Genetic Diversity and Natural Selection of Plasmodium vivax Merozoite Surface Protein 8 in Global Populations.

Plasmodium vivax Merozoite Surface Protein 8 (PvMSP8) is a promising candidate target for the development of multi-component vaccines. Therefore, determining the genetic variation pattern of Pvmsp8 is essential in providing a reference for the rational design of the P. vivax malaria vaccines. This study delves into the genetic characteristics of the Pvmsp8 gene, specifically focusing on samples from the China-Myanmar border (CMB) region, and contrasts these findings with broader global patterns. The study uncovers that Pvmsp8 exhibits a notable level of conservation across different populations, with limited polymorphisms and relatively low nucleotide diversity (0.00023-0.00120). This conservation contrasts starkly with the high polymorphisms found in other P. vivax antigens such as Pvmsp1. A total of 25 haplotypes and 14 amino acid mutation sites were identified in the global populations, and all mutation sites were confined to non-functional regions. The study also notes that most CMB Pvmsp8 haplotypes are shared among Burmese, Cambodian, Thai, and Vietnamese populations, indicating less geographical variance, but differ notably from those found in Pacific island regions or the Panama. The findings underscore the importance of considering regional genetic diversity in P. vivax when developing targeted malaria vaccines. Non departure from neutral evolution were found by Tajima's D test, however, statistically significant differences were observed between the kn/ks rates. The study's findings are crucial in understanding the evolution and population structure of the Pvmsp8 gene, particularly during regional malaria elimination efforts. The highly conserved nature of Pvmsp8, combined with the lack of mutations in its functional domain, presents it as a promising candidate for developing a broad and effective P. vivax vaccine. This research thus lays a foundation for the rational development of multivalent malaria vaccines targeting this genetically stable antigen.

Global One Health index for zoonoses: A performance assessment in 160 countries and territories.

The One Health (OH) approach is used to control/prevent zoonotic events. However, there is a lack of tools for systematically assessing OH practices. Here, we applied the Global OH Index (GOHI) to evaluate the global OH performance for zoonoses (GOHI-Zoonoses). The fuzzy analytic hierarchy process algorithm and fuzzy comparison matrix were used to calculate the weights and scores of five key indicators, 16 subindicators, and 31 datasets for 160 countries and territories worldwide. The distribution of GOHI-Zoonoses scores varies significantly across countries and regions, reflecting the strengths and weaknesses in controlling or responding to zoonotic threats. Correlation analyses revealed that the GOHI-Zoonoses score was associated with economic, sociodemographic, environmental, climatic, and zoological factors. Additionally, the Human Development Index had a positive effect on the score. This study provides an evidence-based reference and guidance for global, regional, and country-level efforts to optimize the health of people, animals, and the environment.

Towards an actionable One Health approach.

BACKGROUND: Despite the increasing focus on strengthening One Health capacity building on global level, challenges remain in devising and implementing real-world interventions particularly in the Asia-Pacific region. Recognizing these gaps, the One Health Action Commission (OHAC) was established as an academic community for One Health action with an emphasis on research agenda setting to identify actions for highest impact. MAIN TEXT: This viewpoint describes the agenda of, and motivation for, the recently formed OHAC. Recognizing the urgent need for evidence to support the formulation of necessary action plans, OHAC advocates the adoption of both bottom-up and top-down approaches to identify the current gaps in combating zoonoses, antimicrobial resistance, addressing food safety, and to enhance capacity building for context-sensitive One Health implementation. CONCLUSIONS: By promoting broader engagement and connection of multidisciplinary stakeholders, OHAC envisions a collaborative global platform for the generation of innovative One Health knowledge, distilled practical experience and actionable policy advice, guided by strong ethical principles of One Health.

Revisiting the antigen markers of vector-borne parasitic diseases identified by immunomics: identification and application to disease control.

