Clonal Dissemination of Plasmid-Mediated Carbapenem and Colistin Resistance in Refugees Living in Overcrowded Camps in North Lebanon

Carbapenem and colistin-resistant bacteria represent a global public health problem. Refugees carrying these bacteria and living in inadequate shelters can spread these microorganisms. The aim of this study was to investigate the intestinal carriage of these bacteria in Syrian refugees in Lebanon. Between June and July 2019, 250 rectal swabs were collected from two refugee camps in North Lebanon. Swabs were cultured on different selective media. Antibiotic susceptibility testing was performed using the disk diffusion method. Carbapenemase-encoding genes and mcr genes were investigated using real-time polymerase chain reaction (RT-PCR) and standard polymerase chain reaction (PCR). Epidemiological relatedness was studied using multilocus sequence typing (MLST). From 250 rectal swabs, 16 carbapenem-resistant, 5 colistin-resistant, and 4 colistin and carbapenem-resistant Enterobacteriaceae were isolated. The isolates exhibited multidrug-resistant phenotypes. Seven Klebsiella pneumoniae isolates harboured the blaOXA-48 gene, and in addition four K. pneumoniae had mutations in the two component systems pmrA/pmrB, phoP/phoQ and co-harboured the blaNDM-1 gene. Moreover, the blaNDM-1 gene was detected in six Escherichia coli and three Enterobacter cloacae isolates. The remaining five E. coli isolates harboured the mcr-1 gene. MLST results showed several sequence types, with a remarkable clonal dissemination. An urgent strategy needs to be adopted in order to avoid the spread of such resistance in highly crowded underserved communities.

Chlamydia pneumoniae-associated pleuropericarditis: a case report and systematic review of the literature

Background Chlamydia pneumoniae is a common cause of atypical community acquired pneumonia (CAP). The diagnostic approach of chlamydial infections remains a challenge. Diagnosis of delayed chlamydial-associated complications, involving complex autoimmune pathophysiological mechanisms, is still more challenging. C. pneumoniae-related cardiac complications have been rarely reported, including cases of endocarditis, myocarditis and pericarditis. Case presentation A 40-year old female was hospitalized for pleuropericarditis following lower respiratory tract infection. The patient had been hospitalized for CAP (fever, dyspnea, chest X-ray positive for consolidation on the left upper lobe) 5 weeks ago and had received ceftriaxone and moxifloxacin. Four weeks after her discharge, the patient presented with fever, shortness of breath and pleuritic chest pain and was readmitted because of pericardial and bilateral pleural effusions (mainly left). The patient did not improve on antibiotics and sequential introduction of colchicine and methylprednisolone was performed. The patient presented impressive clinical and laboratory response. Several laboratory and clinical assessments failed to demonstrate any etiological factor for serositis. Chlamydial IgM and IgG antibodies were positive and serial measurements showed increasing kinetics for IgG. Gold standard polymerase chain reaction of respiratory tract samples was not feasible but possibly would not have provided any additional information since CAP occurred 5 weeks ago. The patient was discharged under colchicine and tapered methylprednisolone course. During regular clinic visits, she remained in good clinical condition without pericardial and pleural effusions relapse. Conclusions C. pneumoniae should be considered as possible pathogen in case of pleuritis and/or pericarditis during or after a lower respiratory tract infection. In a systematic review of the literature only five cases of C. pneumoniae associated pericarditis were identified. Exact mechanisms of cardiovascular damage have not yet been defined, yet autoimmune pathways might be implicated.

A Pioneer Study on a Non-invasive Method for in Ovo Chicken Egg Sexing.

Background: Day-old male chicks culling is one of the world most inhumane problems in the poultry industry. Every year, seven billions of male chicks are being killed in laying-hen hatcheries, due to their higher feed exchange rate, lower management compare to female ones and production costs. This work reports a novel non-invasive method for the gender identification of chicken eggs. Four electrodes were attached onto each egg during the incubation process for the data gathering period of fourteen days. Standard polymerase chain reaction (PCR) based chicken gender determination protocol was applied to the eggs on the last day of the incubation process to get the gender information. Result: A relationship between the collected data and the gender of the egg was built, and it was found to have a reliable connection, indicating that by measure the impedance data of the eggs from day 9 of incubation with the four electrodes setting and applying the self-normalization technique, we can determine the chicken egg gender.Conclusion: This is a pioneer founding, proving that impedance spectroscopy can be use to sexing of chicken eggs, before its life has been formed, relieving the poultry industry from such an ethical burden.

