Laser-assisted dacryocystorhinostomy of acquired dacryops in a dog.

OBJECTIVE: To describe a novel minimally invasive surgical approach to treatment of a nasolacrimal duct cyst in a dog and outcome. ANIMAL STUDIED: A 10-year-old, male entire, Springer Spaniel with a chronic right-sided maxillary swelling and associated right eye epiphora and sneezing. PROCEDURE: Computed tomography with dacryocystography confirmed a cystic structure and destruction of the right maxilla and zygomatic bone with invasion of the nasal cavity. Obstruction of the nasolacrimal duct was confirmed. Rhinoscopy was performed allowing visualization of the ventromedial wall of the cyst. Under direct visualization, dacryocystorhinostomy was performed and the wall of the cyst was ablated with the aid of a thulium laser fiber. RESULTS: No intraoperative complications occurred during the procedure. Full resolution of clinical signs was noted within 2 weeks of treatment. No recurrence had occurred on long-term follow-up at 32 months. CONCLUSIONS: Laser-assisted dacrocystorhinostomy under direct visualization was successful in the treatment and resolution of a nasolacrimal duct cyst (dacryops) in this case.

Fine-scale spatial distribution of deltamethrin resistance and population structure of Anopheles funestus and Anopheles arabiensis populations in Southern Mozambique.

BACKGROUND: Insecticide resistance in malaria vectors can be spatially highly heterogeneous, yet population structure analyses frequently find relatively high levels of gene flow among mosquito populations. Few studies have contemporaneously assessed phenotypic, genotypic and population structure analysis on mosquito populations and none at fine geographical scales. In this study, genetic diversity, population structure, and insecticide resistance profiles of Anopheles funestus and Anopheles arabiensis were examined across mosquito populations from and within neighbouring villages. METHODS: Mosquitoes were collected from 11 towns in southern Mozambique, as well as from different neighbourhoods within the town of Palmeira, during the peak malaria transmission season in 2016. CDC bottle bioassay and PCR assays were performed with Anopheles mosquitoes at each site to determine phenotypic and molecular insecticide resistance profiles, respectively. Microsatellite analysis was conducted on a subsample of mosquitoes to estimate genetic diversity and population structure. RESULTS: Phenotypic insecticide resistance to deltamethrin was observed in An. funestus sensu stricto (s.s.) throughout the area, though a high level of mortality variation was seen. However, 98% of An. funestus s.s. were CYP6P9a homozygous resistant. An. arabiensis was phenotypically susceptible to deltamethrin and 99% were kdr homozygous susceptible. Both Anopheles species exhibited high allelic richness and heterozygosity. Significant deviations from Hardy-Weinberg equilibrium were observed, and high linkage disequilibrium was seen for An. funestus s.s., supporting population subdivision. However, the FST values were low for both anophelines (- 0.00457 to 0.04213), Nm values were high (9.4-71.8 migrants per generation), AMOVA results showed almost 100% genetic variation among and within individuals, and Structure analysis showed no clustering of An. funestus s.s. and An. arabiensis populations. These results suggest high gene flow among mosquito populations. CONCLUSION: Despite a relatively high level of phenotypic variation in the An. funestus population, molecular analysis shows the population is admixed. These data indicate that CYP6P9a resistance markers do not capture all phenotypic variation in the area, but also that resistance genes of high impact are likely to easily spread in the area. Conversely, other strategies, such as transgenic mosquito release programmes will likely not face challenges in this locality.

Retrobulbar respiratory epithelial orbital cyst in a dog: A case report.

