Metatranscriptomics analysis reveals a novel transcriptional and translational landscape during Middle East respiratory syndrome coronavirus infection.

Among all RNA viruses, coronavirus RNA transcription is the most complex and involves a process termed "discontinuous transcription" that results in the production of a set of 3'-nested, co-terminal genomic and subgenomic RNAs during infection. While the expression of the classic canonical set of subgenomic RNAs depends on the recognition of a 6- to 7-nt transcription regulatory core sequence (TRS), here, we use deep sequence and metagenomics analysis strategies and show that the coronavirus transcriptome is even more vast and more complex than previously appreciated and involves the production of leader-containing transcripts that have canonical and noncanonical leader-body junctions. Moreover, by ribosome protection and proteomics analyses, we show that both positive- and negative-sense transcripts are translationally active. The data support the hypothesis that the coronavirus proteome is much vaster than previously noted in the literature.

SARS-CoV-2 in Terms of Medical Microbiology Laboratory.

Coronaviruses (CoV), which are in the Coronaviridae family, cause different severity of gastrointestinal, respiratory and systemic diseases in wild and domestic animals, and can lead to different clinical manifestations, ranging from colds to pneumonia, depending on immunity. To date, seven types of coronavirus have been identified as infectious agents in humans;of these, HCoV 229E, HCoV NL63, HCoV HKU1 and HCoV OC43 typically cause cold symptoms in immunocompetent individuals, while SARS-CoV (Severe Acute Respiratory Syndrome Coronavirus) and MERS-CoV (Middle East Respiratory Syndrome Coronavirus) is zoonotic and cause severe respiratory diseases and deaths. SARS-CoV-2, the causative agent of COVID-19, is the seventh coronavirus identified as an infection agent in humans, which started in December 2019 in Wuhan, Hubei Province of China and was identified as a pandemic in a short time. Since the World Health Organization (WHO) defines SARS-CoV-2-sourced COVID-19 as a pandemic, and because of the increasing number of cases and deaths worldwide, structure of the novel virus and viral diagnosis methods gained importance respectively for vaccine studies and for controlling the outbreak caused by the virus.Copyright © 2020 Ankara Pediatric Hematology Oncology Training and Research Hospital. All rights reserved.

Teaching Medical Microbiology With a Web-Based Course During the COVID-19 Pandemic: Retrospective Before-and-After Study.

BACKGROUND: The COVID-19 pandemic has imposed unprecedented hurdles on health care systems and medical faculties alike. Lecturers of practical courses at medical schools have been confronted with the challenge of transferring knowledge remotely. OBJECTIVE: We sought to evaluate the effects of a web-based medical microbiology course on learning outcomes and student perceptions. METHODS: During the summer term of 2020, medical students at Saarland University, Germany, participated in a web-based medical microbiology course. Teaching content comprised clinical scenarios, theoretical knowledge, and instructive videos on microbiological techniques. Test performance, failure rate, and student evaluations, which included open-response items, for the web-based course were compared to those of the on-site course from the summer term of 2019. RESULTS: Student performance was comparable between both the online-only group and the on-site comparator for both the written exam (n=100 and n=131, respectively; average grade: mean 7.6, SD 1.7 vs mean 7.3, SD 1.8; P=.20) and the oral exam (n=86 and n=139, respectively; average grade: mean 33.6, SD 4.9 vs mean 33.4, SD 4.8; P=.78). Failure rate did not significantly differ between the online-only group and the comparator group (2/84, 2.4% vs 4/120, 3.3%). While lecturer expertise was rated similarly as high by students in both groups (mean 1.47, SD 0.62 vs mean 1.27, SD 0.55; P=.08), students who took the web-based course provided lower scores for interdisciplinarity (mean 1.7, SD 0.73 vs mean 2.53, SD 1.19; P<.001), opportunities for interaction (mean 1.46, SD 0.67 vs mean 2.91, SD 1.03; P<.001), and the extent to which the educational objectives were defined (mean 1.61, SD 0.76 vs mean 3.41, SD 0.95; P<.001). Main critiques formulated within the open-response items concerned organizational deficits. CONCLUSIONS: Web-based courses in medical microbiology are a feasible teaching option, especially in the setting of a pandemic, leading to similar test performances in comparison to on-site courses. The lack of interaction and the sustainability of acquired manual skills warrant further research.

