Laboratory diagnosis of bacterial meningitis by direct detection, serotyping and Next Generation Sequencing: How 10 years of testing in New York State has evolved to improve laboratory diagnosis and public health.

Since 2005, the Wadsworth Center (WC) has provided molecular testing on cerebrospinal fluid (CSF) and whole blood specimens in close collaboration with epidemiologists in New York State and New York City. In this study, we analyzed 10 years of data to demonstrate the significant value of utilizing molecular methods to assess patient specimens for etiologic agents of bacterial meningitis. A comprehensive molecular testing algorithm to detect and serotype/serogroup bacterial agents known to cause bacterial meningitis (Neisseria meningitidis, Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus agalactiae) has evolved, and retrospective specimen testing has been essential for each improvement. Over a ten-year span from 2010 to 2019 the WC received 831 specimens from 634 patients with suspected bacterial meningitis. Real-time PCR was positive for at least one of the agents in 223 (27%) specimens from 183 patients (29%). Of the 223 positives, 146 (66%) were further characterized by real-time PCR into serogroup/serotype. Additionally, examination of 131 paired specimens of CSF and whole blood from the same patients found better detection in CSF, but whole blood is a useful alternative for diagnosis when CSF is not available. For specimens initially PCR-negative, 16S rDNA Sanger sequencing was requested by the submitter for 146 cases resulting in the identification of bacterial agents in an additional 24 (16%) specimens. In a retrospective study, Next Generation Sequencing (NGS) was evaluated for the detection of pathogens in 53 previously tested PCR-negative CSF specimens and identified bacteria in 14 (26%) specimens. This molecular testing algorithm has provided clinicians a diagnosis when culture is negative with the potential to guide therapy. It has also aided public health in determining when antibiotic prophylaxis was needed, augmented surveillance data to yield a fuller picture of community prevalence, and highlighted gaps in the spectrum of agents that cause bacterial meningitis.

Screening for depressive symptoms in cardiovascular patients at a tertiary centre in Trinidad and Tobago: investigation of correlates in the SAD CAT study.

BACKGROUND: This study aimed to screen cardiovascular patients for depressive symptoms at a tertiary centre in Trinidad and Tobago; and to determine any significant associations amongst patients' demographics, comorbidities, and cardiovascular medications with depressive symptoms. METHODS: In this observational, cross-sectional study, patients (n = 1203) were randomly selected from the cardiology outpatient clinics at the Eric Williams Medical Sciences Complex. After meeting selection criteria, informed consent was obtained, and patients were administered a case report form, which included the Patient Health Questionnaire-9 (PHQ-9). Descriptive analyses included frequency, percentage and summary statistics. Inferential analyses included 95% confidence intervals (CIs), independent sample t-test, Fisher's exact test, Chi-square test, and multivariate logistic regression. RESULTS: The study had a 96% respondent rate, whereby the average age was 62 years old. Slightly less than half were male, and 52.5% were female. Over 90 % of the sample had cardiovascular disease (CVD). One-quarter of the sample had a PHQ-9 score of ≥10, with almost one-fifth having no depressive symptoms. Females, lower levels of education and income were all found to be statistically significant at risk for depressive symptoms (all p-values < 0.001). Comorbidities associated with depressive symptoms included hypertension, prior cerebrovascular events, chronic kidney disease, and chronic obstructive pulmonary disease with odds ratios (ORs) and 95% confidence intervals (CIs) of OR 1.988 (CI 1.414-2.797), OR 1.847 (CI 1.251-2.728), OR 1.872 (CI 1.207-2.902) and OR 1.703 (CI 1.009-2.876) respectively. Only the cardiovascular medication of ticagrelor was found to be significantly associated with depressive symptoms (p-value < 0.001). CONCLUSIONS: Twenty-five percent of screened cardiovascular patients displayed significant depressive symptoms with a PHQ-9 ≥ 10. This study also highlights the importance of implementing a multidisciplinary approach to managing cardiovascular disease and screening for depressive symptoms in this subpopulation. Further studies are required to validate these findings. TRIAL REGISTRATION: ClinicalTrials.gov number, NCT03863262 . This trial was retrospectively registered on 20th February 2019.