A 10-month-old female, neutered French Bulldog presented with a history of unilateral right-sided intermittent conjunctivitis and exophthalmos. The patient suffered blunt force trauma to the right eye after a tennis ball impact approximately five months prior to presentation. Examination identified the patient was visual with exophthalmos, lateral strabismus, conjunctival hyperaemia, episcleral congestion, and papilloedema. Magnetic resonance imaging and ultrasonography identified an approximately two centimeter diameter fluid-filled structure directly posterior to the globe leading to displacement of the optic nerve and distortion of the posterior globe wall. Centesis of the lesion demonstrated neutrophilic and macrophagic inflammation with evidence of prior hemorrhage. Within four weeks, the structure had re-filled to its original size and was subsequently excised in its entirety, via a lateral orbitotomy. Histopathologic findings indicated a non-keratinising orbital cyst, the lining of which was consistent with a respiratory epithelial cyst. Recurrence had not occurred seven months' post-surgery. To the author's knowledge, previous reports of retrobulbar respiratory epithelial cysts have not been documented in animals.

Comparing two different schedules of online learning for updated cardiopulmonary resuscitation guidelines in Covid-19 patients: A randomized study.

Background Coronavirus disease 2019 (Covid-19) is an evolving disease with newly generated evidence related to the clinical management of Covid-19 patients. We aimed to compare two online learning schedules for disseminating new cardiopulmonary resuscitation (CPR) guidelines in terms of knowledge gain and acceptability among nurses. Methods In a prospective randomized controlled study, 61 nurses trained in comprehensive cardiopulmonary life support (CCLS) were randomized to synchronous (n=31) and asynchronous learning groups (n= 30). The enhanced training module on CPR (ETMCPR) prepared by a team of experts was used to impart training to the nurses. Baseline data and pre-intervention knowledge of participants were collected using a structured demographic sheet and knowledge questionnaire (25 items) in a google form. Nurses in the synchronous group were provided training using ETMCPR through a licensed Zoom platform, while the nurses in the asynchronous group had access to the uploaded ETMCPR module in the e-learning platform. At the end of the intervention, the knowledge of the nurses was assessed along with their acceptability to the online learning schedule. Results Both schedules of online learning were effective in improving the knowledge scores of the nurses (11.93 [3.26] v. 21.15 [1.90], p=0.01 and 11.71 [3.12] v. 20.32 [1.71], p=0.01). The mean acceptability scores of nurses in the asynchronous group were statistically lower than in the synchronous group (38.93 [2.50] v. 42.5 [3.08], p=0.007). Conclusion Both synchronous and asynchronous schedules of online learning were effective in disseminating updated CPR guidelines; however, nurses in the synchronous group were more satisfied with the learning schedule.

Telemental Health Collaborative Care Medication Management: Implementation and Outcomes.

Introduction: Access to quality mental health medication management (MM) in the United States is limited, even among those with employment-based health insurance. This implementation, feasibility, and outcome study sought to design and evaluate an evidence-based telemental health MM service using a collaborative care model (CoCM). Materials and Methods: CoCM MM was available to adult employees/dependents through their employer benefits, in addition to therapy. Outcomes included Patient Health Questionnaire-9 (PHQ-9) and the Generalized Anxiety Disorder-7 (GAD-7) collected at baseline and throughout participation. This analysis was not deemed to be human subjects research by the Western Institutional Review Board. Results: Over 17 months, 212 people enrolled and completed >2 assessments; the enrollees were 58.96% female with average age of 32.00 years (standard deviation [SD] = 7.38). In people with moderate to severe depression or anxiety, PHQ-9 and GAD-7 scores reduced by an average of 7.27 (SD = 4.80) and 6.71 (SD = 5.18) points after at least 12 ± 4 weeks in the program. At 24 ± 4 weeks, the PHQ-9 and GAD-7 reductions were on average 7.17 (SD = 5.00) and 6.03 (SD = 5.37), respectively. Approximately 65.88% of participants with either baseline depression or anxiety had a response on either the PHQ-9 or GAD-7 at 12 ± 4 weeks and 44.71% of participants experienced remission; at 24 ± 4 weeks, 56.41% had response and 41.03% experienced remission. Conclusions: An evidence-based CoCM telemedicine service within an employee behavioral health benefit is feasible and effective in reducing anxiety and depression symptoms when using measurement-based care. Widespread implementation of a benefit like this could expand access to evidence-based mental health MM.

