A Day Saved is a Life Saved: Direct Antimicrobial Susceptibility Testing from Positively Flagged Blood Culture Bottles and their Concordance with the Routine Method.

BACKGROUND: Sepsis is a major health problem worldwide and is associated with high morbidity and mortality with every hour delay in initiation of therapy. A conventional method of blood culture and Antimicrobial Susceptibility Testing (AST) takes around 48-72 hours. Empirical antibiotics need to be administered until the sensitivity report is made available. It has been estimated that 20-50% of the empirical antibiotics are inappropriate, resulting in prolonged hospital stays, adverse effects, and emergence of drug resistance. Additionally, this also puts an extra financial burden on both the patients and healthcare settings. Performing direct Antimicrobial Sensitivity Testing (dAST) is an important tool to reduce turn-around time (TAT) by at least 18-24 hours, thus reducing morbidity and mortality among critically ill patients. METHODS: Direct AST (dAST) was performed from the positively flagged blood culture bottles received between December, 2021 to May, 2022 from Intensive Care Units (ICUs) on MuellerHinton Agar (MHA) using four drops of withdrawn blood. dAST was performed for six drugs: Ceftriaxone-30 µg (CTR), Piperacillin/Tazobactam-100/10 µg (PIT), Meropenem-10 µg (MRP), Ciprofloxacin-5 µg (CIP), Aztreonam-30 µg (AT), and Colistin (CL). The zone of inhibition was interpreted as per CLSI M100 ed32, 2022 guidelines. A parallel conventional method was also performed to examine for categorical agreement and disagreement. Identification was carried out using MALDI-TOF MS from the colonies that appeared on the dAST plate on the subsequent day. RESULTS: A total of 162 positively flagged blood culture bottles were included in the study. The majority of the Gram-negative organisms were from Enterobacterales (n=109), followed by Acinetobacter spp. (n=28) and Pseudomonas aeruginosa (n=25). Out of the 972 isolate-antimicrobial combinations, overall Categorical Agreement (CA) was seen in 936 (96.3%), whereas disagreement was observed in 36 with minor error (mE) in 21 (2.2%), major error (ME) in 7 (0.7%), and very major error (VME) in 8 (0.8%) when compared to the routine method. Categorical agreement (CA) of > 99% was seen in ceftriaxone (CTR) and ciprofloxacin (CIP). In comparison, the lowest CA was observed with meropenem (MRP) at 92%. Colistin dAST was performed using the E-strip method, and the result obtained was highly convincing, with an overall disagreement of only 1.2%. CONCLUSION: Rapid dAST from positively flagged blood culture bottles proved to significantly reduce the TAT from the time of sample collection to the first availability of antimicrobial susceptibility report with excellent categorical agreement of > 95% using the conventional disc diffusion method. Results obtained were within the acceptance criteria set by U. S. Food and Drug Administration (FDA) guidelines of > 90% categorical agreement for a new method. We were able to obtain excellent concordance for colistin using the E-strip method. Performing dAST not only saves a "day", but its proper implementation would save a "life".

Hit or Miss: The Dilemma of Specimen Selection for Microbiological Diagnosis of Rhino-Orbital-Cerebral- COVID-Associated Mucormycosis (ROCM-CAM).

BACKGROUND: We diagnosed various cases of rhino-orbital-cerebral- COVID-associated Mucormycosis (ROCM-CAM) during India's second wave of COVID-19. This helped formulate novel suggestions for improving laboratory output, applicable anywhere in the world. METHODS: To diagnose ROCM-CAM by microbiological methods, we used direct microscopy and conventional culture on various clinical samples within the shortest turn-around time. DESIGN: Prospective single-center observational study. PARTICIPANTS: Patients with ROCM-CAM. RESULTS: Of 113 suspected cases of ROCM-CAM during May 2021, direct microscopy and culture could confirm the disease in 87.61% and 44.25% of patients, respectively. The highest pathogen isolation was seen from maxillary bone fragments, FESS-guided biopsy from pterygopalatine fossae, nasal turbinates and nasal mucosal biopsy. Direct microscopy could diagnose the disease in almost 40% of patients within 24 hours and 60% within two days. Conventional cultures yielded Rhizopus spp. (86%) as the commonest fungal pathogen followed by Mucor spp. (12%) within 7 days. Deep tissue biopsies are more useful for rapid diagnosis than superficial specimens. Routine fungal cultures can supplement case detection and help prognosticate survivors. CONCLUSION: The management of ROCM is a surgical emergency. The diagnosis of the condition must therefore be prompt and precise. Despite ongoing antifungal therapy, nasal mucosal tissue, FESSguided, and intra-operative tissue biopsies showed the pathogen's highest diagnostic yield. The diagnostic index improved further when multiple (4-5) high-quality specimens were collected. Nasal swabs and crusts, among the most commonly requested specimens worldwide, were found to have an overall low diagnostic potential.

Developing a co-production strategy to facilitate the adoption and implementation of evidence-based colorectal cancer screening interventions for rural health systems: a pilot study.

