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1.
Front Immunol ; 11: 893, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32477363

RESUMO

Clinical immunity to malaria develops after repeated exposure to Plasmodium falciparum parasites. Broadly reactive antibodies against parasite antigens expressed on the surface of infected erythrocytes (variable surface antigens; VSAs) are candidates for anti-malaria therapeutics and vaccines. Among the VSAs, several RIFIN, STEVOR, and SURFIN family members have been demonstrated to be targets of naturally acquired immunity against malaria. For example, RIFIN family members are important ligands for opsonization of P. falciparum infected erythrocytes with specific immunoglobulins (IgG) acquiring broad protective reactivity. However, the global repertoire of human anti-VSAs IgG, its variation in children, and the key protective targets remain poorly understood. Here, we report wheat germ cell-free system-based production and serological profiling of a comprehensive library of A-RIFINs, B-RIFINs, STEVORs, and SURFINs derived from the P. falciparum 3D7 parasite strain. We observed that >98% of assayed proteins (n = 265) were immunogenic in malaria-exposed individuals in Uganda. The overall breadth of immune responses was significantly correlated with age but not with clinical malaria outcome among the study volunteers. However, children with high levels of antibodies to four RIFINs (PF3D7_0201000, PF3D7_1254500, PF3D7_1040600, PF3D7_1041100), STEVOR (PF3D7_0732000), and SURFIN 1.2 (PF3D7_0113600) had prospectively reduced the risk of developing febrile malaria, suggesting that the 5 antigens are important targets of protective immunity. Further studies on the significance of repeated exposure to malaria infection and maintenance of such high-level antibodies would contribute to a better understanding of susceptibility and naturally acquired immunity to malaria.

2.
F1000Res ; 92020.
Artigo em Inglês | MEDLINE | ID: mdl-32399189

RESUMO

Much of the gain in malaria control, in terms of regional achievements in restricting geographical spread and reducing malaria cases and deaths, can be attributed to large-scale deployment of antimalarial drugs, insecticide-treated bed nets, and early diagnostics. However, despite impressive progress, control efforts have stalled because of logistics, unsustainable delivery, or short-term effectiveness of existing interventions or a combination of these reasons. A highly efficacious malaria vaccine as an additional tool would go a long way, but success in the development of this important intervention remains elusive. Moreover, most of the vaccine candidate antigens that were investigated in early-stage clinical trials, selected partly because of their immunogenicity and abundance during natural malaria infection, were polymorphic or structurally complex or both. Likewise, we have a limited understanding of immune mechanisms that confer protection. We reflect on some considerable technological and scientific progress that has been achieved and the lessons learned.


Assuntos
Vacinas Antimaláricas , Malária/prevenção & controle , Antígenos de Protozoários/imunologia , Humanos , Malária/imunologia
3.
Parasit Vectors ; 13(1): 170, 2020 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-32252804

RESUMO

Serine repeat antigen (SERA) is conserved among species of the genus Plasmodium. Sera genes form a multigene family and are generally tandemly clustered on a single chromosome. Although all Plasmodium species encode multiple sera genes, the number varies between species. Among species, the members share similar sequences and gene organization. SERA possess a central papain-like cysteine protease domain, however, in some members, the active site cysteine residue is substituted with a serine. Recent studies implicate this gene family in a number of aspects in parasite biology and induction of protective immune response. This review summarizes the current understanding on this important gene family in several Plasmodium species. The Plasmodium falciparum (Pf)-sera family, for example, consists of nine gene members. Unlike other multigene families in Plasmodium species, Pf-sera genes do not exhibit antigenic variation. Pf-sera5 nucleotide diversity is also low. Moreover, although Pf-sera5 is highly transcribed during the blood stage of malaria infection, and a large amount is released into the host blood following schizont rupture, in malaria endemic countries the sero-positive rates for Pf-SERA5 are low, likely due to Pf-SERA5 binding of host proteins to avoid immune recognition. As an antigen, the N-terminal 47 kDa domain of Pf-SERA5 is a promising vaccine candidate currently undergoing clinical trials. Pf-SERA5 and Pf-SERA6, as well as P. berghei (Pb)-SERA3, and Pb-SERA5, have been investigated for their roles in parasite egress. Two P. yoelii SERA, which have a serine residue at the protease active center, are implicated in parasite virulence. Overall, these studies provide insight that during the evolution of the Plasmodium parasite, the sera gene family members have increased by gene duplication, and acquired various functions that enable the parasite to survive and successfully maintain infection in the host.

