Properties of non-coding mutation hotspots as urinary biomarkers for bladder cancer detection.

Mutations at specific hotspots in non-coding regions of ADGRG6, PLEKHS1, WDR74, TBC1D12 and LEPROTL1 frequently occur in bladder cancer (BC). These mutations could function as biomarkers for the non-invasive detection of BC but this remains largely unexplored. Massively-parallel sequencing of non-coding hotspots was applied to 884 urine cell pellet DNAs: 591 from haematuria clinic patients (165 BCs, 426 non-BCs) and 293 from non-muscle invasive BC surveillance patients (29 with recurrence). Urine samples from 142 non-BC haematuria clinic patients were used to optimise variant calling. Non-coding mutations are readily detectable in the urine of BC patients and undetectable, or present at much lower frequencies, in the absence of BC. The mutations can be used to detect incident BC with 66% sensitivity (95% CI 58-75) at 92% specificity (95% CI 88-95) and recurrent disease with 55% sensitivity (95% CI 36-74) at 85% specificity (95% CI 80-89%) using a 2% variant allele frequency threshold. In the NMIBC surveillance setting, the detection of non-coding mutations in urine in the absence of clinically detectable disease was associated with an increased relative risk of future recurrence (RR = 4.62 (95% CI 3.75-5.48)). As urinary biomarkers, non-coding hotspot mutations behave similarly to driver mutations in BC-associated genes and could be included in biomarker panels for BC detection.

Native tricuspid valve endocarditis presenting as pyrexia of unknown origin.

Native tricuspid valve endocarditis is quite rare without any predisposing factors and poses a diagnostic challenge because of fewer cardiac symptoms and lesser peripheral manifestations. This is a case report of a 25-year-old female who presented with high-grade fever, dry cough, decreased appetite, and weight loss for 1 month with no history of intravenous drug use or evidence of underlying cardiac abnormality and was diagnosed with native tricuspid valve endocarditis.

Duration of stenting with Montgomery T-tubes in severe laryngotracheal stenosis: does it matter?

OBJECTIVE: Laryngotracheal stenosis management remains largely discretionary in surgical practice. Duration of stenting remains variable following open reconstruction procedures in absence of clearly established differences. The current study evaluates successful decannulation after short-term periods compared with longer periods. METHOD: A comparative study over 18 months evaluated differences in successful decannulation between short- and long-term stent groups. Patients with grade II, III or IV laryngotracheal stenosis were placed on Montgomery T-tube for a short-term period (n = 15), and decannulation rates were compared with age-, sex- and diagnosis-matched patients (n = 15) from historical cases with long-term stent placement. RESULTS: Thirty patients were included. There was no difference between the two groups at baseline. Nine patients (30 per cent) were successfully decannulated, and there was no difference in rates of decannulation between the two groups (p = 0.8). Granulations at the proximal end of tube (38.7 per cent), superior migration of tube (16.1 per cent) and dysphonia (12.9 per cent) were common complications. CONCLUSION: Decannulation was not more successful after placing Montgomery T-tubes for longer periods of time when compared with removal within the first three months. Early removal after proper case selection and planning may be considered for a successful outcome in laryngotracheal stenosis.

Impact of coronavirus disease 2019 on ENT clinical practice and training: the resident's perspective.

BACKGROUND: The arrival of the coronavirus disease 2019 pandemic disrupted life suddenly and forcefully, and healthcare systems around the world are still struggling to come to terms with it. This paper reviews the impact of the pandemic on ENT practice and training. METHODS: The present manuscript was developed as a narrative review to examine the role of otorhinolaryngologists in the management of the pandemic, and assess its impact on practice and training in the specialty. RESULTS: Otorhinolaryngologists handle secretions of organs implicated in disease transmission, leaving them particularly vulnerable even while performing simple procedures. Although the pandemic increased skill expectations, it simultaneously reduced learning opportunities for trainees. In addition, attention to emergencies has been delayed. Further, the suspension of elective procedures has affected patients with malignancies. CONCLUSION: While planning service resumption, provisions need to be made for protective equipment and training; improving teleconsultation services will help provide sustainable care during further waves.

