Evaluation of hematological changes, oxidant/antioxidant status and immuno-logical responses in sheep and goats naturally infected with Linguatula serrata.

Linguatula serrata is a worldwide zoonotic food-borne parasite. The parasite is responsible for linguatulosis and poses a concern to human and animal health in endemic regions. This study investigated the hematological changes, oxidant/antioxidant status and immunological responses in goats and sheep naturally infected with L. serrata. Hematological changes, antioxidant enzymes and malondialdehyde (MDA) levels were measured. The level of inter-leukin-2 (IL-2), IL-4, IL-5, IL-10, and tumor necrosis factor alpha (TNF-α) mRNA expression was investigated in lymph nodes. According to the hemogram results, eosinophils were significantly increased in the infected goats and sheep, and Horizontal Gene Transfer (HGT), hematocrit (HCT), and mean corpuscular hemoglobin concentration (MCHC) were significantly decreased. The levels of MDA and the activity of glutathione peroxidase (GPx) were significantly higher in infected animals than in non-infected animals. However, the activity of superoxide dismutase (SOD) and catalase (CAT) was significantly lower in infected animals than in non-infected animals. A comparison of the cytokine mRNA expression in lymph nodes from infected and non-infected animals showed higher cytokine expression in the infected animals. Infection with L. serrata caused microcytic hypochromic and normocytic hypochromic anemia in goats and sheep. The inconsistent results of immunological changes were found in infected goats and sheep. In both animals, oxidative stress occurred and led to an increase in lipid peroxidation. L. serrata created a cytokine microenvironment biased towards the type 2 immune responses.

An Enzyme-Linked Immunosorbent Assay for the Detection of Antibodies to Linguatula serrata in Experimentally Infected Dogs.

BACKGROUND: Linguatula serrata is a causative agent of visceral and nasopharyngeal linguatulosis in humans and animals. The aim of the present study was to investigate the immune response of dogs experimentally infected by L. serrata with ELISA. METHODS: Five puppies were infected by inserting the L. serrata nymphs in their nasal cavities (infected group) in the Department of Parasitology of Shahid Chamran University of Ahvaz, during 2018-2019. Three animals were kept as the non-infected control group. Blood samples were collected from the animals for seven months at approximately monthly intervals for serum preparation. Nasal samples were taken weekly from the fourth month. ELISA was designed and performed on 64 sera (24 negatives, and 40 positives) using somatic (S), and excretory-secretory (ES) antigens. RESULTS: Overall, 100% of the animals were infected with the parasite. Based on the results of ELISA, the ES antigen (sensitivity 95% and specificity 92%) was more preferred than the S antigen (sensitivity 95% and specificity 85%). Female parasites had significant effects on the immune response. There was a significant correlation between the clinical symptoms and the presence of female parasites (P<0.05). CONCLUSION: The results showed a practical method for dogs' experimental infection. ELISA method is suitable for the detection of infection at different stages of development, especially before the maturation stage of the parasite. In this regard, the ES antigen of the parasite was more immunogenic. Therefore, ELISA can be used as a serological method in the early detection and epidemiological studies of infection with L. serrata in dogs.

The Effects of Intensive Blood Pressure Control on Cardiovascular Outcomes Based on 10-Year ASCVD Risk Score: An Analysis of a Clinical Trial.

