A Malaria Fingerprint in the Human Genome?

Organismic and Evolutionary Biolog

Cytokine-dependent and–independent gene expression changes and cell cycle block revealed in Trypanosoma cruzi-infected host cells by comparative mRNA profiling

Background: The requirements for growth and survival of the intracellular pathogen Trypanosoma cruzi within mammalian host cells are poorly understood. Transcriptional profiling of the host cell response to infection serves as a rapid read-out for perturbation of host physiology that, in part, reflects adaptation to the infective process. Using Affymetrix oligonucleotide array analysis we identified common and disparate host cell responses triggered by T. cruzi infection of phenotypically diverse human cell types. Results: We report significant changes in transcript abundance in T. cruzi-infected fibroblasts, endothelial cells and smooth muscle cells (2852, 2155 and 531 genes respectively; fold-change ≥ 2, p-value < 0.01) 24 hours post-invasion. A prominent type I interferon response was observed in each cell type, reflecting a secondary response to secreted cytokine in infected cultures. To identify a core cytokine-independent response in T. cruzi-infected fibroblasts and endothelial cells transwell plates were used to distinguish cytokine-dependent and -independent gene expression profiles. This approach revealed the induction of metabolic and signaling pathways involved in cell proliferation, amino acid catabolism and response to wounding as common themes in T. cruzi-infected cells. In addition, the downregulation of genes involved in mitotic cell cycle and cell division predicted that T. cruzi infection may impede host cell cycle progression. The observation of impaired cytokinesis in T. cruzi-infected cells, following nuclear replication, confirmed this prediction. Conclusion Metabolic pathways and cellular processes were identified as significantly altered at the transcriptional level in response to T. cruzi infection in a cytokine-independent manner. Several of these alterations are supported by previous studies of T. cruzi metabolic requirements or effects on the host. However, our methods also revealed a T. cruzi-dependent block in the host cell cycle, at the level of cytokinesis, previously unrecognized for this pathogen-host cell interaction

Chaperone expression profiles correlate with distinct physiological states of Plasmodium falciparum in malaria patients

<p>Abstract</p> <p>Background</p> <p>Molecular chaperones have been shown to be important in the growth of the malaria parasite <it>Plasmodium falciparum </it>and inhibition of chaperone function by pharmacological agents has been shown to abrogate parasite growth. A recent study has demonstrated that clinical isolates of the parasite have distinct physiological states, one of which resembles environmental stress response showing up-regulation of specific molecular chaperones.</p> <p>Methods</p> <p>Chaperone networks operational in the distinct physiological clusters in clinical malaria parasites were constructed using cytoscape by utilizing their clinical expression profiles.</p> <p>Results</p> <p>Molecular chaperones show distinct profiles in the previously defined physiologically distinct states. Further, expression profiles of the chaperones from different cellular compartments correlate with specific patient clusters. While cluster 1 parasites, representing a starvation response, show up-regulation of organellar chaperones, cluster 2 parasites, which resemble active growth based on glycolysis, show up-regulation of cytoplasmic chaperones. Interestingly, cytoplasmic Hsp90 and its co-chaperones, previously implicated as drug targets in malaria, cluster in the same group. Detailed analysis of chaperone expression in the patient cluster 2 reveals up-regulation of the entire Hsp90-dependent pro-survival circuitries. In addition, cluster 2 also shows up-regulation of <it>Plasmodium </it>export element (PEXEL)-containing Hsp40s thought to have regulatory and host remodeling roles in the infected erythrocyte.</p> <p>Conclusion</p> <p>In all, this study demonstrates an intimate involvement of parasite-encoded chaperones, PfHsp90 in particular, in defining pathogenesis of malaria.</p

In vivo transcriptional profiling of Plasmodium falciparum

BACKGROUND: Both host and pathogen factors contribute to disease outcome in Plasmodium falciparum infection. The feasibility of studying the P. falciparum in vivo transcriptome to understand parasite transcriptional response while it resides in the human host is presented. METHODS: A custom made oligonucleotide array with probes based on the P. falciparum 3D7 laboratory strain chromosome 2 sequence was used to detect in vivo P. falciparum transcripts. This study analyzed transcripts from total RNA derived from small blood samples of P. falciparum infected patients and compared the in vivo expression profile to the in vitro cultivated 3D7 strain transcriptome. RESULTS: The data demonstrated that in vivo transcription can be studied from a small blood sample, despite the abundance of human RNA. The in vivo transcriptome is similar to the 3D7 ring stage transcriptome, but there are significant differences in genes encoding a sexual stage antigen and surface proteins. CONCLUSIONS: Whole genome transcription analysis of P. falciparum can be carried out successfully and further studies in selected patient cohorts may provide insight into parasite in vivo biology and defense against host immunity

Plasmodium falciparum gene expression measured directly from tissue during human infection