COVID-19 appropriate behavior in India: Time to invest for the benefits in future.

The COVID-19 pandemic gave an opportunity to adopt many appropriate changes in the behavior of the people in India. The major gears of those behavior changes were the enforcement by the government, fear, motivation (self and induced), and self-experiences or realizations with time. If those changes are fitted in the Trans-Theoretical Model, Indian people have passed through the "Pre-Contemplation" to "Action" stage of behavior changes during different phases of this pandemic. Frequent hand hygiene, maintaining physical distancing, use of face mask, cough etiquettes, avoid greetings through physical contacts, fear in spitting and urination at public places, refrain from gatherings and avoiding outside food are some of the examples of those appropriate behaviors which were enforced or learnt during the COVID pandemic. The continuous lockdown made people understand the difference between "want" and "need," the importance of local production, and the significance of social media and technology in routine life. The work-from-home strategy gave a chance to appreciate the work--life balance in a more applied way. The first-ever lifetime experience of unbelievable rejuvenating nature because of lack of human play taught people to appreciate nature. Although the current focus is on responding to the pandemic and on coping with its immediate effects, yet this is the time when there is an urgent need to create an enabling environment to support and sustain these COVID-19 appropriate behaviors (maintenance stage) to reap the maximum benefits out of them. Sustaining these appropriate behaviors is also important considering the bimodal distribution of the COVID-19 and possibility of advent of the second wave of COVID-19 in near future.

Development of a PCR-lateral flow assay for rapid detection of Yersinia pestis, the causative agent of plague.

Plague is a zoonotic disease caused by Yersinia pestis, a Gram-negative, rod shaped coccobacillus, which is primarily found in rodents and can be transmitted to humans through flea bite. The disease has three major clinical forms bubonic (by flea bite), pneumonic (by respiratory droplets) and septicemic plague. Y. pestis is classified as a category 'A' agent by NIAID, USA due to its high mortality and easy person to person dissemination. The conventional diagnostic methods available for Y. pestis show cross-reactivity with other enteropathogenic bacteria making its detection difficult. There is a need to develop sensitive and specific molecular assay for accurate detection of Y. pestis. PCR is well suited molecular biology tool for rapid diagnosis of plague but after completion of thermal cycling steps, it requires additional time to analyze amplified product using agarose gel electrophoresis. In the present study, PCR assay coupled with lateral flow strips has been developed for rapid detection of Y. pestis. Lateral flow strips give an alternative to gel electrophoresis and permit easy and rapid detection of PCR products. The PCR was performed with 5' 6-FAM and biotin tagged primers specific for Y. pestis, targeting yihN gene located on chromosome. The PCR product was analyzed using lateral flow strips which yielded result within 2-3 minutes. The analytical sensitivity of PCR-lateral flow (PCR-LF) assay was 1 pg genomic DNA of Y. pestis and 500 copies of target DNA sequence harboured in a recombinant plasmid. The assay could detect Y. pestis DNA extracted from spiked human blood samples containing ≥104 CFU per mL of bacteria. The assay was found to be specific and did not cross react with other closely related bacterial species. The developed assay was highly specific, sensitive and also did not require agarose gel electrophoresis for post amplification analysis.

Development of a PCR Lateral Flow Assay for Rapid Detection of Bacillus anthracis, the Causative Agent of Anthrax.

