Casting a Wide Net around Immunity to Malaria Catches p53.

The mechanisms underlying acquisition of naturally acquired immunity to malaria are poorly understood. In this issue of Immunity, Tran and colleagues (2019) demonstrate that systems immunology is a powerful tool to decipher molecular and cellular components contributing to this immunity.

Deciphering host immunity to malaria using systems immunology.

A century of conceptual and technological advances in infectious disease research has changed the face of medicine. However, there remains a lack of effective interventions and a poor understanding of host immunity to the most significant and complex pathogens, including malaria. The development of successful interventions against such intractable diseases requires a comprehensive understanding of host-pathogen immune responses. A major advance of the past decade has been a paradigm switch in thinking from the contemporary reductionist (gene-by-gene or protein-by-protein) view to a more holistic (whole organism) view. Also, a recognition that host-pathogen immunity is composed of complex, dynamic interactions of cellular and molecular components and networks that cannot be represented by any individual component in isolation. Systems immunology integrates the field of immunology with omics technologies and computational sciences to comprehensively interrogate the immune response at a systems level. Herein, we describe the system immunology toolkit and report recent studies deploying systems-level approaches in the context of natural exposure to malaria or controlled human malaria infection. We contribute our perspective on the potential of systems immunity for the rational design and development of effective interventions to improve global public health.

Monitoring mosquito richness in an understudied area: can environmental DNA metabarcoding be a complementary approach to adult trapping?

Mosquito surveillance programmes are essential to assess the risks of local vector-borne disease outbreaks as well as for early detection of mosquito invasion events. Surveys are usually performed with traditional sampling tools (i.e., ovitraps and dipping method for immature stages or light or decoy traps for adults). Over the past decade, numerous studies have highlighted that environmental DNA (eDNA) sampling can enhance invertebrate species detection and provide community composition metrics. However, the usefulness of eDNA for detection of mosquito species has, to date, been largely neglected. Here, we sampled water from potential larval breeding sites along a gradient of anthropogenic perturbations, from the core of an oil palm plantation to the rainforest on São Tomé Island (Gulf of Guinea, Africa). We showed that (i) species of mosquitoes could be detected via metabarcoding mostly when larvae were visible, (ii) larvae species richness was greater using eDNA than visual identification and (iii) new mosquito species were also detected by the eDNA approach. We provide a critical discussion of the pros and cons of eDNA metabarcoding for monitoring mosquito species diversity and recommendations for future research directions that could facilitate the adoption of eDNA as a tool for assessing insect vector communities.

Natural-Product-Based Solutions for Tropical Infectious Diseases.

About half of the world's population and 80% of the world's biodiversity can be found in the tropics. Many diseases are specific to the tropics, with at least 41 diseases caused by endemic bacteria, viruses, parasites, and fungi. Such diseases are of increasing concern, as the geographic range of tropical diseases is expanding due to climate change, urbanization, change in agricultural practices, deforestation, and loss of biodiversity. While traditional medicines have been used for centuries in the treatment of tropical diseases, the active natural compounds within these medicines remain largely unknown. In this review, we describe infectious diseases specific to the tropics, including their causative pathogens, modes of transmission, recent major outbreaks, and geographic locations. We further review current treatments for these tropical diseases, carefully consider the biodiscovery potential of the tropical biome, and discuss a range of technologies being used for drug development from natural resources. We provide a list of natural products with antimicrobial activity, detailing the source organisms and their effectiveness as treatment. We discuss how technological advancements, such as next-generation sequencing, are driving high-throughput natural product screening pipelines to identify compounds with therapeutic properties. This review demonstrates the impact natural products from the vast tropical biome have in the treatment of tropical infectious diseases and how high-throughput technical capacity will accelerate this discovery process.

Intelligibility of pseudo-whispered speech after total laryngectomy.

