Introduction of double-stranded RNA (dsRNA) into some cells or organisms results in degradation of its homologous mRNA, a process called RNA interference (RNAi). The dsRNAs are processed into short interfering RNAs (siRNAs) that subsequently bind to the RNA-induced silencing complex (RISC), causing degradation of target mRNAs. Because of this sequence-specific ability to silence target genes, RNAi has been extensively used to study gene functions and has the potential to control disease pathogens or vectors. With this promise of RNAi to control pathogens and vectors, this paper reviews the current status of RNAi in protozoans, animal parasitic helminths and disease-transmitting vectors, such as insects. Many pathogens and vectors cause severe parasitic diseases in tropical regions and it is difficult to control once the host has been invaded. Intracellularly, RNAi can be highly effective in impeding parasitic development and proliferation within the host. To fully realize its potential as a means to control tropical diseases, appropriate delivery methods for RNAi should be developed, and possible off-target effects should be minimized for specific gene suppression. RNAi can also be utilized to reduce vector competence to interfere with disease transmission, as genes critical for pathogenesis of tropical diseases are knockdowned via RNAi.
Animals ; Communicable Diseases/*genetics/*parasitology ; Helminths/*genetics/metabolism ; Humans ; Insect Vectors/*genetics/metabolism ; Protozoa/*genetics/physiology ; *RNA Interference ; *Tropical Climate

Introduction of double-stranded RNA (dsRNA) into some cells or organisms results in degradation of its homologous mRNA, a process called RNA interference (RNAi). The dsRNAs are processed into short interfering RNAs (siRNAs) that subsequently bind to the RNA-induced silencing complex (RISC), causing degradation of target mRNAs. Because of this sequence-specific ability to silence target genes, RNAi has been extensively used to study gene functions and has the potential to control disease pathogens or vectors. With this promise of RNAi to control pathogens and vectors, this paper reviews the current status of RNAi in protozoans, animal parasitic helminths and disease-transmitting vectors, such as insects. Many pathogens and vectors cause severe parasitic diseases in tropical regions and it is difficult to control once the host has been invaded. Intracellularly, RNAi can be highly effective in impeding parasitic development and proliferation within the host. To fully realize its potential as a means to control tropical diseases, appropriate delivery methods for RNAi should be developed, and possible off-target effects should be minimized for specific gene suppression. RNAi can also be utilized to reduce vector competence to interfere with disease transmission, as genes critical for pathogenesis of tropical diseases are knockdowned via RNAi.
Animals ; Communicable Diseases/*genetics/*parasitology ; Helminths/*genetics/metabolism ; Humans ; Insect Vectors/*genetics/metabolism ; Protozoa/*genetics/physiology ; *RNA Interference ; *Tropical Climate

OBJECTIVETo compare the sequences of cytochrome C oxidase subunit I gene (COI) in Aedes albopictus from different geographic strains in China and to discuss the differences in susceptibility among different geographic strains to dengue virus (DV).

METHODSCOI was amplified with polymerase chain reaction method and sequenced from its genomic DNA. Molecular phylogenetic trees were constructed with Neighbor-Joining method.

RESULTSSequence length of COI fragment in each geographic strains was 415 bp. The rates of shift and reverse of base pairs in Simao strain were 1.93% and 0.24% respectively. The rate of shift in Mawei and Nanning strains was 0.48%. The analyses of phylogenetic of COI sequences showed that there was close relationship between Simao strain in Yunnan and Mawei strain in Guizhou and between Mawei strain and Nanning strain in Guangxi.

CONCLUSIONSThe susceptibility was widely related to many factors including genetic and environmental ones. COI in Aedes albopictus from different geographic strains in China belonged to the same gene type. There were no direct correlations between COI gene type in different geographic strains and susceptibility to DV.

Aedes ; genetics ; virology ; Animals ; Dengue ; transmission ; Electron Transport Complex IV ; genetics ; Genotype ; Insect Vectors ; Polymerase Chain Reaction

Since Zika virus (ZIKV) has firstly been isolated in 1947, Uganda, outbreaks of Zika fever have been reported in many areas such as in Africa, Southeast Asia and America. Imported cases in China also have been reported. Zika virus belongs to the family Flaviviridae, genus Flavivirus, and include Africa subtype and Asia subtype. It is a mosquito-borne virus primarily transmitted by Aedes aegypti mosquitoes. Sexual transmission, Blood transmission and mother-to-fetus transmission were also reported. Zika virus can go though blood-brain barrier and infect central nervous system. Symptoms are generally mild and self-limited, but recent evidence suggests a possible association between maternal Zika virus infection and adverse fetal outcomes, such as congenital microcephaly, as well as a possible association with Guillain-Barré syndrome. Laboratorial Diagnosis includes nucleic acid detection, Serological test, and isolation of virus. Currently, no vaccine or medication exists to prevent or treat Zika virus infection. Preventive measures against Zika virus infection should be taken through prevention of mosquito bites and surveillance in epidemic area.
Aedes ; physiology ; virology ; Animals ; Humans ; Insect Vectors ; physiology ; virology ; Zika Virus ; genetics ; physiology ; Zika Virus Infection ; transmission ; virology

