Potent transmission-blocking monoclonal antibodies from naturally exposed individuals target a conserved epitope on Plasmodium falciparum Pfs230.

Pfs230 is essential for Plasmodium falciparum transmission to mosquitoes and is the protein targeted by the most advanced malaria-transmission-blocking vaccine candidate. Prior understanding of functional epitopes on Pfs230 is based on two monoclonal antibodies (mAbs) with moderate transmission-reducing activity (TRA), elicited from subunit immunization. Here, we screened the B cell repertoire of two naturally exposed individuals possessing serum TRA and identified five potent mAbs from sixteen Pfs230 domain-1-specific mAbs. Structures of three potent and three low-activity antibodies bound to Pfs230 domain 1 revealed four distinct epitopes. Highly potent mAbs from natural infection recognized a common conformational epitope that is highly conserved across P. falciparum field isolates, while antibodies with negligible TRA derived from natural infection or immunization recognized three distinct sites. Our study provides molecular blueprints describing P. falciparum TRA, informed by contrasting potent and non-functional epitopes elicited by natural exposure and vaccination.

Vaccination with a structure-based stabilized version of malarial antigen Pfs48/45 elicits ultra-potent transmission-blocking antibody responses.

Malaria transmission-blocking vaccines (TBVs) aim to elicit human antibodies that inhibit sporogonic development of Plasmodium falciparum in mosquitoes, thereby preventing onward transmission. Pfs48/45 is a leading clinical TBV candidate antigen and is recognized by the most potent transmission-blocking monoclonal antibody (mAb) yet described; still, clinical development of Pfs48/45 antigens has been hindered, largely by its poor biochemical characteristics. Here, we used structure-based computational approaches to design Pfs48/45 antigens stabilized in the conformation recognized by the most potently inhibitory mAb, achieving >25°C higher thermostability compared with the wild-type protein. Antibodies elicited in mice immunized with these engineered antigens displayed on liposome-based or protein nanoparticle-based vaccine platforms exhibited 1-2 orders of magnitude superior transmission-reducing activity, compared with immunogens bearing the wild-type antigen, driven by improved antibody quality. Our data provide the founding principles for using molecular stabilization solely from antibody structure-function information to drive improved immune responses against a parasitic vaccine target.

Prediction of Long-Term Physical, Mental, and Cognitive Problems Following Critical Illness: Development and External Validation of the PROSPECT Prediction Model.

OBJECTIVES: ICU survivors often suffer from long-lasting physical, mental, and cognitive health problems after hospital discharge. As several interventions that treat or prevent these problems already start during ICU stay, patients at high risk should be identified early. This study aimed to develop a model for early prediction of post-ICU health problems within 48 hours after ICU admission. DESIGN: Prospective cohort study in seven Dutch ICUs. SETTING/PATIENTS: ICU patients older than 16 years and admitted for greater than or equal to 12 hours between July 2016 and March 2020. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Outcomes were physical problems (fatigue or ≥ 3 new physical symptoms), mental problems (anxiety, depression, or post-traumatic stress disorder), and cognitive impairment. Patient record data and questionnaire data were collected at ICU admission, and after 3 and 12 months, of 2,476 patients. Several models predicting physical, mental, or cognitive problems and a composite score at 3 and 12 months were developed using variables collected within 48 hours after ICU admission. Based on performance and clinical feasibility, a model, PROSPECT, predicting post-ICU health problems at 3 months was chosen, including the predictors of chronic obstructive pulmonary disease, admission type, expected length of ICU stay greater than or equal to 2 days, and preadmission anxiety and fatigue. Internal validation using bootstrapping on data of the largest hospital ( n = 1,244) yielded a C -statistic of 0.73 (95% CI, 0.70-0.76). External validation was performed on data ( n = 864) from the other six hospitals with a C -statistic of 0.77 (95% CI, 0.73-0.80). CONCLUSIONS: The developed and externally validated PROSPECT model can be used within 48 hours after ICU admission for identifying patients with an increased risk of post-ICU problems 3 months after ICU admission. Timely preventive interventions starting during ICU admission and follow-up care can prevent or mitigate post-ICU problems in these high-risk patients.

