-
Open Forum Infectious Diseases Nov 2023Scarcity of annotated image data sets of thin blood smears makes expert-level differentiation among species challenging. Here, we aimed to establish a deep learning...
BACKGROUND
Scarcity of annotated image data sets of thin blood smears makes expert-level differentiation among species challenging. Here, we aimed to establish a deep learning algorithm for identifying and classifying malaria parasites in thin blood smears and evaluate its performance and clinical prospect.
METHODS
You Only Look Once v7 was used as the backbone network for training the artificial intelligence algorithm model. The training, validation, and test sets for each malaria parasite category were randomly selected. A comprehensive analysis was performed on 12 708 thin blood smear images of various infective stages of 12 546 malaria parasites, including , , , , , and . Peripheral blood samples were obtained from 380 patients diagnosed with malaria. Additionally, blood samples from monkeys diagnosed with malaria were used to analyze . The accuracy for detecting -infected blood cells was assessed through various evaluation metrics.
RESULTS
The total time to identify 1116 malaria parasites was 13 seconds, with an average analysis time of 0.01 seconds for each parasite in the test set. The average precision was 0.902, with a recall and precision of infected erythrocytes of 96.0% and 94.9%, respectively. Sensitivity and specificity exceeded 96.8% and 99.3%, with an area under the receiver operating characteristic curve >0.999. The highest sensitivity (97.8%) and specificity (99.8%) were observed for trophozoites and merozoites.
CONCLUSIONS
The algorithm can help facilitate the clinical and morphologic examination of malaria parasites.
PubMed: 37937045
DOI: 10.1093/ofid/ofad469 -
Inhibition of malaria and babesiosis parasites by putative red blood cell targeting small molecules.Frontiers in Cellular and Infection... 2024Chemotherapies for malaria and babesiosis frequently succumb to the emergence of pathogen-related drug-resistance. Host-targeted therapies are thought to be less...
BACKGROUND
Chemotherapies for malaria and babesiosis frequently succumb to the emergence of pathogen-related drug-resistance. Host-targeted therapies are thought to be less susceptible to resistance but are seldom considered for treatment of these diseases.
METHODS
Our overall objective was to systematically assess small molecules for host cell-targeting activity to restrict proliferation of intracellular parasites. We carried out a literature survey to identify small molecules annotated for host factors implicated in infection. Alongside , we implemented parasite susceptibility assays also in the zoonotic parasite and the veterinary parasite . We additionally carried out assays to test directly for action on RBCs apart from the parasites. To distinguish specific host-targeting antiparasitic activity from erythrotoxicity, we measured phosphatidylserine exposure and hemolysis stimulated by small molecules in uninfected RBCs.
RESULTS
We identified diverse RBC target-annotated inhibitors with -specific, specific, and broad-spectrum antiparasitic activity. The anticancer MEK-targeting drug trametinib is shown here to act with submicromolar activity to block proliferation of spp. in RBCs. Some inhibitors exhibit antimalarial activity with transient exposure to RBCs prior to infection with parasites, providing evidence for host-targeting activity distinct from direct inhibition of the parasite.
CONCLUSIONS
We report here characterization of small molecules for antiproliferative and host cell-targeting activity for malaria and babesiosis parasites. This resource is relevant for assessment of physiological RBC-parasite interactions and may inform drug development and repurposing efforts.
Topics: Animals; Humans; Parasites; Babesiosis; Malaria; Erythrocytes; Plasmodium; Malaria, Falciparum; Babesia; Antimalarials; Plasmodium falciparum
PubMed: 38572319
DOI: 10.3389/fcimb.2024.1304839 -
Vaccine Jun 2024Recent data indicate increasing disease burden and importance of Plasmodium vivax (Pv) malaria. A robust assay will be essential for blood-stage Pv vaccine development....