4.
Malar J ; 19(1): 155, 2020 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-32295584

RESUMO

BACKGROUND: The malaria parasite Plasmodium falciparum is a protozoan that develops in red blood cells (RBCs) and requires various host factors. For its development in RBCs, nutrients not only from the RBC cytosol but also from the extracellular milieu must be acquired. Although the utilization of host nutrients by P. falciparum has been extensively analysed, only a few studies have reported its utilization of host serum proteins. Hence, the aim of the current study was to comprehensively identify host serum proteins taken up by P. falciparum parasites and to elucidate their role in pathogenesis. METHODS: Plasmodium falciparum was cultured with human serum in vitro. Uptake of serum proteins by parasites was comprehensively determined via shotgun liquid chromatography-mass spectrometry/mass spectrometry and western blotting. The calcium ion concentration in serum was also evaluated, and coagulation activity of the parasite lysate was assessed. RESULTS: Three proteins, vitamin K-dependent protein S, prothrombin, and vitronectin, were selectively internalized under sufficient Ca2+ levels in the culture medium. The uptake of these proteins was initiated before DNA replication, and increased during the trophozoite and schizont stages, irrespective of the assembly/disassembly of actin filaments. Coagulation assay revealed that prothrombin was activated and thereby induced blood coagulation. CONCLUSIONS: Serum proteins were taken up by parasites under culture conditions with sufficient Ca2+ levels. This uptake phenomenon was associated with their pathogenicity.

5.
Malar J ; 19(1): 76, 2020 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-32070358

RESUMO

BACKGROUND: Usage of chloroquine was discontinued from the treatment of Plasmodium falciparum infection in almost all endemic regions because of global spread of resistant parasites. Since the first report in Malawi, numerous epidemiological studies have demonstrated that the discontinuance led to re-emergence of chloroquine-susceptible P. falciparum, suggesting a possible role in future malaria control. However, most studies were cross-sectional, with few studies looking at the persistence of chloroquine recovery in long term. This study fills the gap by providing, for a period of at least 6 years, proof of persistent re-emergence/stable recovery of susceptible parasite populations using both molecular and phenotypic methods. METHODS: Ex vivo drug-susceptibility assays to chloroquine (n = 319) and lumefantrine (n = 335) were performed from 2013 to 2018 in Gulu, Northern Uganda, where chloroquine had been removed from the official malaria treatment regimen since 2006. Genotyping of pfcrt and pfmdr1 was also performed. RESULTS: Chloroquine resistance (≥ 100 nM) was observed in only 3 (1.3%) samples. Average IC50 values for chloroquine were persistently low throughout the study period (17.4-24.9 nM). Parasites harbouring pfcrt K76 alleles showed significantly lower IC50s to chloroquine than the parasites harbouring K76T alleles (21.4 nM vs. 43.1 nM, p-value = 3.9 × 10-8). Prevalence of K76 alleles gradually increased from 71% in 2013 to 100% in 2018. CONCLUSION: This study found evidence of stable persistence of chloroquine susceptibility with the fixation of pfcrt K76 in Northern Uganda after discontinuation of chloroquine in the region. Accumulation of similar evidence in other endemic areas in Uganda could open channels for possible future re-use of chloroquine as an option for malaria treatment or prevention.

6.
Front Immunol ; 10: 2669, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31824483

RESUMO

Clinical manifestation of malaria is mainly due to intra-erythrocytic development of Plasmodium parasites. Plasmodium falciparum merozoites, the invasive form of the blood-stage parasite, invade human erythrocytes in a complex but rapid process. This multi-step progression involves interactions between parasite and human host proteins. Here we show that antibodies against a vaccine antigen, PfGAMA, co-immunoprecipitate with PfMSP10. This interaction was validated as direct by surface plasmon resonance analysis. We then demonstrate that antibodies against PfMSP10 have growth inhibitory activity against cultured parasites, with the region PfMSP10 R1 that is critical for its interaction with PfGAMA being the key target. We also observe that the PfMSP10 R1 region is highly conserved among African field isolates. Lastly, we show that high levels of antibodies against PfMSP10 R1 associate with reduced risk to clinical malaria in children resident in a malaria endemic region in northern Uganda. Put together, these findings provide for the first time the functional context of the important role of PfGAMA/PfMSP10 interaction in erythrocyte invasion and unveil a novel asexual blood-stage malaria vaccine target for attenuating P. falciparum merozoite invasion.

