Stephen Peterson

Sleep continuity and architecture: Associations with pain-inhibitory processes in patients with temporomandibular joint disorder

Recent research suggests bi‐directional interactions between the experience of pain and the process of sleep; pain interferes with the ability to obtain sleep, and disrupted sleep contributes to enhanced pain perception. Our group recently reported, in a controlled experimental study, that sleep fragmentation among healthy adults resulted in subsequent decrements in endogenous pain inhibition. The present report follows up that observation by extending this line of research to a sample of patients experiencing persistent pain. Patients with chronic temporomandibular joint disorder (TMD) pain were studied using polysomnography and psychophysical evaluation of pain responses. We assessed whether individual differences in sleep continuity and/or architecture were related to diffuse noxious inhibitory controls (DNIC), a measure of central nervous system pain inhibition. Among 53 TMD patients, higher sleep efficiency and longer total sleep time were positively associated with better functioning of DNIC (r = 0.42–0.44, p < 0.01; ps < 0.05 for the multivariate analyses). These results suggest the possibility that disrupted sleep may serve as a risk factor for inadequate pain‐inhibitory processing and hint that aggressive efforts to treat sleep disturbance early in the course of a pain condition might be beneficial in reducing the severity or impact of clinical pain.

Improved Sensitivity for Molecular Detection of Bacterial and Candida Infections in Blood

ABSTRACT The rapid identification of bacteria and fungi directly from the blood of patients with suspected bloodstream infections aids in diagnosis and guides treatment decisions. The development of an automated, rapid, and sensitive molecular technology capable of detecting the diverse agents of such infections at low titers has been challenging, due in part to the high background of genomic DNA in blood. PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) allows for the rapid and accurate identification of microorganisms but with a sensitivity of about 50% compared to that of culture when using 1-ml whole-blood specimens. Here, we describe a new integrated specimen preparation technology that substantially improves the sensitivity of PCR/ESI-MS analysis. An efficient lysis method and automated DNA purification system were designed for processing 5 ml of whole blood. In addition, PCR amplification formulations were optimized to tolerate high levels of human DNA. An analysis of 331 specimens collected from patients with suspected bloodstream infections resulted in 35 PCR/ESI-MS-positive specimens (10.6%) compared to 18 positive by culture (5.4%). PCR/ESI-MS was 83% sensitive and 94% specific compared to culture. Replicate PCR/ESI-MS testing from a second aliquot of the PCR/ESI-MS-positive/culture-negative specimens corroborated the initial findings in most cases, resulting in increased sensitivity (91%) and specificity (99%) when confirmed detections were considered true positives. The integrated solution described here has the potential to provide rapid detection and identification of organisms responsible for bloodstream infections.

Comparative Analysis of Two Broad-Range PCR Assays for Pathogen Detection in Positive-Blood-Culture Bottles: PCR–High-Resolution Melting Analysis versus PCR-Mass Spectrometry

ABSTRACT Detection of pathogens in bloodstream infections is important for directing antimicrobial treatment, but current culture-based approaches can be problematic. Broad-range PCR assays which target conserved genomic motifs for postamplification amplicon analysis permit detection of sepsis-causing pathogens. Comparison of different broad-range assays is important for informing future implementation strategies. In this study, we compared positive-blood-culture bottles processed by PCR coupled to high-resolution melting curve analysis (PCR/HRMA) and PCR coupled to electrospray ionization-mass spectrometry (PCR/ESI-MS) to microbiology culture results. Genus-level concordance was 90% (confidence interval [CI], 80 to 96%) for PCR/HRMA and 94% (CI, 85 to 98%) for PCR/ESI-MS. Species-level concordance was 90% (CI, 80 to 96%) for PCR/HRMA and 86% (CI, 75 to 93%) for PCR/ESI-MS. Unlike PCR/HRMA, PCR/ESI-MS was able to resolve polymicrobial samples. Our results demonstrated that the two assays have similar overall concordance rates but may have different roles as potential adjunctive tests with standard blood culture, since each method has different capabilities, advantages, and disadvantages.

The effect of catecholamine depletion by alpha-methyl-para-tyrosine on measures of cognitive performance and sleep in abstinent MDMA users

