Vairimorpha ceranae was the only detected microsporidian species from Iranian honey bee colonies: a molecular and phylogenetic study.

Nosemosis caused by Vairimorpha ceranae is one of the most important threats to honeybee colonies worldwide. This study aimed to determine the prevalence and intensity of Vairimorpha infection in different types of colonies and locations in Iran. In October 2017 and May 2018, 376 colonies from 97 apiaries were selected for each month according to a randomly clustered design. By considering 3-5 colonies for each apiary, 20 adult bees as pooled samples were collected from each colony. In microscopic analysis, 46.52% and 46.1% of samples in May and October showed Vairimorpha spores, respectively. The infection intensities in May and October were 5.94 ± 0.19 (× 106) and 5.86 ± 0.23 (× 106) spores/bee in a pooled sample, respectively. The mean infection intensity ranged from 1.8 to 12.5 (× 106) spores/bee. Statistically, there were no significant differences in the prevalence and intensity of V. ceranae infection between May and October samples. No significant differences were found among the prevalence rates of infection in the types of colonies; however, the intensity was significantly higher in migratory and mountainous colonies in May and only in migratory colonies in October. There was a significant correlation between the prevalence and intensity of V. ceranae infection (r2 = 0.695). PCR analysis showed that the samples were only infected with V. ceranae. No intraspecific variation to V. ceranae was found by direct sequencing of the amplified fragment of 16S rRNA. The obtained sequence was mainly 100% similar to those of V. ceranae isolates from European countries.

Evaluation of bioaccumulation of some heavy metals in liver flukes (Fasciola hepatica and Dicrocoelium dendriticum) and liver samples of sheep.

Different living organisms are used as applicable bioindicators to determine heavy metal pollutions. Recent studies have shown that helminths parasites can be used as efficient environmental sentinels. This study aimed to evaluate Fasciola hepatica and Dicrocoelium dendriticum as bioaccumulators of lead (Pb), chromium (Cr), cadmium (Cd), copper (Cu). For this work. A total of 50 samples (F. hepatica, D. dendriticum, and livers from the infected and uninfected sheep, each of 10 samples) were collected from sheep slaughtered in Tabriz abattoir. One gram of each sample was incinerated and analyzed by Flame Atomic Absorption Spectrometry. The analysis of samples showed that Pb, Cr and Cu values in F. hepatica were higher than those in D. dendriticum, but only the differences of Pb and Cu were significant. The values of heavy metals in F. hepatica were significantly higher than those in the infected livers (except for Cd), while in D. dendriticum, Cr and Cd were only higher. Based on metal levels in livers, it was found that bioconcentration factors (BCFs) of Cr, Pb and Cu for F. hepatica were much more than one, and BCFs of these three metals between two flukes were statistically significant. This study indicated that F. hepatica had a higher bioindicator potential than D. dendriticum to evaluate environmental pollutants by some metals.

Anthelmintic activity of crude powder and crude aqueous extract of Trachyspermum ammi on gastrointestinal nematodes in donkey (Equus asinus): An in vivo study.

ETHNOPHARMACOLOGICAL RELEVANCE: Trachyspermum ammi has been used traditionally as a popular ethnobotanical plant in human and animal parasitic infestations. Few scientific studies have been conducted on in vitro anthelmintic activity of T. ammi against various helminths and there is no study on its in vivo/in vitro anthelmintic properties against equine helminths. AIM OF THE STUDY: The present study aimed to evaluate in vivo anthelmintic activity of crude powder (CP) and crude aqueous extract (CAE) of T. ammi seeds against gastrointestinal nematodes in the donkey. MATERIALS AND METHODS: Thirty donkeys, naturally and severely infected with Strongyle-type nematodes, were randomly divided into six groups and administered orally with graded doses of CP and CAE at single dose as follow: Two groups were treated with 1 and 3 gkg-1 body weight of CP, two groups were treated with CAE at the equivalent dose rate 1 and 3 gkg-1 bw of CP and one group was treated with 200 µgkg-1 bw of Ivermectin 0.2% (Positive Control). One group received no medication (Negative Control). Efficacy was determined by faecal egg count reduction (FECR) test. The faecal samples were taken from the donkeys on days 7, 14 and 28 post-treatments (PT) and the egg per gram (EPG) of faeces was determined for each animal. FECR for donkeys treated with CP, CAE, and Ivermectin 0.2% was calculated on days 7, 14, and 28 PT. In the present study, CAE was analyzed using gas chromatography-mass spectrometry (GC-MS) to determine CAE chemical components. High-performance liquid chromatography (HPLC) using a marker (thymol) was applied to confirm the existence of thymol as an active principle. RESULTS: Based on the FECR findings, the maximum FECR (71.2%) was observed in the group treated with CAE3 on day 14 PT, while it was 66.2% for CP3 on the same day. The effects of different treatments (P < 0.001), time (P < 0.001) and their interactions (P < 0.05) on EPG showed a significant difference. A dose-dependent response in the anthelmintic activity of CP and CAE was observed. GC-MS analysis indicated that thymol was the major component of CAE (Area = 45.01%) with the respective peak at retention times (Rt) 10.41 min. On HPLC analysis of CAE, thymol was found to be the major constituent of CAE (Area = 58.76%) with the same Rt at 13.233 min. CONCLUSION: The present study indicated that both CP and CAE of T. ammi had the dose-dependent anthelmintic effects. Due to the development of drug resistance against synthetic anti-helminthics, T. ammi seems to be a promising alternative for the treatment and prevention of helminths in livestock.