7.
Sci Rep ; 9(1): 12517, 2019 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-31467354

RESUMO

The faecal microbiota plays a critical role in host health, with alterations in the human faecal microbial composition associated with various conditions, particularly diarrhoeal diseases. However, little is known about microbial changes during cryptosporidiosis, one of the most important diarrhoeal diseases caused by protozoa in cattle. In this study, alterations in the faecal microbiota of neonatal calves as a result of Cryptosporidium parvum infection were investigated on a C. parvum-positive farm. Comparisons were made among groups of C. parvum-infected, rotavirus-infected, and the pathogen-negative calves. A specific increase in the abundance of Fusobacterium was observed in the faecal microbiota of C. parvum-infected animals. Diarrhoea severity increased in accordance with the abundance of C. parvum and Fusobacterium. Moreover, the specific increase of Fusobacterium appeared to be a universal feature of C. parvum infection, since neonatal calves from geographically separated areas showed the same result. These observations indicated that the growth of Fusobacterium may be an important aggravating factor of cryptosporidiosis.

8.
Emerg Microbes Infect ; 8(1): 1043-1053, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31287781

RESUMO

The prevalence of nontuberculous mycobacteria (NTM) pulmonary diseases has been increasing worldwide. NTM consist of approximately 200 species and distinguishing between them at the subspecies level is critical to treatment. In this study, we sequenced 63 NTM genomes, 27 of which were newly determined, by hybrid assembly using sequencers from Illumina and Oxford Nanopore Technologies (ONT). This analysis expanded the available genomic data to 175 NTM species and redefined their subgenus classification. We also developed a novel multi-locus sequence typing (MLST) database based on 184 genes from 7547 assemblies and an identification software, mlstverse, which can also be used for detecting other bacteria given a suitable MLST database. This method showed the highest sensitivity and specificity amongst conventional methods and demonstrated the capacity for rapid detection of NTM, 10 min of sequencing of the ONT MinION being sufficient. Application of this methodology could improve disease epidemiology and increase the cure rates of NTM diseases.


Assuntos
Genoma Bacteriano , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas/isolamento & purificação , Genômica , Humanos , Tipagem de Sequências Multilocus , Micobactérias não Tuberculosas/classificação , Micobactérias não Tuberculosas/genética , Filogenia
9.
Malar J ; 18(1): 237, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31307493

RESUMO

BACKGROUND: Basic blue 3 is a promising anti-malarial lead compound based on the π-delocalized lipophilic cation hypothesis. Its derivatives with nitrogen atoms bonded to carbon atoms at the 3- and 7-positions on the phenoxazine ring were previously shown to exert potent antiprotozoal activity against Plasmodium falciparum, Trypanosoma cruzi, Trypanosoma brucei rhodesiense, and Leishmania donovani parasites in vitro. However, compounds with nitrogen modification at the 10-position on the phenoxazine ring were not evaluated. METHODS: Six acylphenoxazine derivatives (ITT-001 to 006) with nitrogen modification at the 10-position on the phenoxazine ring, which were synthesized from basic blue 3, were characterized and evaluated for anti-malarial activity in vitro with an automated haematology analyzer (XN-30) and light microscopy. Intensity of self-fluorescence was measured using a fluorometer. Localization of basic blue 3 was observed by fluorescence microscopy. Cytotoxicity was evaluated using human cell lines, HEK293T and HepG2 cells. Finally, anti-malarial activity was evaluated in a rodent malaria model. RESULTS: All the six derivatives showed anti-malarial efficacy even against chloroquine-, pyrimethamine-, and artemisinin-resistant field isolates similar to the sensitive strains and isolates in vitro. The efficacy of basic blue 3 was the strongest, followed by that of ITT-001 to 004 and 006, while that of ITT-005 was the weakest. Basic blue 3 showed strong self-fluorescence, whereas ITT derivatives had five- to tenfold lower intensity than that of basic blue 3, which was shown by fluorescence microscopy to be selectively accumulated in the plasmodial cytoplasm. In contrast, ITT-003, 004, and 006 exhibited the lowest cytotoxicity in HEK293T and HepG2 cells in vitro and the highest selectivity between anti-malarial activity and cytotoxicity. The in vivo anti-malarial assay indicated that oral administration of ITT-004 was the most effective against the rodent malaria parasite, Plasmodium berghei NK65 strain. CONCLUSIONS: The six ITT derivatives were effective against chloroquine- and pyrimethamine-resistant strains and artemisinin-resistant field isolates as well as the sensitive ones. Among them, ITT-004, which had high anti-malarial activity and low cytotoxicity in vitro and in vivo, is a promising anti-malarial lead compound.