Trends in molecular diagnostics: 12th European Meeting on Molecular Diagnostics, Noordwijk aan Zee, the Netherlands, 12-14 October 2022.

This report provides an overview of the highlights of the 12th European Meeting on Molecular Diagnostics held in Noordwijk aan Zee, The Netherlands, 12-14 October 2022. This 3-day conference covered many relevant topics in the field of molecular diagnostics in humans i.e. oncology, infectious diseases, laboratory medicine, pharmacogenetics, pathology, and preventive medicine. Other relevant topics included quality management, laboratory automation, diagnostic preparedness, and lessons learned from the COVID pandemic. More than 400 participants, the majority coming from European countries, attended the meeting. Besides high-quality scientific presentations, more than 40 diagnostic companies presented their latest innovations, altogether in an informal and inspiring ambiance.

SARS-CoV-2 in Terms of Medical Microbiology Laboratory

Coronaviruses (CoV), which are in the Coronaviridae family, cause different severity of gastrointestinal, respiratory and systemic diseases in wild and domestic animals, and can lead to different clinical manifestations, ranging from colds to pneumonia, depending on immunity. To date, seven types of coronavirus have been identified as infectious agents in humans;of these, HCoV 229E, HCoV NL63, HCoV HKU1 and HCoV OC43 typically cause cold symptoms in immunocompetent individuals, while SARS-CoV (Severe Acute Respiratory Syndrome Coronavirus) and MERS-CoV (Middle East Respiratory Syndrome Coronavirus) is zoonotic and cause severe respiratory diseases and deaths. SARS-CoV-2, the causative agent of COVID-19, is the seventh coronavirus identified as an infection agent in humans, which started in December 2019 in Wuhan, Hubei Province of China and was identified as a pandemic in a short time. Since the World Health Organization (WHO) defines SARS-CoV-2-sourced COVID-19 as a pandemic, and because of the increasing number of cases and deaths worldwide, structure of the novel virus and viral diagnosis methods gained importance respectively for vaccine studies and for controlling the outbreak caused by the virus.

