Falciparum Infected Individuals Research Articles

Persistent carriage of subpatent Plasmodium falciparum parasites associated with clinical malaria in a low transmission area in Senegal

ObjectivesIn low malaria transmission areas, the elimination of the disease has been hampered partly by the existence of a reservoir of subpatent Plasmodium falciparum infections within communities. This reservoir, often undetected, serves as a source of parasites and contributes to ongoing transmission and clinical malaria cases. MethodsThis study, spanning a period of 9 years from June 2014 to December 2022, examined individual variations and long-term subpatent P. falciparum carriage in two matched cohorts of 44 individuals each living in Dielmo village in the Sudanian area of Senegal. Biannual blood samples, collected in June/July and December of each year, underwent P. falciparum diagnosis by quantitative polymerase chain reaction. QGIS and R analytical tools were used to map infections, assess the occurrence and clustering of subpatent and clinical P. falciparum infections, and determine the risk of infection in the vicinity of asymptomatic P. falciparum carriers. ResultsThe point frequency and long-term P. falciparum carriage were significantly higher among the quantitative polymerase chain reaction (qPCR) positive cohort compared to the negative cohort across the 16 cross-sectional surveys analyzed in this study (19.76% vs 10.99%, P-value <0.001). Asymptomatic carriage events in qPCR-positive group were 18.86 ± 1.72% and significantly greater (P-value = 0.001) than in the qPCR-negative group (11.32 ± 1.32%). The relative risk of P. falciparum infection or clinical malaria calculated with a 95% confidence interval significantly increased in the vicinity of infected individuals and was 1.44 (P-value = 0.53) and 2.64 (P-value = 0.04) when at least one individual in the direct (household) or indirect (block of households) vicinity is infected, respectively. The risk increased to 3.64 (P-value <0.001) if at least 1/5 of individuals in the indirect vicinity were P. falciparum-infected. ConclusionsThe study provides a detailed qualitative and quantitative analysis of the asymptomatic P. falciparum reservoir and its temporal and spatial dynamics within two subgroups of P. falciparum-infected and non-infected individuals in Dielmo village. It identified high-risk populations known as “hotpops” and hotspots that could be targeted by innovative interventions to accelerate the elimination of malaria in Dielmo village.

Evaluation of immunophenotypic alterations of peripheral blood lymphocytes and their sub-sets in uncomplicated P. Falciparum infection

BackgroundMalaria is a life-threatening parasitic disease typically transmitted through the bite of an infected Anopheles mosquito. There is ample evidence showing the potential of malaria infection to affect the counts of lymphocyte subpopulations in the peripheral blood, but the extent of alteration might not be consistent in all geographical locations, due to several local factors. Although Ghana is among the malaria-endemic countries, there is currently no available data on the level of alterations that occur in the counts of lymphocyte subpopulations during P. falciparum malaria infection among adults.AimThe study was to determine the immunophenotypic alterations in the level of peripheral blood lymphocytes and their subsets in adults with uncomplicated P. falciparum malaria infection and apparently healthy participants.MethodsThe study was a cross-sectional comparative study conducted in two municipalities of the Volta region of Ghana. Blood samples were collected from study participants and taken through serology (P. falciparum/Pan Rapid Diagnostic Kits), microscopy (Thick and thin blood films) and Haematological (Flow cytometric and Full blood count) analysis.ResultsA total of 414 participants, comprising 214 patients with malaria and 200 apparently healthy individuals (controls) were recruited into this study. Parasite density of the malaria patients ranged from 75/µL to 84,364/µL, with a mean of 3,520/µL. It was also observed that the total lymphocytes slightly decreased in the P. falciparum-infected individuals (Mean ± SD: 2.08 ± 4.93 × 109/L) compared to the control group (Mean ± SD: 2.47 ± 0.80 × 109/L). Again, there was a significant moderate positive correlation between parasite density and haematocrit levels (r = 0.321, p < 0.001). Apart from CD45 + T-cells, more people in the control group had normal values for the lymphocyte subsets measured compared to the malaria patients.ConclusionsFrom the results obtained, there was high parasite density among the malaria patients suggestive of high intensity of infection in the case group. The malaria patients again showed considerable haematological alterations in lymphocyte sub-sets and the parasite density appeared to be strongly associated with CD4 + T-cell reduction. Also, the parasite density significantly associated with decreasing haematocrit levels. This indicates that lymphocyte subset enumeration can be used to effectively support malaria diagnosis.

High Frequency of Deletions in the pfhrp2 and pfhrp3 Genes of Plasmodium falciparum in the Middle Rio Negro Region of the Brazilian Amazon.