Evaluation of oxidative stress and antioxidant status, serum trace mineral levels and cholinesterases activity in cattle infected with Anaplasma marginale.

This study was undertaken to assess the influence of an Anaplasma marginale infection on oxidative stress and antioxidant status, trace elements and cholinesterase as markers of the inflammatory process and biomarkers of oxidative imbalance. An infected group comprised of 35 crossbred Holstein cattle, about 2-3 years old, naturally infected with Anaplasma marginale, were divided into 4 subgroups according to their parasitemia rates (<1%, 1-10%, 10-20%, >20%) and also 10 healthy cattle as control were selected. Blood samples were taken and hematological parameters, activities of antioxidant enzymes including erythrocyte glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), catalase (CAT), glucose-6-phosphate dehydrogenase (G6PD), total antioxidant capacity (TAC), median corpuscularfragility (MCF) as well as acetylcholinesterase (AChE), and serum concentrations of antioxidant trace minerals (copper, iron, zinc, manganese, and selenium) and butyrylcholinesterase (BChE) were determined. In addition, as an index of lipid peroxidation, the level of malondialdehyde (MDA) was measured. The results revealed a significant decrease (P < 0.05) in RBC count, packed cell volume (PCV) and Hb concentration as well as the activities of erythrocyte GSH-Px, SOD, CAT, G6PD, TAC, MCF and AChE and serum concentrations of Cu, Zn, Mn, Se and BchE in the infected cattle. In contrast, significantly increased (P < 0.05) levels of MDA and erythrocyte osmotic fragility as well as serum concentration of iron were recorded in the infected animals. The significant decrease in antioxidant enzyme activities and substantial elevated levels of lipid peroxidation and erythrocyte osmotic fragility associated with the notable increase in parasitemia indicate increased exposure of RBCs to oxidative damage. Furthermore, decrease of cholinesterase in infection by A. marginale can and directly or indirectly lead to increase acetylcholine levels potent anti-inflammatory molecules, thereby inhibiting inflammation.

Semi-nested polymerase chain reaction-based detection of Babesia spp. in small ruminants from Northwest of Iran.

AIM: The present study aimed to detect Babesia ovis and Babesia motasi in the blood samples of sheep and goats from Northwest of Iran by the semi-nested polymerase chain reaction (PCR) technique. MATERIALS AND METHODS: A total of 166 whole blood samples (including 123 sheep and 43 goats) were collected. In the first stage, the PCR was performed to amplify a piece of 18S rRNA gene of Babesia and Theileria genera. Then, semi-nested PCR was carried out on all PCR products to differentiate B. ovis and B. motasi. RESULTS: The PCR indicated that totally, 19 (11.44%) out of 166 samples were positive for Babesia or Theileria spp. The semi-nested PCR showed that 38 samples (22.89%) were positive only for B. ovis. No significant association was found between the infection rate of B. ovis and age, gender and species of animals. CONCLUSION: In the present study, there was no evidence for B. motasi infection in small ruminants from Northwest of Iran. Therefore, B. ovis was the main causative agent of ovine Babesiosis in this region.

Molecular Determination of Fasciola Spp. Isolates from Domestic Ruminants Fecal Samples in the Northwest of Iran.