A CASE OF EGGERTHELLA BACTEREMIA IN A PATIENT WITH COVID-19

SESSION TITLE: COVID-19 Co-Infections SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: We present a case of Eggerthella bacteremia in a patient with COVID-19. CASE PRESENTATION: A 69-year-old woman presented to the emergency room with chief complaint of cough, dyspnea, and malaise. After testing positive with a home COVID-19 test three days earlier, she continued to have worsening respiratory status and was brought in via ambulance. She was found to be tachycardic and hypoxic, requiring high-flow oxygen to maintain saturation in the emergency department. Chest X-ray showed bilateral patchy opacities consistent with multifocal COVID-19 pneumonia, and she was admitted to the intensive care unit for acute hypoxic respiratory failure. COVID-19 drug therapy was initiated, including baricitinib, remdesivir and decadron. Shortly after hospitalization, she began to endorse worsening abdominal pain. Physical exam elicited tenderness to palpation of her right lower quadrant. Abdominal CT scan showed distal ileum fluid collection concerning for possible bowel perforation. She underwent exploratory laparotomy which confirmed perforation, and a small bowel resection with anastomosis was performed. Blood cultures were positive for gram-positive bacilli, which were further identified as Eggerthella species. She required mechanical ventilation for worsening respiratory function post-surgery but remained unresponsive on the ventilator. The patient was administered vancomycin but continued to decline and eventually expired. DISCUSSION: Eggerthella is an anaerobic, gram-positive bacilli present in the gut microflora. Eggerthella infection has most often been reported in intra-abdominal infections. However, cases of bacteremia infection remain sparse. Most infections have been associated with other gastrointestinal processes including Crohn's disease, ulcerative colitis, appendicitis, and diverticulitis abscesses. Our case involved a patient with no significant gastrointestinal history admitted for COVID-19 pneumonia infection on baricitinib complicated by bowel perforation and bacteremia. Bowel perforation is a known risk factor of baricitinib use, and these risks should be discussed with the patient before beginning therapy. Overall mortality for Eggerthella species infection remains high, with some estimates as high as 31%. Much remains unknown about the impact on gut microbiome by SARS-CoV-2, however, early research suggests a higher rate of fungal co-infection in patients with COVID-19. As the literature on COVID-19 expands, more and more unusual pathogens such as Eggerthella may be found to contribute to the morbidity and mortality of patients being treated for COVID-19. CONCLUSIONS: Unusual pathogens such as Eggerthella may complicate a patient's hospital course while undergoing treatment for COVID-19. Reference #1: Alejandra Ugarte-Torres, Mark R Gillrie, Thomas P Griener, Deirdre L Church, Eggerthella lenta Bloodstream Infections Are Associated With Increased Mortality Following Empiric Piperacillin-Tazobactam (TZP) Monotherapy: A Population-based Cohort Study, Clinical Infectious Diseases, Volume 67, Issue 2, 15 July 2018. Reference #2: Gardiner BJ, Tai AY, Kotsanas D, et al. Clinical and microbiological characteristics of Eggerthella lenta bacteremia. J Clin Microbiol. 2015. Reference #3: Lau SK, Woo PC, Fung AM, Chan K-M, Woo GK, Yuen K-Y. Anaerobic, non-sporulating, gram-positive bacilli bacteraemia characterized by 16s rrna gene sequencing. Journal of medical microbiology. 2004. DISCLOSURES: No relevant relationships by Kristin Davis No relevant relationships by Charles Peng

Aspergillus Infections in Intensive Care Units: Before and after the COVID-19 Pandemic

Introduction: Aspergillus species have begun to cause invasive pulmonary aspergillosis (IPA) with increasing frequency in patients with known risk factors in intensive care units (ICU). An international multicenter cohort study (AspICU) established criteria for diagnosis of invasive pulmonary aspergillosis (IPA) in intensive care units. In our study, patients with Aspergillus spp. growth in deep tracheal aspirate (DTA) samples in ICU were evaluated according to AspICU criteria. Materials and Methods: This study is a retrospective study. DTA samples were collected from the Pandemic and Reanimation ICU and performed in the Medical Microbiology Laboratory by separated two periods;pre-pandemic (1 March 2019-31 December 2019) and post-pandemic (1 March 2020-31 December 2020). Cases with Aspergillus spp. growth in the DTA samples in the Pandemic ICU were evaluated as COVID 19 associated pulmonary aspergillosis (CAPA) according to AspICU criteria. Results: While Aspergillus spp. was grown in the DTA of three patients in 2019 and five patients in 2020 in the Reanimation ICU, and 11 patients in the Pandemic ICU. Growths belonging to one patient from both Reanimation (2019) and Pandemic ICUs were considered as colonization. Other growths were interpreted as IPA according to AspICU criteria. When the incidence rates according to 10000 patient days were compared, the incidence rate increased significantly in 2020 (19.1) (p< 0.001) compared to 2019 (3.4);In 2020, it was determined that it increased significantly in the Pandemic ICU (40.4) (p< 0.001) compared to Reanimation ICU (9.2). Conclusion: It should not be forgotten that intensive care patients are also at risk for IPA, especially after viral infections (such as COVID-19, Influenza). Although the incidence of IPA was not very high, it was observed that it tended to increase according to our study. The diagnosis of IPA is problematic, therefore it is necessary to increase awareness and sample diversity and to use biomarkers more widely other than hematology patients.

Evidence showing lipotoxicity worsens outcomes in covid-19 patients and insights about the underlying mechanisms.