This study aimed to assess the intelligibility of so-called 'pseudo-whispered speech' (pseudo-WS), as produced without voice nor pulmonic airstream by some alaryngeal patients prior to rehabilitation. Several perception tests were submitted to three experienced clinicians and three naive listeners, relying on the speech samples of 20 French native speakers: 10 alaryngeal speakers, solely using pseudo-WS when starting speech therapy up to six months after total laryngectomy, and 10 control speakers, recorded in the closest speech mode available, whispered speech (WS). Experts were asked to identify consonants (C) in the /a/+C+/a/ context and to rate intelligibility, unintended additive noise, and fluency on a likert-scale, while naive listeners completed a quantitative test of intelligibility. Intelligibility of WS was found to be high, with scores ranging from 46.33/54 to 53.67/54 (median 52.5, interquartile range 2.33) for the quantitative test, and segmental intelligibility ranging from 68.75% to 94.79% (median 87.5, interquartile range 17.71). Segmental confusion affected voicing in favour of unvoiced consonants, as previously reported in the literature. By contrast, intelligibility of pseudo-WS was found to be poor, with scores ranging from 1/54 (unintelligibility) to 28.33/54 (median 8.66, interquartile range 14.67) for the quantitative test, and segmental intelligibility ranging from 3.13% to 28.13% (median 9.24, interquartile range 14.58). Segmental intelligibility was not uniformly affected: stops, labials and unvoiced consonants were better identified than other categories. Finally, a significant correlation was found between global intelligibility and articulatory precision, while unintended additive noise and fluency seemed to play no role.

Avian blood parasite richness decreases with major histocompatibility complex class I loci number.

Major histocompatibility complex (MHC) genes are among the most polymorphic in the vertebrate genome. The high allele diversity is believed to be maintained primarily by sexual and pathogen-mediated balancing selection. The number of MHC loci also varies greatly across vertebrates, most notably across birds. MHC proteins play key roles in presenting antigens on the cell surface for recognition by T cells, with class I proteins specifically targeting intracellular pathogens. Here, we explore the hypothesis that MHC class I diversity (measured as loci number) coevolves with haemosporidian parasite burden of the host. Using data on 54 bird species, we demonstrate that high-MHC class I diversity is associated with significantly lower richness of Plasmodium, Haemoproteus as well as overall haemosporidian parasite lineages, the former thus indicating more efficient protection against intracellular pathogens. Nonetheless, the latter associations were only detected when MHC diversity was assessed using cloning and not 454 pyrosequencing-based studies, nor across all genotyping methods combined. Our results indicate that high-MHC class I diversity might play a key role in providing qualitative resistance against diverse haemosporidian parasites in birds, but further clarification is needed for the origin of contrasting results when using different genotyping methods for MHC loci quantification.

Diversity, prevalence and host specificity of avian parasites in southern Tunisian oases.

Oases are semi-natural woodlots surrounded by an inhospitable desert environment. This insular-like habitat system is known to support a mixture of sedentary and migratory bird species originating from different areas. However, little is known about the interactions between these birds and parasites. In this study, we investigated the diversity, prevalence and host specificity of avian haemosporidian parasites in southern Tunisian oases in two sedentary and common bird species, the laughing dove Spilopelia senegalensis and hybrid sparrow Passer domesticus × hispaniolensis, in six sites that differ regarding vegetation structure and distance to the coast. Two new Haemoproteus lineages, related to other Haemoproteus transmitted by biting midges, were detected in doves. With regard to sparrows, all detected parasites have previously been reported in other sparrow populations, except for one new Haemoproteus lineage. Our results also showed that densely vegetated sites were characterized by the higher prevalence of Plasmodium but a lower prevalence of Haemoproteus compared with less-vegetated sites. This is the first study aiming to explore avian parasites in the oasis habitat. Gathering data on a larger sample of oases with different sizes and isolation levels will be the next step to better understand factors shaping the transmission dynamics of avian parasites in such ecosystems.

Coevolutionary patterns and diversification of avian malaria parasites in African sunbirds (Family Nectariniidae).

The coevolutionary relationships between avian malaria parasites and their hosts influence the host specificity, geographical distribution and pathogenicity of these parasites. However, to understand fine scale coevolutionary host-parasite relationships, robust and widespread sampling from closely related hosts is needed. We thus sought to explore the coevolutionary history of avian Plasmodium and the widespread African sunbirds, family Nectariniidae. These birds are distributed throughout Africa and occupy a variety of habitats. Considering the role that habitat plays in influencing host-specificity and the role that host-specificity plays in coevolutionary relationships, African sunbirds provide an exceptional model system to study the processes that govern the distribution and diversity of avian malaria. Here we evaluated the coevolutionary histories using a multi-gene phylogeny for Nectariniidae and avian Plasmodium found in Nectariniidae. We then assessed the host-parasite biogeography and the structuring of parasite assemblages. We recovered Plasmodium lineages concurrently in East, West, South and Island regions of Africa. However, several Plasmodium lineages were recovered exclusively within one respective region, despite being found in widely distributed hosts. In addition, we inferred the biogeographic history of these parasites and provide evidence supporting a model of biotic diversification in avian Plasmodium of African sunbirds.