Combined real-time PCR and rpoB gene pyrosequencing for rapid identification of Mycobacterium tuberculosis and determination of rifampin resistance directly in clinical specimens.

Our laboratory has developed a rapid, sensitive, and specific molecular approach for detection in clinical specimens, within 48 h of receipt, of both Mycobacterium tuberculosis complex (MTBC) DNA and mutations within the 81-bp core region of the rpoB gene that are associated with rifampin (RIF) resistance. This approach, which combines an initial real-time PCR with internal inhibition assessment and a pyrosequencing assay, was validated for direct use with clinical specimens. To assess the suitability of real-time PCR for use with respiratory, nonrespiratory, acid-fast bacillus (AFB)-positive and AFB-negative specimens, we evaluated specimens received in our laboratory between 11 October 2007 and 30 June 2009. With culture used as the "gold standard," the sensitivity, specificity, and positive and negative predictive values were determined for 1,316 specimens to be as follows: for respiratory specimens, 94.7%, 99.9%, 99.6%, and 98.6%, respectively; for nonrespiratory specimens, 88.5%, 100.0%, 100.0%, and 96.9%, respectively; for AFB-positive specimens, 99.6%, 100.0%, 100.0%, and 97.7%, respectively; and for AFB-negative specimens, 75.4%, 99.9%, 98.0%, and 98.4%, respectively. PCR inhibition was determined to be minimal in this assay, occurring in 0.2% of tests. The rpoB gene pyrosequencing assay was evaluated in a similar prospective study, in which 148 clinical specimens positive for MTBC DNA by real-time PCR were tested. The final results revealed that the results of direct testing of clinical specimens by the pyrosequencing assay were 98.6% concordant with the results of conventional testing for susceptibility to RIF in liquid culture and that our assay displayed adequate sensitivity for 96.6% of the clinical specimens tested. Used together, these assays provide reliable results that aid with the initial management of patients with suspected tuberculosis prior to the availability of the results for cultured material, and they also provide the ability to predict RIF resistance in Mycobacterium tuberculosis-positive specimens in as little as 48 h from the time of clinical specimen receipt.

Growth efficiency increases as relative growth rate increases in shoots and roots of Eucalyptus globulus deprived of nitrogen or treated with salt.

We used calorimetry to test whether there is a single general relationship between growth and respiration in shoots and roots of Eucalyptus globulus Labill. seedlings when stressed, irrespective of the type or severity of stress. We found that nitrogen (N) deprivation and salt treatment had no effect on the relationship between growth and respiration and little effect on absolute rates of respiration. Carbon-conversion efficiency (epsilonC) ranged from 0.7 to 0.9 for specific growth rates (R(SG)) greater than 0.3 day(-1). Above an R(SG) of 0.1 day(-1), epsilonC decreased gradually with decreasing R(SG) and between an R(SG) of 0- 0.1 day(-1), epsilonC decreased rapidly. We conclude that the relationship between epsilonC and R(SG) is not greatly affected by salt or N-deprivation stresses. Relationships between gross productivity and epsilonC may be generally applicable, in which case they could improve on the "flat-tax" approach to modeling net primary productivity from gross productivity while avoiding the complexity of more explicit models of plant respiration. However, the relationship between gross productivity and epsilonC was sensitive to temperature and the effect of temperature on epsilonC thus requires further investigation. Nitrogen deprivation caused large decreases in leaf area and shoot to root ratio, and mature leaves of N-deprived plants had lower intrinsic water-use efficiencies than leaves of plants well supplied with nutrients. Nitrogen deprivation increased apical dominance and most of the reduction in leaf area was the result of fewer secondary branches, although leaf size was also reduced. Our results suggest that N deprivation reduces productivity primarily by reducing sink size, rather than sink activity, and that apical dominance may be an important mechanism for maintaining adequate epsilonC in resource-limited conditions.