Development and External Validation of a Prediction Model for Quality of Life of ICU Survivors: A Subanalysis of the MONITOR-IC Prospective Cohort Study.

OBJECTIVES: To develop and externally validate a prediction model for ICU survivors' change in quality of life 1 year after ICU admission that can support ICU physicians in preparing patients for life after ICU and managing their expectations. DESIGN: Data from a prospective multicenter cohort study (MONITOR-IC) were used. SETTING: Seven hospitals in the Netherlands. PATIENTS: ICU survivors greater than or equal to 16 years old. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Outcome was defined as change in quality of life, measured using the EuroQol 5D questionnaire. The developed model was based on data from an academic hospital, using multivariable linear regression analysis. To assist usability, variables were selected using the least absolute shrinkage and selection operator method. External validation was executed using data of six nonacademic hospitals. Of 1,804 patients included in analysis, 1,057 patients (58.6%) were admitted to the academic hospital, and 747 patients (41.4%) were admitted to a nonacademic hospital. Forty-nine variables were entered into a linear regression model, resulting in an explained variance ( R2 ) of 56.6%. Only three variables, baseline quality of life, admission type, and Glasgow Coma Scale, were selected for the final model ( R2 = 52.5%). External validation showed good predictive power ( R2 = 53.2%). CONCLUSIONS: This study developed and externally validated a prediction model for change in quality of life 1 year after ICU admission. Due to the small number of predictors, the model is appealing for use in clinical practice, where it can be implemented to prepare patients for life after ICU. The next step is to evaluate the impact of this prediction model on outcomes and experiences of patients.

Ascaris lumbricoides and ticks associated with sensitization to galactose α1,3-galactose and elicitation of the alpha-gal syndrome.

BACKGROUND: IgE to galactose alpha-1,3 galactose (alpha-gal) causes alpha-gal syndrome (delayed anaphylaxis after ingestion of mammalian meat). Development of sensitization has been attributed to tick bites; however, the possible role of other parasites has not been well studied. OBJECTIVE: Our aims were to assess the presence, relative abundances, and site of localization of alpha-gal-containing proteins in common ectoparasites and endoparasites endemic in an area of high prevalence of alpha-gal syndrome, as well as to investigate the ability of ascaris antigens to elicit a reaction in a humanized rat basophil in vitro sensitization model. METHODS: Levels of total IgE, Ascaris-specific IgE, and alpha-gal IgE were measured in sera from patients with challenge-proven alpha-gal syndrome and from controls without allergy. The presence, concentration, and localization of alpha-gal in parasites were assessed by ELISA, Western blotting, and immunohistochemistry. The ability of Ascaris lumbricoides antigen to elicit IgE-dependent reactivity was demonstrated by using the RS-ATL8 basophil reporter system. RESULTS: Alpha-gal IgE level correlated with A lumbricoides-specific IgE level. Alpha-gal protein at 70 to 130 kDa was detected in A lumbricoides at concentrations higher than those found in Rhipicephalus evertsi and Amblyomma hebraeum ticks. Immunohistochemistry was used to localize alpha-gal in tick salivary acini and the helminth gut. Non-alpha-gal-containing A lumbricoides antigens activated RS-ATL8 basophils primed with serum from subjects with alpha-gal syndrome. CONCLUSION: We demonstrated the presence, relative abundances, and site of localization of alpha-gal-containing proteins in parasites. The activation of RS-ATL8 IgE reporter cells primed with serum from subjects with alpha-gal syndrome on exposure to non-alpha-gal-containing A lumbricoides proteins indicates a possible role of exposure to A lumbricoides in alpha-gal sensitization and clinical reactivity.

Asymptomatic School-Aged Children Are Important Drivers of Malaria Transmission in a High Endemicity Setting in Uganda.

