A series of epidemiological explorations has suggested a negative association between national bacillus Calmette–Guérin (BCG) vaccination policy and the prevalence and mortality of coronavirus ...disease 2019 (COVID-19). However, these comparisons are difficult to validate due to broad differences between countries such as socioeconomic status, demographic structure, rural vs. urban settings, time of arrival of the pandemic, number of diagnostic tests and criteria for testing, and national control strategies to limit the spread of COVID-19. We review evidence for a potential biological basis of BCG cross-protection from severe COVID-19, and refine the epidemiological analysis to mitigate effects of potentially confounding factors (e.g., stage of the COVID-19 epidemic, development, rurality, population density, and age structure). A strong correlation between the BCG index, an estimation of the degree of universal BCG vaccination deployment in a country, and COVID-19 mortality in different socially similar European countries was observed (r² = 0.88; P = 8 × 10−7), indicating that every 10% increase in the BCG index was associated with a 10.4% reduction in COVID-19 mortality. Results fail to confirm the null hypothesis of no association between BCG vaccination and COVID-19 mortality, and suggest that BCG could have a protective effect. Nevertheless, the analyses are restricted to coarse-scale signals and should be considered with caution. BCG vaccination clinical trials are required to corroborate the patterns detected here, and to establish causality between BCG vaccination and protection from severe COVID-19. Public health implications of a plausible BCG cross-protection from severe COVID-19 are discussed.
It has been shown that nearly a quarter of the initial predicted gene models in the Plasmodium falciparum genome contain errors. Although there have been efforts to obtain complete cDNA sequences to ...correct the errors, the coverage of cDNA sequences on the predicted genes is still incomplete, and many gene models for those expressed in sexual or mosquito stages have not been validated. Antisense transcripts have widely been reported in P. falciparum; however, the extent and pattern of antisense transcripts in different developmental stages remain largely unknown.
We have sequenced seven bidirectional libraries from ring, early and late trophozoite, schizont, gametocyte II, gametocyte V, and ookinete, and four strand-specific libraries from late trophozoite, schizont, gametocyte II, and gametocyte V of the 3D7 parasites. Alignment of the cDNA sequences to the 3D7 reference genome revealed stage-specific antisense transcripts and novel intron-exon splicing junctions. Sequencing of strand-specific cDNA libraries suggested that more genes are expressed in one direction in gametocyte than in schizont. Alternatively spliced genes, antisense transcripts, and stage-specific expressed genes were also characterized.
It is necessary to continue to sequence cDNA from different developmental stages, particularly those of non-erythrocytic stages. The presence of antisense transcripts in some gametocyte and ookinete genes suggests that these antisense RNA may play an important role in gene expression regulation and parasite development. Future gene expression studies should make use of directional cDNA libraries. Antisense transcripts may partly explain the observed discrepancy between levels of mRNA and protein expression.
Significance The Anopheles gambiae mosquito is a very effective vector of human Plasmodium falciparum malaria. We recently found that the Pfs47 gene allows the parasite to survive, by evading the ...mosquito immune system. In this study, we explored the mechanism of Pfs47 immune evasion. We found that Pfs47 inhibits Jun-N-terminal kinase-mediated activation of apoptosis in invaded mosquito midgut cells by preventing activation of several caspases. Furthermore, the lack of caspase-S2 activation prevents the induction of enzymes that potentiate epithelial nitration, a reaction required for parasites to be “visible” to the mosquito complement-like system. These findings shed new light on how a single parasite gene inactivates the mosquito immune system and allows it to be successfully transmitted to a new host.
The malaria parasite, Plasmodium , must survive and develop in the mosquito vector to be successfully transmitted to a new host. The Plasmodium falciparum Pfs47 gene is critical for malaria transmission. Parasites that express Pfs47 ( NF54 WT ) evade mosquito immunity and survive, whereas Pfs47 knockouts ( KO ) are efficiently eliminated by the complement-like system. Two alternative approaches were used to investigate the mechanism of action of Pfs47 on immune evasion. First, we examined whether Pfs47 affected signal transduction pathways mediating mosquito immune responses, and show that the Jun-N-terminal kinase (JNK) pathway is a key mediator of Anopheles gambiae antiplasmodial responses to P. falciparum infection and that Pfs47 disrupts JNK signaling. Second, we used microarrays to compare the global transcriptional responses of A. gambiae midguts to infection with WT and KO parasites. The presence of Pfs47 results in broad and profound changes in gene expression in response to infection that are already evident 12 h postfeeding, but become most prominent at 26 h postfeeding, the time when ookinetes invade the mosquito midgut. Silencing of 15 differentially expressed candidate genes identified caspase-S2 as a key effector of Plasmodium elimination in parasites lacking Pfs47 . We provide experimental evidence that JNK pathway regulates activation of caspases in Plasmodium -invaded midgut cells, and that caspase activation is required to trigger midgut epithelial nitration. Pfs47 alters the cell death pathway of invaded midgut cells by disrupting JNK signaling and prevents the activation of several caspases, resulting in an ineffective nitration response that makes the parasite undetectable by the mosquito complement-like system.