Several countries are reporting natural populations of P. falciparum with deletions in the pfhrp2/3 genes that can lead to false-negative results in rapid diagnostic tests. To investigate the prevalence of deletion in the pfhrp2/3 genes in the Rio Negro basin in the Brazilian Amazon and identify whether there is clinical differentiation between individuals infected by these parasites, clinical samples collected from 2003 to 2016 were analyzed from symptomatic and asymptomatic P. falciparum-infected individuals. The molecular deletion of pfhrp2 and pfhrp3 genes was evaluated using the protocols recommended by the WHO. From 82 samples used, 28 (34.2%) had a single deletion in pfhrp2, 19 (23.2%) had a single deletion in pfhrp3, 15 (18.3%) had a double deletion (pfhrp2/3), and 20 (24.4%) did not have a deletion in either gene. In total, 29.3% of individuals had an asymptomatic plasmodial infection and were 3.64 times more likely to have parasites with a double deletion (pfhrp2/3) than patients with clinical malaria (p = 0.02). The high prevalence of parasites with pfhrp2/3 deletions shows the need to implement a surveillance program in this area. Deletions in parasites may be associated with the clinical pattern of the disease in this area. More studies must be carried out to elucidate these findings.

Plasmodium falciparum infection in humans and mosquitoes influence natural Anopheline biting behavior and transmission

The human infectious reservoir of Plasmodium falciparum is governed by transmission efficiency during vector-human contact and mosquito biting preferences. Understanding biting bias in a natural setting can help target interventions to interrupt transmission. In a 15-month cohort in western Kenya, we detected P. falciparum in indoor-resting Anopheles and human blood samples by qPCR and matched mosquito bloodmeals to cohort participants using short-tandem repeat genotyping. Using risk factor analyses and discrete choice models, we assessed mosquito biting behavior with respect to parasite transmission. Biting was highly unequal; 20% of people received 86% of bites. Biting rates were higher on males (biting rate ratio (BRR): 1.68; CI: 1.28–2.19), children 5–15 years (BRR: 1.49; CI: 1.13–1.98), and P. falciparum-infected individuals (BRR: 1.25; CI: 1.01–1.55). In aggregate, P. falciparum-infected school-age (5–15 years) boys accounted for 50% of bites potentially leading to onward transmission and had an entomological inoculation rate 6.4x higher than any other group. Additionally, infectious mosquitoes were nearly 3x more likely than non-infectious mosquitoes to bite P. falciparum-infected individuals (relative risk ratio 2.76, 95% CI 1.65–4.61). Thus, persistent P. falciparum transmission was characterized by disproportionate onward transmission from school-age boys and by the preference of infected mosquitoes to feed upon infected people.

Burden of Submicroscopic Plasmodium Infections and Detection of kelch13 Mutant Parasites in Military and Civilian Populations in Papua New Guinea.

Malaria remains a major public health problem in Papua New Guinea (PNG) and an important force health protection issue for both PNG and Australian Defence Forces. To investigate the malaria burden in the military and civilians residing on military bases, a cross-sectional survey was conducted in April 2019 at three military bases in Wewak, Manus Island, and Vanimo, PNG. A total of 1,041 participants were enrolled; 235 military personnel from three bases and 806 civilians from Wewak and Vanimo. Polymerase chain reaction (PCR) revealed an overall high prevalence of Plasmodium infection in both the military and civilians. Among the military, the infection prevalence was significantly higher in Wewak (35.5%) and Vanimo (33.3%) bases than on Manus Island (11.8%). Among civilians, children (<16 years old) had significantly higher odds of being PCR positive than adults (≥16 years old). At Wewak and Vanimo, Plasmodium vivax accounted for 85.4%, 78.2%, and 66.2% of infections in military, children, and adult populations. Overall, 87.3%, 41.3%, and 61.3% of Plasmodium infections in the military, children, and adults, respectively, were detected only by PCR, not by microscopy (submicroscopic [SM] infections). Children had a significantly lower proportion of SM infections than adults and Papua New Guinea Defence Force personnel. Infection status was not associated with hemoglobin levels in these populations at the time of the survey. Mutant kelch13 (C580Y) parasites were identified in 5/68 Plasmodium falciparum-infected individuals. The survey results indicate extensive malaria transmission on these bases, especially in Wewak and Vanimo. More intensified interventions are required to reduce malaria transmission on PNG military bases.

