RAPID IDENTIFICATION OF ETIOLOGICAL AGENTS OF RESPIRATORY TRACT INFECTIONS DURING THE COVID-19 PANDEMIC USING THE MULTIPLEX-PCR METHOD.

Objective. Most respiratory infections have similar symptoms, so it is clinically difficult to determine their etiology. This study aimed to show the importance of molecular diagnostics in identifying the etiological agent of respiratory infections, especially during the coronavirus disease 2019 (COVID-19) pandemic. Methods. A total of 849 samples from patients hospitalized at the University Clinical Center Kragujevac (from January 1 to August 1, 2022) were examined using automated multiplex-polymerase chain reaction (PCR) tests. The BioFire-FilmArray-Respiratory Panel 2.1 test was used for 742 nasopharyngeal swabs [identification of 19 viruses (including SARS-CoV-2) and four bacteria], while the BioFire-FilmArray-Pneumonia Panel was used [identification of 18 bacteria and nine viruses] (BioMerieux, Marcy l'Etoile, France) for 107 tracheal aspirates. The tests were performed according to the manufacturer's instructions, and the results were available within an hour. Results. In 582 (78.4%) samples, the BioFire-FilmArray-Respiratory Panel 2.1 plus test identified at least one pathogen. The rhinovirus (20.6%), SARS-CoV-2 (17.7%), influenza A (17.5%), respiratory syncytial virus (12.4%), and parainfluenza 3 (10.1%) were the most common. Other viruses were found less frequently, and Bordetella parapertussis was detected in one sample. In 85 (79.4%) samples, the BioFire-FilmArray-Pneumonia Panel test identified at least one bacterium or virus. The most prevalent bacteria were Staphylococcus aureus (42.4%), Haemophilus influenzae (41.2%), Streptococcus pneumoniae (36.5%), Moraxella catarrhalis (22.3%), and Legionella pneumophila (2.4%). Among viruses, rhinovirus (36.5%), adenovirus (23.5%), influenza A (11.8%), and the genus Coronavirus (4.7%), were detected. Conclusion. Multiplex-PCR tests improved the implementation of therapeutic and epidemiological measures, preventing the spread of the COVID-19 infection and Legionnaires' disease.Copyright © 2022, Serbian Medical Society. All rights reserved.

Prevalence of multidrug-resistant Gram-negative bacteria from blood cultures and rapid detection of beta-lactamase-encoding genes by multiplex PCR assay

Introduction This study aimed to determine the prevalence of multidrug-resistant Gram-negative bacteria (GNB) from blood cultures in a tertiary-care hospital and the multiplex PCR assay's ability to detect resistance genes. Methods A total of 388 GNB isolates obtained from hospitalized patients between November 2019 and November 2021 were included in the study. Antimicrobial susceptibility testing was done by VITEK 2 system and broth microdilution method. Beta-lactamase-encoding genes were detected by multiplex PCR assays, BioFire-Blood Culture Identification 2 (BCID2) panel (bioMerieux, France). Extended-spectrum beta-lactamases (ESBLs) were detected phenotypically with VITEK AST-GN71 card (bioMerieux, France). The isolates of GNB were classified into multidrug-resistant, extensively-drug-resistant, and pandrug-resistant categories, and their prevalence and distribution in different wards, including coronavirus diseases 2019 (COVID-19) intensive care units (ICU), were calculated. Results Results revealed that all isolates of Acinetobacter baumannii and Pseudomonas aeruginosa were multidrug-resistant as well as 91.6% of Enterobacter cloacae, 80.6% of Proteus mirabilis, and 76.1% of Klebsiella pneumoniae, respectively. In fermentative bacteria, blaOXA-48-like (58.1%), blaNDM (16.1%), blaKPC (9.7%) and blaVIM (6.5%) genes were detected. More than half of Enterobacter cloacae (58.3%) and Klebsiella pneumoniae (53.7%) produced ESBLs. Among non-fermenters, the blaNDM gene was carried by 55% of Pseudomonas aeruginosa and 19.5% of Acinetobacter baumannii. In the COVID-19 ICU, Acinetobacter baumannii was the most common isolate (86.1%). Conclusions This study revealed high proportions of multidrug-resistant blood isolates and various underlying resistance genes in Gram-negative strains. The BCID2 panel seems to be helpful for the detection of the most prevalent resistance genes of fermentative bacteria.Copyright © GERMS 2022.

Clinical influence of multiplex polymerase chain reaction routine uses in urgent pediatric admissions.

