Accuracy of automated analyzers for the estimation of CSF cell counts: A systematic review and meta-analysis.

This systematic review evaluates the evidence for accuracy of automated analyzers that estimate cerebrospinal fluid (CSF) white blood cell counts (WBC) compared to manual microscopy. Inclusion criteria of original research articles included human subjects, English language, and manual microscopy comparator. PUBMED, EMBASE and Cochrane Review databases were searched through 2019 and QUADAS-2 Tool was used for assessment of bias. Data were pooled and analyzed by comparison method, using random effects estimation. Among 652 titles, 554 abstracts screened, 104 full-text review, 111 comparisons from 41 studies were included. Pooled estimates of sensitivity and specificity (n = 7) were 95% (95%-CI 93%-97%) and 84% (95%-CI: 64%-96%), respectively. Pooled R2 estimates (n = 29) were 0.95 (95%-CI: 0.95-0.96); Pooled spearman rho correlation (n = 27) estimates were 0.95 (95% CI 0.95-0.96). Among those comparisons using Bland-Altman analysis (n = 11) pooled mean difference was estimated at 0.98 (95% CI-0.54-2.5). Among comparisons using Passing-Bablok regressions (n = 14) the pooled slope was estimated to be 1.05 (95% CI 1.03-1.07). Q tests of homogeneity were all significant with the exception of the Bland-Altman comparisons (I2 10%, p value 0.35). There is good overall accuracy for CSF WBC by automated hematologic analyzers. These findings are limited by the small sample sizes and inconsistent validation methodology in the reviewed studies.

Prolonged Unconsciousness is Common in COVID-19 and Associated with Hypoxemia.

OBJECTIVE: The purpose of this study was to estimate the time to recovery of command-following and associations between hypoxemia with time to recovery of command-following. METHODS: In this multicenter, retrospective, cohort study during the initial surge of the United States' pandemic (March-July 2020) we estimate the time from intubation to recovery of command-following, using Kaplan Meier cumulative-incidence curves and Cox proportional hazard models. Patients were included if they were admitted to 1 of 3 hospitals because of severe coronavirus disease 2019 (COVID-19), required endotracheal intubation for at least 7 days, and experienced impairment of consciousness (Glasgow Coma Scale motor score <6). RESULTS: Five hundred seventy-one patients of the 795 patients recovered command-following. The median time to recovery of command-following was 30 days (95% confidence interval [CI] = 27-32 days). Median time to recovery of command-following increased by 16 days for patients with at least one episode of an arterial partial pressure of oxygen (PaO2 ) value ≤55 mmHg (p < 0.001), and 25% recovered ≥10 days after cessation of mechanical ventilation. The time to recovery of command-following  was associated with hypoxemia (PaO2 ≤55 mmHg hazard ratio [HR] = 0.56, 95% CI = 0.46-0.68; PaO2 ≤70 HR = 0.88, 95% CI = 0.85-0.91), and each additional day of hypoxemia decreased the likelihood of recovery, accounting for confounders including sedation. These findings were confirmed among patients without any imagining evidence of structural brain injury (n = 199), and in a non-overlapping second surge cohort (N = 427, October 2020 to April 2021). INTERPRETATION: Survivors of severe COVID-19 commonly recover consciousness weeks after cessation of mechanical ventilation. Long recovery periods are associated with more severe hypoxemia. This relationship is not explained by sedation or brain injury identified on clinical imaging and should inform decisions about life-sustaining therapies. ANN NEUROL 2022;91:740-755.

Clinical Significance of Positive Results of the BioFire Cerebrospinal Fluid FilmArray Meningitis/Encephalitis Panel at a Tertiary Medical Center in the United States.