Dynamics of Plasmodium species and genotype diversity in adults with asymptomatic infection in Gabon

ObjectivesWe investigated the diversity and dynamics of Plasmodium infection in serially collected samples from asymptomatic participants of a clinical trial assessing the efficacy and safety of ivermectin in Gabon. We checked whether the baseline sample reflected the P. falciparum genotype and Plasmodium species diversity seen over 7 days of follow-up. MethodsBlood samples were collected at inclusion, every 8 hours until hour 72, daily until day 7, and on day 14. Plasmodium species was determined by qPCR and pfmsp1 length polymorphism was assessed for P. falciparum genotyping. ResultsIn 17/48 (35%) individuals, all pfmsp1 genotypes identified during the assessed period were detected at baseline; in 31/48 (65%), new genotypes were found during follow-up. Additional sampling at hour 24 allowed the identification of all genotypes seen over 7 days in 50% of the individuals. Ivermectin did not impact the genotype dynamics. Mixed Plasmodium spp. infections were detected in 28/49 (57%) individuals at baseline, and detection of non-falciparum infections during follow-up varied. ConclusionsOur results reveal complex intra-host dynamics of P. falciparum genotypes and Plasmodium species and underscore the importance of serial sampling in clinical trials for antimalarial drugs with asymptomatically P. falciparum-infected individuals. This might allow a more accurate identification of genotypes in multiple infections, impacting the assessment of drug efficacy.

Evaluation of a new fusion antigen, cd loop and HAP2-GCS1 domain (cd-HAP) of Plasmodium falciparum Generative Cell Specific 1 antigen formulated with various adjuvants, as a transmission blocking vaccine

BackgroundMalaria is a major global health challenge, and for the elimination and eradication of this disease, transmission-blocking vaccines (TBVs) are a priority. Plasmodium falciparum Generative Cell Specific 1 (PfGCS1), a promising TBV candidate, is essential for gamete fertilization. The HAP2-GCS1 domain of this antigen as well as its cd loop could induce antibodies that partially inhibit transmission of P. falciparum.MethodsIn the current study, a new synthetic fusion antigen containing cd loop and HAP2-GCS1 domain (cd-HAP) of PfGCS1 was evaluated as a transmission blocking vaccine candidate. Initially, the profile of naturally acquired IgG antibodies to the cd-HAP antigen was analysed in Iranian individuals infected with P. falciparum, to confirm that this new fusion protein has the appropriate structure containing common epitopes with the native form of PfGCS1. Then, the immunogenicity of cd-HAP was evaluated in BALB/c mice, using different adjuvant systems such as CpG, MPL, QS-21, and a combination of them (CMQ). Furthermore, the blocking efficacy of polyclonal antibodies induced against these formulations was also assessed by oocyst intensity and infection prevalence in the Standard Membrane Feeding Assay (SMFA).ResultsThe naturally acquired antibodies (dominantly IgG1 and IgG3 subclasses) induced in P. falciparum-infected individuals could recognize the cd-HAP antigen which implies that the new fusion protein has a proper conformation that mimics the native structure of PfGCS1. Concerning the immunogenicity of cd-HAP antigen, the highest IgG levels and titers, by a Th1-type immune profile, and elevated antibody avidity were induced in mice immunized with the cd-HAP antigen formulated with a combination of adjuvants (P < 0.0001). Additionally, cytokine profiling of the immunized mice displayed that a high level of IFN-γ response, a Th1-type immune response, was produced by splenocytes from immunized mice that received cd-HAP antigen in combination with CMQ adjuvants (P < 0.0001). This formulation of cd-HAP antigen with CMQ adjuvants could reduce oocyst intensity and infection prevalence by 82%, evidenced by the SMFA and hold significant implications for future malaria vaccine development.ConclusionAltogether, the results showed that cd-HAP antigen formulated with a combination of the adjuvants (CMQ), could be a promising formulation to develop a PfGCS1-based transmission-blocking vaccine.

Prevalence of mutations in the cysteine desulfurase IscS (Pfnfs1) gene in recurrent Plasmodium falciparum infections following artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) treatment in Matayos, Western Kenya