BACKGROUND: The coronavirus disease 2019 outbreak has prompted some hospitals to implement screening tests upon admission since 2020. FilmArray® Respiratory 2.1 Panel (FilmArray) is a multiplex polymerase chain reaction (PCR) test with high sensitivity and specificity for detecting respiratory pathogens. We aimed to assess the clinical influence of the routine use of FilmArray for pediatric patients, including those without symptoms suggestive of an infection. METHODS: We conducted a single-center retrospective observational study, which investigated patients aged ≤15 years who underwent FilmArray on admission in 2021. We collected the patients' epidemiological information, symptoms, and FilmArray results from their electronic health records. RESULTS: A positive result was observed in 58.6% of patients admitted to the general ward or intensive care unit (ICU) but only in 1.5% of patients in the neonatal ward. Among the patients admitted to the general ward or ICU who tested positive, 93.3% had symptoms suggestive of infections, 44.6% had a sick contact before admission, and 70.5% had siblings. However, 62 (28.2%) out of 220 patients without the four (fever, respiratory, gastrointestinal, and dermal) symptoms also had positive results. Among them, 18 patients with adenovirus and three with respiratory syncytial virus were isolated to private rooms. However, 12 (57.1%) patients were discharged without symptoms suggestive of viral infection. CONCLUSION: Multiplex PCR routine use for all inpatients may lead to excessive management of positive cases because FilmArray cannot quantify microorganisms. Thus, targets for testing should be considered carefully based on patients' symptoms and histories of sick contacts.

RAPID IDENTIFICATION OF ETIOLOGICAL AGENTS OF RESPIRATORY TRACT INFECTIONS DURING THE COVID-19 PANDEMIC USING THE MULTIPLEX-PCR METHOD

Objective. Most respiratory infections have similar symptoms, so it is clinically difficult to determine their etiology. This study aimed to show the importance of molecular diagnostics in identifying the etiological agent of respiratory infections, especially during the coronavirus disease 2019 (COVID-19) pandemic. Methods. A total of 849 samples from patients hospitalized at the University Clinical Center Kragujevac (from January 1 to August 1, 2022) were examined using automated multiplex-polymerase chain reaction (PCR) tests. The BioFire-FilmArray-Respiratory Panel 2.1 test was used for 742 nasopharyngeal swabs [identification of 19 viruses (including SARS-CoV-2) and four bacteria], while the BioFire-FilmArray-Pneumonia Panel was used [identification of 18 bacteria and nine viruses] (BioMérieux, Marcy l'Etoile, France) for 107 tracheal aspirates. The tests were performed according to the manufacturer's instructions, and the results were available within an hour. Results. In 582 (78.4%) samples, the BioFire-FilmArray-Respiratory Panel 2.1 plus test identified at least one pathogen. The rhinovirus (20.6%), SARS-CoV-2 (17.7%), influenza A (17.5%), respiratory syncytial virus (12.4%), and parainfluenza 3 (10.1%) were the most common. Other viruses were found less frequently, and Bordetella parapertussis was detected in one sample. In 85 (79.4%) samples, the BioFire-FilmArray-Pneumonia Panel test identified at least one bacterium or virus. The most prevalent bacteria were Staphylococcus aureus (42.4%), Haemophilus influenzae (41.2%), Streptococcus pneumoniae (36.5%), Moraxella catarrhalis (22.3%), and Legionella pneumophila (2.4%). Among viruses, rhinovirus (36.5%), adenovirus (23.5%), influenza A (11.8%), and the genus Coronavirus (4.7%), were detected. Conclusion. Multiplex-PCR tests improved the implementation of therapeutic and epidemiological measures, preventing the spread of the COVID-19 infection and Legionnaires' disease. © 2022, Serbian Medical Society. All rights reserved.

Long Term Circulation of Sars-Cov-2 Related Lineages in Bats in Cambodia

Intro: Recent evidence shows the Greater Mekong Subregion to be a hotspot for Sarbecoviruses in bats, especially insectivorous Horseshoe bats (genus Rhinolophus). However, prevalence, maintenance, and evolution of these viruses in Rhinolophids is still poorly understood. Sampling efforts are still limited and generally only cover cross-sectional surveillance at single points in time. Following the detection of Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2)-related viruses in Rhinolophus shameli from 2010 in Steung Treng, Cambodia, further active longitudinal surveillance in the same area between 2020-2021 continued the detection of these viruses. Method(s): Live bat capture and sampling has been implemented in several sites located in Stung Treng province. All rectal swabs of bats were tested for the detection of SARS-CoV-2 or Sarbecoviruses by real time RT-PCR. RNA samples from positive RT-PCR bats were then sequenced using a highly multiplexed PCR amplicon approach using new designed primers set guided by the ARTIC Network multiplex PCR primers set (https://artic.network/ncov-2019), on Oxford Nanopore technology. Finding(s): The sarbecoviruses were detected in four Rhinolophus shameli bats, a percentage of similarity ranging at the nucleotide level between 98.8% - 99.1% when compared to two other Cambodian bat sarbecoviruses from 2010 and between 92.4% - 94.5% when compared to human SARS-CoV-2 across the whole genome. Discussion(s): The bat SARS-CoV-2 related virus recently detected in four positive bats in 2020-2021 are genetically homologous with the virus detected in 2010, indicating a geographically/host limited population that is stable over time in the past ten years. Conclusion(s): Overall, our findings indicate further complexity in the diversity and evolution of sarbecoviruses and add intricacy to the search for the origins of the Coronavirus Disease 2019 (COVID-19) pandemic.Copyright © 2023

RAPID IDENTIFICATION OF ETIOLOGICAL AGENTS OF RESPIRATORY TRACT INFECTIONS DURING THE COVID-19 PANDEMIC USING THE MULTIPLEX-PCR METHOD.