Background: During the latter half of the natural 48-h intraerythrocytic life cycle of human Plasmodium falciparum infection, parasites sequester deep in endothelium of tissues, away from the spleen and inaccessible to peripheral blood. These late-stage parasites may cause tissue damage and likely contribute to clinical disease, and a more complete understanding of their biology is needed. Because these life cycle stages are not easily sampled due to deep tissue sequestration, measuring in vivo gene expression of parasites in the trophozoite and schizont stages has been a challenge. Methods: We developed a custom nCounter® gene expression platform and used this platform to measure malaria parasite gene expression profiles in vitro and in vivo. We also used imputation to generate global transcriptional profiles and assessed differential gene expression between parasites growing in vitro and those recovered from malaria-infected patient tissues collected at autopsy. Results: We demonstrate, for the first time, global transcriptional expression profiles from in vivo malaria parasites sequestered in human tissues. We found that parasite physiology can be correlated with in vitro data from an existing life cycle data set, and that parasites in sequestered tissues show an expected schizont-like transcriptional profile, which is conserved across tissues from the same patient. Imputation based on 60 landmark genes generated global transcriptional profiles that were highly correlated with genome-wide expression patterns from the same samples measured by microarray. Finally, differential expression revealed a limited set of in vivo upregulated transcripts, which may indicate unique parasite genes involved in human clinical infections. Conclusions: Our study highlights the utility of a custom nCounter® P. falciparum probe set, validation of imputation within Plasmodium species, and documentation of in vivo schizont-stage expression patterns from human tissues. Electronic supplementary material The online version of this article (doi:10.1186/s13073-014-0110-6) contains supplementary material, which is available to authorized users

Efficacy and Safety of COVID-19 Convalescent Plasma in Hospitalized Patients: A Randomized Clinical Trial

Importance: There is clinical equipoise for COVID-19 convalescent plasma (CCP) use in patients hospitalized with COVID-19. Objective: To determine the safety and efficacy of CCP compared with placebo in hospitalized patients with COVID-19 receiving noninvasive supplemental oxygen. Design, setting, and participants: CONTAIN COVID-19, a randomized, double-blind, placebo-controlled trial of CCP in hospitalized adults with COVID-19, was conducted at 21 US hospitals from April 17, 2020, to March 15, 2021. The trial enrolled 941 participants who were hospitalized for 3 or less days or presented 7 or less days after symptom onset and required noninvasive oxygen supplementation. Interventions: A unit of approximately 250 mL of CCP or equivalent volume of placebo (normal saline). Main outcomes and measures: The primary outcome was participant scores on the 11-point World Health Organization (WHO) Ordinal Scale for Clinical Improvement on day 14 after randomization; the secondary outcome was WHO scores determined on day 28. Subgroups were analyzed with respect to age, baseline WHO score, concomitant medications, symptom duration, CCP SARS-CoV-2 titer, baseline SARS-CoV-2 serostatus, and enrollment quarter. Outcomes were analyzed using a bayesian proportional cumulative odds model. Efficacy of CCP was defined as a cumulative adjusted odds ratio (cOR) less than 1 and a clinically meaningful effect as cOR less than 0.8. Results: Of 941 participants randomized (473 to placebo and 468 to CCP), 556 were men (59.1%); median age was 63 years (IQR, 52-73); 373 (39.6%) were Hispanic and 132 (14.0%) were non-Hispanic Black. The cOR for the primary outcome adjusted for site, baseline risk, WHO score, age, sex, and symptom duration was 0.94 (95% credible interval [CrI], 0.75-1.18) with posterior probability (P[cOR Conclusions and relevance: In this trial, CCP did not meet the prespecified primary and secondary outcomes for CCP efficacy. However, high-titer CCP may have benefited participants early in the pandemic when remdesivir and corticosteroids were not in use. Trial registration: ClinicalTrials.gov Identifier: NCT04364737

Transcriptional Profiling of Plasmodium falciparum Parasites from Patients with Severe Malaria Identifies Distinct Low vs. High Parasitemic Clusters

Background: In the past decade, estimates of malaria infections have dropped from 500 million to 225 million per year; likewise, mortality rates have dropped from 3 million to 791,000 per year. However, approximately 90% of these deaths continue to occur in sub-Saharan Africa, and 85% involve children less than 5 years of age. Malaria mortality in children generally results from one or more of the following clinical syndromes: severe anemia, acidosis, and cerebral malaria. Although much is known about the clinical and pathological manifestations of CM, insights into the biology of the malaria parasite, specifically transcription during this manifestation of severe infection, are lacking. Methods and Findings: We collected peripheral blood from children meeting the clinical case definition of cerebral malaria from a cohort in Malawi, examined the patients for the presence or absence of malaria retinopathy, and performed whole genome transcriptional profiling for Plasmodium falciparum using a custom designed Affymetrix array. We identified two distinct physiological states that showed highly significant association with the level of parasitemia. We compared both groups of Malawi expression profiles with our previously acquired ex vivo expression profiles of parasites derived from infected patients with mild disease; a large collection of in vitro Plasmodium falciparum life cycle gene expression profiles; and an extensively annotated compendium of expression data from Saccharomyces cerevisiae. The high parasitemia patient group demonstrated a unique biology with elevated expression of Hrd1, a member of endoplasmic reticulum-associated protein degradation system. Conclusions: The presence of a unique high parasitemia state may be indicative of the parasite biology of the clinically recognized hyperparasitemic severe disease syndrome