BackgroundMalaria remains a public health concern globally. Resistance to anti-malarial drugs has consistently threatened the gains in controlling the malaria parasites. Currently, artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) are the treatment regimens against Plasmodium falciparum infections in many African countries, including Kenya. Recurrent infections have been reported in patients treated with AL or DP, suggesting the possibility of reinfection or parasite recrudescence associated with the development of resistance against the two therapies. The Plasmodium falciparum cysteine desulfurase IscS (Pfnfs1) K65 selection marker has previously been associated with decreased lumefantrine susceptibility. This study evaluated the frequency of the Pfnfs1 K65 resistance marker and associated K65Q resistant allele in recurrent infections collected from P. falciparum-infected individuals living in Matayos, Busia County, in western Kenya.MethodsArchived dried blood spots (DBS) of patients with recurrent malaria infection on clinical follow-up days after treatment with either AL or DP were used in the study. After extraction of genomic DNA, PCR amplification and sequencing analysis were employed to determine the frequencies of the Pfnfs1 K65 resistance marker and K65Q mutant allele in the recurrent infections. Plasmodium falciparum msp1 and P. falciparum msp2 genetic markers were used to distinguish recrudescent infections from new infections.ResultsThe K65 wild-type allele was detected at a frequency of 41% while the K65Q mutant allele was detected at a frequency of 22% in the recurrent samples. 58% of the samples containing the K65 wild-type allele were AL treated samples and while 42% were DP treated samples. 79% of the samples with the K65Q mutation were AL treated samples and 21% were DP treated samples. The K65 wild-type allele was detected in three recrudescent infections (100%) identified from the AL treated samples. The K65 wild-type allele was detected in two recrudescent DP treated samples (67%) while the K65Q mutant allele was identified in one DP treated (33%) recrudescent sample.ConclusionsThe data demonstrate a higher frequency of the K65 resistance marker in patients with recurrent infection during the study period. The study underscores the need for consistent monitoring of molecular markers of resistance in regions of high malaria transmission.

IgG antibody responses to Anopheles gambiae gSG6-P1 salivary peptide are induced in human populations exposed to secondary malaria vectors in forest areas in Cameroon.

Human IgG antibody response to Anopheles gambiae gSG6-P1 salivary peptide was reported to be a pertinent indicator for assessing human exposure to mosquito bites and evaluating the risk of malaria transmission as well as the effectiveness of vector control strategies. However, the applicability of this marker to measure malaria transmission risk where human populations are mostly bitten by secondary vectors in Africa has not yet been evaluated. In this study, we aimed to investigate whether anti-gSG6-P1 antibodies response could be induced in humans living in forest areas in Cameroon where An. gambiae s.l is not predominant. In October 2019 at the pick of the rainy season, blood samples were collected from people living in the Nyabessang in the forest area in the South region of Cameroon. Malaria infection was determined using thick blood smear microscopy and Rapid Diagnostic Test. The level of IgG Anti-gSG6-P1 response as a biomarker of human exposure to Anopheles bite, was assessed using enzyme-linked immunosorbent assay. Mosquitoes were collected using the human landing catches to assess Anopheles density and for the identification of Anopheles species present in that area. IgG antibody response to the gSG6-P1 salivary peptide was detected in inhabitants of Nyabessang with high inter-individual heterogeneity. No significant variation in the level of this immune response was observed according to age and gender. The concentration of gSG6-P1 antibodies was significantly correlated with the malaria infection status and, Plasmodium falciparum-infected individuals presented a significantly higher level of IgG response than uninfected individuals (p = 0.0087). No significant difference was observed according to the use of insecticide treated nets. Out of the 1,442 Anopheles mosquitoes species collected, 849 (58.9%) were identified as An. paludis, 489 (33.91%) as An. moucheti, 28 (4.44%) as An. nili, 22 (2.08%) as An. gambiae s.l and 10 (0.69%) as An. marshallii. Our findings show that IgG response to An. gambiae gSG6-P1 peptide could be detected in humans exposed predominantly to An. moucheti and An. paludis bites. Taken together, the data revealed the potential of the Anti-gSG6-P1 IgG antibody response to serve as a universal marker to assess human exposure to any Anopheles species.

Pro- and anti-inflammatory cytokine profiles in Plasmodium Falciparum-infected individuals from Baiyeku, Ikorodu, Lagos, Nigeria