Objective. Most respiratory infections have similar symptoms, so it is clinically difficult to determine their etiology. This study aimed to show the importance of molecular diagnostics in identifying the etiological agent of respiratory infections, especially during the coronavirus disease 2019 (COVID-19) pandemic. Methods. A total of 849 samples from patients hospitalized at the University Clinical Center Kragujevac (from January 1 to August 1, 2022) were examined using automated multiplex-polymerase chain reaction (PCR) tests. The BioFire-FilmArray-Respiratory Panel 2.1 test was used for 742 nasopharyngeal swabs [identification of 19 viruses (including SARS-CoV-2) and four bacteria], while the BioFire-FilmArray-Pneumonia Panel was used [identification of 18 bacteria and nine viruses] (BioMerieux, Marcy l'Etoile, France) for 107 tracheal aspirates. The tests were performed according to the manufacturer's instructions, and the results were available within an hour. Results. In 582 (78.4%) samples, the BioFire-FilmArray-Respiratory Panel 2.1 plus test identified at least one pathogen. The rhinovirus (20.6%), SARS-CoV-2 (17.7%), influenza A (17.5%), respiratory syncytial virus (12.4%), and parainfluenza 3 (10.1%) were the most common. Other viruses were found less frequently, and Bordetella parapertussis was detected in one sample. In 85 (79.4%) samples, the BioFire-FilmArray-Pneumonia Panel test identified at least one bacterium or virus. The most prevalent bacteria were Staphylococcus aureus (42.4%), Haemophilus influenzae (41.2%), Streptococcus pneumoniae (36.5%), Moraxella catarrhalis (22.3%), and Legionella pneumophila (2.4%). Among viruses, rhinovirus (36.5%), adenovirus (23.5%), influenza A (11.8%), and the genus Coronavirus (4.7%), were detected. Conclusion. Multiplex-PCR tests improved the implementation of therapeutic and epidemiological measures, preventing the spread of the COVID-19 infection and Legionnaires' disease.Copyright © 2022, Serbian Medical Society. All rights reserved.

Role of antiviral therapy in respiratory infections in children: Analysis of clinical and laboratory observations.

Objective. To evaluate the efficacy of therapy for acute respiratory viral infections (ARVIs) in children with antiviral medications: inosine pranobex (Groprinosin, Gedeon Richter) and Kagocel (Kagocel, Niarmedic Pharma LLC) in comparison with symptomatic treatment without etiotropic agents based on clinical and laboratory parameters. Patients and methods. The clinical and laboratory observation was conducted in an outpatient setting in the pre-COVID-19 period between 2018 and 2020. Acute respiratory infections were diagnosed using licensed testing systems by multiplex polymerase chain reaction (PCR) with detection of nucleic acid viral genomes: influenza, rhinovirus, respiratory syncytial virus, metapneumovirus, parainfluenza, seasonal coronaviruses, adenoviruses, and bocavirus). A total of 151 children aged 3 to 15 years were examined and monitored in dynamics, with 78.7% of positive and 21.3% of negative results detected by PCR in the nasopharyngeal and oropharyngeal swabs. The patients were randomized into three groups depending on the antiviral medication prescribed: group 1 (53 children) received Groprinosin;group 2 (52 children) received Kagocel;group 3 (control, 46 children) received only symptomatic therapy without antiviral agents. Results. The study demonstrated a significant positive effect in patients in group 1 treated with Groprinosin (n = 53). At the end of therapy for both mono- and mixed infections, there were 95.8% of negative results (according to PCR diagnosis, that is, the absence of viral genome). In children in group 2 (n = 52) treated with Kagocel, the absence of viral nucleic acids (NAs) was observed less frequently (in 77.3% of cases). In children in group 3 (n = 46) who did not receive etiotropic antiviral therapy, there were only 40.3% of negative results after the end of treatment, and viral NAs were detected in 59.7% of patients. In this case, a 5-day course of Groprinosin was prescribed, after which the PCR results became negative in all patients. Therefore, children with recurrent respiratory infections, mixed infections, and herpesvirus infections require longer therapy. Additionally, a high frequency of ARVI complications was noted in group 3 (5 (10.9%) patients, where otitis was observed in 1 case, sinusitis - in 2 cases, bronchitis - in 2 cases), whereas 1 (1.8%) patient taking Groprinosin had otitis, and 1 (1.9%) patient taking Kagocel had pneumonia. Conclusion. This study was the first to investigate antibody titers to respiratory viruses in dynamics at 3, 6 and 12 months after the onset of ARVI. It showed that the development of antibodies to respiratory viruses is very unstable and does not occur in all patients. Antibodies almost disappeared by the third month after ARVI and were no longer detectable by the sixth month. After 12 months, patients suffered a new ARVI and developed the corresponding antibodies. This information will be especially relevant in conditions of the rise in the incidence of ARVIs, as well as the COVID-19 pandemic observed in recent years.Copyright © 2022, Voprosy Prakticheskoi Pediatrii. All rights reserved.

