Evaluation of the DPPH Radical Scavenging Activity, Total Phenolic Content and Total Flavonoid Content of Different Solvent Extracts of Catunaregam tomentosa (Blume ex DC) Tirveng Leaves

Catunaregam tomentosa (Blume ex DC) Tirveng is commonly known as Khet in Thailand and Bisa Ular or Badang in Malaysia. The tree is widely distributed in the north-east region of Thailand while in Malaysia the tree usually grows in the open waterfront area at Terengganu. This plant belongs to the Rubiaceae family, and the genus catunaregam has interesting pharmacological activities such as anti-inflammatory, antispasmodic, antidysenteric, antifertility and immunomodulatory. In this study, the leaves were extracted using dichloromethane, ethyl acetate and ethanol. Total phenolic was determined by Folin-Ciocalteau method while total flavonoid was determined by the aluminium chloride calorimetric method. Meanwhile, its antioxidant activity was evaluated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. The ethanolic extract was found to have the highest percentages of phenolic and flavonoid content. Interestingly, ethanolic extract also demonstrated strong DPPH scavenging activity with IC50 at 20.07 ± 0.51µg/mL

A Framework to Spatially Cluster Air Quality Monitoring Stations in Peninsular Malaysia using the Hybrid Clustering Method

Multiple variables must be analyzed in order to assess air quality trends. It turns into a multidimensional issue that calls for dynamic methods. In order to provide an improved spatial cluster distribution with distinct validation, this study set out to illustrate the hybrid cluster method in air quality monitoring stations in Peninsular Malaysia. The Department of Environment, Malaysia (DOE), provided the data set, which covered the two-year period from 2018 to 2019. This study included six air quality pollutants: PM10, PM2.5, SO2, NO2, O3, and CO. Principal component analysis (PCA), a multivariate technique, was used to condense the information found in enormous data tables in order to better comprehend the variables (to reduce dimensionality) prior to grouping the data. The PCA factor scores were then used to produce the AHC. The clusters were validated using discriminant analysis (DA). 36 of 47 stations required additional analysis using AHC, according to the PCA factor scores. Low Polluted Region (LPR = seven stations), Moderate Polluted Region (MPR = 20 stations), and High Polluted Region (HPR = nine stations) were created from AHC and share the same characteristics. The DA results showed 84 % correct classification rate for the clusters. With regard to identifying and categorizing stations according to air quality characteristics, the framework presented here offers an improved method. This illustrates that the hybrid cluster method utilized in this work can produce a new method of pollutant distributions that is helpful in air pollution investigations

A P. falciparum NF54 Reporter Line Expressing mCherry-Luciferase in Gametocytes, Sporozoites, and Liver-Stages

Transgenic malaria parasites expressing fluorescent and bioluminescent proteins are valuable tools to interrogate malaria-parasite biology and to evaluate drugs and vaccines. Using CRISPR/Cas9 methodology a transgenic Plasmodium falciparum (Pf) NF54 line was generated that expresses a fusion of mCherry and luciferase genes under the control of the Pf etramp10.3 gene promoter (line [email protected]). Pf etramp10.3 is related to rodent Plasmodium uis4 and the uis4 promoter has been used to drive high transgene expression in rodent parasite sporozoites and liver-stages. We examined transgene expression throughout the complete life cycle and compared this expression to transgenic lines expressing mCherry-luciferase and GFP-luciferase under control of the constitutive gapdh and eef1a promoters. The [email protected] parasites express mCherry in gametocytes, sporozoites, and liver-stages. While no mCherry signal was detected in asexual blood-stage parasites above background levels, luciferase expression was detected in asexual blood-stages, as well as in gametocytes, sporozoites and liver-stages, with the highest levels of reporter expression detected in stage III-V gametocytes and in sporozoites. The expression of mCherry and luciferase in gametocytes and sporozoites makes this transgenic parasite line suitable to use in in vitro assays that examine the effect of transmission blocking inhibitors and to analyse gametocyte and sporozoite biology

Creation and preclinical evaluation of genetically attenuated malaria parasites arresting growth late in the liver.

Whole-sporozoite (WSp) malaria vaccines induce protective immune responses in animal malaria models and in humans. A recent clinical trial with a WSp vaccine comprising genetically attenuated parasites (GAP) which arrest growth early in the liver (PfSPZ-GA1), showed that GAPs can be safely administered to humans and immunogenicity is comparable to radiation-attenuated PfSPZ Vaccine. GAPs that arrest late in the liver stage (LA-GAP) have potential for increased potency as shown in rodent malaria models. Here we describe the generation of four putative P. falciparum LA-GAPs, generated by CRISPR/Cas9-mediated gene deletion. One out of four gene-deletion mutants produced sporozoites in sufficient numbers for further preclinical evaluation. This mutant, PfΔmei2, lacking the mei2-like RNA gene, showed late liver growth arrest in human liver-chimeric mice with human erythrocytes, absence of unwanted genetic alterations and sensitivity to antimalarial drugs. These features of PfΔmei2 make it a promising vaccine candidate, supporting further clinical evaluation. PfΔmei2 (GA2) has passed regulatory approval for safety and efficacy testing in humans based on the findings reported in this study