Available evidence indicates that the various stages of the malaria parasite life cycle elicit specific immune responses of which the relative levels of pro-inflammatory cytokines are key to disease progression, killing the parasite and mediating disease outcomes. This study investigated T-cell response in malaria. Four hundred and sixty-two participants were screened in a community survey of Plasmodium falciparum malaria in Baiyeku, Lagos, Nigeria. P. falciparum parasitaemia was determined by microscopy while the serum levels of IL-10, IFNγ and TNFα were determined by Enzyme-linked immunosorbent assay (ELISA). A total of 70 (15.2 %) participants were microscopically positive for P. falciparum of which 70% were females, 30% were males while children aged 1-17 years were 65.7%. The geometric mean parasite density (GMPD) was significantly (p=0.001) higher among females than males. The GMPD of participants &lt;5 years of age was also significantly (p=0.001) higher than other age groups. About 46.8% of the participants were underweight (Body Mass Index, BMI &lt; 18.5) and also had the highest parasite intensity. The TNFα, IFNγ and IL-10 levels were significantly (p&lt; 0.05) higher in the infected than the uninfected participants. IFN-γ values were significantly (p=0.014) elevated among the symptomatic than the asymptomatic participants while there was no significant difference (P&gt;0.053) in the levels of TNF-α and IL-10 (P&gt;0.093) between the symptomatic and asymptomatic participants. The prevalence of P. falciparum obtained in this study area which is endemic to malaria is 15.2% suggesting a significant reduction of the disease over time due to awareness of the disease in the community. The levels of pro-inflammatory cytokines (IFN-γ and TNF-α) in this study were lower due to the down-regulatory action of the anti-inflammatory cytokines (IL-10). These findings suggest that higher levels of anti-inflammatory cytokines, especially IL-10 levels may contribute to the pathogenesis of uncomplicated malaria.

Sequence polymorphism within erythrocyte binding domain of EBA175 in Indian and African field isolates

Invasion of erythrocyte by Plasmodium merozoites is mediated by specific molecular interactions between proteins expressed on merozoite surface and the receptors present on erythrocytes. Erythrocyte binding antigen 175 (EBA175) is one such protein that interacts with the sialic acid residues on glycophorin A present on erythrocytes’ surface during invasion. The FII region (PfFII) of EBA175 has been mapped to be critical for binding to erythrocytes. It is reported that antibodies against FII region blocks binding. Polymorphisms in FII region of EBA175 are already reported. The goal of this study was to investigate whether polymorphism in FII region of African P. falciparum field isolates has any effect on erythrocyte binding and also to find whether antibodies raised against FII region from P. falciparum Malayan Camp strain (Camp) can inhibit erythrocyte binding. Genomic DNA of parasites from the blood samples of P. falciparum infected individuals was isolated and PfFII region from these genomic DNA were amplified, cloned and sequenced. Following sequence analysis, we selected three isolates harboring higher PfFII polymorphisms, expressed them on the surface of COS cells as chimeric proteins using secretory signal and transmembrane segments of Herpex simplex virus glycoprotein D (HSVg D) and tested for their erythrocyte binding ability. We further tested the inhibition of erythrocyte binding of these polymorphic FII regions using anti-campPfF2 antibodies. Our results reveal that the polymorphisms in different field isolates included in this study do not have any significant effect on erythrocyte binding and antibodies raised against FII region of camp strain could inhibit erythrocyte binding by all the polymorphic PfFII. This observation strengthens the possibility that PfFII can be a potential candidate vaccine.

Natural infections with different Plasmodium species induce antibodies reactive to a chimeric Plasmodium vivax recombinant protein

BackgroundAs malaria incidence and transmission in a region decreases, it becomes increasingly difficult to identify areas of active transmission. Improved methods for identifying and monitoring foci of active malaria transmission are needed in areas of low parasite prevalence in order to achieve malaria elimination. Serological assays can provide population-level infection history to inform elimination campaigns.MethodsA bead-based multiplex antibody detection assay was used to evaluate a chimeric Plasmodium vivax MSP1 protein (PvRMC-MSP1), designed to be broadly immunogenic for use in vaccine studies, to act as a pan-malaria serological tool based on its ability to capture IgG in plasma samples obtained from naturally exposed individuals. Samples from 236 US travellers with PCR confirmed infection status from all four major Plasmodium species infecting humans, Plasmodium falciparum (n = 181), Plasmodium vivax (n = 38), Plasmodium malariae (n = 4), and Plasmodium ovale (n = 13) were tested for IgG capture using PvRMC-MSP1 as well as the four recombinant MSP1-19 kD isoforms representative of these Plasmodium species.ResultsRegardless of infecting Plasmodium species, a large proportion of plasma samples from infected US travellers provided a high assay signal to the PvRMC-MSP1 chimeric protein, with 115 high responders out of 236 samples assessed (48.7%). When grouped by active infection, 38.7% P. falciparum-, 92.1% of P. vivax-, 75.0% P. malariae-, and 53.4% of P. ovale-infected individuals displayed high assay signals in response to PvRMC-MSP1. It was also determined that plasma from P. vivax-infected individuals produced increased assay signals in response to the PvRMC-MSP1 chimera as compared to the recombinant PvMSP1 for 89.5% (34 out of 38) of individuals. PvRMC-MSP1 also showed improved ability to capture IgG antibodies from P. falciparum-infected individuals when compared to the capture by recombinant PvMSP1, with high assay signals observed for 38.7% of P. falciparum-infected travellers in response to PvRMC-MSP1 IgG capture compared to just 1.1% who were high responders to capture by the recombinant PvMSP1 protein.ConclusionsThese results support further study of designed antigens as an approach for increasing sensitivity or broadening binding capacity to improve existing serological tools for determining population-level exposure to Plasmodium species. Including both broad-reacting and Plasmodium species-specific antigen-coated beads in an assay panel could provide a nuanced view of population-level exposure histories, an extensive IgG profile, and detailed seroestimates. A more sensitive serological tool for detection of P. vivax exposure would aid malaria elimination campaigns in co-endemic areas and regions where P. vivax is the dominant parasite.