Pulmonary function in non-hospitalized adults and children after mild Covid-19

Objective Asesmentof lung function impairment after mild SARS-CoV-2 infection in non-hospitalized children and adults. Additionally focusing on previous and persistent symptoms due to Covid-19 as well as current respiratory tract infection status. Methods Patients aged 6-60 years were recruited by telephone after laboratory-confirmed positive PCR result for SARS-CoV-2. Excluding criteria were hospitalization during Covid-19, pre-existing lung diseases (bronchial asthma, COPD) and smoking within the last five years. Pulmonary function testing was performed 4-12 weeks after infection, including Multiple-breath washout (LCI), spirometry (FEV?, FVC, Tiffeneau-Index) and diffusion capacity testing (DLCO, TLC;Hb corrected). All patients answered a questionnaire regarding previous and persistent symptoms. To gather information about the current infection status, a pharyngeal swab was taken to detect common respiratory bacterial and viral pathogens using a multiplex PCR approach. Patients with abnormalities in pulmonary function were invited to a follow up testing three months later. Results 110 patients, 90 adults and 20 children, were included. 44 adults and 17 children had at least one abnormal value in pulmonary function tests after an average of 7.7 weeks (range 4.3-11.3) to confirmed SARS-Cov-2 infection. Among these 44 adults, 33 reported pulmonary symptoms during Covid-19 and 19 persistent respiratory symptoms. No abnormalities in DLCO were found in adults. At the second pulmonary function testing 12.5 weeks (range 11.0-16.7) on average after the first appointment, improvement was shown in 61,7% ( n=29 of 47) with previous abnormal LCI, in 69,2% (n=9 of 13) with prior abnormal FVC and in 4 of 5 children with abnormal DLCO. No large correlation was detected between impaired pulmonary function and multiplex PCR results. Conclusion Mild lung function impairment was shown at the first appointment, particularly in LCI, but not equally measured in the entirety of lung function tests. Pulmonary function results were not affected by current infection status and partially mismatching with stated persisting symptoms. Within 3 months, most initially abnormal values improved, and self- perceived health status increased. Long term pulmonary function impairment was rarely detected after mild, non-hospitalized Covid-19 course. .

Bacterial and viral upper and lower airways infection in people with cystic fibrosis in stable condition

Background: Bacterial and viral airway infections are adverse factors for prognosis in people with cystic fibrosis (CF). The role of viral infections is unclear. Proper microbiological follow-up is essential, and the correlation between upper (UAW) and lower airway (LAW) microbiology may be important for lung disease management. We aim to evaluate airway microbiology in patients in stable clinical condition. Method(s): Between September 2021 and March 2022 in the Florence CF center, 144 nasal lavage-throat swab paired samples were collected from 72 clinically stable people with CF not chronically colonized by Pseudomonas aeruginosa. The study enrolled 59 children (median age 9, range 2-16) and 13 adults (median age 28, range 18-59). LAW specimens (72)were sampled as throat swab and UAWspecimens (72)were randomly collected by nasal lavage with two methods-Mainz (44) or Ryno-set (28). We performed conventional microbiological analyses on all samples. To screen for respiratory viruses, multiplex polymerase chain reaction (BioFire FilmArray RP 2.1 Plus) was performed. Respiratory symptoms and forced expiratory volume in 1 second (FEV1) valueswere evaluated for all patients. Result(s): Twenty-one (29%) patients tested positive for at least one virus in UAW and LAW specimens. The most frequently identified viruses were human rhinovirus or enterovirus (22%) and respiratory syncytial virus (6%). Two (3%) patients tested positive for SARS-CoV-2. Concordance between sampling methods for viral detection in UAW and LAW specimens was observed in 59 paired samples (82%), including 40 patients with no viral infections and 19 virus positive for both samples. Discordance was described in 13 subjects;10 of 13 did not show viral infection in nasal lavage. Twenty-one percent of positive nasal lavage was performed using Ryno-set and 36% using the Mainz approach. The prevalent bacteriumwas Staphylococcu aureus in UAW (53%) and LAW (69%) cultures, followed by Enterobacteriaceae (UAW 8%, LAW 6%), methicillin-resistant S. aureus (UAW 7%, LAW 6%), P. aeruginosa (UAW 4%, LAW 6%), and other clinically relevant gram-negative bacteria such as Achromobacter xylosoxidans, Stenotrophomonas maltophilia, and Ochrobactrum anthropi (UAW 7%, LAW 13%). Nasal lavage performed with Ryno-set tested positive in 72% of patients, and 64% of Mainz lavage were positive. Mainz nasal lavage showed different S. aureus and P. aeruginosa isolations (48% and 5%, respectively) than the samples collected with Ryno-set technique (61% and 4%, respectively). Concordance between sampling methods for bacterial detection in UAW and LAW was the same with the two methods (53%). Bacterial and viral infections were found in UAWand LAWof stable people with CF, but no clinical correlation was observed. Conclusion(s): The two methods of UAW lavage had slight differences in performance. Virus infection appeared to be less prevalent than bacterial infection in UAWand LAW.We did not find correlations between presence of viruses and respiratory symptoms, but further investigation is needed for a better understanding of the clinical role of viral infection in people with CF.Copyright © 2022, European Cystic Fibrosis Society. All rights reserved

Advances in molecular diagnostic tests for community-acquired pneumonia

The use of a timely and appropriate antibiotic therapy, which requires early and accurate microorganisms' detection in pneumonia. Currently, the identification of microorganisms in pneumonia is limited by the low sensitivity and long response time of standard culture-based diagnostic tools. For this reason, treatment in pneumonia is empirical. An inadequate empirical treatment is related to poor outcomes in patients with pneumonia. The microbiological diagnosis is key to improve the outcomes in patient with pneumonia. Over the past years there was a significant advance in the molecular diagnosis of infectious diseases including pneumonia. Also the impact of the COVID-19 pandemic has impacted the development and application of these new molecular techniques. This review summarizes the advances in molecular diagnosis of community-acquired pneumonia.Copyright © 2022 EDIZIONI MINERVA MEDICA.

