Anticancer Activity of Extremely Effective Recombinant L-Asparaginase from Burkholderia pseudomallei.

L-asparaginase (E.C. 3.5.1.1) purified from bacterial cells is widely used in the food industry, as well as in the treatment of childhood acute lymphoblastic leukemia. In the present study, the Burkholderia pseudomallei L-asparaginase gene was cloned into the pGEX-2T DNA plasmid, expressed in E. coli BL21 (DE3) pLysS, and purified to homogeneity using Glutathione Sepharose chromatography with 7.26 purification fold and 16.01% recovery. The purified enzyme exhibited a molecular weight of ~33.6 kDa with SDS-PAGE and showed maximal activity at 50°C and pH 8.0. It retained 95.1, 89.6%, and 70.2% initial activity after 60 min at 30°C, 40°C, and 50°C, respectively. The enzyme reserved its activity at 30°C and 37°C up to 24 h. The enzyme had optimum pH of 8 and reserved 50% activity up to 24 h. The recombinant enzyme showed the highest substrate specificity towards L-asparaginase substrate, while no detectable specificity was observed for L-glutamine, urea, and acrylamide at 10 mM concentration. THP-1, a human leukemia cell line, displayed significant morphological alterations after being treated with recombinant L-asparaginase and the IC50 of the purified enzyme was recorded as 0.8 IU. Furthermore, the purified recombinant L-asparaginase improved cytotoxicity in liver cancer HepG2 and breast cancer MCF-7 cell lines, with IC50 values of 1.53 and 18 IU, respectively.

A new myxozoan parasite, Myxobolus allami sp. n. (Myxozoa: Myxobolidae) from the intestinal wall of Sparidentex hasta (Valenciennes) in Arabian Gulf.

Myxobolus allami sp. n. is described from the intestinal wall of the silvery black porgy, Sparidentex hasta (Valenciennes), off Saudi Arabian coast of Arabian Gulf. Two of 20 examined fish were found to be infected with irregular-shaped plasmodia 3-8 mm long × 2-3 mm wide. Mature myxospores are subspherical to elliptical in the valvular view and oval in the sutural view, and are 11-13 (12) µm long, 7-8 (7.5) µm wide and 10-12 (10.8) µm thick. Spores have relatively thin valves and mostly (~ 72%) end with short caudal appendages of ~3 µm long. The spores also have two polar capsules, which are oval to elliptical and measure 5-7 (5.7) µm in length and 2-3 (2.7) µm in width. Polar filaments are coiled, with three turns. Transmission electron microscopy revealed that caudal appendages originated from the sutural edge at the posterior pole of the myxospore with density similar to that of its valves. The SSU rRNAgene sequence of the present species does not match any available sequences in GenBank. Phylogenetically, this species is sister to Myxobolus khaliji Zhang, Al-Qurausihy et Abdel-Baki, 2014 within a well-supported clade of Myxobolus-Henneguya with species infecting marine fishes. The combination of molecular data and morphological differences between this and other species of Myxobolus Bütschli, 1882 lead us to propose that the present form be established as a new species, M. allami. The present study also provides more evidence for the idea that caudal appendages cannot be reliably used to distinguish the species of the genera Myxobolus and Henneguya Thélohan, 1892.

Haemonchus contortus Susceptibility and Resistance to Anthelmintics in Naturally Infected Egyptian Sheep.

