Molecular characterization, expression and in-silico analysis of fibrinogen-related protein 1 (frep1) in malaria vector Anopheles stephensi.

BACKGROUND: Fibrinogen-related protein 1 (frep1) is a member of the pattern-recognizing receptor family (PRR) which generates an innate immune response after recognizing the pattern associated molecular pattern (PAMP) that occurs on the surface of microorganisms. The main objective of this study is to characterize frep1 and its in-silico analysis in Anopheles stephensi. METHODS AND RESULT: The DNA was extracted from female Anopheles stephensi. PCR was performed for complete analysis of frep1 using specific primers. The gene sequence of frep1 was identified by Sanger sequencing. The bioinformatics structure analysis approach revealed the presence of 3 exons and 4 introns in the frep1. The sequence of frep1 was submitted to NCBI GeneBank with accession number ON817187.1. Quantitative real-time PCR was performed to analyze frep1 expression. At the developmental stage, frep1 is highly expressed in the L1 stage, egg, and adult female mosquito. In addition, frep1 is highly expressed in the tissue fat body, midgut, and salivary gland. After blood-fed, an upregulation of frep1 at 48 h in the midgut, and downregulation in fat body were observed at different time intervals. CONCLUSION: The genomic data of frep1 is encoded by 12,443 bp. The frep1 has a significant role in the early metamorphosis. Its expression in fat body and midgut suggests it could be important for fat metabolism and post-blood digestion. The conserved domain could be targeted for vector control. Further study is required to elucidate its function against malaria parasites to confirm its agonist role in malaria transmission.

Mosquito gene targeted RNAi studies for vector control.

Vector-borne diseases are serious public health concern. Mosquito is one of the major vectors responsible for the transmission of a number of diseases like malaria, Zika, chikungunya, dengue, West Nile fever, Japanese encephalitis, St. Louis encephalitis, and yellow fever. Various strategies have been used for mosquito control, but the breeding potential of mosquitoes is such tremendous that most of the strategies failed to control the mosquito population. In 2020, outbreaks of dengue, yellow fever, and Japanese encephalitis have occurred worldwide. Continuous insecticide use resulted in strong resistance and disturbed the ecosystem. RNA interference is one of the strategies opted for mosquito control. There are a number of mosquito genes whose inhibition affected mosquito survival and reproduction. Such kind of genes could be used as bioinsecticides for vector control without disturbing the natural ecosystem. Several studies have targeted mosquito genes at different developmental stages by the RNAi mechanism and result in vector control. In the present review, we included RNAi studies conducted for vector control by targeting mosquito genes at different developmental stages using different delivery methods. The review could help the researcher to find out novel genes of mosquitoes for vector control.

Zika virus vertical transmission in mosquitoes: A less understood mechanism.

Zika virus disease is a great concern in different parts of the world, and it has become a Public Health Emergency of International Concern. The global pandemic of ZIKV in 2015 prompted concern among scientific community. Zika is a flavivirus of the family Flaviviridae transmitted by mosquitoes. Natural vertical transmission is an ecological strategy that arboviruses adopt to ensure their survival inside the mosquito vector during harsh conditions or interepidemic periods when horizontal transmission is difficult. ZIKV is vertically transmitted from infected females to its offspring. This review has concluded various studies regarding the vertical transmission ability of different mosquito species for ZIKV. Previously Aedes aegypti was considered to be a major vector, however Aedes albopictus and Culex quinquifasciatus are discovered to have the similar vertical transmission potential. Different studies shown that natural vertical transmission has been detected in mosquito species which are not implicated as possible vectors. It leads to the possibility that many other mosquito species may be potential ZIKV vectors.

Emergence of SARS-CoV-2 New Variants and Their Clinical Significance.

COVID-19 is a respiration-related disease caused by SARS-CoV-2 and was identified in China's Wuhan city. More than 223 countries are affected by the disease worldwide. The new variants of the COVID-19 virus are causing problems, from average to life-threatening pneumonia and acute respiratory distress syndrome (ARDS). Presently, there are 170 vaccine candidates, out of which 10 have been approved by the WHO for vaccination, such as Ad26.COV2.S, Pfizer/BioNTech, COVISHIELD, Covovax, Moderna, KoviVac, and some other vaccines to combat the deadly SARS-CoV-2 infection. From all these vaccines, Pfizer/BioNTech and Moderna are showing the highest efficacy against COVID-19. These vaccines are highly efficient against COVID-19 disease, but their potentiality against new variants remains a question. COVID-19 vaccines are highly effective at preventing severe illnesses, hospitalizations, and death. The antibodies elicited by earlier infection or vaccination are the key for possible protection against SARS-CoV-2. The problem has been exacerbated by new information from Africa on the origins of the novel contagious SARS-CoV-2 strain. These new strains occur due to unique mutations in the spike protein, which modify SARS-CoV-2 transmission and infection capabilities, limiting the efficacy of the COVID-19 vaccination. Hence, there is a need to find a potential vaccine against it.