First detection of porcine circovirus type 3 in Ukraine

To date, there is no information regarding the occurrence of porcine circovirus type 3 (PCV-3) in pigs in Ukraine. Aim. The aim of this work was to study the probable occurrence of the little-studied PCV-3 in pigs with different health status in Dnipropetrovsk, Donetsk, Kyiv, and Kharkiv regions of Ukraine. Methods. Blood, semen, liver, spleen, lung samples and nasal swabs of sows and boars of different ages and with different health status, belonging to farms from Dnipro, Donetsk, Kyiv, and Kharkiv regions of Ukraine, were used for the study. PCV-3 genomic material was detected by the standard polymerase chain reaction using specific primers, flanking a fragment of the rep gene of the virus with the length of 418 bp. To visualize the amplicons, horizontal gel electrophoresis was used and ethidium bromide staining after electrophoresis, followed by photographing the gels using Image Lab 5.2.1 software. Results. DNA of PCV-3 was found in two liver samples and four nasal swabs in two different farms, obtained from clinically healthy pigs, which suggests the possibility of the circulation of this infectious agent at the subclinical level of infection at the farm under investigation. No PCV-3 coinfection with the causative agents of porcine reproductive and respiratory syndrome (PRRS), Aujeszky’s disease, PCV-2, and mycoplasmas was found at this farm. Conclusions. Porcine circovirus type 3 (PCV-3) – (a little-studied causative agent of swine disease) was detected in 6 out of 61 samples, originating from two farms in the Kyiv and Kharkiv regions, obtained from clinically healthy animals) for the first time in Ukraine. This indicates possible circulation of the pathogen among pig farms in Ukraine and demonstrates the need to create and implement a target risk analysis, an extensive survey, as well as to develop control measures of the disease spreading (both organizational and technical preventive). Molecular genetic surveying and subsequent monitoring of PCV-3 among domestic and wild animals, which can cross the borders, will give a possibility to determine the risks of its spreading and related economic and epidemiological consequences. The whole-genome DNA sequencing of the detected virus isolates is planned to determine the relation of Ukrainian strains of the virus to other strains circulating in Europe and other parts of the world. Better understanding the risks, epidemiology and pathology, associated with this new virus for the Ukrainian pig breeding industry, will help to prevent and control its further spread and harmful effects.

Detection of clonotypic DNA in the cerebrospinal fluid as a marker of central nervous system invasion in lymphoma

Diagnosis of parenchymal central nervous system (CNS) invasion and prediction of risk for future CNS recurrence are major challenges in the management of aggressive lymphomas, and accurate biomarkers are needed to supplement clinical risk predictors. For this purpose, we studied a next-generation sequencing (NGS)-based assay that detects tumor-derived DNA for clonotypic immunoglobulin gene rearrangements in the cerebrospinal fluid (CSF) of patients with lymphomas. Used as a diagnostic tool, the NGS-MRD assay detected clonotypic DNA in 100% of CSF samples from 13 patients with known CNS involvement. These included 7 patients with parenchymal brain disease only, whose CSF tested negative by standard cytology and flow cytometry, and 6 historical DNA aliquots collected from patients at median 39 months prior to accession, which had failed to show clonal rearrangements using standard polymerase chain reaction. For risk prognostication, we prospectively collected CSF from 22 patients with newly diagnosed B-cell lymphomas at high clinical risk of CNS recurrence, of whom 8 (36%) had detectable clonotypic DNA in the CSF. Despite intrathecal prophylaxis, a positive NGS-MRD assay in the CSF was associated with 29% cumulative risk of CNS recurrence within 12 months from diagnosis, in contrast with 0% risk among patients with a negative assay (P=0.045). These observations suggest that detection of clonotypic DNA can aid in diagnosis of suspected parenchymal brain recurrence in aggressive lymphoma. Furthermore, the NGS-MRD assay might enhance clinical risk assessment for CNS recurrence among newly diagnosed patients and help select those who might benefit most from novel approaches to CNS-directed prophylaxis.