Tumor agnostic comparison of immunohistochemistry and next-generation sequencing in detecting ALK fusions and assessment of ALK tyrosine kinase inhibitor efficacy

Background: Reliable methods to identify anaplastic lymphoma kinase (ALK) fusions are critical to matching patients to ALK tyrosine kinase inhibitors (TKIs) therapy, on or off trial. Various methods including FISH have been used, but immunohistochemistry (IHC) and next-generation sequencing (NGS) are most commonly employed. Evaluating the concordance of IHC and NGS is key, particularly in non-lung cancers where data is sparse. Method(s): NGS+ (MSK-IMPACT DNA hybrid capture NGS and/or RNA anchored multiplex PCR) and/or IHC+ (clone: D5F3) patients with cancers of any histology were identified as ALK+. ALK IHC was scored as negative (0), equivocal (e: 1+, 2+) or positive (3). Concordance of ALK detection (number of NGS+ and IHC+/total number of patients with NGS and IHC) was calculated. For patients with metastatic disease treated with any ALK TKI in the first-line (1L) setting, progression-free survival (PFS) was reported. Result(s): 347 ALK+ solid tumor patients were identified. As expected, the majority (96%, n=336) had lung cancer, however, 11 patients with 11 unique non-lung cancer histologies were found (3 gastrointestinal, 2 gynecologic, 1 breast, 1 thyroid, 1 primary brain tumor, 1 DLBCL, 1 PEComa, and 1 CUP). 57% had EML4-ALK fusions;36 non-EML4 ALK rearrangements were identified, including four novel fusions (PEKHA7-ALK, ZFPM2-ALK, TRIM24-ALK, ALK-MYO3B). ALK was evaluated by IHC alone in 83 patients (23.9%). The concordance rate between NGS and IHC was 85%. Among discordant cases, 11% (n=28) were IHC+/NGS-, 24% (n=63) were IHCe/NGS-, 3% (n=8) were IHCe/NGS+, and 0.4% (n=1) was IHC-/NGS+. The most frequent ALK TKIs were alectinib (n= 87, 58%) and crizotinib (n= 56, 38%). PFS on 1L ALK TKIs for patients with IHC+/NGS+ (n=134), IHC-/NGS+(n=1), IHC+/NGS- (n=8), IHCe/NGS+ (n=4), IHCe/NGS- (n=1) was 26 months, 26 months, 39 months, 41 months, 9 months respectively. Conclusion(s): In a population including multiple tumor types, NGS and IHC were highly concordant in ALK fusion detection. ALK TKI benefit may be observed in cases with discordant testing, in which only one assay detects a putative ALK fusion. Legal entity responsible for the study: The authors. Funding(s): NIH Cancer Center grant: P30CA008748. Disclosure: M.G. Kris: Financial Interests, Personal, Research Grant: Boehringer Ingelheim, National Lung Cancer Partnership, Pfizer, PUMA, Stand up to Cancer;Financial Interests, Personal, Advisory Role: Ariad, AstraZeneca, Bind Bioscience, Boehringer Ingelheim, Chug Pharma, Clovis, Covidien, Daiichi Sankyo, Esanex, Genentech;Financial Interests, Personal, Invited Speaker: Boehringer Ingelheim, Novartis, Millenium, Pfizer, Roche. A. Drilon: Financial Interests, Personal, Advisory Board: Ignyta/Genentech/Roche, Loxo/Bayer/Lilly, Takeda/Ariad/Millennium, TP Therapeutics, AstraZeneca, Pfizer, Blueprint Medicines, Helsinn, BeiGene, BerGenBio, Hengrui Therapeutics, Exelixis, Tyra Biosciences, Verastem Oncology, MORE Health, AbbVie, 14ner/Elevation Oncology, Remedica Ltd, ArcherDX, Monopteros, Novartis, EMD Serono, Melendi, Liberum, Repare RX, Amgen, Janssen, EcoR1, Monte Rosa;Financial Interests, Personal, Other, CME: Medscape, Onclive, PeerVoice, Physicians Education Resources, Targeted Oncology, Research to Practice, PeerView Institute, Paradigm Medical Communications, WebMD, MJH Life Sciences, Med Learning, Imedex, Answers in CME, Medscape, Clinical Care Options, AiCME;Financial Interests, Personal, Other, CME, Consulting: Axis;Financial Interests, Personal, Other, Consulting: Nuvalent, Merus, EPG Health, mBrace, Harborside Nexus, Ology, TouchIME, Entos, Treeline Bio, Prelude, Applied Pharmaceutical Science, Inc;Financial Interests, Personal, Invited Speaker: Chugai Pharmaceutical, Remedica Ltd, RV More;Financial Interests, Personal, Stocks/Shares: Treeline Biosciences;Financial Interests, Personal, Royalties: Wolters Kluwer;Financial Interests, Personal, Other, stocks: mBrace;Financial Interests, Institutional, Funding, Research funding: Pfizer, Exelixis, GlaxoSmithKline, Teva, Taiho, PharmaMar;Finan ial Interests, Personal, Funding, Research: Foundation Medicine;Non-Financial Interests, Personal, Member: ASCO, AACR, IASLC;Other, Personal, Other, Food/Beverage: Merck, PUMA, Merus;Other, Personal, Other, Other: Boehringer Ingelheim. All other authors have declared no conflicts of interest.Copyright © 2023 European Society for Medical Oncology

