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Coinfections with human pegivirus 1 (HPgV-1) are common in chronic hepatitis C virus (HCV) patients. However, little is known about whether HPgV-1 is affected by direct-acting antivirals during HCV treatment. Metagenomic analysis and reverse transcriptase-quantitative PCR (RT-qPCR) were performed on RNA from the plasma of 88 selected chronic HCV patients undergoing medical treatment. Twenty (23%) of these HCV patients had HPgV-1 coinfections and were followed by RT-qPCR during treatment and follow-up to investigate HPgV-1 RNA titers. Recovered sequences could be assembled to complete HPgV-1 genomes, and most formed a genotype 2 subclade. All HPgV-1 viral genomic regions were under negative purifying selection. Glecaprevir/pibrentasvir treatment in five patients did not consistently lower the genome titers of HPgV-1. In contrast, a one log10 drop of HPgV-1 titers at week 2 was observed in 10 patients during treatment with sofosbuvir-containing regimens, sustained to the end of treatment (EOT) and in two cases decreasing to below the detection limit of the assay. For the five patients treated with ledipasvir/sofosbuvir with the inclusion of pegylated interferon, titers decreased to below the detection limit at week 2 and remained undetectable to EOT. Subsequently, the HPgV-1 titer rebounded to pretreatment levels for all patients. In conclusion, we found that HCV treatment regimens that included the polymerase inhibitor sofosbuvir resulted in decreases in HPgV-1 titers, and the addition of pegylated interferon increased the effect on patients with coinfections. This points to the high specificity of protease and NS5A inhibitors toward HCV and the more broad-spectrum activity of sofosbuvir and especially pegylated interferon. IMPORTANCE: Human pegivirus 1 coinfections are common in hepatitis C virus (HCV) patients, persisting for years. However, little is known about how pegivirus coinfections are affected by treatment with pangenotypic direct-acting antivirals (DAAs) against HCV. We identified human pegivirus by metagenomic analysis of chronic HCV patients undergoing protease, NS5A, and polymerase inhibitor treatment, in some patients with the addition of pegylated interferon, and followed viral kinetics of both viruses to investigate treatment effects. Only during HCV DAA treatment regimens that included the more broad-spectrum drug sofosbuvir could we detect a consistent decline in pegivirus titers that, however, rebounded to pretreatment levels after treatment cessation. The addition of pegylated interferon gave the highest effect with pegivirus titers decreasing to below the assay detection limit, but without clearance. These results reveal the limited effect of frontline HCV drugs on the closest related human virus, but sofosbuvir appeared to have the potential to be repurposed for other viral diseases.
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Antivirales , Bencimidazoles , Coinfección , Infecciones por Flaviviridae , Hepatitis C Crónica , Pegivirus , Sofosbuvir , Humanos , Antivirales/uso terapéutico , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/virología , Bencimidazoles/uso terapéutico , Sofosbuvir/uso terapéutico , Pegivirus/efectos de los fármacos , Coinfección/tratamiento farmacológico , Coinfección/virología , Masculino , Infecciones por Flaviviridae/tratamiento farmacológico , Infecciones por Flaviviridae/virología , Femenino , Genotipo , Quinoxalinas/uso terapéutico , Persona de Mediana Edad , Pirrolidinas , Sulfonamidas/uso terapéutico , ARN Viral/sangre , ARN Viral/genética , Hepacivirus/genética , Hepacivirus/efectos de los fármacos , Genoma Viral , Adulto , Anciano , Carga Viral/efectos de los fármacos , Metagenómica , Ciclopropanos , Ácidos Aminoisobutíricos , Filogenia , Combinación de MedicamentosRESUMEN
BACKGROUND: Appendicitis seems to be a disease of infectious origin, but the detailed pathogenesis is unknown. We aimed to investigate the microbiome of the appendix lumen in patients with and without appendicitis, including a comparison of the subgroups of complicated versus uncomplicated appendicitis. METHODS: This prospective observational cohort study included adult patients undergoing laparoscopic appendectomy for suspected appendicitis. According to histopathologic findings, the investigated groups consisted of patients with and without appendicitis, including subgroups of complicated versus uncomplicated appendicitis based on the surgical report. A swab of the appendix lumen was analyzed for genetic material from bacteria with shotgun metagenomics, and outcomes included analyses of microbiome diversity and differential abundance of bacteria. RESULTS: A total of 53 swabs from patients with suspected appendicitis were analyzed: 42 with appendicitis (16 complicated) and 11 without appendicitis. When comparing patients with and without appendicitis, they were equally rich in bacteria (alpha diversity), but the microbiome composition was dissimilar between these groups (beta diversity) (P < .01). No consistent bacterial species were detected in all patients with appendicitis, but a least 3 genera (Blautia, Faecalibacterium, and Fusicatenibacter) and 2 species, Blautia faecis and Blautia wexlerae, were more abundant in patients without appendicitis. For the subgroups complicated versus uncomplicated appendicitis, both measures for microbiome diversity were similar. CONCLUSION: The appendix microbiome composition of genetic material from bacteria in adult patients with and without appendicitis differed, but the microbiome was similar for patients with complicated versus uncomplicated appendicitis. Trial registration NCT03349814.