CONTEXT.­: The FilmArray Meningitis/Encephalitis (ME) panel is the first US Food and Drug Administration-cleared multiplex polymerase chain reaction panel for the detection of central nervous system infections. While the assay's performance characteristics have been described, the real-world significance of positive results has not been fully characterized. OBJECTIVE.­: To evaluate the clinical significance of positive ME panel results in a tertiary care medical center in New York, New York. DESIGN.­: Four physicians independently performed retrospective clinical assessments of all positive ME panel results at Columbia University Irving Medical Center, including the Children's Hospital of New York, during an 18-month period. Each reviewer determined the likelihood of central nervous system infection for all cases and whether cases fit Brighton diagnostic criteria for meningitis, encephalitis, or meningoencephalitis. RESULTS.­: Among 119 cases, there was 75% positive agreement (95% CI, 54%-89%) between ME panel results and clinical consensus, which varied among panel targets. CONCLUSIONS.­: The ME panel showed good agreement with expert clinical consensus for patients presenting with acute meningitis/encephalitis. Factors contributing to clinically insignificant ME positive results included low pretest probability, traumatic lumbar puncture, specimen contamination, and detection of incidental viral targets such as human herpesvirus 6. Notably, the ME panel detected more than twice the number of cases of bacterial meningitis detected by culture alone, particularly among patients receiving empiric antimicrobial therapy before lumbar puncture. Appropriate test use and contextual interpretation of results are critical to leveraging the advantages of the platform while avoiding potential pitfalls.

Hispanic/Latino-Serving Hospitals Provide Less Targeted Temperature Management Following Out-of-Hospital Cardiac Arrest.

Background Variation exists in outcomes following out-of-hospital cardiac arrest (OHCA), but whether racial and ethnic disparities exist in post-arrest provision of targeted temperature management (TTM) is unknown. Methods and Results We performed a retrospective analysis of a prospectively collected cohort of patients who survived to admission following OHCA from the Cardiac Arrest Registry to Enhance Survival, whose catchment area represents ~50% of the United States from 2013-2019. Our primary exposure was race/ethnicity and primary outcome was utilization of TTM. We built a mixed-effects model with both state of arrest and admitting hospital modeled as random intercepts to account for clustering. Among 96,695 patients (24.6% Black, 8.0% Hispanic/Latino, 63.4% White), a smaller percentage of Hispanic/Latino patients received TTM than Black or White patients (37.5% vs. 45.0 % vs 43.3%, P < .001) following OHCA. In the mixed-effects model, Black patients (Odds Ratio [OR] 1.153, 95% Confidence Interval [CI] 1.102-1.207, P < .001) and Hispanic/Latino patients (OR 1.086, 95% CI 1.017-1.159, P < .001) were slightly more likely to receive TTM compared to White patients, perhaps due to worse admission neurological status. We did find community level disparity as Hispanic/Latino-serving hospitals (defined as the top decile of hospitals that cared for the highest proportion of Hispanic/Latino patients) provided less TTM (OR 0.587, 95% CI 0.474 to 0.742, P < .001). Conclusions Reassuringly, we did not find evidence of intrahospital or interpersonal racial or ethnic disparity in the provision of TTM. However, we did find inter-hospital, community level disparity. Hispanic/Latino-serving hospitals provided less guideline-recommended TTM after OHCA.

The Association Between Magnetic Resonance Imaging Disc Pathology and Provocative Discography at the Lumbar Level.

OBJECTIVE: The objective of this study was to show that degenerative lumbar magnetic resonance imaging findings variably increase discography pain by level. METHODS: Lumbar discography and magnetic resonance imaging of 736 patients were retrospectively reviewed. Univariate/multivariate logistic regressions calculated the odds ratio (OR) (95% confidence interval, P < 0.05). RESULTS: L3-4 multivariate regression OR for a degenerative disc is 9.9; for bulge, 10.9; for annular tear, 38.9; for herniation, 51.5; and for degenerative facet, 2.158. Endplate changes were not significant. L4-5 OR for a degenerative disc is 4.52; for bulge, 13.74, for tear, 19.13; for herniation, 28.65; for endplate edema, 3.47; and fatty change, 3.84. Degenerative facet ORs were not significant. L5-S1 OR for a degenerative disc is 6.86; for bulge, 5.65; for tear, 40.56; and for herniation, 77.98. Endplate changes and degenerative facet OR's were not significant. CONCLUSIONS: Advancing degeneration increases pain at L5-S1 followed by L3-4. Endplate signal is significant only at L4-5.