BACKGROUND: Evidence-based colorectal cancer screening (CRCS) interventions have not been broadly adopted in rural primary care settings. Co-production of implementation strategies through a bundled approach may be promising in closing this gap by helping rural healthcare practitioners select and implement the best fitting CRCS interventions to the local context. This paper describes the process and outcomes of co-development and delivery of the bundled implementation strategy to improve adoption and implementation of CRCS interventions with two rural clinics. METHODS: We used a bundle of implementation strategies with a core focus on academic-clinical partnership development (strategy 1) and Plan-Do-Study-Act cycles (strategy 2) to identify clinical partner interests/preferences on delivery methods and content needed to facilitate intervention identification and implementation that improves CRCS. We also developed an implementation blueprint for each clinic (strategy 3) through an online blueprinting process based on adapted "Putting Public Health Evidence in Action" (PPHEA) training curriculum. Clinic physicians and staff (n = 7) were asked to evaluate the bundled approach based on overall reactions and perceptions of innovation characteristics using 5-point Likert scale. After completing the bundled approach, we collected implementation outcomes and limited intervention effectiveness of the CRCS evidence-based interventions (EBIs) developed through the process. RESULTS: Our co-production strategy yielded a prototype online blueprinting process consisting of 8 distance-learning PPHEA modules that guide selection and implementation of EBIs tailored to CRCS. Modules were delivered to clinic participants with minor adaptations, using PDSA cycle to improve quality of module contents and formats. Overall, participants in both clinics reported positive reactions toward the bundled approach. Both clinics reported improvements in how they perceived the characteristics of the innovation (the bundled approach) to tailor selected CRCS EBIs. As a result of the bundled strategies, each clinic selected and adopted specific EBI(s) with the varying degrees of implementation and CRCS outcomes. CONCLUSIONS: The bundle of implementation strategies used were feasible and acceptable in rural primary care practices to facilitate the use of EBIs to improve CRCS.

Immune profile and responses of a novel dengue DNA vaccine encoding an EDIII-NS1 consensus design based on Indo-African sequences.

The ongoing COVID-19 pandemic highlights the need to tackle viral variants, expand the number of antigens, and assess diverse delivery systems for vaccines against emerging viruses. In the present study, a DNA vaccine candidate was generated by combining in tandem envelope protein domain III (EDIII) of dengue virus serotypes 1-4 and a dengue virus (DENV)-2 non-structural protein 1 (NS1) protein-coding region. Each domain was designed as a serotype-specific consensus coding sequence derived from different genotypes based on the whole genome sequencing of clinical isolates in India and complemented with data from Africa. This sequence was further optimized for protein expression. In silico structural analysis of the EDIII consensus sequence revealed that epitopes are structurally conserved and immunogenic. The vaccination of mice with this construct induced pan-serotype neutralizing antibodies and antigen-specific T cell responses. Assaying intracellular interferon (IFN)-γ staining, immunoglobulin IgG2(a/c)/IgG1 ratios, and immune gene profiling suggests a strong Th1-dominant immune response. Finally, the passive transfer of immune sera protected AG129 mice challenged with a virulent, non-mouse-adapted DENV-2 strain. Our findings collectively suggest an alternative strategy for dengue vaccine design by offering a novel vaccine candidate with a possible broad-spectrum protection and a successful clinical translation either as a stand alone or in a mix and match strategy.

The Mental Health of Health Care Workers in the UK During COVID-19: The Prevalence of Anxiety, Depression, and Stress.

Introduction: The COVID-19 pandemic had a significant impact on the mental health of health care workers (HCWs). Aim: This study investigated the mental health of HCWs working in Leicester, UK during COVID-19. Settings: Two hospital trusts in Leicester, UK. Methods: An online survey was sent to HCWs in two trusts in July 2020. The Generalized Anxiety Disorder-7 scale (GAD-7) and the Patient Health Questionnaire-9 scale (PHQ-9) were used to measure the prevalence rates of anxiety and depression. The Perceived Stress Scale-14 (PSS-14) was used to measure levels of perceived stress. Other questions were used to identify the prevalence of increased alcohol intake and possible risk factors. Statistical Analysis Used: Chi-square test, independent sample t-test, ANOVA, and logistic regression. Results: A total of 1009 HCWs completed the survey. Using a cutoff score of 5 (mild symptoms), for both GAD-7 and PHQ-9, 80.2% of participants had at least one condition and 71.5% had both. Using the cutoff score of 10 (moderate/severe symptoms), 27.2% had at least one condition and 27.25% had both conditions. In addition, 37.5% of those who did not report pre-existing mental health conditions now have at least one condition. About 33.6% of participants reported an increase in alcohol consumption. A number of risk factors were identified: having less social support, not feeling supported at work, and poor pre-existing mental health. Conclusions: The pandemic had a significant impact on mental health of HCWs. Health organizations need to monitor and address these emerging effects.

A 5-year study of dengue seropositivity among suspected cases attending a teaching hospital of North-Western region of India.

