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Objective: To investigate phytochemicals present in the essential oil from aerial parts of eastern red cedar, Juniperus virginiana (J. virginiana) L. (Cupressaceae) and to determine its killing and repellent activities against larvae, pupae, and adults of the Asian malaria mosquito, Anopheles stephensi (Diptera: Culicidae). Methods: J. virginiana essential oil was extracted by hydrodistillation, and its chemical composition was determined by gas chromatography-mass spectrometry. Seven different logarithmic concentrations of J. virginiana essential oils were used in larvicidal and pupicidal assays. J. virginiana essential oils-impregnated bed nets were applied in a designed animal module to test excito-repellent activity against adult mosquitoes. Results: Fourteen constituents corresponding to 99.98% of J. virginiana essential oils were identified. Five main components were terpinen-4-ol (25.21%), camphor (19.89%), E-3-hexen-1-ol (13.30%), γ-terpinene (7.86%), and l-menthone (2.27%). The LC
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Non-invasive therapeutic methods have recently been used in medical sciences. Enzymes have shown high activity at very low concentrations in laboratories and pharmaceutical, enabling them to play crucial roles in different biological phenomena related to living organism, especially human medicine. Recently, using the therapeutic methods based on non-invasive approaches has been emphasized in medical society. Researchers have focused on producing medicines and tools reducing invasive procedures in medical. Collagenases are proteins which catalyze chemical processes and break the peptide bonds in collagen. Collagen may be generated more than the required amount or produced in unsuitable sites or may not degrade after a certain time. In such cases, using an injectable collagenase or its ointment can be helpful in collagen degradation. In both in vitro and in vivo tests, it has been revealed that collagenases have several therapeutic properties in wound healing, burns, nipple pain and some diseases including intervertebral disc herniation, keloid, cellulite, lipoma among others. This review describes the therapeutic application of collagenase in medical sciences and the process for its production using novel methods, paving the way for more effective and safe applications of collagenases.
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Objective: Apical membrane antigen-1 [AMA-1] is one of the most promising bloodstage candidate antigens for production of a malaria vaccine against the Plasmodium parasite. Genetic diversity in protective antigens, which is a common phenomenon in a complex pathogen such as the Plasmodium parasite, is responsible for problems with the development of an effective malaria vaccine. This phenomenon will increase the parasite's ability to evade immune responses. Therefore, malaria vaccine development requires the evaluation of immune responses to different allelic forms of the vaccine candidate antigens
Methods: In this investigation, the two variant forms of PvAMA-1 [PvAMA-1A and B] were expressed in an Escherichia coli M15-pQE30 system using genomic DNA from Iranian individuals with patent Plasmodium vivax infection. The IgG responses of two antigens were evaluated in BALB/c mice with the purified protein emulsified in Freund's adjuvant. In addition, the correct conformation of the recombinant proteins was evaluated by the indirect immunofluorescence antibody test [IFA]
Results: The evaluation of immunogenic responses of two variant forms of PvAMA-1 showed the presence of IgG responses in mice after three immunizations. Cross-reactions were observed. Monitoring of IgG responses showed the persistence up to one year after the last immunization. The antibodies raised against recombinant PvAMA-1s in injected mice recognized the native protein [PvAMA-1] localized on Plasmodium vivax merozoites
Conclusion: The present outcomes confirmed the presence of common epitopes in recombinant forms of the protein that corresponded to native proteins. These emulsified proteins in Freund's adjuvant were immunogenic in BALB/c mice and IgG responses persisted for up to one year. The IgG responses to two PvAMA-1 variants did not differ significantly. The presence of cross-reactive antibodies has implied that one of these two forms of protein could be used in a universal blood-stage vaccine based on the PvAMA-1 antigen