Evaluation of the ESGE recommendations for COVID-19 pre-endoscopy risk-stratification in a high-volume center in Germany

Background and study aims The European Society of Gastrointestinal Endoscopy (ESGE) has defined COVID-19 infection prevention and control strategies within the endoscopy unit. These include pre-endoscopic questionnaire-based risk-stratification as well as pre-procedure viral testing. Real-life data on the effectiveness of these measures are presented here. Patients and methods Data from the outpatient endoscopic unit of the University Hospital Augsburg between July 1, 2020 and December 31, 2020 including the second pandemic wave were reviewed retrospectively. All patients were assessed with a pre-endoscopic risk-stratification questionnaire as well as viral testing using an antigen point-of-care test (Ag-POCT) in conjunction with a standard polymerase chain reaction (PCR) test. Highly elective procedures were postponed. The theoretically expected number of SARS-CoV-2-positive patients was simulated and compared with the actual number. In addition, endoscopy staff was evaluated with a rapid antibody test to determine the number of infections among the personnel. Results In total, 1029 procedures, 591 questionnaires, 591 Ag-POCTs, and 529 standard PCR tests were performed in 591 patients. 247 procedures in 142 patients were postponed. One Ag-POCT was positive but with a negative PCR test, while one PCR test was positive but with a negative Ag-POCT. This was lower than the theoretically expected number of COVID-19-positive patients (n = 15). One of 43 employees (2.3 %) in the outpatient endoscopy unit was seropositive. Conclusions Pre-endoscopic risk management including questionnaire-based risk stratification and viral testing seems to be an effective tool in combination with personal protective equipment for SARS-CoV-2 infection prevention and control within the endoscopy unit even in a high-prevalence setting.

Rapid and Visualized Detection of Virulence-Related Genes of Vibrio cholerae in Water and Aquatic Products by Loop-Mediated Isothermal Amplification

Vibrio cholerae can cause pandemic cholera in humans. The bacterium resides in aquatic environments worldwide. Continuous testing of V. cholerae contamination in water and aquatic products is imperative for food safety control and human health. In this study, a rapid and visualized method was for the first time developed based on loop-mediated isothermal amplification (LAMP) for detection of very important virulence-related genes ace , zot , cri , and nanH for toxins and infection process of V. cholerae . Three pairs of molecular probes targeting each of these genes were designed and synthesized. The one-step LAMP reaction was conducted at 65 o C for 40 min. Positive results were simply inspected by the production of light green color under visible light or green fluorescence under UV light (302 nm). Limit of detection (LOD) of the LAMP method ranged from 1.85-2.06 pg/reaction of genomic DNA or 2.50-4.00×10 2 CFU/reaction for target genes of cell culture of V. cholerae , which was more sensitive than standard polymerase chain reaction (PCR). Inclusivity and exclusivity of the LAMP method were 100% for all target genes. The method showed similar high efficiency to a certain extent in rapid testing of spiked or collected specimens of water and aquatic products. Target genes were detected by the absence from all water samples from various sources. However, high occurrences of nanH gene were observed in intestine samples derived from four species of fish and one species of shellfish, indicating a risk of potentially toxic V. cholerae in commonly consumed aquatic products. The results in this study provide a potential tool for rapid and visualized detection of V. cholerae in water and aquatic products.

Agreement Test of Histopathology in the Diagnosis of Extrapulmonary Tuberculosis with Gold Standard Polymerase Chain Reaction Technique: A Step to Overcome False Diagnosis

BACKGROUND: Tuberculosis (TB) is global health problem which is caused by Mycobacterium tuberculosis (M. tuberculosis) bacteria. One-quarter of the world’s populations is infected by M. tuberculosis and only 10–15% of those develop the disease, while the remaining 85–95% of the population are carrying the bacteria and cannot transmit the disease to the others. M. tuberculosis bacteria affects the lungs, but any organ in the body can be affected by the bacteria. About 15% of M. tuberculosis infections are of in the extrapulmonary type. The diagnosis of extrapulmonary TB (EPTB) is very challenging because most sites are inaccessible and paucibacillary nature of the bacteria in these sites. The need for rapid and more sensitive and specific tests for the diagnosis of EPTB in comparison to culture and histopathology is increasing. The molecular methods for the detection of M. tuberculosis gene(s) in the provided sample are now promising. PATIENT AND METHODS: A cross-sectional descriptive study at AL-Kindy Teaching Hospital at Al-Resaffa part of Baghdad city, Iraq. Data collection has been done in three months duration (July, August, and September) 2015. A total of 74 formalin-fixed paraffin-embedded samples from suspected EPTB cases was collected, both Polymerase Chain reaction test for M. tuberculosis and histopathological examination was done for each sample. RESULTS: A total of 74 patients (18 males, 56 females), mean age 29.72 suspected to had extrapulmonary TB underwent biopsies from different tissue types. The biopsies from the 74 patients were taken from different tissues according to the site of lesion, 49 (66.2%) biopsies were taken from lymph node, 12 biopsies (16.2%) was taken from mass in the axilla, 6 (8.1%) from abscess, 4 (5.4%) from the intestine, 3 (4.1%) from fistula. Of the 74 studied patients 57 (77%) showed positive polymerase chain reaction (PCR) and 17 (23%) showed negative PCR results. Regarding to the histopathological reports of the biopsies, there were 54 (73%) patients had positive histopathological (granuloma) result and 20 (27%) patients had negative results (nongranuloma). The sensitivity of histopathological examination of the biopsies was 91.02%, the specificity 88.2%, and the kappa was 0.748 (p = 0.00) which is mean good agreement between histopathological examination of the biopsies and the polymerase chain reaction test. CONCLUSIONS: The sensitivity, specificity, and the positive predictive value of histopathology examination of biopsies were 91.02%, 88.2%, and 96%, respectively. The kappa was 0.748 (p = 0.00) which is mean good agreement between histopathological examination of the biopsies and the polymerase chain reaction test.

