Lack of K13 mutations in Plasmodium falciparum persisting after artemisinin combination therapy treatment of Kenyan children.

BACKGROUND: Studies in Southeast Asia reported a strong relationship between polymorphisms at the propeller domain of the Kelch 13 (K13) protein encoded by the Plasmodium falciparum k13 (pfk13) gene and delayed parasite clearance after artemisinin treatment. In Africa, P. falciparum remains susceptible and combination therapy regimens which include an artemisinin component display good efficacy. Using quantitative real-time PCR (qPCR), sub-microscopic persistence of P. falciparum has previously been reported in one-third of children treated with artemisinin combination therapy (ACT) in western Kenya. In this study, further investigation was made to evaluate whether these sub-microscopic residual parasites also harbour mutations at the propeller region of pfk13 and whether the mutations, if any, affect treatment outcome. METHODS: The pfk13 propeller domain was genotyped in DNA samples obtained in 2009 from Kenyan children treated with artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP). Paired samples at pre-treatment (day 0) and day of treatment failure (day 28 or 42) for 32 patients with documented recurrent parasitaemia were available for genotyping. Additional day 3 DNA samples were available for 10 patients. RESULTS: No mutation associated with artemisinin resistance in Southeast Asia was observed. Only one DP-treated patient harboured a non-synonymous mutation at codon 578 (A578S) of pfk13-propeller gene in the day 0 sample, but this allele was replaced by the wild-type (A578) form on day 3 and on the day of recurrent parasitaemia. The mutation at amino acid codon 578 showed no association with any phenotype. Polymorphisms in pfk13 were not responsible for parasite persistence and gametocyte carriage in the children treated with ACT. CONCLUSION: This study contributes to the ongoing surveillance of suspected artemisinin resistance parasites in Africa by providing baseline prevalence of k13-propeller mutations in western Kenya with samples collected from a longitudinal study. Clinical Trials Registration NCT00868465

Sensitive Detection of Gene Expression in Mycobacteria under Replicating and Non-Replicating Conditions Using Optimized Far-Red Reporters

Fluorescent reporter proteins have proven useful for imaging techniques in many organisms. We constructed optimized expression systems for several fluorescent proteins from the far-red region of the spectrum and analyzed their utility in several mycobacterial species. Plasmids expressing variants of the Discosoma Red fluorescent protein (DsRed) from the Mycobacterium bovis hsp60 promoter were unstable; in contrast expression from the Mycobacterium smegmatis rpsA promoter was stable. In Mycobacterium tuberculosis expression of several of the far-red reporters was readily visualised by eye and three reporters (mCherry, tdTomato, and Turbo-635) fluoresced at a high intensity. Strains expressing mCherry showed no fitness defects in vitro or in macrophages. Treatment of cells with antibiotics demonstrated that mCherry could also be used as a reporter for cell death, since fluorescence decreased in the presence of a bactericidal compound, but remained stable in the presence of a bacteriostatic compound. mCherry was functional under hypoxic conditions; using mCherry we demonstrated that the PmtbB is expressed early in hypoxia and progressively down-regulated. mCherry and other far-red fluorescent proteins will have multiple uses in investigating the biology of mycobacteria, particularly under non-replicating, or low cell density conditions, as well as providing a novel means of detecting cell death rapidly

Risky sexual behaviours among Ugandan university students: a pilot study exploring the role of adverse childhood experiences, substance use history, and family environment