(±) 3, 4-Methylenedioxymethamphetamine (MDMA) is a popular recreational drug of abuse and a brain serotonin (5-HT) neurotoxin in animals. Growing evidence suggests that humans who use MDMA recreationally can also develop 5-HT neurotoxic injury, although functional consequences have been difficult to identify. Twenty-five abstinent MDMA users and 23 non-MDMA using controls were studied to determine whether pharmacologic depletion of brain catecholamines by alpha-methyl-para-tyrosine (AMPT) would differentially effect MDMA users on measures of cognition and sleep, two processes dually modulated by brain serotonergic and catecholaminergic neurons. During a 5-day in-patient study, all subjects underwent formal neuropsychiatric testing, repeated computerized cognitive testing, and all-night sleep studies. At baseline, MDMA users had performance deficits on tasks of verbal and visuospatial working memory and displayed increased behavioral impulsivity on several computerized tasks, reflecting a tendency to perform quickly at the expense of accuracy. Baseline sleep architecture was also altered in abstinent MDMA users compared to controls. AMPT produced differential effects in MDMA users compared to controls on several cognitive and sleep measures. Differences in cognitive performance, impulsivity, and sleep were significantly correlated with MDMA use. These data extend findings from earlier studies demonstrating cognitive deficits, behavioral impulsivity, and sleep alterations in abstinent MDMA users, and suggest that lasting effects of MDMA lead to alterations in the ability to modulate behaviors reciprocally influenced by 5-HT and catecholamines. More research is needed to determine potential relationships between sleep abnormalities, cognitive deficits and impulsive behavior in abstinent MDMA users.

The IRIDICA BAC BSI Assay: Rapid, Sensitive and Culture-Independent Identification of Bacteria and Candida in Blood.

Bloodstream infection (BSI) and sepsis are rising in incidence throughout the developed world. The spread of multi-drug resistant organisms presents increasing challenges to treatment. Surviving BSI is dependent on rapid and accurate identification of causal organisms, and timely application of appropriate antibiotics. Current culture-based methods used to detect and identify agents of BSI are often too slow to impact early therapy and may fail to detect relevant organisms in many positive cases. Existing methods for direct molecular detection of microbial DNA in blood are limited in either sensitivity (likely the result of small sample volumes) or in breadth of coverage, often because the PCR primers and probes used target only a few specific pathogens. There is a clear unmet need for a sensitive molecular assay capable of identifying the diverse bacteria and yeast associated with BSI directly from uncultured whole blood samples. We have developed a method of extracting DNA from larger volumes of whole blood (5 ml per sample), amplifying multiple widely conserved bacterial and fungal genes using a mismatch- and background-tolerant PCR chemistry, and identifying hundreds of diverse organisms from the amplified fragments on the basis of species-specific genetic signatures using electrospray ionization mass spectrometry (PCR/ESI-MS). We describe the analytical characteristics of the IRIDICA BAC BSI Assay and compare its pre-clinical performance to current standard-of-care methods in a collection of prospectively collected blood specimens from patients with symptoms of sepsis. The assay generated matching results in 80% of culture-positive cases (86% when common contaminants were excluded from the analysis), and twice the total number of positive detections. The described method is capable of providing organism identifications directly from uncultured blood in less than 8 hours. Disclaimer: The IRIDICA BAC BSI Assay is not available in the United States.

Scaling Up HIV Testing in an Academic Emergency Department: An Integrated Testing Model with Rapid Fourth-Generation and Point-of-Care Testing.

Objective. We evaluated two approaches for implementing routine HIV screening in an inner-city, academic emergency department (ED). These approaches differed by staffing model and type of HIV testing technology used. The programmatic outcomes assessed included the total number of tests performed, proportion of newly identified HIV-positive patients, and proportion of newly diagnosed individuals who were linked to care. Methods. This study examined specific outcomes for two distinct, successive approaches to implementing HIV screening in an inner-city, academic ED, from July 2012 through June 2013 (Program One), and from August 2013 through July 2014 (Program Two). Program One used a supplementary staff-only HIV testing model with point-of-care (POC) oral testing. Program Two used a triage-integrated, nurse-driven HIV testing model with fourth-generation blood and POC testing, and an expedited linkage-to-care process. Results. During Program One, 6,832 eligible patients were tested for HIV with a rapid POC oral HIV test. Sixteen patients (0.2%) were newly diagnosed with HIV, of whom 13 were successfully linked to care. During Program Two, 8,233 eligible patients were tested for HIV, of whom 3,124 (38.0%) received a blood test and 5,109 (62.0%) received a rapid POC test. Of all patients tested in Program Two, 29 (0.4%) were newly diagnosed with HIV, four of whom had acute infections and 27 of whom were successfully linked to care. We found a statistically significant difference in the proportion of the eligible population tested—8,233 of 49,697 (16.6%) in Program Two and 6,832 of 46,818 (14.6%) in Program One. These differences from Program One to Program Two corresponded to increases in testing volume (n = 1,401 tests), number of patients newly diagnosed with HIV (n=13), and proportion of patients successfully linked to care (from 81.0% to 93.0%). Conclusion. Integrating HIV screening into the standard triage workflow resulted in a higher proportion of ED patients being tested for HIV as compared with the supplementary staff-only HIV testing model. New rapid fourth-generation testing technology allowed the identification of acute HIV infection and same-visit confirmation of a positive diagnosis.