Assuntos
Antimaláricos/farmacologia , Oxazinas/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Antimaláricos/toxicidade , Células HEK293 , Células Hep G2 , Humanos , Oxazinas/toxicidade , Testes de Toxicidade
10.
J Nat Med ; 73(4): 820-825, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31140017

RESUMO

Two new sarpagine-type indole alkaloids (1 and 2), together with five known alkaloids; 12-methoxy-4-methylvoachalotine (3), 16-demethoxycarbonylvoacamine (4), isositsirikine (5), affinisine (6), affinine (7), were isolated from the bark of Tabernaemontana macrocarpa Jack. The structures of these alkaloids were determined based on spectroscopic data, chemical correlation, and comparison with the literature. 16-Demethoxycarbonylvoacamine (4) showed antiplasmodial activities against Plasmodium falciparum 3D7 and cytotoxic activities against human cell line, HepG2 cells.


Assuntos
Antimaláricos/farmacologia , Alcaloides Indólicos/farmacologia , Extratos Vegetais/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Tabernaemontana/química , Antimaláricos/química , Humanos , Alcaloides Indólicos/química , Alcaloides Indólicos/isolamento & purificação , Estrutura Molecular , Extratos Vegetais/química
11.
Sci Rep ; 9(1): 7274, 2019 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-31086239

RESUMO

The malaria parasite species, Plasmodium vivax infects not only humans, but also African apes. Human specific P. vivax has evolved from a single ancestor that originated from a parasite of African apes. Although previous studies have proposed phylogenetic trees positioning P. vivax (the common ancestor of human and African ape P. vivax) within the assemblages of Asian primate parasites, its position has not yet been robustly confirmed. We determined nearly complete apicoplast genome sequences from seven Asian primate parasites, Plasmodium cynomolgi (strains Ceylonensis and Berok), P. knowlesi P. fragile, P. fieldi, P. simiovale, P. hylobati, P. inui, and an African primate parasite, P. gonderi, that infects African guenon. Phylogenetic relationships of the Plasmodium species were analyzed using newly and previously determined apicoplast genome sequences. Multigene maximum likelihood analysis of 30 protein coding genes did not position P. vivax within the Asian primate parasite clade but positioned it basal to the clade, after the branching of an African guenon parasite, P. gonderi. The result does not contradict with the emerging notion that P. vivax phylogenetically originated from Africa. The result is also supported by phylogenetic analyses performed using massive nuclear genome data of seven primate Plasmodium species.

12.
J Nat Med ; 73(3): 533-540, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30911994

RESUMO

Two new bisindole alkaloids, leucophyllinines A (1) and B (2) consisting of eburnane and quebrachamine-type skeletons were isolated from the bark of Leuconotis eugeniifolia, and their structures were elucidated on the basis of spectroscopic data. Leucophyllinines A and B showed antiplasmodial activities against Plasmodium falciparum 3D7.


Assuntos
Antimaláricos/química , Antimaláricos/farmacologia , Apocynaceae/metabolismo , Alcaloides Indólicos/química , Alcaloides Indólicos/isolamento & purificação , Plasmodium falciparum/efeitos dos fármacos , Humanos , Estrutura Molecular , Casca de Planta/metabolismo
13.
Malar J ; 18(1): 8, 2019 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-30642330