Submucosal resection via a transanal approach for treatment of epithelial rectal tumors - a multicenter study.

OBJECTIVE: To report complications and long-term outcomes after submucosal resections of benign and malignant epithelial rectal masses. STUDY DESIGN: Retrospective multicentric study. SAMPLE POPULATION: Medical records of 93 dogs at 7 referral hospitals. METHODS: Records were reviewed for surgical time, diagnosis, margins, complications, and recurrences. Survival of dogs was evaluated based on tumor types, categorized as benign, carcinoma in situ, and carcinoma. The Kaplan-Meier survival curve and Cox proportional hazards analysis were used to determine the association of a range of variables with recurrence and survival time. RESULTS: Duration of follow up was 708 days (range, 25-4383). Twenty-seven dogs (29%) developed complications. Recurrence was identified in 20/93 (21%), with 12/20 recurrent masses treated with repeat submucosal resection. Median survival was not reached in any group. The 1-,2-, 5-year survival rates for carcinomas were 95%, 89%, and 73% respectively. However, overall survival was longer for benign tumors than carcinomas (P = .001). Recurrence was more likely when complications (P = .032) or incomplete margins (P = .023) were present. Recurrence was associated with an increased risk of death (P = .046). CONCLUSION: Submucosal resection of both benign and malignant rectal masses was associated with a low rate of severe complications and prolonged survival in the 93 dogs described here. CLINICAL SIGNIFICANCE: Submucosal resection is a suitable technique for resection of selected rectal masses.

Comparison of Clinical Outcome between Dexamethasone and Methyl Prednisolone in Treatment of Moderate to Severe COVID-19: A Systematic Review and Meta-Analysis.

OBJECTIVE: Though systemic corticosteroid is used for treatment of COVID 19, questions regarding the appropriate dose, duration and type of corticosteroid use still remain unanswered. This study aimed to address, whether choice of systemic corticosteroid significantly influences the clinical outcome of COVID 19 patients. MATERIALS AND METHODS: Studies reporting the comparison between clinical outcome of dexamethasone and methylprednisolone in treatment of COVID 19 were searched from inception till April, 2021. Random-effect model was used. Odd's ratio (OR) and 95% confidence interval was expressed. I2 statistics used for test of heterogeneity. RESULT: Three studies with 373 patients (160 in dexamethasone group and 213 in methyl prednisolone group) were included. Though, statistically significant reduction in all-cause mortality with methyl prednisolone group in comparison to dexamethasone group (OR=1.80, 95%CI: 1.08 to 3.01, P=0.02) estimated, sub group analysis of observational studies did not support the finding (OR=1.60, 95% CI: 0.88 to 2.92, P=0.12). No significant difference in terms of need for invasive ventilator or intensive care unit (ICU) between the 2 groups. The grade of evidence was very low for both the outcomes. Conclusion and Relevance: In the present context, both dexamethasone and methyl prednisolone are equally effective in the management of COVID 19.

Characteristics of Persons With Secondary Detection of Severe Acute Respiratory Syndrome Coronavirus 2 ≥90 days After First Detection, New Mexico 2020.

The New Mexico Department of Health (NMDOH) conducted a matched case-control study to compare 315 persons (cases) with and 945 persons (controls) without severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) secondary detection (ie, positive SARS-CoV-2 test ≥90 days after first detection as of December 10, 2020). Compared with controls, cases had greater odds of higher SARS-CoV-2 testing frequency (adjusted odds ratio [aOR] = 1.2), being female (aOR = 1.6), being non-Hispanic American Indian/Alaska Native (aOR = 2.3), having diabetes mellitus (aOR = 1.8), and residing and/or working in detention and/or correctional facilities (aOR = 4.7). Diagnostic tools evaluating infectiousness at secondary detection are urgently needed to inform infection control practices.