There is still controversy about whether clinicians should include cardiovascular disease (CVD) risk stratification into the consideration for treatment of hypertension. This was a post hoc analysis of the Systolic Blood Pressure Intervention Trial (SPRINT). A total of 9361 nondiabetic patients without a history of stroke were randomly assigned to the intensive-treatment group (with an SBP target of <120 mm Hg) and the standard-treatment group (with an SBP target of <140 mm Hg). The patients were categorized into four groups based on the Atherosclerotic Cardiovascular Disease (ASCVD) risk score. The groups contained participants with ASCVD < 7.5%, 7.5% ≤ ASCVD <10%, 10% ≤ ASCVD < 15%, and ASCVD ≥ 15%. The incidence of the primary outcome, secondary outcome, and serious adverse events was compared between the two groups. The primary outcome was a composite of nonfatal myocardial infarction (MI), acute coronary syndrome (ACS) not resulting in MI, stroke, acute decompensated heart failure (HF), or death from cardiovascular causes. The secondary outcomes consisted of the individual components of the primary outcome and all-cause death. Intensive blood pressure (BP) control significantly reduced the incidence of primary outcome event in patients with 10% ≤ ASCVD < 15% (hazard ratio (HR) 0.593; 95% confidence interval (CI) 0.361-0.975; P = 0.039) and ASCVD ≥ 15% (HR 0.778; CI 0.644-0.940; P = 0.009). Intensive BP control was also beneficial for the primary prevention of cardiovascular events in patients with an ASCVD risk of 7.5-10% (HR 0.187; 95% CI 0.040-0.862; P = 0.032). However, intensive treatment was associated with higher incidence of hypotension and acute renal failure in participants with ASCVD ≥ 15%. In patients without diabetes mellitus and prior stroke who had a 10-year risk of cardiovascular events above 10% based on the ASCVD risk score, intensive BP control played an important role in the reduction of major cardiovascular events. Additionally, intensive treatment would be beneficial for primary prevention in patients with ASCVD ≥ 7.5% without previous history of any cardiovascular disorders. Trial registration: ClinicalTrials.gov number; the trial is registered with NCT01206062.

Novel mutations in the PHKB gene in an iranian girl with severe liver involvement and glycogen storage disease type IX: a case report and review of literature.

BACKGROUND: Glycogen storage disease (GSD) type IXb is one of the rare variants of GSDs. It is a genetically heterogeneous metabolic disorder due to deficient hepatic phosphorylase kinase activity. Diagnosis of GSD can be difficult because of overlapping manifestations. Mutation analysis of the genes related to each type of GSD is supposed to be problem-solving, however, the presence of novel mutations can be confusing. In this case report, we will describe our experience with a young girl with the diagnosis of GSD and two novel mutations related to GSD type IXb. CASE PRESENTATION: A 3-year- old girl presented with short stature, hepatomegaly, and liver cirrhosis. No specific diagnosis was made based on laboratory data, so liver biopsy and targeted-gene sequencing (TGS) were performed to find out the specific molecular basis of her disease. It was confirmed that the patient carries two novel variants in the PHKB gene. The variant in the PHKB gene was classified as pathogenic. CONCLUSIONS: This is the first reported case of a dual molecular mutation of glycogen storage disease type IXb in the same patient. Two novel variants in PHKB were identified and one of them was a pathogenic split-site mutation. In conclusion, for the first time, identification of the novel variants in this patient expands the molecular and the phenotype basis of dual variants in GSD-IXb.

Clinical and genetic spectrum of glycogen storage disease in Iranian population using targeted gene sequencing.

Glycogen storage diseases (GSDs) are known as complex disorders with overlapping manifestations. These features also preclude a specific clinical diagnosis, requiring more accurate paraclinical tests. To evaluate the patients with particular diagnosis features characterizing GSD, an observational retrospective case study was designed by performing a targeted gene sequencing (TGS) for accurate subtyping. A total of the 15 pediatric patients were admitted to our hospital and referred for molecular genetic testing using TGS. Eight genes namely SLC37A4, AGL, GBE1, PYGL, PHKB, PGAM2, and PRKAG2 were detected to be responsible for the onset of the clinical symptoms. A total number of 15 variants were identified i.e. mostly loss-of-function (LoF) variants, of which 10 variants were novel. Finally, diagnosis of GSD types Ib, III, IV, VI, IXb, IXc, X, and GSD of the heart, lethal congenital was made in 13 out of the 14 patients. Notably, GSD-IX and GSD of the heart-lethal congenital (i.e. PRKAG2 deficiency) patients have been reported in Iran for the first time which shown the development of liver cirrhosis with novel variants. These results showed that TGS, in combination with clinical, biochemical, and pathological hallmarks, could provide accurate and high-throughput results for diagnosing and sub-typing GSD and related diseases.

Novel PRKAG2 variant presenting as liver cirrhosis: report of a family with 2 cases and review of literature.