RESUMO

BACKGROUND: The erythrocytic stage of Plasmodium falciparum parasites in humans is clinically important, as the parasites at this growth stage causes malarial symptoms. Most of the currently available anti-malarial drugs target this stage, but the emergence and spread of parasites resistant to anti-malarial drugs are a major challenge to global eradication efforts; therefore, the development of novel medicines is urgently required. In this study, the in vitro anti-malarial activity of five current anti-malarial drugs (artemisinin, atovaquone, chloroquine, mefloquine, and pyrimethamine) and 400 compounds from the Pathogen Box provided by the Medicines for Malaria Venture on P. falciparum parasites was characterized using the XN-30 analyzer. Furthermore, the outcomes obtained using the analyser were classified according to the parasitaemias of total and each developmental stages. RESULTS: The growth inhibition rate and the half-maximal (50%) inhibitory concentration (IC50) of the five current anti-malarial drugs were calculated from the parasitaemia detected using the XN-30 analyzer. Respective strains and drugs presented strongly fitted sigmoidal curves, and the median SD at all tested concentrations was 1.6, suggesting that the variation in values measured with the analyser was acceptably low for the comparison of drug efficacy. Furthermore, the anti-malarial activity of the 400 compounds from the Pathogen Box was tested, and 141 drugs were found to be effective. In addition, the efficacy was classified into 4 types (Type I, parasites were arrested or killed without DNA replication; Type II, parasites were arrested or killed similar to Type I, and the parasitaemia was apparently decreased; Type III, parasites progressed to trophozoite without sufficient DNA replication; and Type IV, parasites were arrested at late trophozoite or schizont after DNA replication). CONCLUSION: The current study demonstrates that the XN-30 analyzer objectively, reproducibly, and easily evaluated and characterized the anti-malarial efficacy of various compounds. The results indicate the potential of the XN-30 analyzer as a powerful tool for drug discovery and development in addition to its use as an important diagnostic tool.


Assuntos
Antimaláricos/farmacologia , Automação Laboratorial/instrumentação , Descoberta de Drogas/instrumentação , Hematologia/instrumentação , Antimaláricos/isolamento & purificação , Atovaquona/farmacologia , Automação Laboratorial/métodos , Cloroquina/farmacologia , Descoberta de Drogas/métodos , Hematologia/métodos , Humanos , Concentração Inibidora 50 , Malária Falciparum/tratamento farmacológico , Mefloquina/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Esquizontes/efeitos dos fármacos , Trofozoítos/efeitos dos fármacos
14.
Vaccine ; 36(45): 6826-6833, 2018 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-30262245

RESUMO

Acquired antibodies directed towards antigens expressed on the surface of merozoites and infected erythrocytes play an important role in protective immunity to Plasmodium falciparum malaria. P. falciparum erythrocyte membrane protein 1 (PfEMP1), the major parasite component of the infected erythrocyte surface, has been implicated in malaria pathology, parasite sequestration and host immune evasion. However, the extent to which unique PfEMP1 domains interact with host immune response remains largely unknown. In this study, we sought to comprehensively understand the naturally acquired antibody responses targeting different Duffy binding-like (DBL), and Cysteine-rich interdomain region (CIDR) domains in a Ugandan cohort. Consequently, we created a protein library consisting of full-length DBL (n = 163) and CIDR (n = 108) domains derived from 62-var genes based on 3D7 genome. The proteins were expressed by a wheat germ cell-free system; a system that yields plasmodial proteins that are comparatively soluble, intact, biologically active and immunoreactive to human sera. Our findings suggest that all PfEMP1 DBL and CIDR domains, regardless of PfEMP1 group, are targets of naturally acquired immunity. The breadth of the immune response expands with children's age. We concurrently identified 10 DBL and 8 CIDR domains whose antibody responses were associated with reduced risk to symptomatic malaria in the Ugandan children cohort. This study highlights that only a restricted set of specific domains are essential for eliciting naturally acquired protective immunity in malaria. In light of current data, tandem domains in PfEMP1s PF3D7_0700100 and PF3D7_0425800 (DC4) are recommended for extensive evaluation in larger population cohorts to further assess their potential as alternative targets for malaria vaccine development.


