BackgroundMalaria, due to Plasmodium ovale, can be challenging to diagnose due to clinically mild disease and low parasite burden. Two genetically distinct sub-species of P. ovale exist: Plasmodium ovale curtisi (classic) and Plasmodium ovale wallikeri (variant). It is presently unknown if the sub-species causing infection affects performance of malaria diagnostic tests. The aim of this work was to understand how the genetically distinct sub-species, P. o. curtisi and P. o. wallikeri, affect malaria diagnostic tests.Methods Plasmodium ovale-positive whole blood specimens were sub-speciated by PCR and sequencing of 18S rRNA and dhfr-ts. Parasitaemia, morphology, pan-aldolase positivity, 18S copy number, and dhfr-ts sequences were compared between sub-species.ResultsFrom 2006 to 2015, 49 P. ovale isolates were identified, of which 22 were P. o. curtisi and 27 P. o. wallikeri; 80% were identified in the last five years, and 88% were acquired in West Africa. Sub-species did not differ by parasitaemia, 18S copy number, or pan-aldolase positivity. Lack of Schüffner’s stippling was over-represented among P. o. wallikeri isolates (p = 0.02). Several nucleotide polymorphisms between the sub-species were observed, but they do not occur at sites believed to relate to antifolate binding.Conclusions Plasmodium ovale is increasing among travellers to West Africa, although sub-species do not differ significantly by parasitologic features such as parasitaemia. Absence of Schüffner’s stippling may be a feature specific to P. o. wallikeri and is a novel finding.

Although microscopic examination of Giemsa-stained blood smears remains the gold standard for the diagnosis of malaria, molecular detection using PCR is becoming increasingly popular. Due to discrepant PCR and microscopy results, we aimed to optimize our detection assays for Plasmodium malariae and Plasmodium ovale by sequencing the 18S rRNA region and developing a new primer and probe set for real-time quantitative PCR (qPCR). Clinical specimens positive for P. malariae (n ‫؍‬ 15) or P. ovale (n ‫؍‬ 33) underwent amplification and sequencing of the 18S rRNA region. Based on sequence discrepancies between our current primer/probe and clinical isolates, degenerate P. ovale primer and probe were developed to determine if their performance characteristics improved. The reference (gold) standard was microscopy. No 18S sequence heterogeneity was observed among the P. malariae isolates, and the sensitivity and specificity of our current P. malariae qPCR assay were both 100%. Compared to microscopy, the sensitivity and specificity of our current P. ovale qPCR assay were 72.7% and 100%, respectively. Five single nucleotide polymorphisms (SNPs) were identified in P. ovale. The sensitivity of the new P. ovale assay increased to 100% with 100% specificity. We therefore improved the performance characteristics of our P. ovale molecular detection assay through the development of a degenerate primer and probe set which accommodates 18S SNPs among the 2 subspecies of P. ovale. Given the suboptimal sensitivity of rapid diagnostic tests for non-falciparum malaria and the typically low parasitemia of P. malariae and P. ovale, a well-performing confirmatory molecular assay is imperative for clinical laboratories.