Recent data indicate increasing disease burden and importance of Plasmodium vivax (Pv) malaria. A robust assay will be essential for blood-stage Pv vaccine development. Results of the in vitro growth inhibition assay (GIA) with transgenic P. knowlesi (Pk) parasites expressing the Pv Duffy-binding protein region II (PvDBPII) correlate with in vivo protection in the first PvDBPII controlled human malaria infection (CHMI) trials, making the PkGIA an ideal selection tool once the precision of the assay is defined. To determine the precision in percentage of inhibition in GIA (%GIA) and in GIA (antibody concentration that gave 50 %GIA), ten GIAs with transgenic Pk parasites were conducted with four different anti-PvDBPII human monoclonal antibodies (mAbs) at concentrations of 0.016 to 2 mg/mL, and three GIAs with eighty anti-PvDBPII human polyclonal antibodies (pAbs) at 10 mg/mL. A significant assay-to-assay variation was observed, and the analysis revealed a standard deviation (SD) of 13.1 in the mAb and 5.94 in the pAb dataset for %GIA, with a LogGIA SD of 0.299 (for mAbs). Moreover, the ninety-five percent confidence interval (95 %CI) for %GIA or GIA in repeat assays was calculated in this investigation. The error range determined in this study will help researchers to compare PkGIA results from different assays and studies appropriately, thus supporting the development of future blood-stage malaria vaccine candidates, specifically second-generation PvDBPII-based formulations.
Topics: Malaria Vaccines; Plasmodium knowlesi; Protozoan Proteins; Plasmodium vivax; Antigens, Protozoan; Humans; Receptors, Cell Surface; Antibodies, Protozoan; Malaria, Vivax; Antibodies, Monoclonal; Vaccine Development; Animals
PubMed: 38704253
DOI: 10.1016/j.vaccine.2024.04.073 -
Frontiers in Immunology 2024Human malaria, caused by five Plasmodium species (, and ), remains a significant global health burden. While most interventions target , the species associated with high...
Identification of conserved cross-species B-cell linear epitopes in human malaria: a subtractive proteomics and immuno-informatics approach targeting merozoite stage proteins.
Human malaria, caused by five Plasmodium species (, and ), remains a significant global health burden. While most interventions target , the species associated with high mortality rates and severe clinical symptoms, non-falciparum species exhibit different transmission dynamics, remain hugely neglected, and pose a significant challenge to malaria elimination efforts. Recent studies have reported the presence of antigens associated with cross-protective immunity, which can potentially disrupt the transmission of various Plasmodium species. With the sequencing of the Plasmodium genome and the development of immunoinformatic tools, in this study, we sought to exploit the evolutionary history of Plasmodium species to identify conserved cross-species B-cell linear epitopes in merozoite proteins. We retrieved Plasmodium proteomes associated with human malaria and applied a subtractive proteomics approach focusing on merozoite stage proteins. Bepipred 2.0 and Epidope were used to predict B-cell linear epitopes using as the reference species. The predictions were further compared against human and non-falciparum databases and their antigenicity, toxicity, and allergenicity assessed. Subsequently, epitope conservation was carried out using locally sequenced isolates from a malaria-endemic region in western Kenya (n=27) and Kenyan isolates from MalariaGEN version 6 (n=131). Finally, physiochemical characteristics and tertiary structure of the B-cell linear epitopes were determined. The analysis revealed eight epitopes that showed high similarity (70-100%) between falciparum and non-falciparum species. These epitopes were highly conserved when assessed across local isolates and those from the MalariaGEN database and showed desirable physiochemical properties. Our results show the presence of conserved cross-species B-cell linear epitopes that could aid in targeting multiple Plasmodium species. Nevertheless, validating their efficacy and experimentally is essential.
Topics: Animals; Humans; Merozoites; Epitopes, B-Lymphocyte; Kenya; Proteomics; Plasmodium falciparum; Plasmodium vivax; Malaria; Malaria, Falciparum; Plasmodium; Malaria, Vivax
PubMed: 38404581
DOI: 10.3389/fimmu.2024.1352618 -
Malaria Journal Oct 2023Since 2018, no indigenous human malaria cases has been reported in Malaysia. However, during the recent COVID-19 pandemic the World Health Organization is concerned that...
BACKGROUND
Since 2018, no indigenous human malaria cases has been reported in Malaysia. However, during the recent COVID-19 pandemic the World Health Organization is concerned that the pandemic might erode the success of malaria control as there are reports of increase malaria cases in resource limited countries. Little is known how the COVID-19 pandemic has impacted malaria in middle-income countries like Malaysia. Here the public health response to a Plasmodium malariae outbreak occurred in a village in Sabah state, Malaysia, during a COVID-19 movement control order is reported.
METHODS
An outbreak was declared following the detection of P. malariae in July 2020 and active case detection for malaria was performed by collecting blood samples from residents residing within 2 km radius of Moyog village. Vector prevalence and the efficacy of residual insecticides were determined. Health awareness programmes were implemented to prevent future outbreaks. A survey was conducted among villagers to understand risk behaviour and beliefs concerning malaria.