Primary resistance of CCR5-tropic HIV-1 to maraviroc cannot be predicted by the V3 sequence.

OBJECTIVES: Resistance of HIV-1 to CCR5 antagonists can occur without coreceptor switching by mutations in envelope glycoproteins that enable virus entry using the inhibitor-bound form of CCR5. We investigated whether mutations in the V3 region of HIV-1 from subjects naive to maraviroc could be associated with primary resistance to this drug. METHODS: The frequency of CCR5-tropic HIV-1 subtype B isolates harbouring putative V3 maraviroc resistance mutations was assessed among the HIV tropism database of Toulouse University Hospital, France. Phenotypic assessment of maraviroc susceptibility was performed for 14 isolates representative of the main mutation patterns and 14 controls. V3 mutations were reversed or introduced by site-directed mutagenesis. RESULTS: Ninety-three of 951 (9.8%) isolates harboured V3 mutations assumed to be associated with maraviroc resistance. Maraviroc completely blocked virus entry for all but 1 of the 14 isolates harbouring V3 mutations [IC50 8.6 nM; 95% CI (6.6-47.4)], as in the 14 control isolates [IC50 13.4 nM; 95% CI (7.7-50.3)] (P = 0.24). Primary resistance to maraviroc, with a plateau in entry inhibition, was found in one isolate (harbouring a 20F/21I genotype). Site-directed mutagenesis showed that V3 mutations are necessary but not sufficient to induce maraviroc resistance. CONCLUSIONS: The impact of V3 mutations depended on the env context in which they occurred. Simple assessment of the V3 genotype thus cannot accurately predict maraviroc resistance. Rather, phenotypic assessment of virus particles expressing the envelope glycoprotein as a whole is required. This approach revealed that primary resistance of CCR5-tropic HIV-1 subtype B isolates to maraviroc seems uncommon.

Host and habitat specialization of avian malaria in Africa.

Studies of both vertebrates and invertebrates have suggested that specialists, as compared to generalists, are likely to suffer more serious declines in response to environmental change. Less is known about the effects of environmental conditions on specialist versus generalist parasites. Here, we study the evolutionary strategies of malaria parasites (Plasmodium spp.) among different bird host communities. We determined the parasite diversity and prevalence of avian malaria in three bird communities in the lowland forests in Cameroon, highland forests in East Africa and fynbos in South Africa. We calculated the host specificity index of parasites to examine the range of hosts parasitized as a function of the habitat and investigated the phylogenetic relationships of parasites. First, using phylogenetic and ancestral reconstruction analyses, we found an evolutionary tendency for generalist malaria parasites to become specialists. The transition rate at which generalists become specialists was nearly four times as great as the rate at which specialists become generalists. We also found more specialist parasites and greater parasite diversity in African lowland rainforests as compared to the more climatically variable habitats of the fynbos and the highland forests. Thus, with environmental changes, we anticipate a change in the distribution of both specialist and generalist parasites with potential impacts on bird communities.

Analogy between laryngeal gesture in Mongolian Long Song and supracricoid partial laryngectomy.

This article reports the results of a multiparametrical analysis of Mongolian Long Song, characterised by multiple ornamentation and shows the similarities between the laryngeal behaviour observed during these ornamentations and the compensatory gesture produced by patients after supracricoid partial laryngectomy. This study includes (1) a physiological analysis of videofiberscopic laryngeal data from a healthy Mongolian singer and from three non-singer French-speaking clinical patients; and (2) an acoustical analysis (fundamental frequency and intensity). For the singer, the fiberoptic analysis showed two main laryngeal behaviours in producing ornamentations: (1) 'lyrical' vibratos mobilising the entire laryngeal block; (2) 'Mongolian' trills with essentially supraglottic movements, the arytenoids being mobilised independently of the rest of the laryngeal block. Patients demonstrated similar aryepiglottic trilling to fulfil a function of voicing. The acoustic analysis showed that the fundamental frequency and the intensity were in phase for vibrato, contrary to the 'Mongolian' trills which were in opposite phase, underlying a change of laryngeal vibratory mechanisms.