Achieving malaria elimination requires a better understanding of the transmissibility of human infections in different transmission settings. This study aimed to characterize the human infectious reservoir in a high endemicity setting in eastern Uganda, using gametocyte quantification and mosquito feeding assays. In asymptomatic infections, gametocyte densities were positively associated with the proportion of infected mosquitoes (ß = 1.60; 95% CI, 1.32-1.92; P < .0001). Combining transmissibility and abundance in the population, symptomatic and asymptomatic infections were estimated to contribute to 5.3% and 94.7% of the infectious reservoir, respectively. School-aged children (5-15 years old) contributed to 50.4% of transmission events and were important drivers of malaria transmission.

Predicting surgical outcome in posterior retroperitoneoscopic adrenalectomy with the aid of a preoperative nomogram.

BACKGROUND: Posterior retroperitoneoscopic adrenalectomy (PRA) has several advantages over transperitoneal laparoscopic adrenalectomy (TLA) regarding operative time, blood loss, postoperative pain, and recovery. However, it can be a technically challenging procedure. To improve patient selection for PRA, we developed a preoperative nomogram to predict operative time. METHODS: All consecutive patients with tumors of ≤ 7 cm and a body mass index (BMI) of < 35 kg/m2 undergoing unilateral PRA between February 2011 and March 2020 were included in the study. The primary outcome was operative time as surrogate endpoint for surgical complexity. Using ten patient variables, an optimal prediction model was created, with a best subsets regression analysis to find the best one-variable up to the best seven-variable model. RESULTS: In total 215 patients were included, with a mean age of 52 years and mean tumor size of 2.4 cm. After best subsets regression analysis, a four-variable nomogram was selected and calibrated. This model included sex, pheochromocytoma, BMI, and perinephric fat, which were all individually significant predictors. This model showed an ideal balance between predictive power and applicability, with an R2 of 38.6. CONCLUSIONS: A four-variable nomogram was developed to predict operative time in PRA, which can aid the surgeon to preoperatively identify suitable patients for PRA. If the nomogram predicts longer operative time and therefore a more complex operation, TLA should be considered as an alternative approach since it provides a larger working space. Also, the nomogram can be used for training purposes to select patients with favorable characteristics when learning this surgical approach.

Associations between Environmental dust composition and Atopic Dermatitis in urban and rural settings.

BACKGROUND: Environmental exposures are involved in the pathogenesis of the allergic phenotype and in determining which individual triggers a person becomes sensitized to. Atopic dermatitis (AD) may modulate these effects through increased penetration through the skin modifying the immune system and AD may be triggered or intensified by environmental exposures. These exposures and immune-modulating factors may differ in urban and rural environments. OBJECTIVES: To compare house dust composition in urban and rural settings and correlate them with AD outcomes. METHODS: Dust samples were collected from the beds of 156 children aged 6 months to 3 years. 42% of participants had atopic dermatitis. Samples were analyzed for bacterial endotoxin, fungal (ß-1,3-glucan) levels, and house dust mite, cockroach, dog, cat, mouse, and peanut allergen. Exposures were compared in urban and rural environments and in participants with and without AD. RESULTS: Endotoxin but not fungal ß-glucan exposure is higher in the environment of healthy controls than children with AD in both urban and rural settings. House dust mite allergen exposure is high in urban and rural settings with Dermatophagoides detected in 100% of samples. Cat and dog allergen exposure mirrors pet ownership patterns which differ slightly between groups and environments. Mouse allergen exposure is higher in urban homes. CONCLUSION: Environmental endotoxin may be protective against AD in both urban and rural settings. There are marked differences in allergen exposure in urban and rural settings, but these are unlikely to be important protective or risk factors.

Molecular IgE sensitization profiles of urban and rural children in South Africa.