PURPOSE: Susceptibility and resistance of Haemonchus contortus to anthelmintic drugs, including ivermectin, levamisole, and albendazole in naturally infected sheep were investigated. METHODS: Three sets of assays were conducted to detect drug efficacy. Firstly, in vivo estimation of drug resistance to H. contortus was explored in 80 sheep naturally infected with H. contortus. Sheep were divided into four equal groups (20 sheep for each group): the first group was treated with albendazole (5.00 mg/kg BW), the second with levamisole (7.50 mg/kg BW), the third with ivermectin (0.20 mg/kg BW), and the fourth group served as the untreated control. Fecal egg reduction test (FERT) was done at days 7 and 14 after treatment. Secondly, for in vitro egg hatching assay (EHA), H. contortus eggs from naturally infected sheep were collected and treated with 0.0002, 0.002, 0.02, 0.2, and 2.0 µg/mL albendazole. Thirdly, molecular detection of the albendazole resistance gene in adult male H. contortus worms and larvae from infected sheep was carried out using allele-specific PCR. RESULTS: The FECRT results showed that the drug efficacy was 86.84% for albendazole and 100% for both levamisole and ivermectin. The result of EHA showed that eggs did not hatch at 2.0 µg/mL albendazole concentration. Molecular findings showed two forms, including H. contortus homozygous susceptible (SS) and heterozygous (RS) of "ß-tubulin" gene at 200 sites, which were recorded in both single male worms and larvae. CONCLUSION: H. contortus, which was susceptible to levamisole and ivermectin, had developed resistance to albendazole.

Ultrastructural description and phylogeny of a novel microsporidian, Glugea eda n. sp. from the striated fusilier, Caesio striata, in the Red Sea off Saudi Arabia.

Glugea eda n. sp. is described from the mesenteries of the striated fusilier, Caesio striata, collected from the Red Sea coast off Yanbu' al Bahr, Saudi Arabia. Numerous blackish xenomas, ranged from 3 to 5 mm, were found in the body cavity associated with the mesenteries. Mature spores are monomorphic, ellipsoidal with an average size of 5(4-6) µm in length and 2.2 (2-3) µm in width. Observations of the ultrastructure revealed that the development was asynchronous and that the nuclei were isolated throughout the life cycle with uninucleate meronts. Sporoblasts were uninucleated and existed together with sporonts in a fully formed parasitophorous vacuole. The polar filament of the mature spore was isofilar with 24-28 coils, arranged in three rows. Phylogenetic analysis placed the current microsporidia within the clade grouping Glugea species and close to the species described from the Red Sea and Arabian Gulf. The morphometric and molecular comparison with other members of the genus Glugea evidenced the taxonomic novelty of the present form, suggesting that it should be considered as a new species. To the best of our knowledge, the parasite here described represents the first occurrence of microsporidian infection in the fish of the family Caesionidae.

Studies on rodents role as reservoir hosts of leishmaniasis with specical reference to their ectoparasites in Suez Governorate.

The distribution of rodents was studied in three different habitats. Seven rodent species were identified: Rattus norvegicus, R. alexandrinus, R. frugivorous, Mus musculus, Acomys russatus, Meriones sacramenti and Gerbillus pyramidum. The species distribution varied with the habitat type. The highest density of rodents was in July and August and the lowest one was in January. However, some species were collected all the year round. The rodents were investigated for the endo- and ecto-parasites. No Leishmania parasites were found. The ectoparasites were: Xenopsylla cheopis, Leptopsylla segnis and Ctenocephalides felis, Polyplax spinulos, Hyalomma dromedarii (nymph) and Echinolaelaps echidninus and Hemolaelaps glassgowi. Ecto-parasites were on rodents all year-round in domestic habitat and peridomestic habitats. In wild one, ecto-parasites activity was from March to December. The rodents' role as reservoir for L. major was experimentally studied. Rodents inoculated with L. major together with hamster and BALB-c mice developed cutaneous lesions. The active lesions, the rodents' ecological habitats and the presence of insect-vector may pave the way to an epidemic zoonotic leishmaniasis role.

Ecology, biology and susceptibility of Phlebotomus papatasi to Leishmania experimental infection in Suez Governorate.

Zoonotic cutaneous leishmaniasis (ZCL) is endemic in Sinai Peninsula. The sand fly and reservoirs were investigated in Suez G., since new settlements and land reclamation programs are ongoing. The results showed that Phlebotomus papatasi reached its highest density in September. The successfully colonized P. papatasi facilitated its biology and competence study. An autogenous trait was proven within P. papatasi population indicating its ability to survive and breed during adverse conditions. The vector competence was carried out under laboratory condition through feeding on lesion of a L. major experimentally infected hamster and by membrane feeding technique. Both hamsters and BALB-c mice inoculated with L. major developed ZCL lesions.