Background: University students are known to have risky sexual behaviours (RSBs). The severity of the RSB is influenced by many factors, including the family environment, exposure to adverse childhood events (ACEs), and the use of addictive substances. However, there is limited information about the influence of ACEs and the family environment of these students in low-and medium-income countries (LMICs). Therefore, a pilot study was conducted among university students from a LMIC, Uganda. Methods: The present study comprised a cross-sectional online survey among Ugandan students at a public university (N = 316; 75% male; 52.2% aged between 18–22 years). The survey included questions relating to socio-demographic information, family environmental information, the Sexual Risk Survey (SRS), and the Adverse Childhood Experiences-International Questionnaire (ACE-IQ). Results: Over half (53.8%) reported having had sexual intercourse. Males reported over two times higher mean total SRS score compared to females (χ2 = 4.06, p = 0.044). Approximately one-sixth of the sample had drunk alcohol or used illicit psychoactive substances in the past six months (16.1%). Among four regression analysis models, sociodemographic variables predicted the highest variance (13%), followed by family environment variables (10%), and both psychoactive substance use history (past six months) and ACEs individually explained approximately 5% variance in total SRS score, with the final model predicting 33% of the variance in RSB. Conclusions: The present study demonstrated a gender disparity with males involved in more RSB than females, as has been reported in most previous RSB studies. Family environment, sociodemographic factors, substance use, and ACEs all appear to contribute to RSB among university students. These findings will benefit other researchers exploring factors associated with RSB among university students and will help develop interventions to reduce RSB to protect students from unwanted pregnancies, sexually transmitted diseases, and HIV/AIDS

pfk13-Independent Treatment Failure in Four Imported Cases of Plasmodium falciparum Malaria Treated with Artemether-Lumefantrine in the United Kingdom.

We present case histories of four patients treated with artemether-lumefantrine for falciparum malaria in UK hospitals in 2015 to 2016. Each subsequently presented with recurrent symptoms and Plasmodium falciparum parasitemia within 6 weeks of treatment with no intervening travel to countries where malaria is endemic. Parasite isolates, all of African origin, harbored variants at some candidate resistance loci. No evidence of pfk13-mediated artemisinin resistance was found. Vigilance for signs of unsatisfactory antimalarial efficacy among imported cases of malaria is recommended

Plasmodium-associated changes in human odor attract mosquitoes.

Malaria parasites (Plasmodium) can change the attractiveness of their vertebrate hosts to Anopheles vectors, leading to a greater number of vector-host contacts and increased transmission. Indeed, naturally Plasmodium-infected children have been shown to attract more mosquitoes than parasite-free children. Here, we demonstrate Plasmodium-induced increases in the attractiveness of skin odor in Kenyan children and reveal quantitative differences in the production of specific odor components in infected vs. parasite-free individuals. We found the aldehydes heptanal, octanal, and nonanal to be produced in greater amounts by infected individuals and detected by mosquito antennae. In behavioral experiments, we demonstrated that these, and other, Plasmodium-induced aldehydes enhanced the attractiveness of a synthetic odor blend mimicking "healthy" human odor. Heptanal alone increased the attractiveness of "parasite-free" natural human odor. Should the increased production of these aldehydes by Plasmodium-infected humans lead to increased mosquito biting in a natural setting, this would likely affect the transmission of malaria

Molecular quantification of Plasmodium parasite density from the blood retained in used RDTs.

Most malaria-endemic countries are heavily reliant upon rapid diagnostic tests (RDT) for malaria case identification and treatment. RDT previously used for malaria diagnosis can subsequently be used for molecular assays, including qualitative assessment of parasite species present or the carriage of resistance markers, because parasite DNA can be extracted from the blood inside the RDT which remains preserved on the internal components. However, the quantification of parasite density has not previously been possible from used RDT. In this study, blood samples were collected from school-age children in Western Kenya, in the form of both dried blood spots on Whatman filter paper, and the blood spot that is dropped into rapid diagnostic tests during use. Having first validated a robotic DNA extraction method, the parasite density was determined from both types of sample by duplex qPCR, and across a range of densities. The methods showed good agreement. The preservation of both parasite and human DNA on the nitrocellulose membrane inside the RDT was stable even after more than one year's storage. This presents a useful opportunity for researchers or clinicians wishing to gain greater information about the parasite populations that are being studied, without significant investment of resources