Dissemination of Cardiopulmonary Resuscitation Training for Nurses Treating Coronavirus Disease-2019 Patients: A Single-arm Pre-experimental Study.

Introduction: Providing cardiopulmonary resuscitation (CPR) to a coronavirus disease-2019 (COVID-19) patient is challenging for the healthcare providers (HCP). COVID-19 cases have specific CPR requirements, which differ from standard resuscitation efforts. Objective: This paper aims to evaluate whether online learning can be an effective methodology for imparting information on the management of cardiopulmonary arrest in COVID-19 patients due to a novel virus unknown to HCP based on evaluation of knowledge acquisition and satisfaction of the nurses in the new area of medicine and virology. Methodology: In a single-arm, one group before and after design, from a cohort of 160 nurses trained in comprehensive cardiac life support (CCLS) formulated by the Indian Resuscitation Council (IRC), 73 nurses participated in the study. After obtaining informed consent through the email from the nurses, baseline data including demographic profile and knowledge related to CPR in COVID-19 patients were collected. An online intervention spread over 1 week was given using a validated e-learning module. The online intervention was found to be effective (pre- and post-intervention knowledge score 13.65 ± 3.01 vs 19.92 ± 1.94, p = 0.001). The majority of nurses were highly satisfied with the content and the training methodology (37.23 ± 4.70). Conclusion: A well-structured, online study material can be used in imparting knowledge and demonstrating the basic and essential skills to nurses, required for giving CPR to COVID-19 patients. How to cite this article: Joshi P, Das S, Thomas M, Mawar S, Garg R, Shariff A, et al. Dissemination of Cardiopulmonary Resuscitation Training for Nurses Treating Coronavirus Disease-2019 Patients: A Single-arm Pre-experimental Study. Indian J Crit Care Med 2022;26(3):327-330.

E-Cigarettes: Harmful or Harm-Reducing? Evaluation of a Novel Online CME Program for Health Care Providers.

AIM: Patients are asking health care providers about e-cigarettes, vaping, and other electronic nicotine delivery systems (ENDS). Provider advice on ENDS has varied greatly, suggesting a need for evidence-based continuing medical education (CME). SETTING: A novel free online CME course was developed on ENDS risks and benefits, product types (e.g., vape pens, pods), and screening and counseling best practices for adults, adolescents, and different smoker profiles (e.g., daily, social). PARTICIPANTS: From January 2017 through June 2018, 1061 individuals accessed the course: 46% physicians, 7% physician assistants, 7% nurse practitioners, 15% nurses, 4% pharmacists, and 28% allied health/student/other; 41% were international. PROGRAM DESCRIPTION: The course was built from observed online patient-provider interactions. Through video role-plays, expert interviews, and interactive activities, the course engaged learners in the evidence on ENDS. Completers earned 1.5 CME units. PROGRAM EVALUATION: A total of 555 health care providers earned 832.5 CME units. Pre- to post-test scores significantly increased from 57 to 90%; 76% rated the course as above average (41%) or outstanding (35%); 99% indicated the course was free of commercial bias. DISCUSSION: Addressing the growing need for balanced provider education on ENDS, this interactive online CME engaged learners and increased knowledge on devices and evidence-based cessation approaches.

Therapeutic potential of ivermectin as add on treatment in COVID 19: A systematic review and meta-analysis.

The current management of COVID-19 is mostly limited to general supportive care and symptomatic treatment. Ivermectin is a broad-spectrum anti-parasitic drug used widely for the treatment of onchocerciasis and lymphatic filariasis. Apart from its anti-parasitic effect it also exhibits antiviral activity against a number of viruses both in vitro and in vivo. Hence, we conducted this systematic review and meta-analysis to assess the currently available data on the therapeutic potential of ivermectin for the treatment of COVID-19 as add on therapy. A total of 629 patients were included in the 4 studies and all were COVID-19 RT-PCR positive. Among them, 397 patients received ivermectin along with usual therapy. The random effect model showed the overall pooled OR to be 0.53 (95%CI: 0.29 to0.96) for the primary outcome (all-cause mortality) which was statistically significant (P=0.04). Similarly, the random effect model revealed that adding ivermectin led to significant clinical improvement compared to usual therapy (OR=1.98, 95% CI: 1.11 to 3.53, P=0.02).  However, this should be inferred cautiously as the quality of evidence is very low. Currently, many clinical trials are on-going, and definitive evidence for repurposing this drug for COVID-19 patients will emerge only in the future.