While the deployment of insecticide-based strategies dramatically reduced the toll of insect-borne diseases in several regions, it resulted in widespread insecticide resistance in natural populations ...4. ...the development of new strategies to reduce disease transmission is greatly needed. Insect immunity is regulated by several different signaling pathways such as the JNK, JAK-STAT, Toll, IMD, and RNAi, which activate final effectors that limit pathogen development and replication 5,6. ...immune priming and other mechanisms of immune memory that result in long-term enhancement of mosquito immunity have gained attention as important mechanisms to reduce disease transmission 7. Oral infection of fly larvae with Drosophila C virus (DCV) enhanced survival to a lethal challenge with the same virus as adults, although there was no difference in viral load, suggesting that previous exposure to the virus enhanced tolerance to infection in adult flies 35. Granulocyte release microvesicles (HdMv) at the site of recruitment, which mediates complement-like activation and Plasmodium elimination. ...the intensity of the mosquito immune response to Plasmodium can be enhanced by a previous infection.
Efforts to knock out Plasmodium falciparum calcium-dependent protein kinase 1 (PfCDPK1) from asexual erythrocytic stage have not been successful, indicating an indispensable role of the enzyme in ...asexual growth. We recently reported generation of a transgenic parasite with mutant CDPK1 Bansal A, et al. (2016) MBio 7:e02011-16. The mutant CDPK1 (T145M) had reduced activity of transphosphorylation. We reasoned that CDPK1 could be disrupted in the mutant parasites. Consistent with this assumption, CDPK1 was successfully disrupted in the mutant parasites using CRISPR/Cas9. We and others could not disrupt PfCDPK1 in the WT parasites. The CDPK1 KO parasites show a slow growth rate compared with the WT and the CDPK1 T145M parasites. Additionally, the CDPK1 KO parasites show a defect in both male and female gametogenesis and could not establish an infection in mosquitoes. Complementation of the KO parasite with full-length PfCDPK1 partially rescued the asexual growth defect and mosquito infection. Comparative global transcriptomics of WT and the CDPK1 KO schizonts using RNA-seq show significantly high transcript expression of gametocyte-specific genes in the CDPK1 KO parasites. This study conclusively demonstrates that CDPK1 is a good target for developing transmission-blocking drugs.
Plasmodium falciparum malaria originated in Africa and became global as humans migrated to other continents. During this journey, parasites encountered new mosquito species, some of them ...evolutionarily distant from African vectors. We have previously shown that the Pfs47 protein allows the parasite to evade the mosquito immune system of Anopheles gambiae mosquitoes. Here, we investigated the role of Pfs47-mediated immune evasion in the adaptation of P. falciparum to evolutionarily distant mosquito species. We found that P. falciparum isolates from Africa, Asia, or the Americas have low compatibility to malaria vectors from a different continent, an effect that is mediated by the mosquito immune system. We identified 42 different haplotypes of Pfs47 that have a strong geographic population structure and much lower haplotype diversity outside Africa. Replacement of the Pfs47 haplotypes in a P. falciparum isolate is sufficient to make it compatible to a different mosquito species. Those parasites that express a Pfs47 haplotype compatible with a given vector evade antiplasmodial immunity and survive. We propose that Pfs47-mediated immune evasion has been critical for the globalization of P. falciparum malaria as parasites adapted to new vector species. Our findings predict that this ongoing selective force by the mosquito immune system could influence the dispersal of Plasmodium genetic traits and point to Pfs47 as a potential target to block malaria transmission. A new model, the "lock-and-key theory" of P. falciparum globalization, is proposed, and its implications are discussed.