BACKGROUND: Mutations in the PRKAG2 gene encoding the 5' Adenosine Monophosphate-Activated Protein Kinase (AMPK), specifically in its γ2 regulatory subunit, lead to Glycogen storage disease of heart, fetal congenital disorder (PRKAG2 syndrome). These mutations are rare, and their functional roles have not been fully elucidated. PRKAG2 syndrome is autosomal dominant disorder inherited with full penetrance. It is characterized by the accumulation of glycogen in the heart tissue, which is clinically manifested as hypertrophic cardiomyopathy. There is little knowledge about the characteristics of this disease. This study reports a genetic defect in an Iranian family with liver problems using targeted-gene sequencing. CASE PRESENTATION: A 4-year-old girl presented with short stature, hepatomegaly, and liver cirrhosis. As there was no specific diagnosis made based on the laboratory data and liver biopsy results, targeted-gene sequencing (TGS) was performed to detect the molecular basis of the disease. It was confirmed that this patient carried a novel heterozygous variant in the PRKAG2 gene. The echocardiography was a normal. CONCLUSION: A novel heterozygous variant c.592A > T (p.Met198Leu) expands the mutational spectrum of the PRKAG2 gene in this family. Also, liver damage in patients with PRKAG2 syndrome has never been reported, which deserves further discussion.

A Preliminary Survey on Gastrointestinal Parasites of Domestic Ducks in Ahvaz, Southwest Iran.

BACKGROUND: Despite ducks being birds resistant to infection, the favorable habitat of ducks such as subtropical climate or stagnant water is also a perfect place for survival of the parasites. METHODS: This study was conducted from Dec 2014 to Apr 2015 to determine the prevalence of gastrointestinal parasites of domestic ducks in Ahvaz and environs, southwest of Iran. Overall, 41 fresh fecal samples were collected and prepared using formol-ether concentration, modified Ziehl-Neelsen, sheather`s floatation and zinc sulfate sedimentation methods. Light microscopic morphometry was used for identification of helminth eggs and oocysts. RESULTS: 60.97% of ducks were infected with three different nematodes and/or four protozoan parasites. The identified nematodes were Capillaria sp., (50%) Subulura spp. (16.66%) and Echinuria spp. (33.33%). The protozoan oocystes were Cryptosporidium spp. (50%) and coccidian species (%58.33) and included Wenionella philiplevinei, Tyzerria spp. and Isospora. mandari. Mixed infection with two or more parasites was common. Twenty (80%) had single, four (16%) double and one (4%) triple infection. CONCLUSION: This is the first report of coccidian infection in domestic ducks of Iran. Further studies will be necessary on epidemiology and pathogenicity of the parasitic infections in ducks of this area.

Prevalence of Cryptosporidium spp. in ruminants of Lorestan province, Iran.

Cryptosporidiosis is one of the important zoonotic diseases caused by an intracellular protozoan parasite called Cryptosporidium. This study aimed to investigate the prevalence of Cryptosporidium spp. infection on 1,115 ruminants, cattle, sheep and goats, in Lorestan province, Iran. Using formol-ether concentration technique and modified Ziehl-Neelsen staining method afterwards, the overall prevalence of Cryptosporidium spp. infection in ruminants of Lorestan province was 7.17 %. Prevalence of infection was 9.07 % (39 of 430), 5.80 % (20 of 345) and 6.18 % (21 of 340) for cattle, sheep and goats respectively. There was no significant difference between contamination of all examined animals and different geographical and climatic situations (P > 0.05) and diarrhea was not directly associated with Cryptosporidium infection (P > 0.05). In conclusion, the prevalence of cryptosporidiosis in Lorestan province was relatively low, but it should be noticed that this opportunistic parasite is zoonosis and also can make epidemics in ruminants as well as human population in suitable conditions.

Occurrence and Intensity of Anisakid Nematode Larvae in Some Commercially Important Fish Species in Persian Gulf.

BACKGROUND: Anisakid nematodes are common parasites of fish, mammals, fish-eating birds, and reptiles with a worldwide distribution, causing diseases in human, fish and important economic losses. METHODS: A preliminary epidemiological study was carried out on Anisakid nematodes larvae in some commercially important fish species to evaluate the anisakid nematode larvae from greater lizardfish, (Saurida tumbil), Japanese thread fin bream (Nemipterus japonicus), crocodile longtom (Tylosurus crocodilus crocodiles) and longfin trevally (Carangoides armatus) from the Persian Gulf of Iran. RESULTS: The collected larvae were identified mainly as the third larval stage (L3) of Hysterothylacium larval type A, B and C, Anisakis sp., Raphidascaris sp., Pseudoterranova sp. and Philometra sp. (Nematoda: Philometridae). The prevalence of Anisakid larvae infection of examined fishes was 97.2% in N. japonicus, 90.3% in S. tumbil, 20.5% in crocodile longtom and 5.5% in longfin trevally. Anisakis type III for the first time was different from Anisakis type I and Anisakis type II. DISCUSSION: Zoonotic anisakids by high prevalence in edible fish could be a health hazard for people. So health practices should be considered in these areas.