Excessive Spending in a "Free-to-Play" Smartphone Game.

BACKGROUND AND OBJECTIVES: Although gaming disorder is increasingly recognized, there has been limited consideration of the impact of free-to-play games with in-game purchases, also called microtransactions. METHODS: Case report (n = 1). RESULTS: A patient with posttraumatic stress disorder, major depressive disorder in remission, polysubstance use disorders in remission, and opioid use disorder on buprenorphine/naloxone developed gaming disorder (based on International Classification of Diseases, 11th revision) and spent up to 40% of his monthly income on microtransactions within a smartphone game. Treatment consisted of relinquishing access to electronic payment mechanisms, motivational interviewing, and couples therapy. DISCUSSION AND CONCLUSIONS: The case highlights how in-game purchases may cause patients with gaming disorder to experience financial consequences, and the need for further characterization of the clinical ramifications of microtransactions. SCIENTIFIC SIGNIFICANCE: To the best of our knowledge, this is the first report in the medical literature of a patient with gaming disorder developing excessive in-game spending from expenditures on microtransactions. (Am J Addict 2020;29:528-530).

Effectiveness of Convalescent Plasma Therapy in the Treatment of Moderate to Severe COVID 19 Patients: A Systematic Review and Meta-Analysis.

Though Convalescent plasma therapy (CPT) is being used for management of COVID-19, the evidence is still equivocal. So, we carried out this study to evaluate the currently available data to provide evidence about CPT in COVD-19 patients. RCTs and observational studies with sample size with more than 5 were included in the analysis. Out of 196 studies, 12 studies were selected for systematic review and meta-analysis was carried out for 6 studies having a control arm. For dichotomous values, risk ratio (RR) and 95% confidence interval was expressed. MAIN OUTCOMES: All-cause mortality, clinical improvement by day 7 and viral detection by day 7 were the defined outcome measures before starting of data extraction. RESULT: For 6 studies (2 RCTs and 4 observational studies) with 474 patients, the overall pooled RR for all-cause mortality was 0.61 (95%CI: 0.37 to 0.99. P= 0.04). Only RCTs and only observational studies for all-cause mortality showed pooled RR of 0.60 (95% CI: 0.33 to 1.10, P=0.10) and 0.48 (95% CI: 0.17 to 1.36, P= 0.17) respectively. There was risk of bias in the studies due to randomization process and confounding. Sensitivity analysis was carried out only for observational studies. The overall pooled RR for clinical improvement by day 7 and viral detection by day 7 were 1.12 (95%CI: 0.96 to 1.31, P=0.16) and 0.19 (95%CI: 0.09 to 0.60, P < 0.0001). CONCLUSION AND RELEVANCE: Though the review suggests modest utility of CPT in reducing all-cause mortality, improving clinical outcome, and early viral clearance, it should be interpreted cautiously.

A Descriptive Study to Assess the Knowledge, Attitude, Practices, Perceived Barriers, and Support Regarding Palliative Care and End-of-Life Care among Critical Care Nurses of Tertiary Care Medical Institute.