The chemopreventive effect of withaferin A on spontaneous and inflammation-associated colon carcinogenesis models.

Chemopreventive effects and associated mechanisms of withaferin A (WA) against intestinal and colon carcinogenesis remain unknown. We investigated the chemopreventive effect of WA on transgenic adenomatous polyposis coli (APCMin/+) mouse and chemically induced azoxymethane/dextran sodium sulfate (AOM/DSS) models of intestinal and colon carcinogenesis. Oral WA administration (4 and 3 mg/kg) inhibited tumor initiation and progression of intestinal polyps formation in APCMin/+ mice and colon carcinogenesis in the AOM/DSS mouse model. WA-administered mice showed a significant reduction in both number [duodenum, 33% (P > 0.05); jejunum, 32% (P < 0.025); ileum, 43% ( P < 0.001); and colon 59% (P < 0.01] and size of polyps in APCMin/+ mice compared with the respective controls. Similarly, tumor multiplicity was significantly reduced (P < 0.05) in the colon of WA-administered AOM/DSS mice. Pathological analysis showed reduced adenomas and tissue inflammation in WA-administered mouse models. Molecular studies suggested that WA inhibited the expression of inflammatory (interluekin-6, tumor necrosis factor-alpha and cyclooxygenase-2), pro-survival (pAKT, Notch1 and NF-κB) markers in APCMin/+ and AOM/DSS models. The results suggest that WA is a potent agent for preventing colon carcinogenesis and further investigation is required to show clinical utility of the agent.

A natural molecule, urolithin A, downregulates androgen receptor activation and suppresses growth of prostate cancer.

Androgen ablation therapy is the primary therapeutic option for locally advanced and metastatic castration-resistant prostate cancer (CRPC). We investigated therapeutic effect of a dietary metabolite Urolithin A (UroA) and dissected the molecular mechanism in CRPC cells. Treatment with UroA inhibited cell proliferation in both androgen receptor-positive (AR+ ) (C4-2B) and androgen receptor-negative (AR- ) (PC-3) cells however, AR+ CaP cells were more sensitive to UroA treatment as compared with AR- CaP cells. Inhibition of the AR signaling was responsible for the UroA effect on AR+ CaP cells. Ectopic expression of AR in PC-3 cells sensitized them to UroA treatment as compared to the vector-expresseing PC-3 cells, which suggests that AR could be a target of UroA. Similarly, in enzalutamide-resistant C4-2B cells, a downregulation of AR expression also suppressed cell proliferation which was observed with the UroA treatment. Oral administration of UroA significantly suppressed the growth of C4-2B xenografts (P = 0.05) compared with PC-3 xenografts (P = 0.069) without causing toxicity to animals. Immunohistochemistry analysis confirmed in vitro findings such as downregulation of AR/pAKT signaling in UroA-treated C4-2B tumors, which suggests that UroA may be a potent chemo-preventive and therapeutic agent for CRPC.

Salinomycin and other ionophores as a new class of antimalarial drugs with transmission-blocking activity.

The drug target profile proposed by the Medicines for Malaria Venture for a malaria elimination/eradication policy focuses on molecules active on both asexual and sexual stages of Plasmodium, thus with both curative and transmission-blocking activities. The aim of the present work was to investigate whether the class of monovalent ionophores, which includes drugs used in veterinary medicine and that were recently proposed as human anticancer agents, meets these requirements. The activity of salinomycin, monensin, and nigericin on Plasmodium falciparum asexual and sexual erythrocytic stages and on the development of the Plasmodium berghei and P. falciparum mosquito stages is reported here. Gametocytogenesis of the P. falciparum strain 3D7 was induced in vitro, and gametocytes at stage II and III or stage IV and V of development were treated for different lengths of time with the ionophores and their viability measured with the parasite lactate dehydrogenase (pLDH) assay. The monovalent ionophores efficiently killed both asexual parasites and gametocytes with a nanomolar 50% inhibitory concentration (IC50). Salinomycin showed a fast speed of kill compared to that of standard drugs, and the potency was higher on stage IV and V than on stage II and III gametocytes. The ionophores inhibited ookinete development and subsequent oocyst formation in the mosquito midgut, confirming their transmission-blocking activity. Potential toxicity due to hemolysis was excluded, since only infected and not normal erythrocytes were damaged by ionophores. Our data strongly support the downstream exploration of monovalent ionophores for repositioning as new antimalarial and transmission-blocking leads.

