A recombinant Aspergillus oryzae fungus transmitted from larvae to adults of Anopheles stephensi mosquitoes inhibits malaria parasite oocyst development.

The control of malaria parasite transmission from mosquitoes to humans is hampered by decreasing efficacies of insecticides, development of drug resistance against the last-resort antimalarials, and the absence of effective vaccines. Herein, the anti-plasmodial transmission blocking activity of a recombinant Aspergillus oryzae (A. oryzae-R) fungus strain, which is used in human food industry, was investigated in laboratory-reared Anopheles stephensi mosquitoes. The recombinant fungus strain was genetically modified to secrete two anti-plasmodial effector peptides, MP2 (midgut peptide 2) and EPIP (enolase-plasminogen interaction peptide) peptides. The transstadial transmission of the fungus from larvae to adult mosquitoes was confirmed following inoculation of A. oryzae-R in the water trays used for larval rearing. Secretion of the anti-plasmodial effector peptides inside the mosquito midguts inhibited oocyst formation of P. berghei parasites. These results indicate that A. oryzae can be used as a paratransgenesis model carrying effector proteins to inhibit malaria parasite development in An. stephensi. Further studies are needed to determine if this recombinant fungus can be adapted under natural conditions, with a minimal or no impact on the environment, to target mosquito-borne infectious disease agents inside their vectors.

Serine proteases activity in miracidia of Fasciola hepatica and effects of chemical and herbal inhibitors.

Fasciolosis is a zoonotic parasitic disease caused by the trematode Fasciola hepatica. The proteases are essential for the survival of parasites. The present study was aimed to determine serine proteases activities in miracidia of F. hepatica and evaluate the effects of pH and different inhibitors on the serine proteases activities. Adult F. hepatica helminths were removed from naturally infected livers of the slaughtered cattle and crushed. The eggs were incubated at 28.00 ˚C for 16 days. The released miracidia were homogenized and total proteolytic activity of the extract of miracidia at different pH values were evaluated. Serine proteases activities were determined using specific substrates. The inhibitory effects of chemical and herbal inhibitors on the enzymes were also assessed. The extract of miracidia hydrolyzed azocasein with optimum activity at pH 8.00. The optimum pH effect on serine proteases activities was found at alkaline pH. Phenylmethylsulfonyl fluoride and Bowman-Birk inhibitors inhibited and decreased the proteases activities in the miracidia extract. It was concluded that there were proteases activities in miracidia of F. hepatica which were inhibited by chemical and herbal inhibitors.

Administration of Nicotine Exacerbates the Quinine-induced Structural and Functional Alterations of Testicular Tissue in Adult Rats: An Experimental Study.

PURPOSE: In this study the role of nicotine (NCT) administration on the intensity of rat testicular tissue alterations induced by quinine (QU) was evaluated. MATERIALS AND METHODS: Forty adult Wistar rats were divided into four groups. Control (CON), NCT administrated (4 mg/kg) (NCT), QU treated (25 mg/kg for 7 days) (QU), and nicotine with quinine received (NCT+QU). After 28 days, serum testosterone and malondialdehyde (MDA) levels were measured. Testes and epididymides samples were prepared for determining tissue MDA levels, histomorphometry, microscopic indices of spermatogenesis, immunohistochemistry of p53 and sperm analysis. RESULTS: Testosterone levels were decreased significantly (P = .0004) in treated groups compared to CON group. Serum MDA levels were increased significantly (P = .0004) in NCT and QU groups compared to CON group. Tissue MDA levels were increased significantly (P = .0012) in NCT+QU group in comparison to CON group. These parameters were changed significantly in NCT+QU group compared to QU group. Seminiferous tubules diameter decreased significantly (P < .0001) in treated groups compared to CON group and in NCT+QU group compared to QU group. The height of germinal epithelium decreased significantly (P = .0001) in NCT and NCT+QU groups compared to CON and QU groups. The number of Sertoli cells, spermatocytes, and spermatids decreased significantly in treated groups compared to CON group. The number of spermatogonia decreased significantly (P = .0017) in NCT and NCT+QU groups compared to CON group. The number of Sertoli cells, spermatogonia, and spermatocytes decreased significantly in NCT+QU group compared to QU group. All indices of spermatogenesis decreased in treated groups compared to CON group. The lowest mean of these indices was observed in NCT+QU group. The sperm viability decreased significantly (P < .0001) in treated groups compared to CON group. Sperm count and motility decreased significantly in NCT and NCT+QU groups compared to CON group. All experimental groups showed the over-expression of p53 compared to CON group. CONCLUSION: The administration of nicotine could be involved in the exacerbation of testicular tissue alterations related to quinine therapy.

The Effects of pH and Temperature on Cysteine Protease (Cathepsin B) Activity in Miracidia and Eggs of Fasciola hepatica.

BACKGROUND: Fascioliasis is a worldwide zoonotic disease caused by the trematodes Fasciola hepatica in humans and animals. Proteases are essential for the survival of parasites and have important activities such as penetration, tissue migration, and egg hatching. This study was conducted to analyze cysteine protease of the miracidia and eggs of F. hepatica, and to assess the effects of pH and temperature on the proteases activity and stability. METHODS: Adults F. hepatica were isolated from infected livers and were morphologically identified in 2018. Eggs collected from the adults and incubated in distilled water at 28 °C for 16 d to produce miracidia. The extract was collected from miracidia and eggs. A substrate for cathepsin B (Z-Arg-Arg-Pna) was used to assess the enzyme activity at different (2-12) pH levels. After homogenization, protein level was measured with Bradford method. Estimation of optimum temperature and pH was performed in the temperature range of 10-90 ° C and pH values from 2-12. RESULTS: The highest activity of the miracidia and eggs enzyme extracts for Z-Arg-Arg-pNA was at pH 4. The miracidia extract was most stable at neutral pH and the eggs extract was most stable in acidic pH. The optimum temperature activity for both stages was 40 °C. These proteases were stable up to 40 °C. CONCLUSION: Upon the importance of pH and temperature in the life cycle of F. hepatica, the current findings can be used for induction of some modifications in pH and preventing the activity of the enzymes for decrement of the efficacy of miracidia penetration into the intermediate snails and egg hatching of this zoonotic parasite.