INTRODUCTION: Protein microarray is a promising immunomic approach for identifying biomarkers. Based on our previous study that reviewed parasite antigens and recent parasitic omics research, this article expands to include information on vector-borne parasitic diseases (VBPDs), namely, malaria, schistosomiasis, leishmaniasis, babesiosis, trypanosomiasis, lymphatic filariasis, and onchocerciasis. AREAS COVERED: We revisit and systematically summarize antigen markers of vector-borne parasites identified by the immunomic approach and discuss the latest advances in identifying antigens for the rational development of diagnostics and vaccines. The applications and challenges of this approach for VBPD control are also discussed. EXPERT OPINION: The immunomic approach has enabled the identification and/or validation of antigen markers for vaccine development, diagnosis, disease surveillance, and treatment. However, this approach presents several challenges, including limited sample size, variability in antigen expression, false-positive results, complexity of omics data, validation and reproducibility, and heterogeneity of diseases. In addition, antigen involvement in host immune evasion and antigen sensitivity/specificity are major issues in its application. Despite these limitations, this approach remains promising for controlling VBPD. Advances in technology and data analysis methods should continue to improve candidate antigen identification, as well as the use of a multiantigen approach in diagnostic and vaccine development for VBPD control.

PRL2 regulates neutrophil extracellular trap formation which contributes to severe malaria and acute lung injury.

Excessive host immune responses contribute to severe malaria with high mortality. Here, we show that PRL2 in innate immune cells is highly related to experimental malaria disease progression, especially the development of murine severe malaria. In the absence of PRL2 in myeloid cells, Plasmodium berghei infection results in augmented lung injury, leading to significantly increased mortality. Intravital imaging revealed greater neutrophilic inflammation and NET formation in the lungs of PRL2 myeloid conditional knockout mice. Depletion of neutrophils prior to the onset of severe disease protected mice from NETs associated lung injury, and eliminated the difference between WT and PRL2 CKO mice. PRL2 regulates neutrophil activation and NET accumulation via the Rac-ROS pathway, thus contributing to NETs associated ALI. Hydroxychloroquine, an inhibitor of PRL2 degradation alleviates NETs associated tissue damage in vivo. Our findings suggest that PRL2 serves as an indicator of progression to severe malaria and ALI. In addition, our study indicated the importance of PRL2 in NET formation and tissue injury. It might open a promising path for adjunctive treatment of NET-associated disease.

Polymorphism of Drug Resistance Genes dhfr and dhps in Plasmodium falciparum Isolates among Chinese Migrant Workers Who Returned from Ghana in 2013.

In 2013, an epidemic of falciparum malaria involving over 820 persons unexpectedly broke out in Shanglin County, Guangxi Zhuang Autonomous Region, China, after a large number of migrant workers returned from Ghana, where they worked as gold miners. Herein, we selected 146 isolates randomly collected from these patients to investigate the resistance characteristics of the parasite to sulfadoxine-pyrimethamine (SP) by screening mutations in the dhfr and dhps genes. All 146 isolates were successfully genotyped for dhps, and only 137 samples were successfully genotyped for dhfr. In the dhfr gene, point mutations occurred at three codons: 51 (83.2%, 114/137), 59 (94.9%, 130/137), and 108 (96.4%, 132/137). In the dhps gene, mutations occurred at four codons: 436 (36.3%, 53/146 for S436A, 0.7%, 1/146 for S436Y), 437 (95.2%, 139/146), 540 (3.4%, 5/146), and 613 (2.7%, 4/146). All 146 isolates had mutations in at least one codon, either within dhfr or dhps. Quadruple mutation I51R59N108/G437 (41.1%, 60/146) of partial or low resistance level was the most prevalent haplotype combination. Quintuple I51R59N108/G437E540 accounted for 2.1% (3/146). Sextuple I51R59N108/A436G437S613 was also found and accounted for 1.4% (2/146). A chronological assay incorporating two sets of resistance data from the studies of Duah and Amenga-Etego provided an overview of the resistance trend from 2003 to 2018. During this period, the results we obtained generally coincided with the total development tendency of SP resistance. It can be concluded that Plasmodium falciparum samples collected from Chinese migrant workers from Ghana presented prevalent but relatively partial or low resistance to SP. A chronological assay incorporating two sets of data around 2013 indicates that our results possibly reflect the SP resistance level of Ghana in 2013 and that the possibility of increased resistance exists. Therefore, reasonable drug use and management should be strengthened while also maintaining a continuous screening of resistance to SP. These findings also underscore the need to strengthen the prevention of malaria importation from overseas and focus on preventing its reintroduction and transmission in China.

Molecular Identification and Genetic Characterization of Public Health Threatening Ticks - Chongming Island, China, 2021-2022.