Role of antiviral therapy in respiratory infections in children: Analysis of clinical and laboratory observations.

Objective. To evaluate the efficacy of therapy for acute respiratory viral infections (ARVIs) in children with antiviral medications: inosine pranobex (Groprinosin, Gedeon Richter) and Kagocel (Kagocel, Niarmedic Pharma LLC) in comparison with symptomatic treatment without etiotropic agents based on clinical and laboratory parameters. Patients and methods. The clinical and laboratory observation was conducted in an outpatient setting in the pre-COVID-19 period between 2018 and 2020. Acute respiratory infections were diagnosed using licensed testing systems by multiplex polymerase chain reaction (PCR) with detection of nucleic acid viral genomes: influenza, rhinovirus, respiratory syncytial virus, metapneumovirus, parainfluenza, seasonal coronaviruses, adenoviruses, and bocavirus). A total of 151 children aged 3 to 15 years were examined and monitored in dynamics, with 78.7% of positive and 21.3% of negative results detected by PCR in the nasopharyngeal and oropharyngeal swabs. The patients were randomized into three groups depending on the antiviral medication prescribed: group 1 (53 children) received Groprinosin;group 2 (52 children) received Kagocel;group 3 (control, 46 children) received only symptomatic therapy without antiviral agents. Results. The study demonstrated a significant positive effect in patients in group 1 treated with Groprinosin (n = 53). At the end of therapy for both mono- and mixed infections, there were 95.8% of negative results (according to PCR diagnosis, that is, the absence of viral genome). In children in group 2 (n = 52) treated with Kagocel, the absence of viral nucleic acids (NAs) was observed less frequently (in 77.3% of cases). In children in group 3 (n = 46) who did not receive etiotropic antiviral therapy, there were only 40.3% of negative results after the end of treatment, and viral NAs were detected in 59.7% of patients. In this case, a 5-day course of Groprinosin was prescribed, after which the PCR results became negative in all patients. Therefore, children with recurrent respiratory infections, mixed infections, and herpesvirus infections require longer therapy. Additionally, a high frequency of ARVI complications was noted in group 3 (5 (10.9%) patients, where otitis was observed in 1 case, sinusitis - in 2 cases, bronchitis - in 2 cases), whereas 1 (1.8%) patient taking Groprinosin had otitis, and 1 (1.9%) patient taking Kagocel had pneumonia. Conclusion. This study was the first to investigate antibody titers to respiratory viruses in dynamics at 3, 6 and 12 months after the onset of ARVI. It showed that the development of antibodies to respiratory viruses is very unstable and does not occur in all patients. Antibodies almost disappeared by the third month after ARVI and were no longer detectable by the sixth month. After 12 months, patients suffered a new ARVI and developed the corresponding antibodies. This information will be especially relevant in conditions of the rise in the incidence of ARVIs, as well as the COVID-19 pandemic observed in recent years.Copyright © 2022, Voprosy Prakticheskoi Pediatrii. All rights reserved.

Diagnosis of SARS-CoV-2 infection: available tests and strategy in the Intensive Care Unit.

Since the beginning of the COVID-19 pandemic, considerable improvement have been made in the technical aspects and in our understanding of diagnostic strategy for SARS-CoV-2 infection. The RT-PCR is still the cornerstone of diagnostic testing for SARS-CoV-2 infection but the clinician should be aware of its imperfect sensibility in particular for mild or moderate COVID-19, and order a second test and/or a CT-scan if needed. Results of all tests should be interpreted taking into account the pretest probability of COVID-19, which depends on the local incidence of the disease and on the patient's symptoms. New molecular testing approaches like multiplex PCR, the use of antigen test and serologic testing now complete the available diagnostic procedures.Copyright © SRLF 2021.

Viral Identification Using Multiplex Polymerase Chain Reaction Testing Does Not Reduce Antibiotic Prescribing in Paediatric Intensive Care Units.

PCR tests for viral identification, performed on nasopharyngeal secretions, have experienced a major boom in the last few years. Their use is very frequent, but their indications are still not well defined, especially in Paediatric Intensive Care Units (PICU). These tests are used for the microbiological diagnosis of lower respiratory infections but can be used in other situations. The aim of the study was to investigate the effect of viral identification on antibiotic therapy management. We conducted a single-centre retrospective study from 1 October 2017 to 31 December 2019. This study included all consecutive FilmArray® Respiratory Panel tests performed in patients hospitalised in a PICU. Patients were identified using the microbiology laboratory prospective database and data were extracted from the medical record. 544 tests corresponding to 408 patients were included. The main reasons for testing were pneumonia (34%) and bronchiolitis (24%). In 70% of cases, at least one virus was identified, with Human Rhinovirus (56%) and Respiratory Syncytial Virus (28%) being the two predominant. Bacterial co-infection was present in 25% of cases. Viral identification was not associated with reduced antibiotic therapy. On multivariate analysis, antibiotic management was significantly associated with clinical gravity, CRP value or radiology findings regardless of virus identification. Viral identification has an epidemiological value, but antibiotic prescription relies on other factors.