Cerebrospinal Analysis in Patients With COVID-19.

BACKGROUND: Assessment of the impact of cerebrospinal fluid (CSF) analysis including investigation for the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential for the optimization of patient care. METHODS: In this case series, we review patients diagnosed with SARS-CoV-2 undergoing lumbar puncture (LP) admitted to Columbia University Irving Medical Center (New York, NY, USA) from March 1 to May 26, 2020. In a subset of patients, CSF SARS-CoV-2 quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) testing is performed. RESULTS: The average age of 27 patients who underwent LP with definitive SARS-CoV-2 (SD) was 37.5 (28.7) years. CSF profiles showed elevated white blood cell counts and protein in 44% and 52% of patients, respectively. LP results impacted treatment decisions in 10 (37%) patients, either by change of antibiotics, influence in disposition decision, or by providing an alternative diagnosis. CSF SARS-CoV-2 qRT-PCR was performed on 8 (30%) patients, with negative results in all samples. CONCLUSIONS: Among patients diagnosed with SARS-CoV-2, CSF results changed treatment decisions or disposition in over one-third of our patient cohort. CSF was frequently abnormal, though CSF SARS-CoV-2 qRT-PCR was negative in all samples. Further studies are required to define whether CSF SARS-CoV-2 testing is warranted in certain clinical contexts.

Prevalence and Characteristics of Neuroinfectious Disease Inquiries Within the Emerging Infections Network: A 22-Year Retrospective Study.

BACKGROUND: To monitor emerging infectious diseases, the Centers for Disease Control and Prevention and the Infectious Disease Society of America established the Emerging Infections Network (EIN), allowing infectious disease specialists to post inquiries about clinical cases. We describe the frequency and characteristics of neuroinfectious disease-related inquiries. METHODS: The EIN listserv was retrospectively reviewed from February 1997 to December 2019 using search terms associated with neurologic diseases. We recorded case summaries, disease type (ie, meningitis, encephalitis), inquiry type (diagnostic approach, result interpretation, management decisions), unique patient populations, exposures, pathogens, ultimate diagnosis, and change in clinical care based on responses. RESULTS: Of 2348 total inquiries, 285 (12.1%) related to neuroinfectious diseases. The majority involved meningitis (99, 34.7%) or encephalitis (56, 19.6%). One hundred fifteen inquiries (40%) related to management, 34 (12%) related to diagnostic workup, and 22 (8%) related to result interpretation. Eight (2.8%) specifically involved results of cerebrospinal fluid polymerase chain reaction testing. Sixty-three (22.1%) involved immunosuppressed patients (29 human immunodeficiency virus-positive cases [46%]). The most common pathogens were Treponema pallidum (19, 6.7%) and Cryptococcus neoformans (18, 6.3%). In 74 (25%) inquiries, patients had neurologic symptoms without a clear infection, 38 (51.3%) of which included noninfectious neurologic etiologies in the differential diagnosis. CONCLUSIONS: This study demonstrates the significant challenges of diagnosis and management of neuroinfectious diseases within the field of infectious diseases. It also highlights the importance of curated forums to guide the approach of difficult cases, in particular instances that mimic infectious diseases. Finally, the EIN listserv may assist in identifying areas for research and training to address these complexities.

Positive Predictive Values of Lumbar Spine Magnetic Resonance Imaging Findings for Provocative Discography.