What is already known about this topic?: Although ticks and tick-borne diseases are prevalent throughout China, there remains a knowledge gap regarding their biology and potential risk of distribution to human and animal populations on Chongming Island. The island, being China's third largest and a crucial component in the ecological preservation of the Yangtze Delta region, has yet to be comprehensively studied in this context. What is added by this report?: In this study, employing molecular methodologies, a significant prevalence of Haemaphysalis (H.) longicornis and H. flava ticks - widely recognized for their high pathogenicity - is reported from Chongming Island. Additionally, the identification of two previously unreported species on the island, namely, H. doenitzi and H. japonica, expands our understanding of both the range and evolution of tick species. What are the implications for public health practice?: The populations of humans and animals in nearly all 18 towns on Chongming Island are potentially at risk for transmission of tick-borne infectious agents. As a result, there is a pressing necessity for public health alerts, proactive tick surveillance, and effective screening of suspected clinical cases of tick-borne diseases within the Chongming population.

Tackling global health security by building an academic community for One Health action.

BACKGROUND: One Health approach is crucial to tackling complex global public health threats at the interface of humans, animals, and the environment. As outlined in the One Health Joint Plan of Action, the international One Health community includes stakeholders from different sectors. Supported by the Bill & Melinda Gates Foundation, an academic community for One Health action has been proposed with the aim of promoting the understanding and real-world implementation of One Health approach and contribution towards the Sustainable Development Goals for a healthy planet. MAIN TEXT: The proposed academic community would contribute to generating high-quality scientific evidence, distilling local experiences as well as fostering an interconnected One Health culture and mindset, among various stakeholders on different levels and in all sectors. The major scope of the community covers One Health governance, zoonotic diseases, food security, antimicrobial resistance, and climate change along with the research agenda to be developed. The academic community will be supported by two committees, including a strategic consultancy committee and a scientific steering committee, composed of influential scientists selected from the One Health information database. A workplan containing activities under six objectives is proposed to provide research support, strengthen local capacity, and enhance global participation. CONCLUSIONS: The proposed academic community for One Health action is a crucial step towards enhancing communication, coordination, collaboration, and capacity building for the implementation of One Health. By bringing eminent global experts together, the academic community possesses the potential to generate scientific evidence and provide advice to local governments and international organizations, enabling the pursuit of common goals, collaborative policies, and solutions to misaligned interests.

China's application of the One Health approach in addressing public health threats at the human-animal-environment interface: Advances and challenges.

Background: Due to emerging issues such as global climate change and zoonotic disease pandemics, the One Health approach has gained more attention since the turn of the 21st century. Although One Health thinking has deep roots and early applications in Chinese history, significant gaps exist in China's real-world implementation at the complex interface of the human-animal-environment. Methods: We abstracted the data from the global One Health index study and analysed China's performance in selected fields based on Structure-Process-Outcome model. By comparing China to the Belt & Road and G20 countries, the advances and gaps in China's One Health performance were determined and analysed. Findings: For the selected scientific fields, China generally performs better in ensuring food security and controlling antimicrobial resistance and worse in addressing climate change. Based on the SPO model, the "structure" indicators have the highest proportion (80.00%) of high ranking and the "outcome" indicators have the highest proportion (20.00%) of low ranking. When compared with Belt and Road countries, China scores above the median in almost all indicators (16 out of 18) under the selected scientific fields. When compared with G20 countries, China ranks highest in food security (scores 72.56 and ranks 6th), and lowest in climate change (48.74, 11th). Conclusion: Our results indicate that while China has made significant efforts to enhance the application of the One Health approach in national policies, it still faces challenges in translating policies into practical measures. It is recommended that a holistic One Health action framework be established for China in accordance with diverse social and cultural contexts, with a particular emphasis on overcoming data barriers and mobilizing stakeholders both domestically and globally. Implementation mechanisms, with clarified stakeholder responsibilities and incentives, should be improved along with top-level design.

Increased interleukin-6 is associated with long COVID-19: a systematic review and meta-analysis.