BACKGROUND: Allergens can act as disease-triggering factors in atopic dermatitis (AD) patients. The aim of the study was to elucidate the molecular IgE sensitization profile in children with and without AD living in urban and rural areas of South Africa. METHODS: Specific IgE reactivity was assessed in 166 Black South African children aged 9-38 months using a comprehensive panel of microarrayed allergens. According to clinical characterization children fell in four groups, urban AD cases (n = 32), urban controls (non-AD, n = 40), rural cases (n = 49) and rural controls (non-AD, n = 45). RESULTS: IgE reactivity to at least one of the allergens was detected in 94% of urban and 86% of rural AD children. House dust mite (HDM; 81% urban, 74% rural AD) and animal-derived allergens (50% urban, 31% rural AD) were the most frequently recognized respiratory allergens, whereas IgE to pollen allergens was almost absent. Urban AD children showed significantly higher frequency of IgE reactivity (50%) to mouse lipocalin, Mus m 1, than rural AD children (12%). The most frequently recognized food allergens were from egg (63% urban, 43% rural AD), peanut (31% vs 41%), and soybean (22% vs 27%), whereas milk sensitization was rare. α-gal-specific IgE almost exclusively occurred in rural children (AD: 14%, non-AD: 49%). CONCLUSION: Molecular allergy diagnosis detects frequent IgE sensitization to HDM, animal but not pollen allergens and to egg, peanut, and soy, but not milk allergens in African AD children. Urban AD children reacted more often to Mus m 1, whereas α-gal sensitization is more common in rural children likely due to parasite exposure.

Flexible modelling of risk factors on the incidence of pneumonia in young children in South Africa using piece-wise exponential additive mixed modelling.

INTRODUCTION: Recurrent episodes of pneumonia are frequently modeled using extensions of the Cox proportional hazards model with the underlying assumption of time-constant relative risks measured by the hazard ratio. We aim to relax this assumption in a study on the effect of factors on the evolution of pneumonia incidence over time based on data from a South African birth cohort study, the Drakenstein child health study. METHODS: We describe and apply two models: a time-constant and a time-varying relative effects model in a piece-wise exponential additive mixed model's framework for recurrent events. A more complex model that fits in the same framework is applied to study the continuously measured seasonal effects. RESULTS: We find that several risk factors (male sex, preterm birth, low birthweight, lower socioeconomic status, lower maternal education and maternal cigarette smoking) have strong relative effects that are persistent across time. When time-varying effects are allowed in the model, HIV exposure status (HIV exposed & uninfected versus HIV unexposed) shows a strong relative effect for younger children, but this effect weakens as children grow older, with a null effect reached from about 15 months. Weight-for-length at birth shows a time increasing relative effect. We also find that children born in the summer have a much higher risk of pneumonia in the 3-to-8-month age period compared with children born in winter. CONCLUSION: This work highlights the usefulness of flexible modelling tools in recurrent events models. It avoids stringent assumptions and allows estimation and visualization of absolute and relative risks over time of key factors associated with incidence of pneumonia in young children, providing new perspectives on the role of risk factors such HIV exposure.

Environmental factors associated with allergy in urban and rural children from the South African Food Allergy (SAFFA) cohort.

BACKGROUND: The prevalence of allergic diseases differs in urban and rural populations. OBJECTIVE: We sought to assess associations between environmental and dietary factors with allergic diseases in urban and rural South African children. METHODS: Toddlers aged 12 to 36 months were assessed for food allergen and aeroallergen sensitization, atopic dermatitis, allergic rhinitis, asthma, and challenge-proved food allergy. Information was collected on family history of allergic diseases, household size, socioeconomic status, delivery mode, antibiotic and probiotic use, exposure to fermented and unpasteurized milk, antihelminth treatment, sunlight exposure, pet and farm animal exposure, cigarette smoke, and household cooking and heating fuels. Antenatal exposures to pets, livestock, and cigarette smoke were assessed. A subsection completed questions on consumption of fruits and vegetables, fast foods, soft drinks/fruit juices, and fried/microwaved meat. RESULTS: Risk and protective factors differed between urban and rural settings. Exposure to farm animals in infants and their mothers during pregnancy was protective against allergic outcomes in the rural population. Consumption of unpasteurized milk is uncommon in this group of rural children and is unlikely to be an important factor in rural protection. In urban children birth by cesarean section is associated with food allergy, and consumption of fermented milk products is associated with reduced asthma and atopic dermatitis. In both cohorts antenatal maternal smoking and environmental smoking exposure were predominantly associated with asthma, and consumption of fast foods and fried meats were associated with allergy. CONCLUSION: In this rural environment exposure to livestock is the strongest protective factor. In urban communities, where animal contact is rare, risk factors include cesarian section, and protective factors include consumption of fermented milk products. Modifiable risk factors urgently require interventions to prevent increasing allergy rates in countries undergoing rapid urbanization.