Rice stripe virus (RSV) transmitted by the small brown planthopper causes severe rice yield losses in Asian countries. Although viral nuclear entry promotes viral replication in host cells, whether this phenomenon occurs in vector cells remains unknown. Therefore, in this study, we systematically evaluated the presence and roles of RSV in the nuclei of vector insect cells. We observed that the nucleocapsid protein (NP) and viral genomic RNAs were partially transported into vector cell nuclei by utilizing the importin α nuclear transport system. When blocking NP nuclear localization, cytoplasmic RSV accumulation significantly increased. In the vector cell nuclei, NP bound the transcription factor YY1 and affected its positive regulation to FAIM. Subsequently, decreased FAIM expression triggered an antiviral caspase-dependent apoptotic reaction. Our results reveal that viral nuclear entry induces completely different immune effects in vector and host cells, providing new insights into the balance between viral load and the immunity pressure in vector insects.
Animals ; Cell Nucleus ; Hemiptera/metabolism* ; Insect Vectors/genetics* ; Insecta ; Nucleocapsid Proteins/metabolism* ; Oryza ; Plant Diseases ; Tenuivirus/metabolism* ; Virus Replication

microRNAs (miRNAs) are an extensive class of -22-nucleotide (nt) endogenous noncoding RNAs regulating life activities ofmetazoans through binding to 3'-untranslated regions (3'-UTRs) of their target genes. This work aimed to identify yan gene in the silkworm, reveal its expression profile and confirm if it is one target of bmo-miR-7 and, as such, have potential for contributing to better understanding of the molecular mechanisms involved in the metamorphosis of silkworm. Based on homolog searching and PCR amplification, we cloned the coding sequence (CDS) of Bmyan, which encodes 476 amino acid residues and contains SAM-PNT and ETs domains. Quantitative PCR (q-PCR), RT-PCR and microarray data revealed high expression of Bmyan in the head, body wall and ovary of day-3 fifth instar larval silkworm, low or no expression in other tissues. It was lowly expressed in the early larval stages, but highly expressed from late spinning to day 4 pupa. The 3'-UTR of Bmyan was obtained by rapid-amplification of cDNA ends (3'RACE) and predicted to contain two potential recognition sites of bmo-miR-7. The luciferase reporter vector containing the 3'-UTR of Bmyan was constructed and co-transfected into BmE cell line with the mimic of bmo-miR-7 and the decreased relative activity of luciferase showed that Bmyan is one target of bmo-miR-7. This work helps further functional analysis of bmo-miR-7 and Bmyan in the silkworm.
3' Untranslated Regions ; Animals ; Bombyx ; genetics ; Cell Line ; Cloning, Molecular ; Female ; Genetic Vectors ; Insect Proteins ; genetics ; Larva ; Metamorphosis, Biological ; MicroRNAs ; genetics ; Pupa

A flavivirus, Culex flavivirus, was first isolated from Chinese mosquitoes with high sequences similarities to those of flaviviruses found in America and Japan. In this study, a total of 48 pools of field-collected mosquitoes were sampled from Dandong of Liaoning Province, China during July to September of 2011. Six isolated viruses showing cytopathic effect (CPE) in C6/C36 cells were tested by reverse transcription polymerase chain reaction(RT-PCR)using Flavivirus genus--specific primers and Culex flavivirus-specific primers and the positive PCR-product was sequenced and compared with the sequences of 10 isolates from GenBank. Phylogenetic tree of NS5 and enevelop genes of flavivirus were constructed. The GenBank accession numbers of NS5 gene were JQ409188, JQ409186, JQ409187, JQ409191, JQ409189 and JQ409190. The GenBank accession numbers of envelope gene were JQ065883, JQ065882, JQ065881, JQ065879,JQ065877 and JQ065878.
Animals ; Base Sequence ; Cell Line ; China ; Culex ; classification ; virology ; Flavivirus ; classification ; genetics ; isolation & purification ; Insect Vectors ; virology ; Molecular Sequence Data ; Phylogeny ; Viral Proteins ; genetics