High susceptibility to severe malaria among patients with A blood group versus those with O blood group: A cross-sectional study in the Democratic Republic of the Congo.

This study aimed to assess the association of severe malaria infection with the ABO blood groups among acute febrile patients at the General Hospital of Rungu, in the Democratic Republic of the Congo. This cross-sectional study was conducted between August and October 2018. Plasmodium falciparum-infected individuals were categorized as severe malaria and uncomplicated malaria. A total of 400 febrile patients were enrolled. The majority (n = 251; 62.8%) was positive P. falciparum in microscopy test, of whom 180 (71.7%) had uncomplicated malaria and 71 (28.3%) severe malaria; 32.3%, 18.3%, 2.8%, and 46.6% were found to be blood group of A, B, AB, and O, respectively. In the multivariate analysis using the logistic regression models, severe malaria was high among patients with A blood group compared to those with O blood group (45.8% vs. 13.7%; adjusted odds ratio: 5.3 [95% confidence interval: 2.7–10.5]; P < 0.001). This survey demonstrates that patients with A blood group had a high susceptibility to severe malaria compared to those with O blood group.

In vitro activity of eravacycline, a novel synthetic halogenated tetracycline, against the malaria parasite Plasmodium falciparum

ObjectivesEravacycline is a novel synthetic halogenated tetracycline derivative with a broad antibacterial spectrum. Antibiotics, including tetracyclines, have been used for prophylaxis and, more rarely, for the treatment of malaria for several decades. The rise in drug-resistant malaria parasites renders the search for new treatment candidates urgent. We determined the in vitro potency of eravacycline against Plasmodium falciparum and investigated the apicoplast as a potential drug target. MethodsFour tetracyclines, including eravacycline, tetracycline, tigecycline, and doxycycline, and the lincosamide clindamycin, were tested in 3-day and 6-day in vitro susceptibility assays of P. falciparum laboratory strain 3D7 and/or of clinical isolates obtained from 33 P. falciparum infected individuals from Gabon in 2018. Assays with isopentenyl pyrophosphate substitution were performed to investigate whether apicoplast-encoded isoprenoid biosynthesis is inhibited by these antibiotics. ResultsEravacycline showed the highest activity of all tetracyclines tested in clinical isolates in the 3-day and 6-day assays. Substitution of isopentenyl pyrophosphate in vitro using the laboratory strain 3D7 reversed the activity of eravacycline and comparator antibiotics, indicating the apicoplast to be the main target organelle. ConclusionsThese results demonstrate the potential of novel antibiotics, and eravacycline, as candidate antimalarial therapies.

P. falciparum and P. vivax Orthologous Coiled-Coil Candidates for a Potential Cross-Protective Vaccine.

Over the last four decades, significant efforts have been invested to develop vaccines against malaria. Although most efforts are focused on the development of P. falciparum vaccines, the current availability of the parasite genomes, bioinformatics tools, and high throughput systems for both recombinant and synthetic antigen production have helped to accelerate vaccine development against the P. vivax parasite. We have previously in silico identified several P. falciparum and P. vivax proteins containing α-helical coiled-coil motifs that represent novel putative antigens for vaccine development since they are highly immunogenic and have been associated with protection in many in vitro functional assays. Here, we selected five pairs of P. falciparum and P. vivax orthologous peptides to assess their sero-reactivity using plasma samples collected in P. falciparum- endemic African countries. Pf-Pv cross-reactivity was also investigated. The pairs Pf27/Pv27, Pf43/Pv43, and Pf45/Pv45 resulted to be the most promising candidates for a cross-protective vaccine because they showed a high degree of recognition in direct and competition ELISA assays and cross-reactivity with their respective ortholog. The recognition of P. vivax peptides by plasma of P. falciparum infected individuals indicates the existence of a high degree of cross-reactivity between these two Plasmodium species. The design of longer polypeptides combining these epitopes will allow the assessment of their immunogenicity and protective efficacy in animal models.