Can microplastics facilitate the emergence of infectious diseases?

Plastic pollution is a major environmental problem. Small plastic particles (called microplastics) have been reported to have pernicious effects on human and wildlife health, by altering physiological functions (e.g., immunity, metabolism) and interfering with commensal microorganisms. However, in addition to these direct toxic effects, we suggest that microplastic pollution might also exert deleterious effects, modifying (i) the exposure to pathogens (e.g., multi-drug resistant bacteria) and (ii) the dynamics of vector-borne diseases. Therefore, we argue that microplastics should be considered as a ubiquitous environmental hazard, potentially promoting the (re)emergence of infectious diseases. The implementation of multi- and interdisciplinary research projects are crucial to properly evaluate if microplastic pollution should be added to the current list of global health threats.

Clinical characteristics of singers attending a phoniatric outpatient clinic.

Purpose: Studies suggest that singers are over-represented in voice clinics and present a high risk of developing voice disorders. This retrospective study aims to describe the characteristics of 78 singers consulting a phoniatrician.Methods: In their medical files, data related to age, gender, occupational status, singing training, musical style, voice complaint, diagnosis, voice-quality grading (GRBAS) and treatment were gathered.Results: The patients were mostly female singers (87%). Non-professional singers (semi-professional included) represented 64%, professional singers 25% and students of singing 11%. The majority of singers were choristers (27%) and 22% were classical-style/oratorio-style singers. Two-thirds of the population had intensive vocal activity in speech or singing. Vocal endurance, somatosensory signs and difficulties with high pitches were the most frequent symptoms. Among the patients, 79% presented with singing-voice disorders with 85% of these having vocal fold lesions. Generally, their speaking voices were preserved. Vocal-folds nodules were the most prevalent pathology (37%) followed by sulcus (26%) and voice therapy was the main treatment.Conclusions: This study emphasizes the fact that singers have specific voice complaints related to their voice usage. The high occurrence of sulcus and other congenital-lesion suspicions, unusual in the general population consulting an ENT phoniatrician, seems to be rather specific for singers in agreement with the literature.

Plasmodium relictum infection and MHC diversity in the house sparrow (Passer domesticus).

Antagonistic coevolution between hosts and parasites has been proposed as a mechanism maintaining genetic diversity in both host and parasite populations. In particular, the high level of genetic diversity usually observed at the major histocompatibility complex (MHC) is generally thought to be maintained by parasite-driven selection. Among the possible ways through which parasites can maintain MHC diversity, diversifying selection has received relatively less attention. This hypothesis is based on the idea that parasites exert spatially variable selection pressures because of heterogeneity in parasite genetic structure, abundance or virulence. Variable selection pressures should select for different host allelic lineages resulting in population-specific associations between MHC alleles and risk of infection. In this study, we took advantage of a large survey of avian malaria in 13 populations of the house sparrow (Passer domesticus) to test this hypothesis. We found that (i) several MHC alleles were either associated with increased or decreased risk to be infected with Plasmodium relictum, (ii) the effects were population specific, and (iii) some alleles had antagonistic effects across populations. Overall, these results support the hypothesis that diversifying selection in space can maintain MHC variation and suggest a pattern of local adaptation where MHC alleles are selected at the local host population level.

Metabarcoding options to study eukaryotic endoparasites of birds.

There is growing interest in the study of avian endoparasite communities, and metabarcoding is a promising approach to complement more conventional or targeted methods. In the case of eukaryotic endoparasites, phylogenetic diversity is extreme, with parasites from 4 kingdoms and 11 phyla documented in birds. We addressed this challenge by comparing different primer sets across 16 samples from 5 bird species. Samples consisted of blood, feces, and controlled mixes with known proportions of bird and nematode DNA. Illumina sequencing revealed that a 28S primer set used in combination with a custom blocking primer allowed detection of various plasmodiid parasites and filarioid nematodes in the blood, coccidia in the feces, as well as two potentially pathogenic fungal groups. When tested on the controlled DNA mixes, these primers also increased the proportion of nematode DNA by over an order of magnitude. An 18S primer set, originally designed to exclude metazoan sequences, was the most effective at reducing the relative number of avian DNA sequences and was the only one to detect Trypanosoma in the blood. Expectedly, however, it did not allow nematode detection and also failed to detect avian malaria parasites. This study shows that a 28S set including a blocking primer allows detection of several major and very diverse bird parasite clades, while reliable amplification of all major parasite groups may require a combination of markers. It helps clarify options for bird parasite metabarcoding, according to priorities in terms of the endoparasite clades and the ecological questions researchers wish to focus on.