BACKGROUND: Coronavirus disease 2019 (COVID-19) can involve persistence, sequelae, and other clinical complications that last weeks to months to evolve into long COVID-19. Exploratory studies have suggested that interleukin-6 (IL-6) is related to COVID-19; however, the correlation between IL-6 and long COVID-19 is unknown. We designed a systematic review and meta-analysis to assess the relationship between IL-6 levels and long COVID-19. METHODS: Databases were systematically searched for articles with data on long COVID-19 and IL-6 levels published before September 2022. A total of 22 published studies were eligible for inclusion following the PRISMA guidelines. Analysis of data was undertaken by using Cochran's Q test and the Higgins I-squared (I2) statistic for heterogeneity. Random-effect meta-analyses were conducted to pool the IL-6 levels of long COVID-19 patients and to compare the differences in IL-6 levels among the long COVID-19, healthy, non-postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (non-PASC), and acute COVID-19 populations. The funnel plot and Egger's test were used to assess potential publication bias. Sensitivity analysis was used to test the stability of the results. RESULTS: An increase in IL-6 levels was observed after SARS-CoV-2 infection. The pooled estimate of IL-6 revealed a mean value of 20.92 pg/ml (95% CI = 9.30-32.54 pg/ml, I2 = 100%, P < 0.01) for long COVID-19 patients. The forest plot showed high levels of IL-6 for long COVID-19 compared with healthy controls (mean difference = 9.75 pg/ml, 95% CI = 5.75-13.75 pg/ml, I2 = 100%, P < 0.00001) and PASC category (mean difference = 3.32 pg/ml, 95% CI = 0.22-6.42 pg/ml, I2 = 88%, P = 0.04). The symmetry of the funnel plots was not obvious, and Egger's test showed that there was no significant small study effect in all groups. CONCLUSIONS: This study showed that increased IL-6 correlates with long COVID-19. Such an informative revelation suggests IL-6 as a basic determinant to predict long COVID-19 or at least inform on the "early stage" of long COVID-19.

Immunogenicity and antigenicity of a conserved fragment of the rhoptry-associated membrane antigen of Plasmodium vivax.

BACKGROUND: Plasmodium vivax rhoptry-associated membrane antigen (RAMA) is a glycophosphatidylinositol-anchored membrane protein currently under consideration as a malaria vaccine candidate. Immunoglobulin G (IgG) antibodies induced by P. vivax RAMA (PvRAMA) have been proved to persist over 12 months in the sera of people infected with P. vivax. It has also been shown that through stimulation of peripheral blood mononuclear cells with PvRAMA in vitro, the antigen can induce CD4+ T cells to produce interleukin-10. However, the genetic diversity of the RAMA gene in isolates of P. vivax (pvrama) and the immunogenicity of PvRAMA in animals remain unclear. METHODS: Genomic DNA was extracted from blood samples (n = 25) of patients in Jiangsu Province, China with imported infections of P. vivax from endemic countries in South and Southeast Asia. The extract genomic DNA was used as templates to amplify the P. vivax rama gene (pvrama) by PCR, and the PCR products were then sequenced and analyzed by the DnaSP, MEGA, and GeneDoc software packages. Recombinant PvRAMA (rPvRAMA) protein was expressed and purified, and then used to immunize mice. Levels of total IgG and different IgG subclasses of rPvRAMA-immunized mice were evaluated by enzyme-linked immunosorbent assay. Also, spleen cells of rPvRAMA-immunized mice were stimulated with rPvRAMA in vitro and levels of T cells were measured by flow cytometry. RESULTS: The average pairwise nucleotide diversity (π) of the pvrama gene was 0.00190, and the haplotype diversity (Hd) was 0.982. The C-terminal of PvRAMA showed lower haplotype diversity compared to the N-terminal and was completely conserved at amino acid sites related to erythrocyte binding. To further characterize immunogenicity of the C-terminal of PvRAMA, mice were immunized with rPvRAMA antigen. The rPvRAMA protein induced antibody responses, with the end-point titer ranging from 1:10,000 to 1:5,120,000. IgG1 was the predominant IgG subclass in rPvRAMA-immunized mice, followed by IgG2b. In addition, levels of CD4+ and CD8+ T cells in the rPvRAMA-stimulated group were significantly higher than those in the phosphate-buffered saline-stimulated group (normal control group). CONCLUSIONS: The high conservation at specific amino acid sites and the high immunogenicity of the C-terminal of PvRAMA indicate the presence of conserved epitopes able to generate broadly reactive humoral and cellular immune responses. These findings support the potential of PvRAMA to serve as a vaccine candidate against P. vivax infection.

Assessment of performance for a key indicator of One Health: evidence based on One Health index for zoonoses in Sub-Saharan Africa.