Acquired antibody responses against merozoite surface protein-119 antigen during Plasmodium falciparum and P.vivax infections in South Indian city of Mangaluru.

Merozoite surface protein-1 (MSP-1) of malaria parasites has been extensively studied as a malaria vaccine candidate and the antibody response to this protein is an important indicator of protective immunity to malaria. Mangaluru city and its surrounding areas in southwestern India are endemic to malaria with Plasmodium vivax being the most widespread and prevalent species although P. falciparum also frequently infects. However, no information is available on the level of protective immunity in this population. In this regard, a prospective hospital-based study was performed in malarial patients to assess antibody responses against the 19-kDa C-terminal portion of P. vivax and P. falciparum MSP-1 (MSP-119). Serum samples from 51 healthy endemic controls and 267 infected individuals were collected and anti-MSP-119 antibody levels were analyzed by ELISA. The possible association between the antibody responses and morbidity parameters such as malarial anemia and thrombocytopenia was investigated. Among the 267 infected cases, 144 had P. vivax and 123 had P. falciparum infections. Significant levels of anti-MSP-119 antibody were observed both in P. vivax (123/144; 85.4%) and P. falciparum (108/123; 87.9%) infected individuals. In both type of infections, the major antibody isotypes were IgG1 and IgG3. The IgG levels were found to be increased in patients with severe anemia and thrombocytopenia. The antibody levels were also higher in infected individuals who had several previous infections, although antibodies produced during previous infections were short lived. The predominance of cytophilic anti-MSP-119 IgG1 and IgG3 antibodies suggests the possibility of a dual role of Pv MSP-119 and Pf MSP-119 during malarial immunity and pathogenesis.

Recombinant proteins of Plasmodium malariae merozoite surface protein 1 (PmMSP1): Testing immunogenicity in the BALB/c model and potential use as diagnostic tool.

BackgroundPlasmodium malariae is the third most prevalent human malaria-causing species and has a patchy, but ample distribution in the world. Humans can host the parasite for years without presenting significant symptoms, turning its diagnosis and control into a difficult task. Here, we investigated the immunogenicity of recombinant proteins of P. malariae MSP1.MethodsFive regions of PmMSP1 were expressed in Escherichia coli as GST-fusion proteins and immunized in BALB/c mice. The specificity, subtyping, and affinity of raised antibodies were evaluated by enzyme-linked immunosorbent assays. Cellular immune responses were analyzed by lymphoproliferation assays and cytokine levels produced by splenocytes were detected by cytometry.ResultsWe found that N-terminal, central regions, and PmMSP119 are strongly immunogenic in mice. After three doses, the induced immune responses remained high for 70 days. While antibodies induced after immunization with N-terminal and central regions showed similar affinities to the target antigens, affinities of IgG against PmMSP119 were higher. All proteins induced similar antibody subclass patterns (predominantly IgG1, IgG2a, and IgG2b), characterizing a mixed Th1/Th2 response. Further, autologous stimulation of splenocytes from immunized mice led to the secretion of IL2 and IL4, independently of the antigen used. Importantly, IgG from P. malariae-exposed individuals reacted against PmMSP1 recombinant proteins with a high specificity. On the other hand, sera from P. vivax or P. falciparum-infected individuals did not react at all against recombinant PmMSP1 proteins.ConclusionRecombinant PmMSP1 proteins are very useful diagnostic markers of P. malariae in epidemiological studies or in the differential diagnosis of malaria caused by this species. Immunization with recombinant PmMSP1 proteins resulted in a significant humoral immune response, which may turn them potential component candidates for a vaccine against P. malariae.

Subclinical Malaria and Reticulocyte Count in Apparently Healthy Female Undergraduate Students in Rivers State University, Port Harcourt, Nigeria