Memory CD8+ T cell compartment associated with delayed onset of Plasmodium falciparum infection and better parasite control in sickle-cell trait children.

OBJECTIVES: Study of individuals with protection from Plasmodium falciparum (Pf) infection and clinical malaria, including individuals affected by the sickle-cell trait (HbAS), offers the potential to identify cellular targets that could be translated for therapeutic development. We previously reported the first involvement of cellular immunity in HbAS-associated relative protection and identified a novel subset of memory-activated NK cells that was enriched in HbAS children and associated with parasite control. We hypothesised that other memory cell subsets might distinguish the baseline profile of HbAS children and children with normal haemoglobin (HbAA). METHODS: Subsets of memory T cells and NK cells were analysed by flow cytometry in paired samples collected from HbAS and HbAA children, at baseline and during the first malaria episode of the ensuing transmission season. Correlations between cell frequencies and features of HbAS-mediated protection from malaria were determined. RESULTS: HbAS children displayed significantly higher frequency of memory CD8+ T cells at baseline than HbAA children. Baseline frequency of memory CD8+ T cells correlated with features of HbAS-mediated protection from malaria. Exploration of memory CD8+ T cell subsets revealed that central memory CD8+ T cell frequency was higher in HbAS children than in HbAA children. CONCLUSION: This study shows that HbAS children develop a larger memory CD8+ T cell compartment than HbAA children, and associates this compartment with better control of subsequent onset of infection and parasite density. Our data suggest that central memory CD8+ T cells may play an important role in the relative protection against malaria experienced by HbAS individuals, and further work to investigate this is warranted.

Th22 cells are efficiently recruited in the gut by CCL28 as an alternative to CCL20 but do not compensate for the loss of Th17 cells in treated HIV-1-infected individuals.

Gut CD4+ T cells are incompletely restored in most HIV-1-infected individuals on antiretroviral therapy, notably Th17 cells, a key subset in mucosal homeostasis. By contrast, gut Th22 cells are usually restored at normal frequencies. Th22 cells display a CCR6+CCR10+ phenotype and could thus respond to CCL20- and CCL28-mediated chemotaxis, while Th17 cells, which express CCR6 but not CCR10, depend on CCL20. Herein, we found that CCL28 is normally expressed by duodenal enterocytes of treated HIV-1-infected individuals, while CCL20 expression is blunted. Ex vivo, we showed that Th22 cells contribute to the reduction of CCL20 production by enterocytes through an IL-22- and IL-18-dependent mechanism. Th22 cells preferentially migrate via CCL20- rather than CCL28-mediated chemotaxis when both chemokines are available in the microenvironment. However, when the CCL20/CCL28 ratio drops, as in treated HIV-1-infected individuals, Th22 cells can migrate via the CCR10-CCL28 axis, as an alternative to CCR6-CCL20. This could explain the better reconstitution of gut Th22 compared with Th17 cells on antiretroviral therapy. Lastly, we assessed the relationships between the frequencies of gut Th17 and Th22 cells and inflammatory markers related to microbial translocation, and showed that Th22 cells do not compensate for the loss of Th17 cells in treated HIV-1-infected individuals.

Perceptual Judgment of Voice Quality in Nondysphonic French Speakers: Effect of Task-, Speaker- and Listener-Related Variables.