Assuntos
Anticorpos Antiprotozoários/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Fatores Etários , Formação de Anticorpos/imunologia , Criança , Feminino , Humanos , Malária/imunologia , Malária/prevenção & controle , Vacinas Antimaláricas/uso terapêutico , Masculino , Plasmodium falciparum/metabolismo , Plasmodium falciparum/patogenicidade , Estudos Prospectivos , Uganda
15.
Parasitol Int ; 67(5): 601-604, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29803940

RESUMO

The asexual blood stages of the Plasmodium falciparum parasite are responsible for inducing the clinical symptoms and the most severe presentations of malaria infection that causes frequent mortality and morbidity in tropical and subtropical areas of the world, making the blood stages of infection a key target of new malaria treatment and prevention strategies. Progress towards the development of more effective treatment and prevention strategies has been hindered by the limited availability of infection models that permit the sequential analysis of blood stage parasites in vitro followed by in vivo analysis to confirm therapeutic benefits. To advance a model for in vitro and in vivo analysis of blood stage parasites, we examined nine laboratory strains of P. falciparum to determine which strains could adapt to growth in vivo in splenectomized squirrel monkeys (Saimiri sciureus). Only one of the nine laboratory strains tested, the FCB strain, adapted to in vivo growth. Morphological analysis show that the adapted ring-stage parasites have a different morphology from original parasites cultured in vitro, and more often they were found to localize at the edge of the infected red blood cell. No remarkable differences were observed for both trophozoites and schizonts. The adapted strain can be cultured back in vitro similar to the original parasite, indicating that the adapted parasite can develop both in vitro and in vivo. This squirrel monkey-adapted P. falciparum parasite is expected to be suitable and is advantageous for the research and development of vaccines and antimalarial drugs.


Assuntos
Adaptação Fisiológica , Plasmodium falciparum/genética , Saimiri/parasitologia , Animais , Modelos Animais de Doenças , Genoma de Protozoário , Laboratórios , Parasitemia , Plasmodium falciparum/fisiologia , Baço/parasitologia
16.
Malar J ; 17(1): 165, 2018 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-29661200

RESUMO

BACKGROUND: The erythrocytic stage, where malaria parasites proliferate in human blood, is clinically significant as this causes the symptoms and illness of malaria. Experimental rodent models of malaria at the erythrocytic stage are used for the development of anti-malarial drugs and for biological analysis. An automated haematology analyzer XN-30 was developed for detection of infected red blood cells (iRBCs) in human blood samples and measurement of their parasitaemia in approximately 1 min through flow cytometry analysis. Additionally, the analyzer simultaneously measured other haematological parameters in these samples. It is inferred that the analyzer would also allow easy and rapid measurement of parasitaemia in mice and provide important clues on the mouse haematological state during infection and treatment. RESULTS: The XN-30 analyzer is a simple and rapid tool to detect iRBCs in mouse blood samples infected with rodent malarial parasites, with three-dimensional analysis permitting the precise measurement of parasitaemia (referred herein as the 'XN-30 system'). The XN-30 analyzer allowed not only the detection of iRBCs but also the monitoring of RBC, white blood cell, and platelet counts, as well as haematocrit, mean corpuscular volume and mean platelet volume values in the mouse blood sample. For anti-malarial drug development, aside from demonstrating possible efficacy in mouse models, XN-30 analyzer could provide a first glimpse of the safety profile of the drug. CONCLUSIONS: The XN-30 system is a powerful tool that can be utilized for the in vivo screening, development, and evaluation of anti-malarial drugs as well as for pre-clinical pharmacology and/or toxicity tests in rodent models.


Assuntos
Eritrócitos/parasitologia , Testes Hematológicos/métodos , Malária Falciparum/diagnóstico , Parasitemia/diagnóstico , Plasmodium falciparum/isolamento & purificação , Animais , Feminino , Citometria de Fluxo , Testes Hematológicos/instrumentação , Malária Falciparum/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Parasitemia/parasitologia
17.
Emerg Infect Dis ; 24(4): 718-726, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29553316

RESUMO

Because ≈90% of malaria cases occur in Africa, emergence of artemisinin-resistant Plasmodium falciparum in Africa poses a serious public health threat. To assess emergence of artemisinin-resistant parasites in Uganda during 2014-2016, we used the recently developed ex vivo ring-stage survival assay, which estimates ring-stage-specific P. falciparum susceptibility to artemisinin. We conducted 4 cross-sectional surveys to assess artemisinin sensitivity in Gulu, Uganda. Among 194 isolates, survival rates (ratio of viable drug-exposed parasites to drug-nonexposed controls) were high (>10%) for 4 isolates. Similar rates have been closely associated with delayed parasite clearance after drug treatment and are considered to be a proxy for the artemisinin-resistant phenotype. Of these, the PfKelch13 mutation was observed in only 1 isolate, A675V. Population genetics analysis suggested that these possibly artemisinin-resistant isolates originated in Africa. Large-scale surveillance of possibly artemisinin-resistant parasites in Africa would provide useful information about treatment outcomes and help regional malaria control.


