iranian isolates showed significantly higher variation in Haemonchus contortus compared to the other species. CONCLUSION: Both the ITS1 and ITS2 sequences are convenient targets to have species-specific identification of Iranian bursate nematodes. On the other hand the IGS region may be a less suitable molecular target.

Epidemiological study of gastrointestinal helminthes of canids in chaharmahal and bakhtiari province of iran.

BACKGROUND: The present study was carried out to describe the epidemiological aspects of gastrointestinal helminthic infections of canids in Chaharmahal and Bakhtiari Province, the central western part of Iran. METHODS: Forty nine canid species including, dogs, jackals, foxes and wolves were included in this study. The contents of their alimentary canal were inspected in order to isolate and identify the parasitic helminthes of this system. To identify the worms, the Soulsbey and Anderson identification key and light microscopy were used. RESULTS: Based on necropsy findings, 35 (71.4%) of examined animals were infected with at least one helminth. The prevalence of identified worms was as follows: Mesocestoides lineatus (55.1%), Joyeuxiella echinorinchoides (26.5%), Taenia hydatigena (12.2%), T. multiceps (8.2%), T. ovis (2%), Dipylidium caninum (2%) and Spirura spp. (2%). No significant difference was noticed between the sampling areas, age and helminth infection. Only a significant difference was observed for prevalence of T. multiceps in wolf (25%), dog (21.4%), jackal and fox (0%), respectively (P < 0.05). CONCLUSION: The canids in Chaharmahal and Bakhtiari harbor several parasites that some kind of them have zoonotic importance and may pose a threat to community health specially in rural areas.

Prevalence of parasites and associated risk factors in domestic pigeons (Columba livia domestica) and free-range backyard chickens of Sistan region, east of Iran.

This study was carried out on free-range backyard chickens and domestic pigeons (Columba livia domestica) from December 2010 to November 2011 to determine the prevalence, intensity and species of internal and external parasites in Sistan region, east of Iran. Of the total of 59 (27 males and 32 females) free-range backyard chickens and 46 (26 males and 20 females) domestic pigeons inspected, 55 (93.22 %) and 39 (84.78 %) were infected respectively. Ten species of free-range backyard chickens parasites were collected from alimentary canals, body, head and neck, comprising of 3 species of nematodes, 4 species of cestodes and 3 species of ectoparasites as follows: Ascaridia galli (16.94 %), Heterakis gallinarum (23.72 %), Subulura brumpti (67.79 %), Raillietina tetragona (35.59 %), Raillietina echinobothrida (27.11 %), Raillietina cesticillus (15.25 %), Choanotaenia infundibulum (40.67 %), Argas persicus (16.94 %), Menopen gallinae (55.93 %) and Menacanthus stramineus (33.89 %). The domestic pigeons were infected with seven species of parasites including 2 species of nematodes, 2 species of cestodes and 3 species of ectoparasites as follows: Ascaridia colombae (15.21 %), Hadjelia truncata (17.39 %), Raillietina tetragona (26.08 %), Raillietina echinobothrida (28.26 %), Argas reflexus (13.04 %), Menopen gallinae (32.60 %), Columbicola Columba (41.30 %). This is the first survey to determine the prevalence and intensity of parasites among free-range backyard chicken and domestic pigeon species in Sistan region.

Application of polyaniline for the reduction of toxic Cr(VI) in water.

Various oxidation states of polyaniline, as film and powder, were used for reduction of highly toxic Cr(VI) to less toxic Cr(III) ion. The effects of various parameters such as synthesis method, physical and oxidation state of polyaniline, film thickness, solution pH and initial Cr(VI) concentration on the kinetics and efficiency of reduction process were investigated. Results showed that a very broad concentration range of Cr(VI) solutions (10-10(5)ppb by emeraldine base and 10-10(6.5)ppb by leucoemeraldine) can be efficiently (>98%) reduced by polyaniline as film or powder. Cr(VI) solutions with concentrations higher than these ranges were caused to the overoxidation and degradation of polymer. Decreasing of solution pH, increased the kinetics and performance of reduction process, but lowered the Cr(VI) concentration ranges caused the overoxidation and degradation of the polymer. Higher reduction efficiencies were obtained for thicker electrochemically synthesized polyaniline films and for thinner chemically prepared polyaniline free standing films. The ability or capacity of various forms of polyaniline in Cr(VI) reduction was evaluated as the milligrams of Cr(VI) reduced by one gram of various forms of polyaniline.