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Objective: The apical membrane antigen-1 [AMA-1] is considered as a promising candidate for development of a malaria vaccine against Plasmodium parasites. The correct conformation of this protein appears to be necessary for the stimulation of parasite-inhibitory responses, and these responses, in turn, seem to be antibody-mediated. Therefore, in the present investigation, we expressed the Plasmodium vivax AMA-1 [PvAMA-1] ectodomain in Escherichia coli [E. coli], purified it using standard procedures and characterized it to determine its biological activities for it to be used as a potential target for developing a protective and safe vivax malaria vaccine
Materials and Methods: In this experimental investigation, the ectodomain of PvAMA-1 antigen [GenBank accession no. JX624741] was expressed in the E. coli M15- pQE30 expression system and purified with immobilized-metal affinity chromatography. The correct conformation of the recombinant protein was evaluated by Western blotting and indirect immunofluorescence antibody [IFA] test. In addition, the immunogenic properties of PvAMA-1 were evaluated in BALB/c mice with the purified protein emulsified in Freund's adjuvant
Results: In the present study, the PvAMA-1 ectodomain was expressed at a high-level [65 mg/L] using a bacterial system. Reduced and non-reduced sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE] as well as Western blot analysis confirmed the appropriate conformation and folding of PvAMA-1. The evaluation of immunogenic properties of PvAMA-1 showed that both T helper-1 and 2 cells [Th1 and Th2] responses were present in mice after three immunizations and persisted up to one year after the first immunization. Moreover, the antibodies raised against the recombinant PvAMA-1 in injected mice could recognize the native protein localized on P. vivax parasites
Conclusion: We demonstrate that our recombinant protein had proper conformation and folding. Also, there were common epitopes in the recombinant forms corresponding to native proteins. These results; therefore, indicate that the expressed PvAMA-1 has the potential to be used as a vivax malaria vaccine
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Asthma is caused by the combination of different factors. Current concepts of asthma pathogenesis emphasize on gene-environment interactions. Mega-genome scanning projects revealed that different Single Nucleotide Polymorphisms [SNPs] are related to asthma susceptibility. rs7216389-T is one of them that is related to childhood asthma and its effect on childhood asthma severity has been proved in different nations, however no study has been performed in Eastern Mediterranean and Middle East countries yet. To perform population genetic studies, a rapid and high-throughput screening method is necessary. High-resolution melting analysis is a rapid, powerful and accurate method, which is suitable for this type of studies. Therefore, it has been decided to develop a high-resolution melting method for rs7216389 locus genotyping in Iranian asthmatic children. In the current study, a high-resolution melting analysis method based on SYBR Green-I was developed to check the frequency of rs7216389-T mutation in Iranian asthmatic children for the first time. Second and third classes of intercalating dyes are commonly used for high-resolution melting method. However, in this study, SYBR Green-I was used for rs7216389 locus genotyping for the first time. Our results for 60 samples showed that SYBR Green-I has good efficacy for rs7216389 locus genotyping through high-resolution melting method in comparison with PCR-RFLP and sequencing. Comparison of our results based on HRM analysis with PCR-RFLP showed that our developed method is rapid, accurate, high-throughput and economic to study the rs7216389 locus in asthmatic children and it is applicable for other similar population genetic studies
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Because of the lack of an effective and economical control strategy against malaria [the most devastating infectious disease in developing countries] Transmission-Blocking Vaccines [TBVs] concept has been raised in recent years, promising a more efficient way to malaria control. TBVs aim at interfering and/or blocking pathogen development within the vector, halting transmission to non-infected vertebrate host. Aminopeptidase N [APN] is one of the most potent proteins in parasite development in Anopheles malaria vectors, which is strongly co-localized with human malaria parasites in the mosquito midgut epithelium. Therefore, Aminopeptidase N is one of the best choices for a new TBV. In this study for the first time we used 3'-RACE to amplify APN gene in Anopheles stephensi [An.stephensi], a major malaria vector in Iran, Indian subcontinent up to China by using different sets of primers including exon junction, conserved and specific region primers. Full length of APN was sequenced stepwise, which could be applied in designing a new regional TBV and act as an essential component of malaria elimination program in An. stephensi distribution areas. Primers design and method modification should be set up exactly in approach based amplifications. From results we came to this conclusion that that 3'-RACE could be applied to amplified key regions which are be-yond reach