Preoperative risk stratification in endometrial cancer (ENDORISK) by a Bayesian network model: A development and validation study.

BACKGROUND: Bayesian networks (BNs) are machine-learning-based computational models that visualize causal relationships and provide insight into the processes underlying disease progression, closely resembling clinical decision-making. Preoperative identification of patients at risk for lymph node metastasis (LNM) is challenging in endometrial cancer, and although several biomarkers are related to LNM, none of them are incorporated in clinical practice. The aim of this study was to develop and externally validate a preoperative BN to predict LNM and outcome in endometrial cancer patients. METHODS AND FINDINGS: Within the European Network for Individualized Treatment of Endometrial Cancer (ENITEC), we performed a retrospective multicenter cohort study including 763 patients, median age 65 years (interquartile range [IQR] 58-71), surgically treated for endometrial cancer between February 1995 and August 2013 at one of the 10 participating European hospitals. A BN was developed using score-based machine learning in addition to expert knowledge. Our main outcome measures were LNM and 5-year disease-specific survival (DSS). Preoperative clinical, histopathological, and molecular biomarkers were included in the network. External validation was performed using 2 prospective study cohorts: the Molecular Markers in Treatment in Endometrial Cancer (MoMaTEC) study cohort, including 446 Norwegian patients, median age 64 years (IQR 59-74), treated between May 2001 and 2010; and the PIpelle Prospective ENDOmetrial carcinoma (PIPENDO) study cohort, including 384 Dutch patients, median age 66 years (IQR 60-73), treated between September 2011 and December 2013. A BN called ENDORISK (preoperative risk stratification in endometrial cancer) was developed including the following predictors: preoperative tumor grade; immunohistochemical expression of estrogen receptor (ER), progesterone receptor (PR), p53, and L1 cell adhesion molecule (L1CAM); cancer antigen 125 serum level; thrombocyte count; imaging results on lymphadenopathy; and cervical cytology. In the MoMaTEC cohort, the area under the curve (AUC) was 0.82 (95% confidence interval [CI] 0.76-0.88) for LNM and 0.82 (95% CI 0.77-0.87) for 5-year DSS. In the PIPENDO cohort, the AUC for 5-year DSS was 0.84 (95% CI 0.78-0.90). The network was well-calibrated. In the MoMaTEC cohort, 249 patients (55.8%) were classified with <5% risk of LNM, with a false-negative rate of 1.6%. A limitation of the study is the use of imputation to correct for missing predictor variables in the development cohort and the retrospective study design. CONCLUSIONS: In this study, we illustrated how BNs can be used for individualizing clinical decision-making in oncology by incorporating easily accessible and multimodal biomarkers. The network shows the complex interactions underlying the carcinogenetic process of endometrial cancer by its graphical representation. A prospective feasibility study will be needed prior to implementation in the clinic.

Rural and urban food allergy prevalence from the South African Food Allergy (SAFFA) study.