Background Gestational helminth infections are correlated to adverse outcomes including maternal anemia; as such, treatment is recommended. However, little published high-quality data exist around the efficacy, safety, and tolerability of anti-helminthics in pregnancy. We therefore conducted a systematic review and synthesized the available data on maternal outcomes following gestational treatment of intestinal nematodes to help guide clinical decision-making. Methods Five electronic databases were searched for studies reporting the efficacy, safety, or tolerability of anti-helminthic drugs for gestational treatment of intestinal nematodes. Studies were systematically screened, followed by data extraction. Trial quality was assessed using the GRADE approach. We conducted a narrative synthesis followed by meta-analyses using random-effects models as appropriate. Data were summarized using qualitative and quantitative measures for specific parasitic infections as well as efficacy and safety of anti-parasitic agents. Outcomes of interest included: maternal anemia, minor adverse outcomes, pregnancy loss, pre-mature delivery, prevalence of infection, and cure rate. Results 23 studies were included. Gestational treatment with albendazole had cure rates up to 90% for hookworm and Ascaris, but only 50% for Trichuris. Mebendazole had an overall cure rate of ≤70% for Ascaris, hookworm, and Trichuris. Pooled relative risk reduction of hookworm prevalence at delivery with albendazole compared to placebo was 90% (95%CI 0.09-0.15, n=2, I2=0%). Rate of pregnancy loss and hemoglobin concentration did not differ between albendazole or mebendazole versus placebo, and rates of pre-term delivery were similar in albendazole-treated pregnant women versus controls. Ivermectin demonstrated a cure rate of 29% for hookworm and 56% for Trichuris in pregnant women. No serious adverse events were attributable to any drug studied. Conclusions With increased international travel and migration of vulnerable populations, practitioners will encounter nematode infections in pregnant patients. Our analysis supports that albendazole in pregnancy has high cure rates for STHs and is safe for the mother.

Background: Rapid diagnostic tests (RDT) and real-time PCR (qPCR) assays are sensitive for diagnosing malaria, but because they detect antigen and DNA, respectively, positivity may not reflect active infection. Performance characteristics of RDT and qPCR in Plasmodium falciparum positive specimens were evaluated over time to elucidate duration of positivity following conversion to microscopy negative. Methods:Specimens from patients with at least one specimen that was positive for P. falciparum by microscopy, and at least one specimen that was negative for P. falciparum within a 1-month period were identified. Survival distributions of the diagnostic tests over time were compared. Performance characteristics for each test were calculated.Results: Ninety specimens were included, with 48 initially positive for P. falciparum, and 42 subsequently negative. Of 42 specimens that converted to microscopy-negative following an initial positive, 26 (61.9 %) and 41 (97.6 %) were positive by qPCR and RDT, respectively. Survival curves of microscopy versus qPCR, as well as microscopy vs RDT differed significantly (p = 0.0002 and p < 0.0001, respectively). Compared to microscopy, sensitivity of qPCR was 100.0 % (95 % CI 90.8-100.0 %), and that of RDT was 100.0 % (95 % CI 90.8-100.0 %). Conclusions:Due to slow clearance of circulating antigen and DNA from bloodstream, RDT and qPCR have low positive predictive value for clinically relevant asexual parasitaemia in post-treatment specimens. Thus, microscopy remains the only available malaria diagnostic that can reliably distinguish true asexual parasitaemia from prolonged clearance of antigen and nucleic acid in a convalescing patient.

Pseudomonas aeruginosa is a common respiratory pathogen in cystic fibrosis (CF) patients which undergoes adaptations during chronic infection towards reduced virulence, which can facilitate bacterial evasion of killing by host cells. However, inflammatory cytokines are often found to be elevated in CF patients, and it is unknown how chronic P. aeruginosa infection can be paradoxically associated with both diminished virulence in vitro and increased inflammation and disease progression. Thus, we investigated the relationship between the stimulation of inflammatory cell death pathways by CF P. aeruginosa respiratory isolates and the expression of key inflammatory cytokines. We show that early respiratory isolates of P. aeruginosa from CF patients potently induce inflammasome signaling, cell death, and expression of IL-1β by macrophages, yet little expression of other inflammatory cytokines (TNF, IL-6 and IL-8). In contrast, chronic P. aeruginosa isolates induce relatively poor macrophage inflammasome signaling, cell death, and IL-1β expression but paradoxically excessive production of TNF, IL-6 and IL-8 compared to early P. aeruginosa isolates. Using various mutants of P. aeruginosa, we show that the premature cell death of macrophages caused by virulent bacteria compromises their ability to express cytokines. Contrary to the belief that chronic P. aeruginosa isolates are less pathogenic, we reveal that infections with chronic P. aeruginosa isolates result in increased cytokine induction due to their failure to induce immune cell death, which results in a relatively intense inflammation compared with early isolates.