Subject(s)
Malaria , Insect Proteins , Anopheles , Communicable Disease Control , Insect VectorsABSTRACT
Calpastatin is an endogenous inhibitor of calpain [calcium-dependent cysteine protease]. Calpastatin activity is highly related to the rate of protein turnover and rate of meat tenderization. In order to characterize the structure of calpastatin in Iranian Afshari breed of sheep, intron 6 and partial exon 7 of the L domain were amplified and sequenced. A fragment of approximately 1.5 kb was identified. In this study, an Afshari calpastatin gene fragment that encoded L Domain amino acids was detected. Hence by detection of such conserved mutations, it is possible to use these polymorphisms in Marker-Assisted Selection [MAS]
Subject(s)
Animals , Sheep/genetics , Sequence AnalysisABSTRACT
Anopheles culicifacies is a main malaria vector in southeastern part of Iran, bordering Afghanistan and Pakistan. So far, resistance to DDT, dieldrin, malathion and partial tolerance to pyrethroids has been reported in An. stephensi, but nothing confirmed on resistance status of An. culicifacies in Iran. In current study, along with WHO routine susceptibility test with DDT [4%], dieldrin [0.4%], malathion [5%], permethrin [0.25%], lambadacyhalothrin [0.1%], and deltamethrin 0.025, we cloned and sequenced segment VI of domain II [SII6] in voltage-gated sodium channel [vgsc] gene of An. Culicifacies specimens collected in Sistan and Baluchistan province [Iran]. A 221-bp amplified fragment showed 91% and 93% similarity with exon I and exon II of An. gambiae. The size of intron II in An. culicifacies is 62 bp, while in An. gambiae is 57 bp. The major difference within An. culicifacies specimens and also with An. gambiae is in position 29 of exon I, which led to substitution of Leu to His amino acid. This data will act as first report on partial sequence of vgsc gene and its polymorphism in An. culicifacies. A Leu to His amino acid substitution detected upstream the formerly known knockdown resistance [kdr] mutation site could be an indication for other possible mutations related to insecticide resistance. However, the result of WHO susceptibility test carried out in Baluchistan of Iran revealed a level of tolerance to DDT and dieldrin, but almost complete susceptibility to pyrethroids in An. culicifacies. We postulate that the molecular diagnostic tool developed for detection and identification of kdr-related mutations in An. culicifacies, could be useful in monitoring insecticide resistance in Iran and neighbouring countries such as Pakistan and Afghanistan. A phylogenetic tree also constructed based on the sequence of exon I and II, which readily separated An. culicifacies populations from An. stephensi, An. fluviatilis and An. gambiae
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Anopheles pulcherrimus Theobald has a wide distribution in western Asia and is a potential vector of malaria in Iran. We have examined the rDNA-ITS2 [internal transcribed spacer 2] region of An. pulcherrimus specimens collected during the two peaks of activity [May-June and October-November] from Sistan and Baluchistan province, southeastern Iran. There were no consistent differences between specimens originated from different ecological areas. Total amplified fragment is 490 bp, which is within the range of the records repeated from other anophelines. ITS2 was 350 bp long in all individuals examined with identical sequence in different populations. Sequence analysis revealed its differences with two other important malaria vectors in the region, An. culicifacies and An. fluviatilis. However, based on ITS2-derived phylogenetic tree, the nearest taxa to An. pulcherrimus is a new species related to An. Culicifacies and called species X in An. culicifacies species complex. These data may provide a better understanding on dynamics of malaria transmission in southeastern corner of Iran and neighboring countries. Moreover, the extent of the genetic variation in these mainly sympatric species could result in designing and application of species-specific diagnostic tools, which can facilitate the management of malaria control program in the region