PURPOSE: The purpose of this study was to calculate the positive predictive value (PPV) of lumbar spine magnetic resonance imaging (MRI) findings for a painful disc using provocative discography. MATERIALS AND METHODS: Lumbar spine discography records and prediscography MRIs of 736 patients (2457 discs) who underwent discography for diagnostic purposes from 2003 to 2007 were retrospectively reviewed in an Institutional Review Board-exempt and Health Insurance Portability and Accountability Act-compliant protocol. Each level was identified as having high-intensity zone (HIZ) disc, disc protrusion, disc extrusion, or combination (any herniation type), disc bulge, disc degeneration, and spondylolisthesis. Statistical analysis used a 2 × 2 contingency table of significant discography results for each of the MRI variables to calculate P value and PPV with a confidence interval from a binomial distribution. RESULTS: An HIZ disc has a PPV of 0.71 (0.65-0.76, P = 4.31E - 44) for a provocative discography. A disc protrusion has a PPV of 0.79 (0.73-0.83, P = 2.68E - 53). A disc extrusion has a PPV of 0.93 (0.79-0.98, P = 1.34E - 14), a bulge of 0.43 (0.37-0.48, P = 0.002), and a degenerative disc of 0.32 (0.28-0.35, P = 0.08), and spondylolisthesis has a PPV of 0.67 (0.59-0.73, P = 1.70E - 20). A herniation of either type (extrusion or protrusion) has a PPV of 0.80 (0.75-0.84, P = 5.86E - 69). CONCLUSIONS: Disc herniations and HIZ discs have high predictive value in identifying a pain generator. An extruded disc herniation has the highest PPV for discogenic pain.

HIV-specific T-cell Responses and Generalized Activation in HIV-1 Infected Long-term Non-progressors and Progressors from South India.

BACKGROUND: Anti-viral cytokine expressions by cytotoxic T-cells and lower activation rates have been reported to correlate with suppressed HIV replication in long-term non-progressors (LTNP). Immune mechanisms underlying disease non-progression in LTNP might vary with HIV-1 subtype and geographical locations. OBJECTIVE: This study evaluates cytokine expression and T-cells activation in relation to disease non-progression in LTNP. METHODS: HIV-1 Subtype C infected LTNP (n=20) and progressors (n=15) were enrolled and flowcytometry assays were performed to study HIV-specific CD8 T-cells expressing IL-2, IFN-γ, TNF-α and MIP-1ß against gag and env peptides. CD4+ T-cell activation was evaluated by surface expression of HLADR and CD38. RESULTS: Proportions of cytokines studied did not differ significantly between LTNP and progressors, while contrasting correlations with disease progression markers were observed in LTNP. CD4+ T-cell activation rates were significantly lower in LTNP compared to progressors which indicate the potential role of T-cell activation rates in disease non-progression in LTNP. CONCLUSION: LTNP and progressors showed similar CD8+ T-cell responses, but final conclusions can be drawn only by comparing multiple immune factors in larger LTNP cohort with HIV-1 infected individuals at various levels of disease progression. A possible role of HIV-1 subtype variation and ethnic differences in addition to host-genetic and viral factors cannot be ruled out.

Incretin-based therapy in type 2 diabetes: An evidence based systematic review and meta-analysis.

Incretin based therapies such as dipeptidyl peptidase-4 inhibitors (DPP-4i) and glucagon-like peptide-1 receptor agonists (GLP-1Ra) are increasingly used for the treatment of Type 2 diabetes mellitus. In clinical practice and in previously performed clinical trials, these agents are often used in combination with other oral anti-diabetic agents (OADs) and Insulin. Prior meta-analytic reviews however do not adequately address the impact of background therapy and active comparator arms. Accordingly, we aimed to further investigate the efficacy of incretin based therapies by updating existing reviews by including clinical trial evidence after 2008; estimating the pooled effect of incretin therapies on glycemic efficacy and weight-loss, stratified by comparator therapy (placebo, mono-therapy, etc.), estimating the impact of background OADs and within class (GLP-1Ra or DPP-4i) comparative efficacy, on glycemia control. 82 randomized controlled trials after 2008 with glycemic control and weight loss as primary end-points were included. Both DPP-4i and GLP-1Ra reduced HbA1c, but only GLP-1Ra caused weight loss when compared to either active comparator drugs or placebo. GLP-1Ra were more effective than DPP-4i in glycemia lowering. Long acting GLP-1Ra were more effective in HbA1c lowering than short-acting agents but with similar weight loss effect. The effect of DPP-4i incretin glycemic efficacy was not modified by background therapy used in the study.