Targeted metabolomics identifies high performing diagnostic and prognostic biomarkers for COVID-19

Research exploring the development and outcome of COVID-19 infections has led to the need to find better diagnostic and prognostic biomarkers. This cross-sectional study used targeted metabolomics to identify potential COVID-19 biomarkers that predicted the course of the illness by assessing 110 endogenous plasma metabolites from individuals admitted to a local hospital for diagnosis/treatment. Patients were classified into four groups (≈ 40 each) according to standard polymerase chain reaction (PCR) COVID-19 testing and disease course: PCR−/controls (i.e., non-COVID controls), PCR+/not-hospitalized, PCR+/hospitalized, and PCR+/intubated. Blood samples were collected within 2 days of admission/PCR testing. Metabolite concentration data, demographic data and clinical data were used to propose biomarkers and develop optimal regression models for the diagnosis and prognosis of COVID-19. The area under the receiver operating characteristic curve (AUC; 95% CI) was used to assess each models’ predictive value. A panel that included the kynurenine: tryptophan ratio, lysoPC a C26:0, and pyruvic acid discriminated non-COVID controls from PCR+/not-hospitalized (AUC = 0.947; 95% CI 0.931–0.962). A second panel consisting of C10:2, butyric acid, and pyruvic acid distinguished PCR+/not-hospitalized from PCR+/hospitalized and PCR+/intubated (AUC = 0.975; 95% CI 0.968–0.983). Only lysoPC a C28:0 differentiated PCR+/hospitalized from PCR+/intubated patients (AUC = 0.770; 95% CI 0.736–0.803). If additional studies with targeted metabolomics confirm the diagnostic value of these plasma biomarkers, such panels could eventually be of clinical use in medical practice.

Targeted Metabolomics Identifies High Performing Diagnostic and Prognostic Biomarkers for COVID-19

Research exploring the development and outcome of COVID-19 infections has led to the need to find better diagnostic and prognostic biomarkers. This cross-sectional study used targeted metabolomics to identify potential COVID-19 biomarkers that predicted the course of the illness by assessing 110 endogenous plasma metabolites from individuals admitted to a local hospital for diagnosis/treatment. Patients were classified into four groups (≈ 40 each) according to standard polymerase chain reaction (PCR) COVID-19 testing and disease course: PCR-/controls (i.e., non-COVID controls), PCR+/not-hospitalized, PCR+/hospitalized, and PCR+/intubated. Blood samples were collected within 2 days of admission/PCR testing. Metabolite concentration data, demographic data and clinical data were used to propose biomarkers and develop optimal regression models for the diagnosis and prognosis of COVID-19. The area under the receiver operating characteristic curve (AUC; 95%CI) was used to assess each models’ predictive value. A panel that included the kynurenine: tryptophan ratio, LysoPC a C26:0, and pyruvic acid discriminated non-COVID controls from PCR+/not-hospitalized (AUC = 0.947; 95%CI 0.931–0.962). A second panel consisting of C10:2, butyric acid, and pyruvic acid distinguished PCR+/not-hospitalized from PCR+/hospitalized and PCR+/intubated (AUC = 0.975; 95%CI 0.968–0.983). Only LysoPC a C28:0 differentiated PCR+/hospitalized from PCR+/intubated patients (AUC = 0.770; 95%CI 0.736–0.803). If additional studies with targeted metabolomics confirm the diagnostic value of these plasma biomarkers, such panels could eventually be of clinical use in medical practice.