INTRODUCTION: The role of nurses working in critical care setup is one of the key members for optimal provision of the palliative care and end-of-life care (EOLC). This study aims to assess knowledge, attitude, practices, and perceived barriers regarding palliative and EOLC among critical care nurses. METHODS: A cross-sectional study was conducted among 386 critical care nurses of a tertiary care center, selected through convenience sampling technique. Data related to knowledge, attitude, practices, and perceived barriers were collected using a standardized tool. The collected data were analyzed using SPSS version 24. RESULTS: We observed that mean knowledge, attitude, and practice scores of critical care nurses were 9.83 ± 2.50 of 20, 104.91 ± 13.04 of 150, and 17.61 ± 4.36 of 27, respectively. One of the major barriers in providing palliative and EOLC was being called for attending to other newly admitted patients. The mean knowledge score of nurses was found to be significantly higher among female nurses (P = 0.016). Nurses who had additional training pertaining to palliative and EOLC had significantly higher mean knowledge (P = 0.001) and attitude scores (P = 0.02) than those who did not have such training. The mean practice (P = 0.02) and attitude score (P = 0.002) was significantly higher among postgraduate nurses as compared to nurses with lower qualification. The mean attitude score was significantly higher among nurses of lower age group (P = 0.04) along with a negative correlation of age of nurses with attitude (P = 0.01). Nurses' practice was positively correlated to the attitude (P = 0.001) and knowledge (P = 0.01). CONCLUSION: There is a considerable gap between critical care nurses' knowledge and practices, in spite of having a good attitude regarding palliative and EOLC. This necessitates the strengthening of both preservice and in-service education for nurses.

Dye Coupled Aptamer-Captured Enzyme Catalyzed Reaction for Detection of Pan Malaria and P. falciparum Species in Laboratory Settings and Instrument-Free Paper-Based Platform.

Malaria diagnosis methods offering species-specific information on the causative parasites, along with their flexibility to use in different resource settings, have great demand for precise treatment and management of the disease. Herein, we report the detection of pan malaria and P. falciparum species using a dye-based reaction catalyzed by the biomarker enzymes Plasmodium lactate dehydrogenase ( PLDH) and Plasmodium falciparum glutamate dehydrogenase ( PfGDH), respectively, through instrument-based and instrument-free approaches. For the detection, two ssDNA aptamers specific to the corresponding PLDH and PfGDH were used. The aptamer-captured enzymes were detected through a substrate-dependent reaction coupled with the conversion of resazurin (blue, ∼λ605nm) to resorufin (pink, ∼λ570nm) dye. The reaction was monitored by measuring the fluorescence intensity at λ660nm for resorufin, absorbance ratio (λ570nm/λ605nm), and change in color (blue to pink). The detection approach could be customized to a spectrophotometer-based method and an instrument-free device. For both the approaches, the biomarkers were captured from the serum samples with the help of aptamer-coated magnetic beads prior to the analysis to exclude potential interferences from the serum. In the instrument-free device, a medical syringe (5 mL) prefabricated with a magnet was used for in situ separation of the enzyme-captured beads from the reaction supernatant. The converted dye in the supernatant was then efficiently adsorbed over a DEAE cellulose-treated paper wick assembled in the syringe hose. The biomarkers could be detected by both qualitative and quantitative format following the color and pixel intensity, respectively, developed on the paper surface. The developed method and technique offered detection of the biomarkers within a clinically relevant dynamic range, with the limit of detection values in the picomolar level. Flexible detection capability, low cost, interference-free detections, and portable nature (for instrument-free devices) are the major advantages offered by the developed approaches.

Simultaneous Quantification of Plasmodium Antigens and Host Factor C-Reactive Protein in Asymptomatic Individuals with Confirmed Malaria by Use of a Novel Multiplex Immunoassay.