Aims: The study was aimed at determining subclinical malaria and estimating reticulocyte count in apparently healthy female undergraduate students of Rivers State University, Port Harcourt.&#x0D; Study Design: This is a non-randomized, comparative case-control study.&#x0D; Place and Duration of Study: The study was conducted using female students residing at the hostels of Rivers State University, Port Harcourt. Analysis was carried out at the Haematology Laboratory, Department of Medical Laboratory Science, Rivers State University, Port Harcourt, Nigeria, between July and August, 2018.&#x0D; Methodology: For the subjects used in this study, a total of 32 students (32%) that were diagnosed of having Plasmodium falciparum malaria infection were used as test subjects, while a total of 68 students (68%) that were diagnosed to be Plasmodium falciparum negative, and without malaria, were used as control. Thick and thin blood films examination using Giemsa staining technique was used to detect and calculate the malaria parasite density while a thin blood film examination using new methylene blue staining technique was used to evaluate the reticulocyte count in the blood.&#x0D; Results: The reticulocyte count of test subjects (subjects with Plasmodium falciparum malaria) was 0.15 ± 0.04% and that of control subjects (subjects without any malaria parasite) was 0.31 ± 0.08%. The test subjects had significantly lower reticulocyte count (p ˂ 0.0001) than the control subjects. The age range “15-19” years had the highest malaria parasite density of 0.52 ± 0.18%, while “25-29” years had the least parasite density of 0.33 ± 0.24. There was no statistical variation in malaria parasite density according to age ranges (p = 0.13; p ˃ 0.05). However, the age range of “15-19” years had the lowest reticulocyte count as most of the female students within this age group were diagnosed to have been infected with malaria parasite.&#x0D; Conclusion: This study revealed that reticulocyte counts of malaria (Plasmodium falciparum) infected individuals decreased when compared to those without malaria parasite and this decrease was statistically significant. There was no statistical significant variation in malaria parasite density irrespective of age ranges. Prophylaxis for malaria in such settings would be an efficient means of preventing infectious reservoirs and higher rates of subclinical malaria infection.

The Search for Plasmodium falciparum histidine-rich protein 2/3 Deletions in Zambia and Implications for Plasmodium falciparum histidine-rich protein 2-Based Rapid Diagnostic Tests.

We attempted to identify Plasmodium falciparum histidine-rich protein 2/3 (pfhrp2/3) deletions among rapid diagnostic test (RDT)-negative but PCR- or microscopy-positive P. falciparum-infected individuals in areas of low transmission (Choma District, 2009-2011) and high transmission (Nchelenge District, 2015-2017) in Zambia. Through community-based surveys, 5,167 participants were screened at 1,147 households by P. falciparum histidine-rich protein 2 (PfHRP2)-based RDTs. Slides were made and dried blood spots were obtained for molecular analysis. Of 28 samples with detectable P. falciparum DNA, none from Nchelenge District were pfhrp2/3 negative. All eight samples from Choma District had detectable pfhrp3 genes, but pfhrp2 was undetectable in three. DNA concentrations of pfhrp2-negative samples were low (< 0.001 ng/μL). These findings suggest that PfHRP2-based RDTs remain effective tools for malaria diagnosis in Nchelenge District, but further study is warranted to understand the potential for pfhrp2/3 deletions in southern Zambia where malaria transmission declined over the past decade.

Heterogeneity in the acquisition of naturally acquired antibodies to cell-traversal protein for ookinetes and sporozoites (CelTOS) and thrombospondin-related adhesion protein (TRAP) of Plasmodium falciparum in naturally infected patients from unstable malaria areas in Iran

Currently, there is no subunit malaria vaccine capable of providing long-lasting protection, and a vaccine based on a single-antigen has shown moderate to unsatisfactory efficacies in clinical trials. As in malaria elimination and eradication strategies, the primary objective is reduction in disease and death due to P. falciparum, in the present investigation, for the first time, we attempted to determine and compare the naturally acquired immune responses to two well-recognized sporozoite antigens, cell-traversal protein for ookinetes and sporozoites (CelTOS) and thrombospondin-related adhesion protein (TRAP), in P. falciparum-infected individuals (n = 204) in low malaria transmission settings of Iran using ELISA. Besides, the profile of IgG isotype responses, the avidity of IgG, IgG1, and IgG3, and the association of anti-PfCelTOS and -PfTRAP antibodies with host age were evaluated. Positive antibody responses to PfCelTOS and PfTRAP antigens were detected in 16.2% and 31.9% of Iranian P. falciparum-infected individuals, respectively, indicating significantly lower immune response to PfCelTOS than PfTRAP (P <0.0001, McNemar's test). Also, among the positive samples for anti-PfCelTOS (n = 33) and -PfTRAP (n = 65) total IgG, the cytophilic IgG1 and IgG3 antibodies were predominant. A significant proportion of the examined positive responders had high- and intermediate-avidity for IgG (93.9%, 87.7%), IgG1 (96.3%, 87.7%), and IgG3 (76%, 78.7%) antibodies to both PfCelTOS and PfTRAP antigens, respectively, with no correlation with age (P >0.05; Spearman’s correlation test). In conclusion, the present data suggests the acquisition of heterogenic immune responses to both antigens in the same patients naturally infected with P. falciparum from settings of low malaria transmission intensity in Iran in which their role in protection to malaria needs further study.