BACKGROUND: Zoonoses are public health threats that cause severe damage worldwide. Zoonoses constitute a key indicator of One Health (OH) and the OH approach is being applied for zoonosis control programmes of zoonotic diseases. In a very recent study, we developed an evaluation system for OH performance through the global OH index (GOHI). This study applied the GOHI to evaluate OH performance for zoonoses in sub-Saharan Africa. METHODS: The framework for the OH index on zoonoses (OHIZ) was constructed including five indicators, 15 subindicators and 28 datasets. Publicly available data were referenced to generate the OHIZ database which included both qualitative and quantitative indicators for all sub-Sahara African countries (n = 48). The GOHI algorithm was used to estimate scores for OHIZ. Indicator weights were calculated by adopting the fuzzy analytical hierarchy process. RESULTS: Overall, five indicators associated with weights were generated as follows: source of infection (23.70%), route of transmission (25.31%), targeted population (19.09%), capacity building (16.77%), and outcomes/case studies (15.13%). Following the indicators, a total of 37 sub-Sahara African countries aligned with OHIZ validation, while 11 territories were excluded for unfit or missing data. The OHIZ average score of sub-Saharan Africa was estimated at 53.67/100. The highest score was 71.99 from South Africa, while the lowest score was 40.51 from Benin. It is also worth mentioning that Sub-Sahara African countries had high performance in many subindicators associated with zoonoses, e.g., surveillance and response, vector and reservoir interventions, and natural protected areas, which suggests that this region had a certain capacity in control and prevention or responses to zoonotic events. CONCLUSIONS: This study reveals that it is possible to perform OH evaluation for zoonoses in sub-Saharan Africa by OHIZ. Findings from this study provide preliminary research information in advancing knowledge of the evidenced risks to strengthen strategies for effective control of zoonoses and to support the prevention of zoonotic events.

Towards a global One Health index: a potential assessment tool for One Health performance.

BACKGROUND: A One Health approach has been increasingly mainstreamed by the international community, as it provides for holistic thinking in recognizing the close links and inter-dependence of the health of humans, animals and the environment. However, the dearth of real-world evidence has hampered application of a One Health approach in shaping policies and practice. This study proposes the development of a potential evaluation tool for One Health performance, in order to contribute to the scientific measurement of One Health approach and the identification of gaps where One Health capacity building is most urgently needed. METHODS: We describe five steps towards a global One Health index (GOHI), including (i) framework formulation; (ii) indicator selection; (iii) database building; (iv) weight determination; and (v) GOHI scores calculation. A cell-like framework for GOHI is proposed, which comprises an external drivers index (EDI), an intrinsic drivers index (IDI) and a core drivers index (CDI). We construct the indicator scheme for GOHI based on this framework after multiple rounds of panel discussions with our expert advisory committee. A fuzzy analytical hierarchy process is adopted to determine the weights for each of the indicators. RESULTS: The weighted indicator scheme of GOHI comprises three first-level indicators, 13 second-level indicators, and 57 third-level indicators. According to the pilot analysis based on the data from more than 200 countries/territories the GOHI scores overall are far from ideal (the highest score of 65.0 out of a maximum score of 100), and we found considerable variations among different countries/territories (31.8-65.0). The results from the pilot analysis are consistent with the results from a literature review, which suggests that a GOHI as a potential tool for the assessment of One Health performance might be feasible. CONCLUSIONS: GOHI-subject to rigorous validation-would represent the world's first evaluation tool that constructs the conceptual framework from a holistic perspective of One Health. Future application of GOHI might promote a common understanding of a strong One Health approach and provide reference for promoting effective measures to strengthen One Health capacity building. With further adaptations under various scenarios, GOHI, along with its technical protocols and databases, will be updated regularly to address current technical limitations, and capture new knowledge.

Could China's journey of malaria elimination extend to Africa?

World Health Organization (WHO) certified China malaria-free on June 30, 2021, which brightens the goal of global malaria elimination efforts. China contributed its unique innovations to the global community: Artemisinin, discovered by Tu Youyou, has saved millions of lives globally; the "1-3-7" norm developed in 2012, has been adapted in the local contexts of countries in the Southeast Asia and Africa. How to the targets of Global Technical Strategy for Malaria (GTS) 2016-2030. By looking into the malaria control phase, towards elimination phase from 1960 to 2011 in sub-Saharan Africa and China, we found that the gap in malaria burden will widen unless the interventions in Africa are enhanced. It is imperative to identify the key China-Africa cooperation areas on malaria control and elimination, so that synergized efforts could be pooled together to help African countries achieve the elimination goal. The practices from China malaria control and elimination efforts could be leveraged to fast-track malaria elimination efforts in Africa, which makes it possible that the China's journey of malaria elimination extends to Africa.