Malaria rapid diagnostic tests (RDTs) primarily detect Plasmodium falciparum antigen histidine-rich protein 2 (HRP2) and the malaria-conserved antigen lactate dehydrogenase (LDH) for P. vivax and other malaria species. The performance of RDTs and their utility is dependent on circulating antigen concentration distributions in infected individuals in a population in which malaria is endemic and on the limit of detection of the RDT for the antigens. A multiplexed immunoassay for the quantification of HRP2, P. vivax LDH, and all-malaria LDH (pan LDH) was developed to accurately measure circulating antigen concentration and antigen distribution in a population with endemic malaria. The assay also measures C-reactive protein (CRP) levels as an indicator of inflammation. Validation was conducted with clinical specimens from 397 asymptomatic donors from Myanmar and Uganda, confirmed by PCR for infection, and from participants in induced blood-stage malaria challenge studies. The assay lower limits of detection for HRP2, pan LDH, P. vivax LDH, and CRP were 0.2 pg/ml, 9.3 pg/ml, 1.5 pg/ml, and 26.6 ng/ml, respectively. At thresholds for HRP2, pan LDH, and P. vivax LDH of 2.3 pg/ml, 47.8 pg/ml, and 75.1 pg/ml, respectively, and a specificity ≥98.5%, the sensitivities for ultrasensitive PCR-confirmed infections were 93.4%, 84.9%, and 48.9%, respectively. Plasmodium LDH (pLDH) concentration, in contrast to that of HRP2, correlated closely with parasite density. CRP levels were moderately higher in P. falciparum infections with confirmed antigenemia versus those in clinical specimens with no antigen. The 4-plex array is a sensitive tool for quantifying diagnostic antigens in malaria infections and supporting the evaluation of new ultrasensitive RDTs.

Correction to: Performance of an ultra-sensitive Plasmodium falciparum HRP2-based rapid diagnostic test with recombinant HRP2, culture parasites, and archived whole blood samples.

Following publication of the original article [1], the authors flagged an error concerning a reference to a product in the Methods section.

Operational Performance of a Plasmodium falciparum Ultrasensitive Rapid Diagnostic Test for Detection of Asymptomatic Infections in Eastern Myanmar.

In the Greater Mekong Subregion in Southeast Asia, malaria elimination strategies need to target all Plasmodium falciparum parasites, including those carried asymptomatically. More than 70% of asymptomatic carriers are not detected by current rapid diagnostic tests (RDTs) or microscopy. An HRP2-based ultrasensitive RDT (uRDT) developed to improve the detection of low-density infections was evaluated during prevalence surveys within a malaria elimination program in a low-transmission area of eastern Myanmar. Surveys were conducted to identify high-prevalence villages. Two-milliliter venous blood samples were collected from asymptomatic adult volunteers and transported to the laboratory. Plasmodium parasites were detected by RDT, uRDT, microscopy, ultrasensitive qPCR (uPCR), and multiplex enzyme-linked immunosorbent assay (ELISA). The sensitivity, specificity, and predictive positive and negative values of RDT and uRDT were calculated compared to uPCR and ELISA. Parasite and antigen concentrations detected by each test were defined using uPCR and ELISA, respectively. A total of 1,509 samples, including 208 P. falciparum-positive samples were analyzed with all tests. The sensitivity of the uRDT was twofold higher than that of RDT, 51.4% versus 25.2%, with minor specificity loss, 99.5% versus 99.9%, against the combined reference (uPCR plus ELISA). The geometric mean parasitemia detected by uRDT in P. falciparum monospecific infections was 3,019 parasites per ml (95% confidence interval [95% CI], 1,790 to 5,094; n = 79) compared to 11,352 parasites per ml (95% CI, 5,643 to 22,837; n = 38) by RDT. The sensitivities of uRDT and RDT dropped to 34.6% and 15.1%, respectively, for the matched tests performed in the field. The uRDT performed consistently better than RDT and microscopy at low parasitemias. It shows promising characteristics for the identification of high-prevalence communities and warrants further evaluation in mass screening and treatment interventions.

Performance of an ultra-sensitive Plasmodium falciparum HRP2-based rapid diagnostic test with recombinant HRP2, culture parasites, and archived whole blood samples.