Suicidal behaviours among Ugandan university students: a cross-sectional study

Background: Suicide remains the leading cause of death among university students often resulting from multiple physical and psychological challenges. Moreover, suicidal behaviours among students appear to have increased due to the COVID-19 pandemic according to some studies. Objective: To explore the prevalence and associated factors for suicidal ideation, suicide plans, and suicide attempts among university students in Uganda. Methods: Cross-sectional study data were collected from May to September 2021 from 540 undergraduate university students in south-western Uganda (363 males, mean age 23.3 years). Questions from the General Health Questionnaire (GHQ-28) were used to assess suicidal ideation, while other bespoke questions were used to assess suicide plans and attempts. The survey also investigated the suicide attempt/plan method, location of the suicidal activity, and reason for not enacting the suicide plan. Three independent regression analyses were used to determine the factors associated with different forms of suicidal behaviours. Results: The prevalence of past-year suicidal behaviours was 31.85% for suicidal ideation, 8.15% for suicide plans, and 6.11% for suicide attempts. Having a chronic physical medical condition increased the likelihood of having all forms of suicidal behaviours. Suicidal ideation was associated with having difficulty paying university tuition fees. However, being in the fifth year of university education, and feeling satisfied with current academic grades reduced the likelihood of suicidal ideation. Individuals feeling satisfied with academic performance appeared to be a protective factor against having suicide plans. Suicide attempts were associated with having a history of sexual abuse and having difficulty paying university tuition fees. The most common method used for attempted suicide was a drug overdose, and the most common location for attempted suicide was their homes. Conclusion: University students have prevalent suicide behaviours especially among students with a chronic physical medical condition, a history of sexual abuse, and problems paying university tuition fees. Based on the present study, for students at risk, universities should provide appropriate interventions such as life skills education and suicide prevention techniques

Prevalence and factors associated with depression and suicidal ideation during the COVID-19 pandemic among university students in Uganda: a cross-sectional study

Background: The COVID-19 pandemic has negatively impacted psychosocial well-being and mental health of students across the world. Although students are vulnerable to depression and suicidal ideation, few studies have been conducted in Uganda. This study aimed to determine the prevalence of depression and suicidal ideation, and associated factors among undergraduate university students in Uganda. Methods: A cross-sectional study was conducted among undergraduates [N = 540; 363 males; mean age = 23.3 (± 2.64) years] recruited from four universities using an online questionnaire that explored sociodemographic factors, depression, and other associated factors. The Patient Health Questionnaire (PHQ-9) was used to assess depression, and Item 9 was used to assess suicidal ideation in the past 2 weeks. Multivariable logistic regression analyses were performed to determine the factors associated with depression and suicidal ideation. Results: The prevalence of moderate to severe depression was 20% (n = 108) (cut-off: 10/27 based on the PHQ-9), and the prevalence of past-2-week suicidal ideation was 13.89% (n = 75) (cut-off: 1/3 based on the PHQ-9 Item 9). About half of the individuals who screened positive for depression had suicidal ideation. Factors associated with depression were: having relationship issues [adjusted odds ratio (aOR) = 1.79, 95% confidence interval (CI) = 1.13–2.81, p = 0.012], and having a history of sexual abuse (aOR = 2.06, 95% CI = 1.10–3.84, p = 0.023). Factors associated with reducing the risk of depression were: satisfaction with current academic performance (aOR = 0.50, 95% CI = 0.32–0.79, p = 0.003), and being in the fifth year of academic study (aOR = 0.16, 95% CI = 0.03–0.73, p = 0.018). Factors associated with suicidal ideation were: smoking cigarettes and/or marijuana (aOR = 4.83, 95% CI = 1.10–21.12, p = 0.037), and having financial tuition constraints (aOR = 1.85, 95% CI = 1.08–3.16, p = 0.024), However, satisfaction with current academic performance reduced the likelihood of suicidal ideation (aOR = 0.40, 95% CI = 0.23–0.70, p = 0.001). Conclusion: Approximately one-fifth of undergraduate university students were moderately to severely depressed, especially those who had relationship issues and those with a history of sexual abuse. Suicidal ideation was common among smokers and those having financial tuition constraints. Therefore, it is recommended that the university authorities implement measures to provide psychological support for the students with problems concerning financial tuition constraints, relationships, and sexual abuse. Also, all students with depression should be screened for suicidality