PURPOSE: Several perceptual scales have been developed to assess voice quality in dysphonic voices, among which the Grade Roughness Breathiness Asthenia Strain and a Rate of Dysphonia scale is probably the most frequently used. However, this clinical tool has not been properly validated with a normophonic population yet. The aim of the present study was to provide a first set of reference data gathered from a normal population, to serve as a basis of comparison for vocologists and laryngologists working with French-speaking patients. A second goal was to investigate the influence on this normal voice dataset, of variables known to affect perceptual judgments of pathological voice. MATERIAL AND METHODS: Sustained vowels and sentences produced by 80 healthy, normophonic French native speakers were perceptually assessed by a panel of 18 raters (nine students, nine experts) using the Grade Roughness Breathiness Asthenia Strain and a Rate of Dysphonia scale. RESULTS: The average overall grade was close to 1 on the (0 to 3) scale, questioning the notion of "normal" voice as opposed to dysphonic voice. Rating reliability as well as perceptual scores were affected by task-, speaker-, and listener-related factors: speech stimuli led to better rating reliability and were judged less severely than voice stimuli; experts were slightly more reliable and less severe than students; older speakers were unanimously considered as more dysphonic. Multiple interactions between these factors were observed, confirming the multidimensional nature of voice quality.

An Analytically and Diagnostically Sensitive RNA Extraction and RT-qPCR Protocol for Peripheral Blood Mononuclear Cells.

Reliable extraction and sensitive detection of RNA from human peripheral blood mononuclear cells (PBMCs) is critical for a broad spectrum of immunology research and clinical diagnostics. RNA analysis platforms are dependent upon high-quality and high-quantity RNA; however, sensitive detection of specific responses associated with high-quality RNA extractions from human samples with limited PBMCs can be challenging. Furthermore, the comparative sensitivity between RNA quantification and best-practice protein quantification is poorly defined. Therefore, we provide herein a critical evaluation of the wide variety of current generation of RNA-based kits for PBMCs, representative of several strategies designed to maximize sensitivity. We assess these kits with a reverse transcription quantitative PCR (RT-qPCR) assay optimized for both analytically and diagnostically sensitive cell-based RNA-based applications. Specifically, three RNA extraction kits, one post-extraction RNA purification/concentration kit, four SYBR master-mix kits, and four reverse transcription kits were tested. RNA extraction and RT-qPCR reaction efficiency were evaluated with commonly used reference and cytokine genes. Significant variation in RNA expression of reference genes was apparent, and absolute quantification based on cell number was established as an effective RT-qPCR normalization strategy. We defined an optimized RNA extraction and RT-qPCR protocol with an analytical sensitivity capable of single cell RNA detection. The diagnostic sensitivity of this assay was sufficient to show a CD8+ T cell peptide epitope hierarchy with as few as 1 × 104 cells. Finally, we compared our optimized RNA extraction and RT-qPCR protocol with current best-practice immune assays and demonstrated that our assay is a sensitive alternative to protein-based assays for peptide-specific responses, especially with limited PBMCs number. This protocol with high analytical and diagnostic sensitivity has broad applicability for both primary research and clinical practice.

A novel population of memory-activated natural killer cells associated with low parasitaemia in Plasmodium falciparum-exposed sickle-cell trait children.

OBJECTIVES: The sickle-cell trait phenotype is associated with protection from malaria. Multiple molecular mechanisms have been proposed to explain this protection, but the role of the host immune system has been poorly investigated. We hypothesised that cellular immunity to malaria may develop differently in sickle-cell trait children (HbAS) and children with normal haemoglobin (HbAA) repeatedly exposed to Plasmodium falciparum (Pf). METHODS: Paired samples collected prior to the Pf transmission season and during the first malaria episode of the ensuing transmission season from HbAS and HbAA children were analysed by multiplex bead-based assay and comprehensive multi-dimensional flow cytometry profiling. RESULTS: Cellular immune profiles were enriched in HbAS relative to HbAA children before the start of the Pf transmission season, with a distinct NK subset. These cells were identified as a novel subset of memory-activated NK cells characterised by reduced expression of the ecto-enzyme CD38 as well as co-expression of high levels of HLA-DR and CD45RO. The frequency of this NK subset before the transmission season was negatively correlated with parasite density quantified during the first malaria episode of the ensuing transmission season. Functional assessment revealed that these CD38dim CD45RO+ HLA-DR+ NK cells represent a important source of IFN-γ. CONCLUSION: Our data suggest that this novel memory-activated NK cell subset may contribute to an accelerated and enhanced IFN-γ-mediated immune response and to control of parasite density in individuals with the sickle-cell trait. This distinct cellular immune profile may contribute to predispose HbAS children to a relative protection from malaria.