Identification of the Recombinant Plasmodium vivax Surface-Related Antigen as a Possible Immune Evasion Factor Against Human Splenic Fibroblasts by Targeting ITGB1.

Plasmodium vivax-infected erythrocytes can enter the spleen and evade spleen clearance to establish chronic infections. However, the mechanism underlying P. vivax immune evasion in the spleen is still unclear. Human splenic fibroblasts (HSF), also known as barrier cells, play an essential role in the immune function of spleen. A hypothesis holds that P. vivax-infected erythrocytes induce spleen structural remodeling to form barrier cells. Subsequently, these infected erythrocytes can selectively cytoadhere to these barrier cells to escape spleen clearance. In this work, we found that P. vivax surface-related antigen (PvSRA; PlasmoDB ID: PVX_084970), an exported protein on infected erythrocyte membrane, could bind with HSF. Considering the above hypothesis, we speculated that PvSRA might be involved in P. vivax immune evasion by changing HSF cell performance. To investigate this speculation, RNA sequencing, protein microarray, and bioinformatics analysis technologies were applied, and in vitro validations were further performed. The results showed that the recombinant PvSRA attracted HSF migration and interacted with HSF by targeting integrin ß1 (ITGB1) along with changes in HSF cell performance, such as focal adhesion, extracellular matrix, actin cytoskeleton, and cell cycle. This study indicated that PvSRA might indeed participate in the immune evasion of P. vivax in the spleen by changing HSF function through PvSRA-ITGB1 axis.

A Molecular Investigation of Malaria Infections From High-Transmission Areas of Southern Togo Reveals Different Species of Plasmodium Parasites.

Malaria particularly burdens people in poor and neglected settings across the tropics of Africa. Meanwhile, a large proportion of the Togo population have poor understanding of malaria epidemiology and parasites. This study carried out a molecular survey of malaria cases in southern Togo during 2017-2019. We estimated Plasmodium species infection rates and microscopic examination compliance with nested PCR results. Sensitivity and specificity analyses were performed in conjunction with predictive values. Also, phylogenetic characterization of species of malaria parasites was assessed. Plasmodium genus-specific nested PCR identified 565 positive cases including 536/611 (87.8%) confirmed cases from the microscopy-positive group and 29/199 (14.6%) diagnosed malaria cases from the microscopy-negative group. Our findings revealed a disease prevalence (69.8%) higher than that reported (25.5-55.1%) for the country. The diagnostic test had 94.9% sensitivity and 69.4% specificity, i.e., it missed 120 of the people who had malaria and about one-third of the people tested positive for the disease, which they did not have, respectively. In conjunction, the test showed 87.7% positive predictive value and 85.4% negative predictive value, which, from a clinical perspective, indicates the chance that a person with a positive diagnostic test truly has the disease and the probability that a person with a negative test does not have the disease, respectively. Further species-specific nested PCR followed by analysis of gene sequences confirmed species of malaria parasites and indicated infection rates for Plasmodium falciparum (Pf), 95.5% (540/565); P. ovale (Po), 0.5% (3/565); and P. malariae (Pm), 0.4% (2/565). In addition, 20 cases were coinfection cases of Pf-Po (15/565) and Pf-Pm (5/565). This study publicly reports, for the first time, a molecular survey of malaria cases in Togo and reveals the presence of other malaria parasites (Po and Pm) other than Pf. These findings might provide answers to some basic questions on the malaria scenario and, knowledge gained could help with intervention deployment for effective malaria control in Togo.

Genetic Diversity and Natural Selection of Plasmodium vivax Duffy Binding Protein-II From China-Myanmar Border of Yunnan Province, China.