The impact of a private-public partnership delivery system on the HIV continuum of care in a South Indian city.

We characterized the impact of a Private-Public Partnership (PPP) on the continuum of HIV care (e.g., treatment initiation, ART effectiveness and loss to follow-up) among adults enrolled at a private hospital/ART link center in the southern state of Karnataka, India from 2007 through 2012. Data on 2326 adults in care were compiled using an electronic database supplemented with medical chart abstraction. Survival methods with staggered entries were used to analyze time to ART initiation and loss to follow-up as well as associated factors. Mixed effects linear regression models were used to assess ART effectiveness. The mean age of adults in care was 36 years; 40% were male. The majority were married, had less than primary education, and less than 45 US dollars (3000 Indian Rupee) monthly income. The mean CD4 at presentation was 527 cells/mm3. The median time from ART eligibility to initiation was 5 and 2 months for before and after the PPP, respectively (p < 0.001). Becoming eligible after PPP was associated with more rapid treatment initiation (Hazard Ratio: [95% Confidence Interval] 1.49 [1.11, 1.99]). Moreover, among the 1639 persons lost to follow-up, more rapid loss was observed before the PPP (12.77 months) vs. after (13.37 months) (p = 0.25) and there was a significant interaction between ART status and calendar time before and after the PPP (p < 0.001). Being on treatment was associated with a lower likelihood of becoming lost before the PPP (HR: [95% CI] 0.33 [0.27, 0.42]), but this association was reversed after the PPP (HR: [95% CI] 1.77 [1.54, 2.04]), p-value for interaction <0.001. Treatment response measured by CD4 was comparable before and after the PPP (p = 0.088). Our findings suggest that PPP models of ART delivery may improve HIV treatment initiation and loss to follow-up without compromising the effectiveness of treatment. Efforts to expand these system-level interventions should be considered with on-going evaluation.

Barriers to and Facilitators of Stocking Healthy Food Options: Viewpoints of Baltimore City Small Storeowners.

Receptivity to strategies to improve the food environment by increasing access to healthier foods in small food stores is underexplored. We conducted 20 in-depth interviews with small storeowners of different ethnic backgrounds as part of a small-store intervention trial. Store owners perceived barriers and facilitators to purchase, stock, and promote healthy foods. Barriers mentioned included customer preferences for higher fat and sweeter taste and for lower prices; lower wholesaler availability of healthy food; and customers' lack of interest in health. Most store owners thought positively of taste tests, free samples, and communication interventions. However, they varied in terms of their expectations of the effect of these strategies on customers' healthy food purchases. The findings reported add to the limited data on motivating and working with small-store owners in low-income urban settings.

Stable patients and patients with advanced disease: consensus definitions to support sustained scale up of antiretroviral therapy.

OBJECTIVE: As guidelines are evolving towards recommending starting antiretroviral therapy (ART) in all HIV-positive individuals irrespective of clinical and immunological status, HIV programmes will be challenged to manage an increasingly diverse set of patient needs. To support global guideline recommendations for differentiated service delivery, WHO developed consensus definitions for two distinct patient populations: patients presenting with advanced disease and patients who are stable on ART. METHODS: An expert panel consisting of 73 respondents from 28 countries across all six WHO regions supported the development of these definitions. The panel included clinicians, researchers, programme managers, technical advisors and patient group representatives. RESULTS: Patients presenting with advanced disease at presentation to care were defined as CD4 count <200 CD4 cells/mm(3) or WHO Stage III & IV defining illness. Patients stable on ART were defined as those who were receiving ART for at least 1 year with no adverse drug reactions requiring regular monitoring, no current illnesses or pregnancy, a good understanding of lifelong adherence, and evidence of treatment success. Treatment success was defined as two consecutive undetectable viral load measures or, in the absence of viral load monitoring, rising CD4 counts or CD4 counts above 200 cells/mm(3) and an objective adherence measure. CONCLUSIONS: Patients who are stable on ART should be offered a less intensive care package that can lead to improved outcomes while saving resources, including less frequent clinic visits, out-of-clinic drug refills and reduced laboratory monitoring. This will allow for clinic resources to be directed towards reducing morbidity and mortality among patients presenting with advanced disease.