RESULTS
A total of 5254 blood samples collected from 19 villages. Among them, 19 P. malariae cases were identified, including the index case, which originated from a man who returned from Indonesia. His return from Indonesia and healthcare facilities visit coincided with the movement control order during COVID-19 pandemic when the healthcare facilities stretched its capacity and only serious cases were given priority. Despite the index case being a returnee from a malaria endemic area presenting with mild fever, no malaria test was performed at local healthcare facilities. All cases were symptomatic and uncomplicated except for a pregnant woman with severe malaria. There were no deaths; all patients recovered following treatment with artemether-lumefantrine combination therapy. Anopheles balabacensis and Anopheles barbirostris were detected in ponds, puddles and riverbeds. The survey revealed that fishing and hunting during night, and self-treatment for mild symptoms contributed to the outbreak. Despite the index case being a returnee from a malaria-endemic area presenting with mild fever, no malaria test was performed at local healthcare facilities.
CONCLUSION
The outbreak occurred during a COVID-19 movement control order, which strained healthcare facilities, prioritizing only serious cases. Healthcare workers need to be more aware of the risk of malaria from individuals who return from malaria endemic areas. To achieve malaria elimination and prevention of disease reintroduction, new strategies that include multisectoral agencies and active community participation are essential for a more sustainable malaria control programme.
Topics: Male; Animals; Female; Humans; Malaysia; Plasmodium malariae; Antimalarials; Public Health; Pandemics; Mosquito Vectors; Plasmodium knowlesi; Artemether; Artemether, Lumefantrine Drug Combination; COVID-19; Malaria; Disease Outbreaks; Anopheles
PubMed: 37789320
DOI: 10.1186/s12936-023-04693-1 -
Acta Tropica Dec 2023
PubMed: 37777354
DOI: 10.1016/j.actatropica.2023.107029 -
Royal Society Open Science Jan 2024Disease surveillance aims to collect data at different times or locations, to assist public health authorities to respond appropriately. Surveillance of the simian...
Disease surveillance aims to collect data at different times or locations, to assist public health authorities to respond appropriately. Surveillance of the simian malaria parasite, , is sparse in some endemic areas and the spatial extent of transmission is uncertain. Zoonotic transmission of has been demonstrated throughout Southeast Asia and represents a major hurdle to regional malaria elimination efforts. Given an arbitrary spatial prediction of relative disease risk, we develop a flexible framework for surveillance site selection, drawing on principles from multi-criteria decision-making. To demonstrate the utility of our framework, we apply it to the case study of malaria surveillance site selection in western Indonesia. We demonstrate how statistical predictions of relative disease risk can be quantitatively incorporated into public health decision-making, with specific application to active human surveillance of zoonotic malaria. This approach can be used in other contexts to extend the utility of modelling outputs.
PubMed: 38204787
DOI: 10.1098/rsos.230641 -
Acta Tropica Jul 2024Natural human infections by Plasmodium cynomolgi and P. inui have been reported recently and gain the substantial attention from Southeast Asian countries. Zoonotic...
BACKGROUND
Natural human infections by Plasmodium cynomolgi and P. inui have been reported recently and gain the substantial attention from Southeast Asian countries. Zoonotic transmission of non-human malaria parasites to humans from macaque monkeys occurred through the bites of the infected mosquitoes. The objective of this study is to establish real-time fluorescence loop-mediated isothermal amplification (LAMP) assays for the detection of zoonotic malaria parasites by combining real-time fluorescent technology with the isothermal amplification technique.
METHODS
By using 18S rRNA as the target gene, the primers for P. cynomolgi, P. coatneyi and P. inui were newly designed in the present study. Four novel real-time fluorescence LAMP assays were developed for the detection of P. cynomolgi, P. coatneyi, P. inui and P. knowlesi. The entire amplification process was completed in 60 min, with the assays performed at 65 °C. By using SYTO-9 as the nucleic acid intercalating dye, the reaction was monitored via real-time fluorescence signal.
RESULTS
There was no observed cross-reactivity among the primers from different species. All 70 field-collected monkey samples were successfully amplified by real-time fluorescence LAMP assays. The detection limit for P. cynomolgi, P. coatneyi and P. knowlesi was 5 × 10 copies/µL. Meanwhile, the detection limit of P. inui was 5 × 10 copies/µL.