BACKGROUND: As malaria endemic countries shift from control to elimination, the proportion of low density Plasmodium falciparum infections increases. Current field diagnostic tools, such as microscopy and rapid diagnostic tests (RDT), with detection limits of approximately 100-200 parasites/µL (p/µL) and 800-1000 pg/mL histidine-rich protein 2 (HRP2), respectively, are unable to detect these infections. A novel ultra-sensitive HRP2-based Alere™ Malaria Ag P.f RDT (uRDT) was evaluated in laboratory conditions to define the test's performance against recombinant HRP2 and native cultured parasites. RESULTS: The uRDT detected dilutions of P. falciparum recombinant GST-W2 and FliS-W2, as well as cultured W2 and ITG, diluted in whole blood down to 10-40 pg/mL HRP2, depending on the protein tested. uRDT specificity was 100% against 123 archived frozen whole blood samples. Rapid test cross-reactivity with HRP3 was investigated using pfhrp2 gene deletion strains D10 and Dd2, pfhrp3 gene deletion strain HB3, and controls pfhrp2 and pfhrp3 double deletion strain 3BD5 and pfhrp2 and pfhrp3 competent strain ITG. The commercial Standard Diagnostics, Inc. BIOLINE Malaria Ag P.f RDT (SD-RDT) and uRDT detected pfhrp2 positive strains down to 49 and 3.13 p/µL, respectively. The pfhrp2 deletion strains were detected down to 98 p/µL by both tests. CONCLUSION: The performance of the uRDT was variable depending on the protein, but overall showed a greater than 10-fold improvement over the SD-RDT. The uRDT also exhibited excellent specificity and showed the same cross-reactivity with HRP3 as the SD-RDT. Together, the results support the uRDT as a more sensitive HRP2 test that could be a potentially effective tool in elimination campaigns. Further clinical evaluations for this purpose are merited.

A new highly sensitive enzyme-linked immunosorbent assay for the detection of Plasmodium falciparum histidine-rich protein 2 in whole blood.

BACKGROUND: The detection of submicroscopic infections in low prevalence settings has become an increasingly important challenge for malaria elimination strategies. The current field rapid diagnostic tests (RDTs) for Plasmodium falciparum malaria are inadequate to detect low-density infections. Therefore, there is a need to develop more sensitive field diagnostic tools. In parallel, a highly sensitive laboratory reference assay will be essential to evaluate new diagnostic tools. Recently, the highly sensitive Alere™ Malaria Ag P.f ELISA (HS ELISA) was developed to detect P. falciparum histidine-rich protein 2 (HRP2) in clinical whole blood specimens. In this study, the analytical and clinical performance of the HS ELISA was determined using recombinant P. falciparum HRP2, P. falciparum native culture parasites, and archived highly pedigreed clinical whole blood specimens from Karen village, Myanmar and Nagongera, Uganda. RESULTS: The HS ELISA has an analytical sensitivity of less than 25 pg/mL and shows strong specificity for P. falciparum HRP2 when tested against P. falciparum native culture strains with pfhrp2 and pfhrp3 gene deletions. Additionally, the Z'-factor statistic of 0.862 indicates the HS ELISA as an excellent, reproducible assay, and the coefficients of variation for inter- and intra-plate testing, 11.76% and 2.51%, were acceptable. Against clinical whole blood specimens with concordant microscopic and PCR results, the HS ELISA showed 100% (95% CI 96.4-100) diagnostic sensitivity and 97.9% (95% CI 94.8-99.4) diagnostic specificity. For P. falciparum positive specimens with HRP2 concentrations below 400 pg/mL, the sensitivity and specificity were 100% (95% CI 88.4-100) and 88.9% (95% CI 70.8-97.6), respectively. The overall sensitivity and specificity for all 352 samples were 100% (CI 95% 96-100%) and 97.3% (CI 95% 94-99%). CONCLUSIONS: The HS ELISA is a robust and reproducible assay. The findings suggest that the HS ELISA may be a useful tool as an affordable reference assay for new ultra-sensitive HRP2-based RDTs.