BACKGROUND: Food sensitization and challenge-proved food allergy (FA) have not been compared in urban and rural settings. OBJECTIVE: We sought to determine and compare the prevalence of food sensitization and challenge-proved IgE-mediated FA in urban and rural South African toddlers aged 12 to 36 months. METHODS: This cross-sectional study of unselected children included 1185 participants in urban Cape Town and 398 in the rural Eastern Cape. All participants completed a questionnaire and underwent skin prick tests (SPTs) to egg, peanut, cow's milk, fish, soya, wheat, and hazelnut. Participants with SPT responses of 1 mm or greater to 1 or more foods and not tolerant on history underwent an open oral food challenge. RESULT: The prevalence of FA was 2.5% (95% CI, 1.6% to 3.3%) in urban children, most commonly to raw egg white (1.9%), followed by cooked egg (0.8%), peanut (0.8%), cow's milk (0.1%), and fish (0.1%). Urban sensitization (SPT response ≥1 mm) to any food was 11.4% (95% CI, 9.6% to 13.3%) and 9.0% (95% CI, 7.5% to 10.8%) at an SPT response of 3 mm or greater. Sensitization in rural cohorts was significantly lower than in the urban cohort (1-mm SPT response, 4.5% [95% CI, 2.5% to 6.6%]; 3-mm SPT response, 2.8% [95% CI, 1.4% to 4.9%]; P < .01). In the rural black African cohort 0.5% (95% CI, 0.1% to 1.8%) of children had food allergy, all to egg. This is significantly lower than the prevalence of the urban cohort overall (2.5%) and urban black African participants (2.9%; 95% CI, 1.5% to 4.3%; P = .006). CONCLUSION: FA prevalence in Cape Town is comparable with rates in industrialized middle-income countries and is significantly greater than in rural areas. Further analysis will describe and compare environmental exposures and other risk factors in this cohort.

Nutrition and allergic diseases in urban and rural communities from the South African Food Allergy cohort.

This study describes and compares allergic diseases and sensitization in urban and rural children in the SAFFA study cohort as well as infant feeding patterns and nutritional status. We assessed the relationship between nutritional status, breastfeeding, complementary feeding patterns, and atopic diseases including aeroallergen and food allergen sensitization, self-reported atopic dermatitis, allergic rhinitis, asthma, and challenge-proven food allergy (FA). METHODOLOGY: A total of 1185 urban and 398 rural toddlers aged 12-36 months were screened for food sensitization (FS) and FA using skin prick testing and oral food challenges. Of these, 535 and 347, respectively, were additionally screened for aeroallergen sensitization. Information was collected on infant feeding practices, and anthropometric measurements and clinical signs for atopy were documented. RESULTS: Markedly higher rates of allergy (asthma 9.0% vs 1.0%, eczema 25.6% vs 2.0%, rhinitis 25.3% vs 3.3%, and FA 2.5% vs 0.5%) exist in urban vs rural children. 13.1% unselected urban South African children were sensitized to aeroallergens compared to 3.8% of their rural counterparts and 9.0% to any food compared to 0.5%. Exclusive breastfeeding duration was longer, and there was a later introduction of allergenic foods in rural communities. Obesity rates were similar between the two groups, but rural children were more likely to be stunted. Being overweight was associated with asthma in urban but not rural settings. In the urban cohort, children with FS and allergy were thinner than their peers. CONCLUSION: Allergy and sensitization rates are significantly higher in unselected urban South African toddlers than their rural counterparts. Risk and protective factors for allergy and atopy may differ between urban and rural settings.

Predictive values of alpha-gal IgE levels and alpha-gal IgE: Total IgE ratio and oral food challenge-proven meat allergy in a population with a high prevalence of reported red meat allergy.