Bacillus anthracis, the causative agent of anthrax is one of the most potent listed biological warfare agents. The conventional microbiological methods of its detection are labor intensive and time consuming, whereas molecular assays are fast, sensitive and specific. PCR is one of the most reliable diagnostic tools in molecular biology. The combination of PCR with lateral flow strips can reduce the diagnostic/detection time. It gives an alternative to gel electrophoresis and offers easy and clear interpretation of results. In the present study, a PCR Lateral flow (PCR-LF) assay targeting cya gene present on pXO1 plasmid of B. anthracis has been developed. The forward and reverse primers were tagged with 6-carboxyflourescein (6-FAM) and biotin, respectively, at 5' end. The dual labeled PCR products were detected using lateral flow (LF) strips developed in this study. The PCR-LF assay could detect ≥ 5 pg of genomic DNA and ≥ 500 copies of target DNA harboured in a recombinant plasmid. The assay was able to detect as few as 103 and 10 CFU/mL of B. anthracis Sterne cells spiked in human blood after 6 and 24 h of enrichment, respectively.

Development of a set of three real-time loop-mediated isothermal amplification (LAMP) assays for detection of Bacillus anthracis, the causative agent of anthrax.

Bacillus anthracis, the causative agent of anthrax is a Gram-positive, non-motile, spore forming bacterium. Its spores can persist in soil and water for years and can also be aerosolized. A rapid, sensitive and specific method to detect B. anthracis is important for clinical management and preventing spread of anthrax. Loop-mediated isothermal amplification (LAMP) assay is a rapid technique that amplifies target DNA in isothermal conditions with high sensitivity and specificity. In this study, a LAMP assay set targeting a chromosomal and two plasmid markers was developed. The individual assays of the LAMP set targeting pXO1 plasmid (lef), pXO2 plasmid (capB), and chromosome (BA5345) sequences could detect 10, 250, and 100 fg of genomic DNA and 10, 100, and 50 copies of the DNA targets harboured in recombinant plasmids, respectively. The lef and capB LAMP assays could detect ≥ 1 × 103 CFU per mL of bacteria in spiked human blood samples, while BA5345 LAMP assay could detect ≥ 1 × 104 CFU of bacteria per mL of spiked blood. The amplification was monitored in real-time by turbidimeter, and visual detection was also accomplished under normal and UV light after adding SYBR Green 1 dye on completion of the reaction. The assay set was found to be highly sensitive and did not cross-react with the closely related Bacillus spp. and other bacterial strains used in the study.

Physical Frailty Phenotype and Depression are Associated with More Severe Disease in Older Subjects Presenting with Acute Coronary Syndrome.

OBJECTIVE: The primary objective was to determine the occurrence of frailty in elderly patients presenting with the acute coronary syndrome (ACS). The secondary objective was to study the association between the deficits in health with the severity of ACS at presentation among them. METHODS: A cross-sectional study conducted in the Departments of Medicine, Community Medicine and Biochemistry in a tertiary care teaching hospital, Delhi, India between November 2014 and April 2016. Patients (≥60 years age) presenting with any one of the spectra of ACS (STEMI, UA, NSTEMI) and giving informed written consent were assessed for frailty and health deficits using questionnaires. ACS assessed by ECG within 24 h and other relevant investigations. Appropriate statistical tests of significance like the Chi-square test were used and correlation coefficients were analyzed. A value of P < 0.05 was considered significant. RESULTS: Seven risk factors apart from old age were studied, in which smoking and dyslipidemia played a major role. 44% of the subjects were frail with the range of frailty scores between 3 and 5. Every one-unit increase in hemoglobin was associated with a reduction in the odds (OR 0.72) for being frail. No association was noted between the severity of ACS and established risk factors like smoking, hypertension, diabetes, family history of CAD, increased waist circumference, dyslipidemia, and male gender. On multivariable linear regression, presence of frailty and depression were associated with severe disease. CONCLUSIONS: Nearly one in two patients presenting with ACS were found frail. Depression and frailty were associated with poorer ejection fraction and severe disease. Correction of anemia and improvement of low-normal hemoglobin levels could reduce frailty and in-turn improve outcomes in ACS.