Analysis of drug resistance gene in Mycoplasma pneumoniae and 13 pathogens in bronchoalveolar lavage fluid of children with Mycoplasma pneumoniae pneumonia. TT -.

Objective To investigate drug resistance gene in Mycoplasma pneumoniae (MP) and the distribution of 13 respiratory pathogens in bronchoalveolar lavage fluid(BALF) of children with Mycoplasma pneumoniae pneumonia(MPP). Methods A total of 100 BALF of children with MPP in Peking University Third Hospital and Peking University First Hospital from January 2018 to January 2019 were collected. Fluorogenic quantitative PCR was used to detect nucleic acid and it's drug resistance gene of MP and multiple PCR method was adopted to detect influenza A virus, influenza A virus - H1 N1, influenza A virus - H3 N2, influenza B, human parainfluenza virus, adenovirus, human bocavirus, human rhino virus, Chlamydia pneumoniae, human metapneumovirus, MP, human corona virus, and respiratory syncytial virus gene, and the results were compared by using Chi square test. Results In 100 BALF samples, MP and drug resistance gene were detected by fluorogenic quantitative PCR. Totally, 83 cases (83. 00%) were MP positive and 78 cases (93. 98%) were drug resistant. All of them had the point mutations A2063G in V region of 23S rRNA domain. A total of 13 kinds of respiratory pathogens were detected by multiplex PCR method, and 89 cases (89. 00%) were positive. Totally, 79 cases (79. 00%) were MP positive, of which 74 cases (74. 00%) detected only MP, and 5 cases (5. 00%) detected MP combined with other pathogens. Other pathogens were detected in 10 cases (10. 00%). The virus detection rate of 0-4 years old group was higher than that of > 4-6 years old group (P - 0. 042) and > 6 years old group (P =0. 002), and the differences were statistically significant. Conclusions MP can be detected in most BALF samples of MPP children, the drug resistance phenomenon is serious, and the main point mutation is A2063G. There were other respiratory pathogens and 2 or 3 pathogens were detected in a small number of BALF samples.Copyright © 2022 Authors. All rights reserved.

The usefulness of a nasopharyngeal multiplex polymerase chain reaction system to screen pediatric patients with preoperative fever.

BACKGROUND: When children have a preoperative fever, anesthesiologists must help determine whether to postpone or proceed with surgery, as fever may be a sign of upper respiratory tract infection (URTI). Such infections are a known risk factor for perioperative respiratory adverse events (PRAEs), which are still one of the prime causes of anesthetic mortality and morbidity in pediatric patients. Since the COVID-19 pandemic, preoperative assessments have become drastically more complex as hospitals strive to balance practicality and safety. In our facility, if pediatric patients presented with preoperative fever, we used the FilmArray® Respiratory Panel 2.1 to determine whether to postpone or proceed with surgery. METHODS: This is a single-center retrospective observational study evaluating the efficacy of the FilmArray® Respiratory Panel 2.1 as a preoperative screening test. This study included pediatric patients scheduled for elective surgeries between March 2021 and February 2022. FilmArray was used if a patient had a preoperative fever (determined by axillary temperature, ≥38°C for <1-year-old, ≥37.5°C for ≥1-year-old) between hospital admission and before surgery. We excluded patients if they had apparent symptoms of URTI. RESULTS: In the FilmArray positive group, 11 of 25 (44%) cases developed subsequent symptoms after surgery was canceled. No patients in the negative group developed symptoms. The proportion of the development of subsequent symptoms between the FilmArray positive and negative groups was statistically significant (p < .001, odds ratio: 29.6, 95% confidence interval: [3.80-1356.01]). CONCLUSIONS: Our retrospective observational study revealed that 44% of the FilmArray positive group subsequently developed symptoms, and no PRAEs were observed in the FilmArray negative group. We suggest that FilmArray could be useful as a screening test for pediatric patients with preoperative fever.

Present status of microfluidic PCR chip in nucleic acid detection and future perspective

Polymerase chain reaction (PCR) amplifies specific fragment of DNA molecules and has been extensively applied in fields of pathogens and gene mutation detection, food safety and clinical diagnosis which on the other hand, holds the drawbacks of large size instrument, high heat dissipation etc. It has been demonstrated that microfluidics technique coupling with PCR reaction exhibits characteristics of integration, automatization, miniaturization, and portability. Meanwhile, various designed fabrication of microchip could contribute to diverse applications. In this review, we summarized major works about a variety of microfluidic chips equipped with several kinds of PCR techniques (PCR, RT-PCR, mPCR, dPCR) and detection methods like fluorescence, electrochemistry, and electrophoresis detection. The development and application of PCR-based microfluidic chip in pathogen and gene mutation detection, diseases prevention and diagnosis, DNA hybridization and low-volume sample treatment were also discussed. Copyright © 2022 Elsevier B.V.