To improve the expression level of tmAMP1m gene from Tenebrio molitor in Escherichia coli, we studied the effects of expression level and activity of the fusion protein HIS-TmAMP1m by conditions, such as culture temperature, inducing time and the final concentration of inductor Isopropyl beta-D-thiogalactopyranoside (IPTG). We analyzed the optimum expression conditions by Tricine-SDS-PAGE electrophoresis, meanwhile, detected its antibacterial activity by using agarose cavity diffusion method. The results suggest that when inducing the recombinant plasmid with a final IPTG concentration of 0.1 mmol/L at 37 degrees C for 4 h, there was the highest expression level of fusion protein HIS-TmAMP1m in Escherichia coli. Under these conditions, the expression of fusion protein accounted for 40% of the total cell lysate with the best antibacterial activity. We purified the fusion protein HIS-TmAMPlm with nickel-nitrilotriacetic acid (Ni-NTA) metal-affinity chromatography matrices. Western blotting analysis indicates that the His monoclonal antibody could be specifically bound to fusion protein HIS-TmAMPlm. After expression by inducing, the fusion protein could inhibit the growth of host cell transformed by pET30a-tmAMP1m. The fusion protein HIS-TmAMP1m had better stability and remained higher antibacterial activities when incubated at 100 degrees C for 10 h, repeated freeze thawing at -20 degrees C, dissolved in strong acid and alkali, or treated by organic solvents and protease. Moreover, the minimum inhibitory concentration results demonstrated that the fusion protein HIS-TmAMP1m has a good antibacterial activity against Staphylococcus aureus, Staphylococcus sp., Corynebacterium glutamicum, Bacillus thuringiensis, Corynebacterium sp. This study laid the foundation to promote the application of insect antimicrobial peptides and further research.
Animals ; Anti-Infective Agents ; pharmacology ; Antimicrobial Cationic Peptides ; biosynthesis ; genetics ; pharmacology ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Insect Proteins ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology ; Tenebrio ; chemistry

In order to construct a novel fusion protein PTD-maxadilan (PTD-MAX) that can enter the blood-brain barrier (BBB) efficiently, a new gene encoding PTD-MAX was synthesized and cloned into the expression vector pKYB. The recombinant vector pKYB-PTD-MAX was transformed into Escherichia coli ER2566. The expression of fusion protein consisting of PTD-MAX, intein and chitin binding domain was induced by IPTG and the target PTD-MAX protein was purified using Intein Mediated Purification with an Affinity Chitin-binding Tag system. The molecular weight of PTD-MAX determined by the laser flight mass spectrometry was coherent with its theoretical value. The results of the experiment in vivo indicated that the recombinant PTD-MAX can permeate into BBS and inhibitory effects on the food intake were more significantly than maxadilan (P<0.05). The preparation of PTD-MAX lay the foundation for its further application.
Animals ; Base Sequence ; Blood-Brain Barrier ; metabolism ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Insect Proteins ; biosynthesis ; genetics ; pharmacokinetics ; Mice ; Molecular Sequence Data ; Protein Structure, Tertiary ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacokinetics ; Vasodilator Agents ; metabolism ; pharmacokinetics

Anopheles fluviatilis James (Diptera: Culicidae) is one of the known malaria vectors in south and southeastern Iran. Earlier ITS2 sequences analysis of specimens from Iran demonstrated only a single genotype that was identical to species Y in India, which is also the same as species T. We identified 2 haplotypes in the An. fluviatilis populations of Iran based on differences in nucleotide sequences of D3 domain of the 28S locus of ribosomal DNA (rDNA). Comparison of sequence data from 44 Iranian specimens with those publicly available in the Genbank database showed that all of the 28S-D3 sequences from Kazeroun and Khesht regions in Fars Province were identical to the database entry representing species U in India. In other regions, all the individuals showed heterozygosity at the single nucleotide position, which identifies species U and T. It is argued that the 2 species may co-occur in some regions and hybridize; however, the heterozygosity in the 28S-D3 locus was not reflected in ITS2 sequences and this locus for all individuals was identical to species T. This study shows that in a newly diverged species, like members of An. fluviatilis complex, a single molecular marker may not be sufficiently discriminatory to identify all the taxa over a vast geographical area. In addition, other molecular markers may provide more reliable information for species discrimination.
Animals ; Anopheles/classification/*genetics ; Base Sequence ; DNA, Ribosomal Spacer/genetics ; *Genetic Variation ; Insect Vectors/classification/genetics ; Iran ; Molecular Sequence Data ; Phylogeny ; RNA, Ribosomal, 28S/genetics ; Sequence Alignment ; Sequence Analysis, DNA