BACKGROUND: Fasciola species are the main causes for fascioliasis with great financial losses and are among the most important food/water-borne parasites worldwide. The basic proceedings such as epidemiology and effective control of fascioliasis rely mainly on precise identification of Fasciola species. The present study was conducted to determine the Fasciola species in ruminant fecal samples from East Azerbaijan Province in Iran. METHODS: Overall, 2012 fecal samples were collected and processed initially for microscopic examination of Fasciola eggs in 2014-15. Then, recovered eggs were subjected to molecular identification. A fragment of 618 bp of the 28S rRNA gene pertaining to Fasciola genus was amplified under PCR. The amplified fragment was restricted by fast digest Ava II enzyme in order to a Restriction Fragment Length Polymorphism. RESULTS: Based on microscopic examination, 72 samples were infected, from which, 10 and 62 cases pertained to cattle and sheep samples respectively. Based on RFLP, the PCR products restricted by the Ava II restriction enzyme produced 529 bp fragments only. According to the positive controls, all restriction patterns were related to Fasciola hepatica, while no restriction patterns were linked to F. gigantica. CONCLUSION: Based on PCR-RFLP, F. hepatica was dominant species in animals of the studied areas and no evidence of F. gigantica was observed. Therefore, further field studies to verify these results are suggested.

Morphological and molecular discrimination of fasciola species isolated from domestic ruminants of urmia city, iran.

BACKGROUND: The trematodes of the genus Fasciola (the liver flukes) are among the well-known instances of food-borne parasites worldwide. Differentiation of Fasciola species is important because of their different transmission and epidemiological characteristics. The current study was undertaken to discriminate Fasciola species in the domestic ruminants of Urmia city, Iran. METHODS: Adult flukes were isolated from the naturally infected livers of the slaughtered water buffaloes and sheep. The flukes were initially identified based on morphological and morphometric parameters. A 618-bp-long fragment of the 28SrRNA gene of Fasciola was amplified by polymerase chain reaction (PCR). The amplified fragment was digested by DraII or AvaII enzymes for a restriction fragment length polymorphism (RFLP) analysis and sequenced for the phylogenetic tree construction. RESULTS: Based on the morphometric examination, the flukes belonged to F. hepatica, F. gigantica and an intermediate Fasciola form. The PCR-RFLP analysis was able to differentiate F. hepatica from F. gigantica. While the phylogenetic reconstruction justified, to some extent, the morphological diagnosis, it failed to segregate F. hepatica from F. gigantica identified in this and the previous studies. CONCLUSION: To resolve fully the problem of taxonomy and evolution in Fasciola species, employing a broad range of molecular and morphological approaches is necessary. This is crucial for epidemiological surveys and successful clinical management of their infection.

Seasonal and Geographic Distribution of Cercarial Infection in Lymnaea gedrosiana (Pulmunata: Lymnaeidae) In North West Iran.

BACKGROUND: Trematodes are a diverse group of endoparasites which require molluscan and vertebrate animals as intermediate and definitive hosts in their life cycle. The present study was carried out to determine the diversity and geographic distribution of infection with trematodes'cercariae in the snail Lymnaea gedrosiana from north-west Iran. METHODS: A total number of 6759 Lymnaeidae snails were collected from 28 snail habitats; of these L. gedrosiana was the prevalent snail (74.37%) which examined for cercarial infection by shedding method. RESULTS: The overall infection rate was 8.03%. The most frequent trematodes cercariae in the snail were xiphidiocercariae (81.98%), furcocercariae (32.26%), echinostome cercariae (5.19%), and monostome cercariae (1.24%). The highest infection rate in L. gedrosiana (100%) was with echinostome cercariae from Golestaneh in autumn. CONCLUSION: Due to the important role of pond snails in transmission of cercariae to fish as a source of zoonotic diseases, it is essential to estimate the distribution and abundance of the snails and the rate of their infection with different trematodes' cercariae, and establish control programs in each region.

Molecular study for detecting the prevalence of Fasciola gigantica in field-collected snails of Radix gedrosiana (Pulmonata: Lymnaeidae) in northwestern Iran.

Fasciolosis is an important disease in veterinary medicine worldwide, and is a cause of great economic loss in livestock husbandry in Iran. This study was aimed to determine prevalence of Fasciola gigantica infection in field-collected snails of Radix gedrosiana in northwestern Iran. The snails were collected from 28 perennial and seasonal freshwater habitats from May to December 2010 and identified. A fragment of 618 bp of 28s rRNA gene was amplified by polymerase chain reaction (PCR). The PCR products were subjected to restriction fragment length polymorphism (RFLP) using DraII and AvaII enzymes. PCR-RFLP patterns revealed that 3.12% of the snails were infected with F. gigantica. It was also found that the infected snails had a limited distribution over the water bodies located in the central part of the region. It was concluded that PCR-RFLP was a reliable approach to detect Fasciola infection in pond snails and may be useful to establish control measures for livestock and humans' fasciolosis in the region.