Plasmodium transmission blocking activities of Vernonia amygdalina extracts and isolated compounds.

BACKGROUND: Medicinal plants are a validated source for discovery of new leads and standardized herbal medicines. The aim of this study was to assess the activity of Vernonia amygdalina leaf extracts and isolated compounds against gametocytes and sporogonic stages of Plasmodium berghei and to validate the findings on field isolates of Plasmodium falciparum. METHODS: Aqueous (Ver-H2O) and ethanolic (Ver-EtOH) leaf extracts were tested in vivo for activity against sexual and asexual blood stage P. berghei parasites. In vivo transmission blocking effects of Ver-EtOH and Ver-H2O were estimated by assessing P. berghei oocyst prevalence and density in Anopheles stephensi mosquitoes. Activity targeting early sporogonic stages (ESS), namely gametes, zygotes and ookinetes was assessed in vitro using P. berghei CTRPp.GFP strain. Bioassay guided fractionation was performed to characterize V. amygdalina fractions and molecules for anti-ESS activity. Fractions active against ESS of the murine parasite were tested for ex vivo transmission blocking activity on P. falciparum field isolates. Cytotoxic effects of extracts and isolated compounds vernolide and vernodalol were evaluated on the human cell lines HCT116 and EA.hy926. RESULTS: Ver-H2O reduced the P. berghei macrogametocyte density in mice by about 50% and Ver-EtOH reduced P. berghei oocyst prevalence and density by 27 and 90%, respectively, in An. stephensi mosquitoes. Ver-EtOH inhibited almost completely (>90%) ESS development in vitro at 50 µg/mL. At this concentration, four fractions obtained from the ethylacetate phase of the methanol extract displayed inhibitory activity >90% against ESS. Three tested fractions were also found active against field isolates of the human parasite P. falciparum, reducing oocyst prevalence in Anopheles coluzzii mosquitoes to one-half and oocyst density to one-fourth of controls. The molecules and fractions displayed considerable cytotoxicity on the two tested cell-lines. CONCLUSIONS: Vernonia amygdalina leaves contain molecules affecting multiple stages of Plasmodium, evidencing its potential for drug discovery. Chemical modification of the identified hit molecules, in particular vernodalol, could generate a library of druggable sesquiterpene lactones. The development of a multistage phytomedicine designed as preventive treatment to complement existing malaria control tools appears a challenging but feasible goal.

A user friendly method to assess Anopheles stephensi (Diptera: Culicidae) vector fitness: fecundity.

Fecundity, bloodmeal size, and survival are among the most important parameters in the overall fitness of mosquitoes. Impact of an intervention that affects fecundity can be assessed by directly counting the eggs laid by exposed mosquitoes, which is usually done manually. We have developed a macroinstruction, which can be used to count thousands of Anopheles stephensi Liston eggs in a few minutes, to provide an alternative and adaptable method to egg counting as a measure of fecundity. The macro was developed using a scanner and a computer running AxioVision Rel. 4.8 software, a freely accessible software compatible with Windows XP/7/Vista. Using this semiautomated method, it is possible to reduce time, avoid human error and bias, and obtain improved consistency in studies measuring mosquito fecundity.

Systematic relationships of Vuilletia and Senegathrips (Thysanoptera, Phlaeothripinae) from galls on the West African shrub Guiera senegalensis.

Widespread and common across much of the drier areas of western Africa, the woody shrub Guiera senegalensis (Combretaceae) is the sole member of its genus. Similarly widespread is Vuilletia houardi, a thrips species that induces galls on this shrub, and is recorded from Mali, Senegal, Gambia and northern Nigeria (Pitkin & Mound 1973). Moreover, large numbers of galls, together with their included thrips, have now been studied from Burkina Faso. Some galls (Figs 1, 2) are invaded by Senegathrips coutini, a species whose biology is not known but that is possibly a predator. Moreover, Liothrips africana also sometimes breeds within these galls, but is possibly using these only as a convenient shelter. A re-description and line-drawings of V. houardi was provided by zur Strassen (1958), but no modern diagnosis of this genus, nor of Senegathrips, is available, the objective here being to provide formal diagnoses for these two monotypic genera.