Incretin-Based Therapy for Diabetes: What a Cardiologist Needs to Know.

Incretin-based therapies are effective glucose-lowering drugs that have an increasing role in the treatment of type 2 diabetes because of their efficacy, safety, and ease of use. Both glucagon-like peptide-1 receptor agonists and dipeptidyl peptidase-4 inhibitors are commonly used for glycemic control as adjuncts to metformin, other oral antiglycemic agents, or insulin. Glucagon-like peptide-1 receptor agonists may have additional effects, such as weight loss, that may be advantageous in obese patients. There is a large body of evidence from randomized controlled clinical trials supporting the cardiovascular safety of dipeptidyl peptidase-4 inhibitors and some glucagon-like peptide-1 receptor agonists, at least in the short term. However, concerns have been raised, particularly regarding their safety in patients with heart failure. In this review, the authors provide a brief but practical evidence-based analysis of the use of incretin-based agents in patients with diabetes, their efficacy, and cardiovascular safety.

TB as a cause of hospitalization and in-hospital mortality among people living with HIV worldwide: a systematic review and meta-analysis.

INTRODUCTION: Despite significant progress in improving access to antiretroviral therapy over the past decade, substantial numbers of people living with HIV (PLHIV) in all regions continue to experience severe illness and require hospitalization. We undertook a global review assessing the proportion of hospitalizations and in-hospital deaths because of tuberculosis (TB) in PLHIV. METHODS: Seven databases were searched to identify studies reporting causes of hospitalizations among PLHIV from 1 January 2007 to 31 January 2015 irrespective of age, geographical region or language. The proportion of hospitalizations and in-hospital mortality attributable to TB was estimated using random effects meta-analysis. RESULTS: From an initial screen of 9049 records, 66 studies were identified, providing data on 35,845 adults and 2792 children across 42 countries. Overall, 17.7% (95% CI 16.0 to 20.2%) of all adult hospitalizations were because of TB, making it the leading cause of hospitalization overall; the proportion of adult hospitalizations because of TB exceeded 10% in all regions except the European region. Of all paediatric hospitalizations, 10.8% (95% CI 7.6 to 13.9%) were because of TB. There was insufficient data among children for analysis by region. In-hospital mortality attributable to TB was 24.9% (95% CI 19.0 to 30.8%) among adults and 30.1% (95% CI 11.2 to 48.9%) among children. DISCUSSION: TB remains a leading cause of hospitalization and in-hospital death among adults and children living with HIV worldwide.

Impact of Changes in Urine Culture Ordering Practice on Antimicrobial Utilization in Intensive Care Units at an Academic Medical Center.