Comparative Assessment of Revascularization Versus Drug Management in Coronary Artery Disease (CAD) Associated with Left Ventricular Dysfunction (EF < 40%) - A 12 Month Study with FDG PET and SPECT MPI Analyses.

AIM OF THE STUDY: Left Ventricular (LV) function and myocardial viability is the key predictor of prognosis after myocardial infarction. Management of ischemic cardiomyopathy (revascularization and or drugs alone) is the objective of this study. METHODOLOGY: 72 patients were assigned to revascularization and medical management group based on the inclusion criteria Follow up was done upto 12 months with advanced imaging techniques (FDG PET and SPECT MPI analyses). RESULTS: Subjects with significant viable myocardium, revascularization resulted in significant improvement in heart failure symptoms. The mean NYHA functional class improved from 2.9 ± 0.3 to 2.3 ± 0.5(mean ± SD) after 6 months of revascularization (p < 0.01). This improvement in functional class was maintained after 12 months of revascularization (2.0 ± 0.4 (mean ± SD). Subjects on medical management with a baseline NYHA functional class 2.7 ± 0.5, at 6 months of follow, there was no significant change in functional class (2.8 ± 0.3) (p<0.24). However at 12 months follow up functional class had dropped to 3.0 + 0.3, which was significant as compared to baseline (p <0.03). CONCLUSION: coronary revascularization has a protective effect on patients with ischemic coronary who have viable myocardium and reversible myocardial ischemia as assessed by 18F-FDG PET and SPECT MPI Imaging.

Development of a pair of real-time loop mediated isothermal amplification assays for detection of Yersinia pestis, the causative agent of plague.

Yersinia pestis, the causative agent of plague mainly infects rodents, while humans are the accidental host. The conventional diagnostic methods available for Y. pestis exhibit cross-reactivity with other enteropathogenic bacteria which makes its detection difficult. Rapid and reliable point-of-care detection of Y. pestis is essential for timely initiation of medical treatment. In the present study, a pair of loop mediated isothermal amplification (LAMP) assays has been developed for rapid detection of Y. pestis. Two sets of LAMP primers, each containing 6 primers were specifically designed targeting caf1 and 3a genes located on pFra plasmid and chromosome of Y. pestis, respectively. Isothermal amplification was accomplished at 65 °C for 40 min for caf1 target, and at 63 °C for 50 min for 3a choromosomal target. The analytical sensitivity of the assay for the caf1 and 3a targets was found to be 500 fg and 100 fg genomic DNA of Y. pestis, respectively. The caf1 and 3a LAMP assays detected as few as 100 copies of caf1 and 10 copies of 3a gene targets harboured in the respective recombinant plasmids. The amplified products were detected visually under visible and UV light using SYBR Green 1 dye. The assay pair was found to be highly specific as it did not cross-react with closely related and other bacterial species.

Evolving epidemiology of poliovirus serotype 2 following withdrawal of the serotype 2 oral poliovirus vaccine.

Although there have been no cases of serotype 2 wild poliovirus for more than 20 years, transmission of serotype 2 vaccine-derived poliovirus (VDPV2) and associated paralytic cases in several continents represent a threat to eradication. The withdrawal of the serotype 2 component of oral poliovirus vaccine (OPV2) was implemented in April 2016 to stop VDPV2 emergence and secure eradication of all serotype 2 poliovirus. Globally, children born after this date have limited immunity to prevent transmission. Using a statistical model, we estimated the emergence date and source of VDPV2s detected between May 2016 and November 2019. Outbreak response campaigns with monovalent OPV2 are the only available method to induce immunity to prevent transmission. Yet our analysis shows that using monovalent OPV2 is generating more paralytic VDPV2 outbreaks with the potential for establishing endemic transmission. A novel OPV2, for which two candidates are currently in clinical trials, is urgently required, together with a contingency strategy if this vaccine does not materialize or perform as anticipated.