Reverse Transcription-PCR–Electrospray Ionization Mass Spectrometry for Rapid Detection of Biothreat and Common Respiratory Pathogens

ABSTRACT Electrospray ionization mass spectrometry (ESI-MS) analysis of reverse transcription (RT)-PCR amplicons from human respiratory samples allows for broad pathogen identification approximately 8 h after collection. We investigated the performance characteristics of a high-throughput RT-PCR-coupled ESI-MS assay for distinguishing biothreat (BT) agents from common bacterial, fungal, and viral respiratory pathogens in bronchoalveolar lavage (BAL) fluid specimens from subjects with suspected respiratory infections. In a retrospective case series, 202 BAL fluid specimens were collected at the Johns Hopkins Hospital between August 2010 and February 2011 from patients with suspected acute respiratory infections. Samples were processed using standard bacterial, viral, and fungal testing in the clinical microbiology laboratory as part of routine care and then were blindly spiked with either water or nucleic acids from BT organisms (Bacillus anthracis, Yersinia pestis, Francisella tularensis, Brucella spp., Burkholderia spp., and Rickettsia prowazekii) and tested by RT-PCR–ESI-MS. The sensitivities and specificities of RT-PCR–ESI-MS versus standard clinical methods were as follows: for mock BT DNA, 98.5% sensitivity (95% confidence interval [CI], 94.2 to 99.7%) and 100% specificity (95% CI, 93.1 to 100.0%); for bacterial pathogens, 81.8% sensitivity (95% CI, 74.3 to 87.6%) and 73.6% specificity (95% CI, 64.2 to 81.4%); for viral pathogens, 93.3% sensitivity (95% CI, 66.0 to 99.7%) and 97.3% specificity (95% CI, 89.7 to 99.5%); for fungal pathogens, 42.6% sensitivity (95% CI, 29.5 to 56.7%) and 97.8% specificity (95% CI, 91.8 to 99.6%). Our data suggest that RT-PCR–ESI-MS is a useful adjunct to standard culture protocols for rapid detection of both BT and common respiratory pathogens; further study is required for assay validation, especially for fungal detection, and potential implementation.

An emergency department registration kiosk can increase HIV screening in high risk patients

We evaluated the feasibility and the patient acceptability of integrating a kiosk into routine emergency department (ED) practice for offering HIV testing. The work was conducted in four phases: phase 1 was a baseline, in which external testing staff offered testing at the bedside; phase 2 was a pilot assessment of a prototype kiosk; phase 3 was a pilot implementation and phase 4 was the full implementation with automated login. Feasibility was assessed by the proportion of offering HIV tests, acceptance, completion and result reporting. During the study period, the number of ED patients and eligible patients for screening were similar in the three main phases. However, the number and proportion of patients offered testing of those eligible for screening increased significantly from phase 1 (32%) to phase 3 (37%) and phase 4 (40%). There were slightly higher prevalences of newly diagnosed HIV with kiosk versus bedside testing (phase 1, 0%; phase 3, 0.2%; phase 4, 0.5%). Compared to patients tested at the bedside, patients tested via the kiosk were significantly younger, more likely to be female, to be black, and to report high risk behaviours. ED-based HIV screening via a registration-based kiosk was feasible, yielded similar proportions of testing, and increased the proportion of engagement of higher-risk patients in testing.

Microbial Typing by Machine Learned DNA Melt Signatures

There is still an ongoing demand for a simple broad-spectrum molecular diagnostic assay for pathogenic bacteria. For this purpose, we developed a single-plex High Resolution Melt (HRM) assay that generates complex melt curves for bacterial identification. Using internal transcribed spacer (ITS) region as the phylogenetic marker for HRM, we observed complex melt curve signatures as compared to 16S rDNA amplicons with enhanced interspecies discrimination. We also developed a novel Naïve Bayes curve classification algorithm with statistical interpretation and achieved 95% accuracy in differentiating 89 bacterial species in our library using leave-one-out cross-validation. Pilot clinical validation of our method correctly identified the etiologic organisms at the species-level in 59 culture-positive mono-bacterial blood culture samples with 90% accuracy. Our findings suggest that broad bacterial sequences may be simply, reliably and automatically profiled by ITS HRM assay for clinical adoption.

A case-control study evaluating RT-PCR/ESI-MS technology compared to direct fluorescent antibody and xTAG RVP PCR

Abstract Waste nasopharyngeal swabs (N = 244) were evaluated by the reverse-transcriptase polymerase chain reaction/electrospray ionization mass spectrometry PLEX-ID Broad Respiratory Virus Surveillance Kit version 2.5 compared to direct fluorescent antibody and xTAG Respiratory Virus Panel for percent agreement, sensitivity, and specificity. Sensitivity and specificity were 91% (111/122) and 95.1% (116/122), respectively. Sensitivity by virus, except parainfluenza, was 92.9–100%, and specificity was 99–100%.