We recently characterized Pfs47, a protein expressed on the surface of sexual stages and ookinetes of Plasmodium falciparum, as a malaria transmission-blocking vaccine (TBV) target. Mice immunization ...induced antibodies that conferred strong transmission-reducing activity (TRA) at a concentration of 200 μg/mL. Here, we sought to optimize the Pfs47 vaccine to elicit higher titers of high-affinity antibodies, capable of inducing strong TRA at a lower concentration. We report the development and evaluation of a Pfs47-based virus-like particle (VLP) vaccine generated by conjugating our 58 amino acid Pfs47 antigen to Acinetobacter phage AP205-VLP using the SpyCatcher:SpyTag adaptor system. AP205-Pfs47 complexes (VLP-P47) formed particles of ~22 nm diameter that reacted with polyclonal anti-Pfs47 antibodies, indicating that the antigen was accessible on the surface of the particle. Mice immunized with VLP-P47 followed by a boost with Pfs47 monomer induced significantly higher antibody titers, with higher binding affinity to Pfs47, than mice that received two immunizations with either VLP-P47 (VLP-P47/VLP-P47) or the Pfs47 monomer (P47/P47). Purified IgG from VLP-P47/P47 mice had strong TRA (83-98%) at concentrations as low as 5 μg/mL. These results indicate that conjugating the Pfs47 antigen to AP205-VLP significantly enhanced antigenicity and confirm the potential of Pfs47 as a TBV candidate.
The involvement of reactive oxygen species (ROS) in mosquito immunity against bacteria and Plasmodium was investigated in the malaria vector Anopheles gambiae. Strains of An. gambiae with higher ...systemic levels of ROS survive a bacterial challenge better, whereas reduction of ROS by dietary administration of antioxidants significantly decreases survival, indicating that ROS are required to mount effective antibacterial responses. Expression of several ROS detoxification enzymes increases in the midgut and fat body after a blood meal. Furthermore, expression of several of these enzymes increases to even higher levels when mosquitoes are fed a Plasmodium berghei-infected meal, indicating that the oxidative stress after a blood meal is exacerbated by Plasmodium infection. Paradoxically, a complete lack of induction of catalase mRNA and lower catalase activity were observed in P. berghei-infected midguts. This suppression of midgut catalase expression is a specific response to ookinete midgut invasion and is expected to lead to higher local levels of hydrogen peroxide. Further reduction of catalase expression by double-stranded RNA-mediated gene silencing promoted parasite clearance by a lytic mechanism and reduced infection significantly. High mosquito mortality is often observed after P. berghei infection. Death appears to result in part from excess production of ROS, as mortality can be decreased by oral administration of uric acid, a strong antioxidant. We conclude that ROS modulate An. gambiae immunity and that the mosquito response to P. berghei involves a local reduction of detoxification of hydrogen peroxide in the midgut that contributes to limit Plasmodium infection through a lytic mechanism.
Traffic congestion has several causes, including insufficient road capacity, unrestricted demand and improper scheduling of traffic signal phases. A great variety of efforts have been made to ...properly program such phases. Some of them are based on traditional transportation assumptions, and others are adaptive, allowing the system to learn the control law (signal program) from data obtained from different sources. Reinforcement Learning (RL) is a technique commonly used in previous research. However, properly determining the states and the reward is key to obtain good results and to have a real chance to implement it. This paper proposes and implements a traffic signal control system (TSCS), detailing its development stages: (a) Intelligent Transportation System (ITS) architecture design for the TSCS; (b) design and development of a system prototype, including an RL algorithm to minimize the vehicle queue at intersections, and detection and calculation of such queues by adapting a computer vision algorithm; and (c) design and development of system tests to validate operation of the algorithms and the system prototype. Results include the development of the tests for each module (vehicle queue measurement and RL algorithm) and real-time integration tests. Finally, the article presents a system simulation in the context of a medium-sized city in a developing country, showing that the proposed system allowed reduction of vehicle queues by 29%, of waiting time by 50%, and of lost time by 50%, when compared to fixed phase times in traffic signals.
Several approaches from different perspectives have been used to solve problems with traffic accidents (TA), which mainly affect low- and middle-income countries. Conditions of certain cities, ...regarding road infrastructure, enforcement of traffic safety regulations, and motor vehicle numbers, influence the increase in TAs. Therefore, medium-sized cities in developing countries (context of interest), which commonly have worrying conditions, are a relevant scenario. One of the approaches to reduce TAs has been the use of data analysis through Machine Learning (ML); however, these techniques require a large amount of data, and medium-sized cities commonly do not have enough. Techniques such as Naturalistic Driving (ND) can be applied as a data collection method. This work proposes an intelligent collision risk detection system (ICDRS) using ND and ML to improve sustainability and safety of transportation in medium-sized cities. The ICRDS design considered the limitations of the context of interest and uses two data collection devices in the vehicle. The ICRDS validation included the design and execution of tests using ND. This validation identified if the collected data in a certain time interval contained high-risk collision events (sudden acceleration, sudden braking, aggressive left or right turn, aggressive left or right lane change). The system implementation results were satisfactory. The developed ML algorithm obtained an average value 0.98 in all the metrics. Two data sets of driving on routes were collected. In addition, the performed tests were able to identify city areas with high accident rates.
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