Dengue virus infection is estimated to cause infection in approximately 390 million people globally each year, of which 96 million develop clinical disease. Dengue serotype 2 (DEN-2) is the most prevalent serotype over the past 50 years in India, but serotypes 3 and 4 have appeared in some epidemics as well. A retrospective study was conducted in a teaching hospital, western India, between January 2014 and December 2018. The records of dengue serological test were analyzed. In total, 40 randomly selected nonstructural protein 1 (NS1) antigen-positive samples were analyzed by a reverse transcription-polymerase chain reaction. The demographic data, that is, age and sex, along with geographic location and platelet count level, were recorded from the Serology laboratory register and Hospital Information System. In total, 14.85% (735/4948) samples tested positive for dengue serology. Most of the laboratory-confirmed dengue cases, 34.97% (257/735), were observed in the 21-30 years of age group. The most common serotype detected in the tested samples was DEN-3 in 55% cases (22/40, 13 monoinfection and 9 coinfection with DEN-1 and DEN-2). The present study gives an insight into the trend of dengue seropositivity among suspected cases in the western part of Rajasthan, India. This study showed a higher seroprevalence of dengue infection as well as a gradual increase in the seroprevalence in this part of India.

Seroprevalence of brucellosis in Western Rajasthan: A study from a tertiary care centre.

Brucellosis is a significant bacterial zoonotic disease with a high seroprevalence in low-to-middle-income countries where there is a significant contact of humans with animals. This prospective study was conducted to observe the seroprevalence of brucellosis in 75 symptomatic patients with pyrexia of unknown origin (PUO) and 75 high-risk individuals (10 veterinarians, 15 milkmen and 50 healthy contacts of symptomatic patients) with possible exposure to brucellosis. Serum samples collected from these patients and individuals were subjected to rose Bengal test (RBT) and ELISA for the detection of IgM and IgG antibodies. RBT was positive in 50 samples (40 PUO and 10 high risk), of which 25 (33.3%) PUO cases had detectable IgM antibodies, whereas IgG antibodies were detected in 20 patients. None of samples from high-risk individuals was reactive for IgM/IgG antibodies by ELISA. The overall seroprevalence in this study was 16.7% (33.3% in PUO patients). Being a predominantly livestock rearing area, brucellosis is a significant health-care problem in this part of India with this entity being linked to 33% of PUO cases.

UTILITY OF ULTRASOUND VERSUS GENE EXPRESSION CLASSIFIER IN THYROID NODULES WITH ATYPIA OF UNDETERMINED SIGNIFICANCE.

OBJECTIVE: Thyroid nodules with fine-needle aspiration (FNA) cytology categorized as atypia of undetermined significance (AUS) often undergo additional diagnostic analysis with the Afirma Gene Expression Classifier (GEC), which classifies these as either high probability of being benign (GEC-B) or suspicious for malignancy (GEC-S). Our goal was to assess the clinical validity and utility of GEC in the evaluation of AUS cytology and evaluate the performance of ultrasonography (USG) for predicting malignancy in this subset. METHODS: We conducted a study with a retrospective cohort of patients from January 2012 to January 2014 who had FNA of thyroid nodules >1 cm in size with AUS cytology. RESULTS: Cleveland Clinic Florida has an overall prevalence of AUS of 5%. A total of 119 cases with nodules >1 cm in size were reported as AUS. Forty-eight (40.3%) had a GEC performed after the first FNA (AUS-1), and 27 of these were GEC-S. Of those 27, 21 went for surgery and 14 (66.6%) had thyroid cancer on histopathology. The remaining 71 with AUS-1 were sent for a second FNA: 19 nodules were benign and did not undergo further evaluation, while the remaining 52 were reported as AUS for the second consecutive time (AUS-2). AUS-2 samples were sent for GEC. Of these 52 AUS-2, 38 (73.1%) were reported as GEC-S. Thirty-five went for surgery and 32 (91.4%) had confirmed malignancy on histopathology. Positive predictive value (PPV) was 91.4% for AUS-2 and 66.6% for AUS-1. Moreover, AUS-2 nodules that were hypoechoic and solid on USG showed a PPV of 92% for malignancy. CONCLUSION: In our practice, the diagnostic accuracy to predict malignancy with GEC for AUS-1 nodules was poor (PPV, 66.6%). The PPV of GEC testing was markedly higher at 91.4% performed after two consecutive AUS cytologies. AUS-2 nodules that were solid and hypoechoic on USG also had a high probability to be malignant (PPV, 92%). We recommend repeat FNA on AUS-1 nodules rather than proceeding directly to GEC testing. Also, we suggest that among AUS-2 nodules, surgery can be recommended when USG shows solid and hypoechoic features with GEC testing reserved for the remainder. ABBREVIATIONS: AUS = atypia of undetermined significance FNA = fine-needle aspiration GEC = gene expression classifier GEC-B = GEC-benign GEC-S = GEC-suspicious for malignancy NPV = negative predictive value PPV = positive predictive value USG = ultrasonography.