We compared three hospitalized patient cohorts and conducted mechanistic studies to determine if lipotoxicity worsens COVID-19. Cohort-1 (n = 30) compared COVID-19 patients dismissed home to those requiring intensive-care unit (ICU) transfer. Cohort-2 (n = 116) compared critically ill ICU patients with and without COVID-19. Cohort-3 (n = 3969) studied hypoalbuminemia and hypocalcemia's impact on COVID-19 mortality. Patients requiring ICU transfer had higher serum albumin unbound linoleic acid (LA). Unbound fatty acids and LA were elevated in ICU transfers, COVID-19 ICU patients and ICU non-survivors. COVID-19 ICU patients (cohort-2) had greater serum lipase, damage-associated molecular patterns (DAMPs), cytokines, hypocalcemia, hypoalbuminemia, organ failure and thrombotic events. Hypocalcemia and hypoalbuminemia independently associated with COVID-19 mortality in cohort-3. Experimentally, LA reacted with albumin, calcium and induced hypocalcemia, hypoalbuminemia in mice. Endothelial cells took up unbound LA, which depolarized their mitochondria. In mice, unbound LA increased DAMPs, cytokines, causing endothelial injury, organ failure and thrombosis. Therefore, excessive unbound LA in the circulation may worsen COVID-19 outcomes.

Computed tomography and [18F]-FDG PET imaging provide additional readouts for COVID-19 pathogenesis and therapies evaluation in non-human primates.

Non-human primates (NHPs) are particularly relevant as preclinical models for SARS-CoV-2 infection and nuclear imaging may represent a valuable tool for monitoring infection in this species. We investigated the benefit of computed X-ray tomography (CT) and [18F]-FDG positron emission tomography (PET) to monitor the early phase of the disease in a large cohort (n = 76) of SARS-CoV-2 infected macaques. Following infection, animals showed mild COVID-19 symptoms including typical lung lesions. CT scores at the acute phase reflect the heterogeneity of lung burden following infection. Moreover, [18F]-FDG PET revealed that FDG uptake was significantly higher in the lungs, nasal cavities, lung-draining lymph nodes, and spleen of NHPs by 5 days postinfection compared to pre-infection levels, indicating early local inflammation. The comparison of CT and PET data from previous COVID-19 treatments or vaccines we tested in NHP, to this large cohort of untreated animals demonstrated the value of in vivo imaging in preclinical trials.

Distribution of anaerobic bacteria and their sensitivity pattern to several antibiotics at the clinical microbiology laboratory of school of medicine, universitas Indonesia, Jakarta in 2019-2020

Background and Objectives: Anaerobic bacteria are a common cause of endogenous infections, some of which can be life threatening. These bacteria are not easily cultured and isolated and often cannot even found from infected sites. Delayed or inappropriate treatment of these microorganisms can lead to failure in eradicating these infections. The purpose of this study was to determine the diversity of anaerobic bacteria at present and their pattern of sensitivity to several antibiotics. Materials and Methods: A retrospective study was conducted over a period of two years on various specimens. Specimens derived from body fluids are inoculated on a BacT/Alert (bioMérieux). Anaerobic isolates were identified by Gram staining and continued identification using Vitek 2® automated system. Antibiotic sensitivity examination was carried out using ATBTM ANA (bioMérieux). Results: A total of 440 specimens were received in microbiology laboratory for anaerobic culture from patients with multiple infections from 13 hospitals in Jakarta. Our research was able to identify 18 species on anaerobic bacteria, consisting 52.5% Gram positive and 47.5% Gram negative bacteria. The most common bacteria found were Clostridium perfringens (15%) from Gram positive and Provetella bivia (10%) from Gram negative. The sensitivity pattern shows that antibiotic piperacilline-tazobactam is 100% effective against anaerobic bacteria, while metronidazole as the drug of choice is only 75% effective. Against Gram positive, several antibiotics such as piperacilline-tazobactam, ticarcilin-clavunic acid, cefoxitin, cefotetan, imipenem and chloramphenicol were 100% effective, however metronidazole occupied the lowest position (61.9%). Meanwhile against Gram negative antibiotics piperacilline-tazobactam is 100% effective and chloramphenicol in the second position (94.75%). Conclusion: Clostridium perfringens and Provetella bivia are the most common bacteria found. The antibiotics piperacilline-tazobactam is 100% effective against both Gram positive and negative. The accuracy of specimen management, isolation, identification and sensitivity examination will determine the successful microbiological investigations. [ FROM AUTHOR] Copyright of Iranian Journal of Microbiology is the property of Tehran University of Medical Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)

An Automated Dashboard to Improve Laboratory COVID-19 Diagnostics Management.