BACKGROUND: Severe meat allergy with anaphylaxis may be caused by sensitization to alpha-gal. Levels of alpha-gal sensitization that correlate with high risk of meat allergy are currently unknown. We have identified an area with a high prevalence of reported red meat allergy which offered the opportunity to evaluate the diagnostic value of IgE antibody tests. METHODS: To determine levels of alpha-gal IgE and alpha-gal:total IgE ratio in a large cohort of subjects with challenge-proven meat allergy compared with control subjects from the same environment, we conducted fieldwork assessing 131 participants who reported adverse reactions to meat, and 26 control subjects, by questionnaires, IgE sensitization to alpha-gal and oral food challenge to beef sausage. RESULTS: Eighty-four participants were diagnosed with alpha-gal allergy. Alpha-gal IgE ranged between 0.7 and 344.5 kU/L. Alpha-gal:total IgE ratio ranged from 0.1% to 67.6%. Logistic regression analysis showed both alpha-gal IgE and alpha-gal:total IgE ratio strongly correlated with meat allergy, with AUC of 0.95. The values giving the best correct classification were IgE of 2.00 kU/L and ratio of 0.75%. The value above which there is a 95% probability of meat allergy is IgE>5.5 kU/L and ratio of 2.12%. CONCLUSION: Alpha-gal allergy in a population with a high prevalence of reported red meat allergy showed a more rapid onset of symptoms than previously described and a high prevalence of isolated subjective gastrointestinal manifestations. Cutoff values are described for levels of sensitization to alpha-gal IgE and alpha-gal:total IgE ratio that are highly likely to result in clinically significant meat allergy.

A logistic model for the remediation of filamentous bulking in a biological nutrient removal wastewater treatment plant.

Biological nutrient removal (BNR) systems across the globe frequently experience bulking and foaming episodes, which present operational challenges such as poor sludge settling due to excessive filamentous bacteria. A full-scale BNR plant treating primarily domestic wastewater was monitored over a period of 1 year to investigate filamentous bacterial growth response under various plant operating parameters. Identification of filamentous bacteria by conventional microscopy and fluorescent in situ hybridisation indicated the dominance of Eikelboom Type021N, Thiothrix spp., Eikelboom Type 1851 and Eikelboom Type 0092. A cumulative logit model (CLM) was applied to elucidate significant relationships between the filamentous bacteria and plant operational parameters. The model could predict the potential abundance of dominant filamentous bacteria in relation to wastewater treatment plant operational parameters. Data obtained from the model corroborated with previous findings on the dominance of most filaments identified, except for Type 0092, which exhibited some unique traits. With further validation, the model could be successfully applied for identifying specific parameters which could contribute towards filamentous bulking, thus, providing a useful tool for regulating specific filamentous growth in full-scale wastewater treatment plants.

AlbuMAX supplemented media induces the formation of transmission-competent P. falciparum gametocytes.

Asexual blood stage culture of Plasmodium falciparum is routinely performed but reproducibly inducing commitment to and maturation of viable gametocytes remains difficult. Culture media can be supplemented with human serum substitutes to induce commitment but these generally only allow for long-term culture of asexual parasites and not transmission-competent gametocytes due to their different lipid composition. Recent insights demonstrated the important roles lipids play in sexual commitment; elaborating on this we exposed ring stage parasites (20-24 hours hpi) for one day to AlbuMAX supplemented media to trigger induction to gametocytogenesis. We observed a significant increase in gametocytes after AlbuMAX induction compared to serum. We also tested the transmission potential of AlbuMAX inducted gametocytes and found a significant higher oocyst intensity compared to serum. We conclude that AlbuMAX supplemented media induces commitment, allows a more stable and predictable production of transmittable gametocytes than serum alone.

Susceptibility of Anopheles gambiae to Natural Plasmodium falciparum Infection: A Comparison between the Well-Established Anopheles gambiae s.s Line and a Newly Established Ugandan Anopheles gambiae s.s. Line.

Much of our understanding of malaria transmission comes from mosquito feeding assays using Anopheles mosquitoes from colonies that are well adapted to membrane feeding. This raises the question whether results from colony mosquitoes lead to overestimates of outcomes in wild Anopheles mosquitoes. We successfully established an Anopheles colony using progeny of wild Anopheles gambiae s.s. mosquitoes (Busia mosquitoes) and directly compared their susceptibility to infection with Plasmodium falciparum with the widely used An. gambiae s.s. mosquitoes (Kisumu mosquitoes) using gametocyte-infected Ugandan donor blood. The proportion of infectious feeds did not differ between Busia (71.8%, 23/32) and Kisumu (68.8%, 22/32, P = 1.00) mosquitoes. When correcting for random effects of donor blood, we observed a 23% higher proportion of infected Busia mosquitoes than infected Kisumu mosquitoes (RR, 1.23; 95% CI, 1.10-1.38, P < 0.001). This study suggests that feeding assays with Kisumu mosquitoes do not overestimate outcomes in wild An. gambiae s.s. mosquitoes, the mosquito species most relevant to malaria transmission in Uganda.