Expression of full-length Plasmodium falciparum P48/45 in P. berghei blood stages: A method to express and evaluate vaccine antigens

Contains fulltext : 195592.pdf (publisher's version ) (Open Access

The effect of dosage on the protective efficacy of whole-sporozoite formulations for immunization against malaria

Abstract Immunization with Plasmodium sporozoites, either attenuated or administered under the cover of an antimalarial drug, can induce strong protection against malaria in pre-clinical murine models, as well as in human trials. Previous studies have suggested that whole-sporozoite (WSpz) formulations based on parasites with longer liver stage development induce higher protection, but a comparative analysis of four different WSpz formulations has not been reported. We employed a rodent model of malaria to analyze the effect of immunization dosage on the protective efficacy of WSpz formulations consisting of (i) early liver arresting genetically attenuated parasites (EA-GAP) or (ii) radiation-attenuated sporozoites (RAS), (iii) late arresting GAP (LA-GAP), and (iv) sporozoites administered under chemoprophylaxis, that are eliminated upon release into the bloodstream (CPS). Our results show that, unlike all other WSpz formulations, EA-GAP fails to confer complete protection against an infectious challenge at any immunization dosage employed, suggesting that a minimum threshold of liver development is required to elicit fully effective immune responses. Moreover, while immunization with RAS, LA-GAP and CPS WSpz yields comparable, dosage-dependent protection, protection by EA-GAP WSpz peaks at an intermediate dosage and markedly decreases thereafter. In-depth immunological analyses suggest that effector CD8+ T cells elicited by EA-GAP WSpz immunization have limited developmental plasticity, with a potential negative impact on the functional versatility of memory cells and, thus, on protective immunity. Our findings point towards dismissing EA-GAP from prioritization for WSpz malaria vaccination and enhance our understanding of the complexity of the protection elicited by these WSpz vaccine candidates, guiding their future optimization

The Plasmodium falciparum male gametocyte protein P230p, a paralog of P230, is vital for ookinete formation and mosquito transmission

Contains fulltext : 196551.pdf (publisher's version ) (Open Access

The use of transgenic parasites in malaria vaccine research

Introduction Transgenic malaria parasites expressing foreign genes, for example fluorescent and luminescent proteins, are used extensively to interrogate parasite biology and host-parasite interactions associated with malaria pathology. Increasingly transgenic parasites are also exploited to advance malaria vaccine development. Areas covered We review how transgenic malaria parasites are used, in vitro and in vivo, to determine protective efficacy of different antigens and vaccination strategies and to determine immunological correlates of protection. We describe how chimeric rodent parasites expressing P. falciparum or P. vivax antigens are being used to directly evaluate and rank order human malaria vaccines before their advancement to clinical testing. In addition, we describe how transgenic human and rodent parasites are used to develop and evaluate live (genetically) attenuated vaccines. Expert commentary Transgenic rodent and human malaria parasites are being used to both identify vaccine candidate antigens and to evaluate both sub-unit and whole organism vaccines before they are advanced into clinical testing. Transgenic parasites combined with in vivo pre-clinical testing models (e.g. mice) are used to evaluate vaccine safety, potency and the durability of protection as well as to uncover critical protective immune responses and to refine vaccination strategies.</p

OX40 Stimulation Enhances Protective Immune Responses Induced After Vaccination With Attenuated Malaria Parasites

Host-parasite interactio

The Plasmodium falciparum male gametocyte protein P230p, a paralog of P230, is vital for ookinete formation and mosquito transmission

Abstract Two members of 6-cysteine (6-cys) protein family, P48/45 and P230, are important for gamete fertility in rodent and human malaria parasites and are leading transmission blocking vaccine antigens. Rodent and human parasites encode a paralog of P230, called P230p. While P230 is expressed in male and female parasites, P230p is expressed only in male gametocytes and gametes. In rodent malaria parasites this protein is dispensable throughout the complete life-cycle; however, its function in P. falciparum is unknown. Using CRISPR/Cas9 methodology we disrupted the gene encoding Pfp230p resulting in P. falciparum mutants (PfΔp230p) lacking P230p expression. The PfΔp230p mutants produced normal numbers of male and female gametocytes, which retained expression of P48/45 and P230. Upon activation male PfΔp230p gametocytes undergo exflagellation and form male gametes. However, male gametes are unable to attach to red blood cells resulting in the absence of characteristic exflagellation centres in vitro. In the absence of P230p, zygote formation as well as oocyst and sporozoite development were strongly reduced (>98%) in mosquitoes. These observations demonstrate that P230p, like P230 and P48/45, has a vital role in P. falciparum male fertility and zygote formation and warrants further investigation as a potential transmission blocking vaccine candidate