OBJECTIVE To assess antimicrobial utilization before and after a change in urine culture ordering practice in adult intensive care units (ICUs) whereby urine cultures were only performed when pyuria was detected. DESIGN Quasi-experimental study SETTING A 700-bed academic medical center PATIENTS Patients admitted to any adult ICU METHODS Aggregate data for all adult ICUs were obtained for population-level antimicrobial use (days of therapy [DOT]), urine cultures performed, and bacteriuria, all measured per 1,000 patient days before the intervention (January-December 2012) and after the intervention (January-December 2013). These data were compared using interrupted time series negative binomial regression. Randomly selected patient charts from the population of adult ICU patients with orders for urine culture in the presence of indwelling or recently removed urinary catheters were reviewed for demographic, clinical, and antimicrobial use characteristics, and pre- and post-intervention data were compared. RESULTS Statistically significant reductions were observed in aggregate monthly rates of urine cultures performed and bacteriuria detected but not in DOT. At the patient level, compared with the pre-intervention group (n=250), in the post-intervention group (n=250), fewer patients started a new antimicrobial therapy based on urine culture results (23% vs 41%, P=.002), but no difference in the mean total DOT was observed. CONCLUSION A change in urine-culture ordering practice was associated with a decrease in the percentage of patients starting a new antimicrobial therapy based on the index urine-culture order but not in total duration of antimicrobial use in adult ICUs. Other drivers of antimicrobial use in ICU patients need to be evaluated by antimicrobial stewardship teams. Infect.

High prevalence of hepatitis delta virus among patients with chronic hepatitis B virus infection and HIV-1 in an intermediate hepatitis B virus endemic region.

We conducted a study to investigate HIV and hepatitis delta virus (HDV) coinfection among patients with chronic hepatitis B virus (HBV) infection and the triple infection's (HIV/HBV/HDV) clinical implications in India, an intermediate HBV endemic region, with an estimated HIV-positive population of 2.5 million. A total of 450 patients (men: 270; women: 180) with chronic HBV infections and 135 healthy volunteers were screened for HIV and HDV. The incidence of the triple infection was low (4 [0.8%]) compared with dual infections of HIV-1/HBV (7 [1.5%]) and HBV/HDV (22[4.8%]). Among 21- to 40-year-olds, HBV/HDV coinfection (45.8%) and HBV/HDV/HIV-1 triple infection was predominant (75%). Among 11 patients coinfected with HIV-1/HBV, 4 (36%) were tri-infected and were also associated with chronic hepatitis and cirrhosis. The HDV coinfection was higher among patients coinfected with HBV/HIV-1, despite the declining trend in HDV infection among HIV-negative patients, as previously reported. Thus, it is important to assess the impact of HIV, chronic HBV, and HDV tri-infection in India.

Intermolecular interaction between Cry2Aa and Cyt1Aa and its effect on larvicidal activity against Culex quinquefasciatus.

The Cyt1Aa protein of Bacillus thuringiensis susbp. israelensis elaborates demonstrable toxicity to mosquito larvae, but more importantly, it enhances the larvicidal activity of this species Cry proteins (Cry11Aa, Cry4Aa, and Cry4Ba) and delays the phenotypic expression of resistance to these that has evolved in Culex quinquefasciatus. It is also known that Cyt1Aa, which is highly lipophilic, synergizes Cry11Aa by functioning as a surrogate membrane-bound receptor for the latter protein. Little is known, however, about whether Cyt1Aa can interact similarly with other Cry proteins not primarily mosquitocidal; for example, Cry2Aa, which is active against lepidopteran larvae, but essentially inactive or has very low toxicity to mosquito larvae. Here we demonstrate by ligand binding and enzyme-linked immunosorbent assays that Cyt1Aa and Cry2Aa form intermolecular complexes in vitro, and in addition show that Cyt1Aa facilitates binding of Cry2Aa throughout the midgut of C. quinquefasciatus larvae. As Cry2Aa and Cry11Aa share structural similarity in domain II, the interaction between Cyt1Aa and Cry2Aa could be a result of a similar mechanism previously proposed for Cry11Aa and Cyt1Aa. Finally, despite the observed interaction between Cry2Aa and Cyt1Aa, only a 2-fold enhancement in toxicity resulted against C. quinquefasciatus. Regardless, our results suggest that Cry2Aa could be a useful component of mosquitocidal endotoxin complements being developed for recombinant strains of B. thuringiensis subsp. israelensis and B. sphaericus aimed at improving the efficacy of commercial products and avoiding resistance.