Plasmodium falciparum gametocyte production correlates with genetic markers of parasite replication but is not influenced by experimental exposure to mosquito biting.

BACKGROUND: Plasmodium blood-stage parasites balance asexual multiplication with gametocyte development. Few studies link these dynamics with parasite genetic markers in vivo; even fewer in longitudinally monitored infections. Environmental influences on gametocyte formation, such as mosquito exposure, may influence the parasite's investment in gametocyte production. METHODS: We investigated gametocyte production and asexual multiplication in two Plasmodium falciparum infected populations; a controlled human malaria infection (CHMI) study and a 28-day observational study in naturally infected individuals in Burkina Faso with controlled mosquito exposure. We measured gene transcript levels previously related to gametocyte formation (ap2-g, surfin1.2, surfin13.1, gexp-2) or inhibition of asexual multiplication (sir2a) and compared transcript levels to ring-stage parasite and mature gametocyte densities. FINDINGS: Three of the five markers (ap2-g, surfin1.2, surfin13.1) predicted peak gametocytaemia in the CHMI study. An increase in all five markers in natural infections was associated with an increase in mature gametocytes 14 days later; the effect of sir2a on future gametocytes was strongest (fold change = 1.65, IQR = 1.22-2.24, P = 0.004). Mosquito exposure was not associated with markers of gametocyte formation (ap2-g P = 0.277; sir2a P = 0.499) or carriage of mature gametocytes (P = 0.379). INTERPRETATION: All five parasite genetic markers predicted gametocyte formation over a single cycle of gametocyte formation and maturation in vivo; sir2a and ap2-g were most closely associated with gametocyte growth dynamics. We observed no evidence to support the hypothesis that exposure to Anopheles mosquito bites stimulates gametocyte formation. FUNDING: This work was funded by the Bill & Melinda Gates Foundation (INDIE OPP1173572), the European Research Council fellowship (ERC-CoG 864180) and UKRI Medical Research Council (MR/T016272/1) and Wellcome Center (218676/Z/19/Z).

Dynamics of asymptomatic Plasmodium falciparum and Plasmodium vivax infections and their infectiousness to mosquitoes in a low transmission setting of Ethiopia: a longitudinal observational study.

OBJECTIVE: A 15-month longitudinal study was conducted to determine the duration and infectivity of asymptomatic qPCR-detected Plasmodium falciparum and Plasmodium vivax infections in Ethiopia. METHOD: Total parasite and gametocyte kinetics were determined by molecular methods; infectivity to Anopheles arabiensis mosquitoes by repeated membrane feeding assays. Infectivity results were contrasted with passively recruited symptomatic malaria cases. RESULTS: For P. falciparum and P. vivax infections detected at enrolment, median durations of infection were 37 days (95% confidence interval [CI], 15-93) and 60 days (95% CI, 18-213), respectively. P. falciparum and P. vivax parasite densities declined over the course of infections. From 47 feeding assays on 22 asymptomatic P. falciparum infections, 6.4% (3/47) were infectious and these infected 1.8% (29/1579) of mosquitoes. No transmission was observed in feeding assays on asymptomatic P. vivax mono-infections (0/56); one mixed-species infection was highly infectious. Among the symptomatic cases, 4.3% (2/47) of P. falciparum and 73.3% (53/86) of P. vivax patients were infectious to mosquitoes. CONCLUSION: The majority of asymptomatic infections were of short duration and low parasite density. Only a minority of asymptomatic individuals were infectious to mosquitoes. This contrasts with earlier findings and is plausibly due to the low parasite densities in this population.