Plasmodium falciparum Infection Does Not Affect Human Immunodeficiency Virus Viral Load in Coinfected Rwandan Adults

Background: Plasmodium falciparum infection has been reported to increase human immunodeficiency virus (HIV) viral load (VL), which can facilitate HIV transmission. We prospectively studied the impact of mild P falciparum coinfection on HIV VL in Rwanda. Methods: We measured plasma HIV VL at presentation with malaria infection and weekly for 4 weeks after artemether-lumefantrine treatment in Rwandan adults infected with HIV with P falciparum malaria. Regression analyses were used to examine associations between malaria infection and HIV VL changes. Samples with detectable virus underwent genotypic drug-resistance testing. Results: We enrolled 28 HIV-malaria coinfected patients and observed 27 of them for 5 weeks. Three patients (11%) were newly diagnosed with HIV. Acute P falciparum infection had no significant effect on HIV VL slope over 28 days of follow-up. Ten patients with VL <40 copies/mL at enrollment maintained viral suppression throughout. Seventeen patients had a detectable VL at enrollment including 9 (53%) who reported 100% adherence to ARVs; 3 of these had detectable genotypic drug resistance. Conclusions: Unlike studies from highly malaria-endemic areas, we did not identify an effect of P falciparum infection on HIV VL; therefore, malaria is not likely to increase HIV-transmission risk in our setting. However, routine HIV testing should be offered to adults presenting with acute malaria in Rwanda. Most importantly, we identified a large percentage of patients with detectable HIV VL despite antiretroviral (ARV) therapy. Some of these patients had HIV genotypic drug resistance. Larger studies are needed to define the prevalence and factors associated with detectable HIV VL in patients prescribed ARVs in Rwanda

Reduced paediatric hospitalizations for malaria and febrile illness patterns following implementation of community-based malaria control programme in rural Rwanda

<p>Abstract</p> <p>Background</p> <p>Malaria control is currently receiving significant international commitment. As part of this commitment, Rwanda has undertaken a two-pronged approach to combating malaria via mass distribution of long-lasting insecticidal-treated nets and distribution of antimalarial medications by community health workers. This study attempted to measure the impact of these interventions on paediatric hospitalizations for malaria and on laboratory markers of disease severity.</p> <p>Methods</p> <p>A retrospective analysis of hospital records pre- and post-community-based malaria control interventions at a district hospital in rural Rwanda was performed. The interventions took place in August 2006 in the region served by the hospital and consisted of mass insecticide treated net distribution and community health workers antimalarial medication disbursement. The study periods consisted of the December–February high transmission seasons pre- and post-rollout. The record review examined a total of 551 paediatric admissions to identify 1) laboratory-confirmed malaria, defined by thick smear examination, 2) suspected malaria, defined as fever and symptoms consistent with malaria in the absence of an alternate cause, and 3) all-cause admissions. To define the impact of the intervention on clinical markers of malaria disease, trends in admission peripheral parasitaemia and haemoglobin were analyzed. To define accuracy of clinical diagnoses, trends in proportions of malaria admissions which were microscopy-confirmed before and after the intervention were examined. Finally, to assess overall management of febrile illnesses antibiotic use was described.</p> <p>Results</p> <p>Of the 551 total admissions, 268 (48.6%) and 437 (79.3%) were attributable to laboratory-confirmed and suspected malaria, respectively. The absolute number of admissions due to suspected malaria was smaller during the post-intervention period (N = 150) relative to the pre-intervention period (N = 287), in spite of an increase in the absolute number of hospitalizations due to other causes during the post-intervention period. The percentage of suspected malaria admissions that were laboratory-confirmed was greater during the pre-intervention period (80.4%) relative to the post-intervention period (48.1%, prevalence ratio [PR]: 1.67; 95% CI: 1.39 – 2.02; chi-squared p-value < 0.0001). Among children admitted with laboratory-confirmed malaria, the risk of high parasitaemia was higher during the pre-intervention period relative to the post-intervention period (age-adjusted PR: 1.62; 95% CI: 1.11 – 2.38; chi-squared p-value = 0.004), and the risk of severe anaemia was more than twofold greater during the pre-intervention period (age-adjusted PR: 2.47; 95% CI: 0.84 – 7.24; chi-squared p-value = 0.08). Antibiotic use was common, with 70.7% of all children with clinical malaria and 86.4% of children with slide-negative malaria receiving antibacterial therapy.</p> <p>Conclusion</p> <p>This study suggests that both admissions for malaria and laboratory markers of clinical disease among children may be rapidly reduced following community-based malaria control efforts. Additionally, this study highlights the problem of over-diagnosis and over-treatment of malaria in malaria-endemic regions, especially as malaria prevalence falls. More accurate diagnosis and management of febrile illnesses is critically needed both now and as fever aetiologies change with further reductions in malaria.</p