Acute flaccid myelitis diagnosed after IVIG: a diagnostic dilemma

Case Report: A 4-year-old African American male presented to an outside emergency department (ED) following sudden inability to move left upper extremity. Past medical history was unremarkable and routine vaccinations were up to date. Radiograph of affected extremity ruled out fractures and patient was discharged to follow up with primary care physician. Two days later mother brought him to our ED due to persistent left upper extremity paralysis, poor appetite, and subjective fever. On exam his left arm was warm and tender to dull and sharp touch;he had definite loss of active movement, hypotonia and absence of deep tendon reflexes. The patient had winging of left scapula and could not shrug left shoulder. MRI of cervical and thoracic spine showed enlargement of spinal cord from C2-C6 level with gray matter hyperintensity, slightly asymmetric to the left. Laboratory studies showed leukocytosis (14 000/mcL) and CSF studies showed pleocytosis of 89 WBC/mcL (93.3% mononuclear cells and 6.7% polymorphonuclear cells), 0 RBCs, normal glucose and protein, and a negative CSF meningoencephalitis multiplex PCR panel. Due to high suspicion of demyelinating or autoimmune condition he was treated with high dose steroids and IVIG. Subsequently neuromyelitis optica was ruled out as aquaporin-4 receptor antibodies (AB) and myelin oligodendrocyte glycoprotein AB were normal. CSF myelin basic protein and oligoclonal bands were absent ruling out demyelinating disorders. CSF arboviruses IgM and West Nile IgM were negative. He showed minimal improvement in left upper extremity movement but repeat spinal cord MRI one week later showed improved cord thickness with less hyperintensity. Respiratory multiplex PCR was negative including enteroviruses. Repeat CSF studies after IVIG showed increased IgG index and IgG synthesis suggestive of recent spinal cord infection, consistent with acute flaccid myelitis (AFM). Pre-IVIG blood PCR was invalid for enteroviruses due to PCR inhibitors found in the sample. Blood post-IVIG was negative for mycoplasma IgM, West Nile IgM, and arboviruses IgM. Enterovirus panel titers (post-IVIG) were positive for coxsackie A (1:32), coxsackie B type 4 (1:80) and 5 (1:320), echovirus type 11 (1:160) and 30 (1:80) as well as positive for poliovirus type 1 and 3. These titers could not distinguish acute infection from patient's immunity or false-positives as a result of IVIG. He was discharged with outpatient follow-up visits with neurology, infectious disease, occupational and physical therapy, showing only mild improvement after discharge. Discussion(s):With the anticipated resurgence of AFM after the peak of COVID-19 pandemic, our case illustrates the need to consider this diagnostic possibility in patients with flaccid paralysis. It is important to remember CSF IgG synthesis is not affected by IVIG. In addition when treatment plans include IVIG, appropriate samples should be collected before IVIG to facilitate accurate work-up for infectious diseases. Copyright © 2023 Southern Society for Clinical Investigation.

Evaluation of a Rapid Multiplex PCR Diagnostic Device for the Detection of Microorganisms from Nasopharyngeal and Throat Swab Specimens in the Near Patient Setting

Background. Limited availability of multiplex molecular tests in the near-patient setting can impact the rapid diagnosis and treatment of patients experiencing symptoms of respiratory tract infections, including pharyngitis. The BioFire Respiratory/ Sore Throat (R/ST) Panel (bioMerieux, Salt Lake City, UT), designed for use with the BioFire SpotFire System, is a PCR-based sample-to-answer diagnostic test that identifies four bacteria and 10 viruses (including SARS-CoV-2) from nasopharyngeal swab (NPS) or throat swab (TS) specimens in about 16 minutes. This study evaluated the performance of an Investigational Use Only (IUO) version of the BioFire R/ST Panel in the near patient setting. Methods. NPS and TS specimens were prospectively enrolled from symptomatic consented/assented volunteers of all ages, or obtained as residual leftover specimens. Enrollment was conducted between December 2020 and September 2021 at five study sites in the US and UK (adult and pediatric emergency departments or urgent care clinics) with testing performed by personnel representative of the intended users (non-laboratory professionals). Several analytes that were not circulating during the COVID-19 pandemic were supplemented with archived specimens of known analyte composition. Performance was determined for both sample types by comparison to FDA cleared multiplex PCR tests or culture and PCR followed by sequencing of isolates (Streptococcus from throat swabs). Results. A total of 1131 NPS and 452 TS prospectively collected specimens and 542 NPS and 128 TS archived specimens were tested with the BioFire R/ST Panel. For NPS specimens (prospective and archived) overall positive percent agreement (PPA) was 98.7% and negative percent agreement (NPA) was 99.1%, and for TS specimens (prospective and archived) overall PPA was 95.9% and NPA was 99.2%. Conclusion. The BioFire R/ST Panel is a sensitive, specific, and robust test for rapid detection of a wide range of organisms in NPS and TS specimens in the nearpatient setting. This test is expected to aid in the timely diagnosis and appropriate management of pharyngitis and other respiratory infections.