Malaria incidence has declined dramatically over the past decade and China was certified malaria-free in 2021. However, the presence of malaria in border areas and the importation of cases of malaria parasites are major challenges for the consolidation of the achievements made by China. Plasmodium vivax Duffy binding protein (PvDBP) performs a significant role in erythrocyte invasion, and is considered a promising P. vivax vaccine. However, the highly polymorphic region of PvDBP (PvDBP-II) impedes the development of blood-stage vaccine against P. vivax. In this study, we investigated the genetic diversity and natural selection of PvDBP-II among 124 P. vivax isolates collected from the China-Myanmar border (CMB) in Yunnan Province, China, during 2009-2011. To compare genetic diversity, natural selection, and population structure with CMB isolates, 85 pvdbp-II sequences of eastern Myanmar isolates were obtained from GenBank. In addition, global sequences of pvdbp-II were retrieved from GenBank to establish genetic differentiation relationships and networks with the CMB isolates. In total, 22 single nucleotide polymorphisms reflected in 20 non-synonymous and two synonymous mutations were identified. The overall nucleotide diversity of PvDBP-II from the 124 CMB isolates was 0.0059 with 21 haplotypes identified (Hd = 0.91). The high ratio of non-synonymous to synonymous mutations suggests that PvDBP-II had evolved under positive selection. Population structure analysis of the CMB and eastern Myanmar isolates were optimally grouped into five sub-populations (K = 5). Polymorphisms of PvDBP-II display that CMB isolates were genetically diverse. Mutation, recombination, and positive selection promote polymorphism of PvDBP-II of P. vivax population. Although low-level genetic differentiation in eastern Myanmar was identified along with the more effective malaria control measures, the complexity of population structure in malaria parasites has maintained. In conclusion, findings from this study advance knowledge of the understanding of the dynamic of P. vivax population, which will contribute to guiding the rational design of a PvDBP-II based vaccine.

Genetic diversity and immunogenicity of the merozoite surface protein 1 C-terminal 19-kDa fragment of Plasmodium ovale imported from Africa into China.

BACKGROUND: Merozoite surface protein 1 (MSP1) plays an essential role in erythrocyte invasion by malaria parasites. The C-terminal 19-kDa region of MSP1 has long been considered one of the major candidate antigens for a malaria blood-stage vaccine against Plasmodium falciparum. However, there is limited information on the C-terminal 19-kDa region of Plasmodium ovale MSP1 (PoMSP119). This study aims to analyze the genetic diversity and immunogenicity of PoMSP119. METHODS: A total of 37 clinical Plasmodium ovale isolates including Plasmodium ovale curtisi and Plasmodium ovale wallikeri imported from Africa into China and collected during the period 2012-2016 were used. Genomic DNA was used to amplify P. ovale curtisi (poc) msp119 (pocmsp119) and P. ovale wallikeri (pow) msp119 (powmsp119) genes by polymerase chain reaction. The genetic diversity of pomsp119 was analyzed using the GeneDoc version 6 programs. Recombinant PoMSP119 (rPoMSP119)-glutathione S-transferase (GST) proteins were expressed in an Escherichia coli expression system and analyzed by western blot. Immune responses in BALB/c mice immunized with rPoMSP119-GST were determined using enzyme-linked immunosorbent assay. In addition, antigen-specific T cell responses were assessed by lymphocyte proliferation assays. A total of 49 serum samples from healthy individuals and individuals infected with P. ovale were used for the evaluation of natural immune responses by using protein microarrays. RESULTS: Sequences of pomsp119 were found to be thoroughly conserved in all the clinical isolates. rPoMSP119 proteins were efficiently expressed and purified as ~ 37-kDa proteins. High antibody responses in mice immunized with rPoMSP119-GST were observed. rPoMSP119-GST induced high avidity indexes, with an average of 92.57% and 85.32% for rPocMSP119 and rPowMSP119, respectively. Cross-reactivity between rPocMSP119 and rPowMSP119 was observed. Cellular immune responses to rPocMSP119 (69.51%) and rPowMSP119 (52.17%) induced in rPocMSP119- and rPowMSP119-immunized mice were found in the splenocyte proliferation assays. The sensitivity and specificity of rPoMSP119-GST proteins for the detection of natural immune responses in patients infected with P. ovale were 89.96% and 75%, respectively. CONCLUSIONS: This study revealed highly conserved gene sequences of pomsp119. In addition, naturally acquired humoral immune responses against rPoMSP1 were observed in P. ovale infections, and high immunogenicity of rPoMSP119 in mice was also identified. These instructive findings should encourage further testing of PoMSP119 for rational vaccine design.