An alternative method for producing Toxocara canis second stage larvae from a paratenic host (pigeon) for mRNA extraction purpose.

Toxocara canis is a prevalent zoonotic parasite which can cause serious disease in puppies and humans. Excretory-secretory and coating antigens of the second stage larvae (L2) are the best targets for performing immunodiagnostic and also immunoprophylactic tests. Various hatching methods have been described to bring out L2 from the resistant infective egg shell; but these methods are difficult to do and have had different results when performed by different practitioners. In this study, second stage larvae were obtained from the viscera of pigeons (a paratenic host) which were infected with infective eggs. Infective Toxocara canis eggs were given to ten pigeons and live larvae were recovered from their excised livers and lungs by using the Baermann's apparatus in the next days. Two in vitro methods for larvae hatching were also performed including a so-called physiological hatching method according to Ponce-Macotela et al. (J Parasitol 175:382-385, 2010), and a mechanical hatching method according to Alcântara-Neves and Santos (J Exp Parasitol 119:349-351, 2008) and their results were compared with the in-vivo method. Results show that averagely 36.2 % of fed larvae recovered from livers and 0.15 % from lungs. Average larvae recovery in the first day after infection (24.2 %) was significantly lower than subsequent days (39 %). Maximum larvae recovered in day 3 (55 %). In-vitro methods we carried out did not have acceptable results and only a few larvae did hatch using these methods.

Molecular detection of Sarcocystis species in slaughtered sheep by PCR-RFLP from south-western of Iran.

Sarcocystis spp. are the cyst forming protozoan parasites that prevalent in livestock all around the world. In the presented work, we examined 40 macroscopic and 40 microscopic sarcocysts from Khouzestan and Lorestan provinces, south-western Iran, utilizing PCR-RFLP based on amplification of 18S rRNA gene. Using AvaI, HindII, TaqI and EcoRI restriction enzymes the results represented Sarcocystis gigantea in both macroscopic and microscopic cysts. This result supports the importance of molecular investigations on characterization of Sarcocystis species when we aimed to assess the reliable control and preventive programs.

Isolation and identification of excretory-secretory and somatic antigens from the Oestrus ovis larvae by SDS-PAGE and immunoblotting.

Oestrus ovis is an economically important parasite of small ruminants and a zoonotic parasite with many reports of ophthalmomyiasis in human from Iran and other countries. The aim of the peresent study was the isolation and identification of excretory-secretory (ES) and somatic (S) antigens of O. ovis second and third stage larvae (L2, L3) collected from Arabi sheep breeds located in southwest of Iran. Positive sera were prepared by marking the sheep, taking blood sample and direct observation of the parasite in the head. Somatic antigens of the larvae (SL2, SL3) were prepared by sonication. Larval excretory-secretory antigens (ESL2, ESL3) were prepared by incubation the larvae in RPMI-1640 RPMI medium. Electrophoretic protein profiles of ESL2 two, ESL3 seven, SL2 eight, SL3 fifteen bands (from 79.0 to below 14.4 KDa) were shown. In immunoblotting with positive sera, four common bands in SL2 and SL3 at 58, 42.0, 29.0 and 28.0 kDa, one specific band in SL3 at 47.0 kDa and one band in ESL2, at 28.0 kDa, and three bands in ESL3 at 58.0, 42.0, 29.0 and 28.0 kDa were recognized. Among the profiles, the 28 kDa protein was the most common antigenic component. Nevertheless, the antigenic proteins 29, 58 kDa were a common protein in electrophoretic patterns of both S and ES proteins of L2 and L3 but, 42.0 kDa antigen the only one detected in immunoblot but not in S and ES protein profiles of the larvae. Therefore, the antigens 29.0, 42.0 and 58.0 kDa can be used for further studies of protective effects and serological diagnostic methods.