CONCLUSION
This is the first report of the detection of four zoonotic malaria parasites by real-time fluorescence LAMP approaches. It is an effective, rapid and simple-to-use technique. This presented platform exhibits considerable potential as an alternative detection for zoonotic malaria parasites.
Topics: Animals; Nucleic Acid Amplification Techniques; Malaria; RNA, Ribosomal, 18S; Molecular Diagnostic Techniques; Plasmodium; Sensitivity and Specificity; Zoonoses; Humans; DNA Primers; Fluorescence; Macaca; Monkey Diseases
PubMed: 38740319
DOI: 10.1016/j.actatropica.2024.107249 -
Acta Tropica Mar 2024Combining the advantages of PCR and LAMP, we described a new technique, namely PCR-LAMP, for malaria diagnosis. The whole process of DNA amplification can be completed...
Combining the advantages of PCR and LAMP, we described a new technique, namely PCR-LAMP, for malaria diagnosis. The whole process of DNA amplification can be completed in 35 min. This hybrid amplification technique markedly improved the sensitivity of detection compared to the classic single PCR or LAMP assay alone. PCR-LAMP assay had a detection limit of 1 copy/µL for P. knowlesi and P. ovale, 0.1 copy/µL for P. vivax, P. falciparum and P. malariae, respectively. To facilitate the endpoint detection, xylenol orange was added. Positive samples were indicated in orange while negative reactions were violet. The inclusion of xylenol orange into the LAMP reaction mix significantly reduces the post-amplification workload. Without relying on the use of specific instruments, the color changes of the amplicons could be visualized directly through the naked eye. In conclusion, PCR-LAMP poses the potential to be developed as a new malaria molecular diagnosis tool.
Topics: Humans; Sensitivity and Specificity; Plasmodium; Malaria; Nucleic Acid Amplification Techniques; Malaria, Falciparum; Malaria, Vivax; Polymerase Chain Reaction; Plasmodium falciparum; Plasmodium vivax; Phenols; Sulfoxides; Molecular Diagnostic Techniques
PubMed: 38199452
DOI: 10.1016/j.actatropica.2024.107120 -
Usefulness of serial testing for the diagnosis of malaria in cases of fever upon return from travel.Journal of Travel Medicine Apr 2024When malaria is suspected in case of fever after travel in endemic areas, the current recommendation is to repeat the malaria test at 24-hour intervals, with up to two...
BACKGROUND
When malaria is suspected in case of fever after travel in endemic areas, the current recommendation is to repeat the malaria test at 24-hour intervals, with up to two additional tests, as long as the test result is negative. A retrospective analysis was conducted to investigate the appropriateness of this recommendation by determining the proportion of tests with negative result at first and subsequently with a positive one at second or third attempt.
METHODS
A retrospective study was conducted at the Centre for Primary Care and Public Health, Lausanne, covering a period of 15 years. All patients tested once for malaria were included. Testing included microscopy thick and thin films as well as malaria rapid diagnostic test used in combination. The main outcome measure was the proportion of patients with a first negative test result, subsequently positive on second or third test over the total patients with suspected malaria assessed. Demographic, travel, clinical, and laboratory variables were collected from patients' records to identify potential predictors of an initially negative and then positive test result.
RESULTS
Four thousand nine hundred seventy-two patients were included. Of those, 4557 (91.7%) had definitive negative test results, and 415 (8.3%) had a positive result on the first test [332/415 (80%) Plasmodium falciparum, 40/415 (9.6%) P. vivax, 21/415 (5.1%) P. ovale, 12/415 (2.9%) P. vivax/ovale, 9/415 (2.2%) P. malariae and 1/415 (0.2%) P. knowlesi], and 3/4972 (0.06%) had a positive result on the second test after a first negative result, 1/4972(0.02%) had a positive test result after 2 negative results, all with P. falciparum. One of the four patients that were positive after their initial negative test was pregnant. The very small number of patients with an initially negative test result and secondarily positive did not allow for risk factor analysis.
CONCLUSIONS
The current recommendation of serial malaria testing is not supported by the present study, a fortiori for those who do not present with a strong clinical or laboratory predictor of malaria.
Topics: Pregnancy; Female; Humans; Retrospective Studies; Malaria; Malaria, Falciparum; Malaria, Vivax; Travel; Fever
PubMed: 38431851
DOI: 10.1093/jtm/taae030