Assuntos
Antimaláricos/farmacologia , Artemisininas/farmacologia , Resistência a Medicamentos , Malária Falciparum/epidemiologia , Malária Falciparum/parasitologia , Plasmodium falciparum/efeitos dos fármacos , Antimaláricos/uso terapêutico , Artemisininas/uso terapêutico , Pré-Escolar , Estudos Transversais , Feminino , Genótipo , História do Século XXI , Humanos , Malária Falciparum/história , Malária Falciparum/mortalidade , Masculino , Mutação , Fenótipo , Plasmodium falciparum/genética , Taxa de Sobrevida , Uganda/epidemiologia , Sequenciamento Completo do Genoma
18.
Sci Rep ; 8(1): 5052, 2018 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-29567995

RESUMO

The malaria parasite Plasmodium falciparum proliferates in the blood stream where the host immune system is most active. To escape from host immunity, P. falciparum has developed a number of evasion mechanisms. Serine repeat antigen 5 (SERA5) is a blood stage antigen highly expressed at late trophozoite and schizont stages. The P47 N-terminal domain of SERA5, the basis of SE36 antigen of the blood stage vaccine candidate under clinical trials, covers the merozoite surface. Exploring the role of the P47 domain, screening of serum proteins showed that vitronectin (VTN) directly binds to 20 residues in the C-terminal region of SE36. VTN co-localized with P47 domain in the schizont and merozoite stages. Phagocytosis assay using THP-1 cells demonstrated that VTN bound to SE36 prevented engulfment of SE36-beads. In addition, several serum proteins localized on the merozoite surface, suggesting that host proteins camouflage merozoites against host immunity via binding to VTN.


Assuntos
Antígenos de Protozoários/genética , Malária Falciparum/genética , Plasmodium falciparum/genética , Vitronectina/genética , Animais , Antígenos/genética , Antígenos/metabolismo , Antígenos de Protozoários/metabolismo , Interações Hospedeiro-Parasita/genética , Interações Hospedeiro-Parasita/imunologia , Humanos , Imunidade/genética , Malária Falciparum/imunologia , Malária Falciparum/parasitologia , Merozoítos/genética , Merozoítos/imunologia , Merozoítos/patogenicidade , Camundongos , Fagocitose/imunologia , Plasmodium falciparum/imunologia , Plasmodium falciparum/patogenicidade , Ligação Proteica/genética , Vitronectina/metabolismo
20.
Malar J ; 17(1): 59, 2018 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-29391022

RESUMO

BACKGROUND: The automated haematology analyzer XN-30 (Sysmex, Kobe, Japan) easily and rapidly detects malarial parasites in clinical blood samples using flow cytometry. The XN-30 analyzer is able to distinguish each developmental stage by measuring DNA content and cell size. Thus, it was expected to be capable of quantifying the developmental stages of cultured falciparum parasite. To achieve this requirement, a modified algorithm was tested for its validity and reliability using in vitro cultured falciparum parasite. RESULTS: The XN-30 analyzer automatically measured each developmental stage as well as total parasitaemia. Comparison of the parasitaemia obtained using the XN-30 analyzer equipped with the modified algorithm with that obtained using microscopy examination of Giemsa-stained smears revealed the greater sensitivity and reproducibility of the former. The XN-30 analyzer also detected free merozoites and purified gametocytes. CONCLUSIONS: The XN-30 analyzer allows the precise recognition and enumeration of total and each developmental stages of cultured falciparum parasites, and permits the sensitive and reproducible calculation of parasitaemia. The results indicate the potential of the XN-30 analyzer for basic research on malarial biology, anti-malarial drug discovery, and evaluation of drug efficacy.


Assuntos
Citometria de Fluxo/métodos , Malária Falciparum/diagnóstico , Parasitemia/diagnóstico , Plasmodium falciparum/fisiologia , Automação Laboratorial/métodos , Técnicas de Cultura , Eritrócitos/parasitologia , Humanos , Malária Falciparum/parasitologia , Merozoítos/isolamento & purificação , Merozoítos/fisiologia , Parasitemia/parasitologia , Parasitologia/métodos , Plasmodium falciparum/isolamento & purificação
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