Background: In response to the COVID-19 pandemic, our microbial diagnostic laboratory located in a university hospital has implemented several distinct SARS-CoV-2 RT-PCR systems in a very short time. More than 148,000 tests have been performed over 12 months, which represents about 405 tests per day, with peaks to more than 1,500 tests per days during the second wave. This was only possible thanks to automation and digitalization, to allow high throughput, acceptable time to results and to maintain test reliability. An automated dashboard was developed to give access to Key Performance Indicators (KPIs) to improve laboratory operational management. Methods: RT-PCR data extraction of four respiratory viruses-SARS-CoV-2, influenza A and B and RSV-from our laboratory information system (LIS), was automated. This included age, gender, test result, RT-PCR instrument, sample type, reception time, requester, and hospitalization status etc. Important KPIs were identified and the visualization was achieved using an in-house dashboard based on the open-source language R (Shiny). Results: The dashboard is organized into three main parts. The "Filter" page presents all the KPIs, divided into five sections: (i) general and gender-related indicators, (ii) number of tests and positivity rate, (iii) cycle threshold and viral load, (iv) test durations, and (v) not valid results. Filtering allows to select a given period, a dedicated instrument, a given specimen, an age range or a requester. The "Comparison" page allows a custom charting of all the available variables, which represents more than 182 combination. The "Data" page, gives the user an access to the raw data in tables format, with possibility of filtering, allowing for a deeper analysis and data download. Informations are updated every 4 h. Conclusions: By giving a rapid access to a huge number of up-to-date information, represented using the most relevant visualization types, without the burden of timely data extraction and analysis, the dashboard represents a reliable and user-friendly tool for operational laboratory management. The dashboard represents a reliable and user-friendly tool improving the decision-making process, resource planning and quality management.

Novel hybrid antiviral VTRRT-13V2.1 against SARS-CoV2 main protease: retro-combinatorial synthesis and molecular dynamics analysis.

The COVID-19 pandemic caused by SARS-CoV-2 has now emerged as a global health problem and is responsible for high mortality and morbidity. The SARS-CoV-2 main protease (Mpro) emerged as a promising drug target because of its essential role in the processing of polyproteins, which is translated from viral RNA. The present study reports a designed novel hybrid antiviral molecule (VTRRT-13.V2.1) against SARS-CoV2 main protease. A series of different combinations of hybrid antiviral were generated from nonspecific antiviral molecules currently used to control COVID-19. To enhance the specificity of the designed hybrid antiviral molecule, the core pocket region of the active site of Mpro protein was targeted. In-silico screening, molecular mechanics, molecular dynamics simulation (MDS) analysis identified a hybrid VTRRT-13.V2 molecule. Retrosynthetic analysis and combinatorial synthesis generated 1000 analogs of VTRRT-13.V2 molecules. Docking, molecular mechanics, and MDS analysis selected VTRRT-13.V2.1 as a possible inhibitor for SARS-CoV2 main protease. Comparative analysis of all the results showed that VTRRT-13.V2.1 have the highest docking Glide score (-12.28 kcal/mol) and best binding energy (-52.23 kcal/mol) as compared to the other hybrid constructs such as VTRRT-13.V2 (-9.47 and -47.36 kcal/mol), VTRRT-13 (-8.9 and -47.55 kcal/mol), and current antiviral investigated. The mutational sensitivity screening showed that binding residues of Mpro are not present in mutation hotspots. It was also observed that VTRRT-13.V2.1 does not have any human off-targets. SARS-CoV2 main protease is essential for the survival of this virus; hence, a designed novel hybrid antiviral molecule (VTRRT-13.V2.1) might be useful to control the infection of COVID-19 infection.