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Apendicectomía , Apendicitis , Apéndice , Humanos , Apendicitis/microbiología , Apendicitis/cirugía , Estudios Prospectivos , Adulto , Femenino , Masculino , Apéndice/microbiología , Apéndice/cirugía , Apéndice/patología , Persona de Mediana Edad , Microbiota , Laparoscopía , Adulto Joven , AncianoRESUMEN
Objectives: To determine if vancomycin-resistant Enterococcus faecium (VREfm) carriers carry the same VREfm clone after a minimum follow-up of 365â days. For those carrying the same clone, we investigated the genomic evolution per year per genome. Methods: We used WGS results to assign VREfm clones to each isolate and determine clone shifts. Finally, we calculated distance in core-genome MLST alleles, and the number of SNPs between consecutive VREfm isolates from patients carrying the same VREfm clone. Results: In total, 44.2% of patients carried the same VREfm clone, and the genomic evolution was 1.8 alleles and 2.6 SNPs per genome per year. Conclusions: In our population of long-term carriers, we calculated a molecular clock of 2.6 SNPs.
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Background: Vaginal dysbiosis covers imbalances in the vaginal microbiota, defined by altered composition of bacteria, viruses, and fungi and is associated with euploid pregnancy losses, premature birth, infertility, or bacterial vaginosis. A large proportion of women who have vaginal dysbiosis do not experience any symptoms. Antibiotics are the traditional treatment, recently combined with local probiotics in some cases. Vaginal Microbiota Transplantation (VMT) with eubiotic vaginal bacterial microbiota after antibiotic eradication of pathogens has successfully been performed in a case study with five patients, but no VMT has been performed without the use of antibiotics. Methods: This is a proof of concept case study. The patient was found to have vaginal dysbiosis at the RPL clinic at Copenhagen University Hospital Hvidovre, Denmark on the 23rd of June 2021. She was offered and accepted to receive experimental treatment in the form of a VMT as a compassionate use case. VMT is the transfer of cervicovaginal secretions (CVS) from a healthy donor with a Lactobacillus-dominant vaginal microbiome to a recipient with a dysbiotic vaginal microbiome. CVS is a mixture of e.g., mucus, bacteria, metabolites present in the vaginal canal. Potential donors were thoroughly screened for the absence of STIs, and the most suitable donor sample for the specific patient in this study was determined via an in vitro microbiome competition assay. Findings: A 30-year-old patient with one livebirth and a complicated pregnancy history of two stillbirths and 1 s trimester pregnancy loss in gestational weeks 27 (2019), 17 (2020) and 23 (2020) respectively with complaints of vaginal irritation and discharge that had aggravated in all her pregnancies. Her vaginal microbiome composition showed a 90% dominance of Gardnerella spp. After one VMT there was a complete shift in microbiome composition to 81.2% L. crispatus and 9% L. jensenii with a concurrent resolvement of vaginal symptoms. Single nucleotide polymorphism-analysis confirmed her microbiome to be of donor origin and it remain stable now 1.5 years after the VMT. Five months after the VMT she became pregnant and has successfully delivered a healthy baby at term. Interpretation: Here we report a successful VMT with confirmed donor strain engraftment followed by a successful pregnancy and delivery after a series of late pregnancy losses/stillbirths. Findings suggest that VMT is a potential treatment for severe vaginal dysbiosis. Further, larger studies are required. Funding: The study was partially funded (i.e., analysis costs) by Freya Biosciences Aps, Fruebjergvej, 2100 Copenhagen, Denmark.
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In Denmark, vaccination against Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2) has been with the Pfizer-BioNTech (BTN162b2) or the Moderna (mRNA-1273) mRNA vaccines. Patients with chronic hepatitis C virus (HCV) infection followed in our clinic received mRNA vaccinations according to the Danish roll-out vaccination plan. To monitor HCV infection, RNA was extracted from patient plasma and RNA sequencing was performed on the Illumina platform. In 10 of 108 HCV patient samples, full-length or traces of SARS-CoV-2 spike mRNA vaccine sequences were found in blood up to 28 days after COVID-19 vaccination. Detection of mRNA vaccine sequences in blood after vaccination adds important knowledge regarding this technology and should lead to further research into the design of lipid-nanoparticles and the half-life of these and mRNA vaccines in humans.
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COVID-19 , Hepatitis C Crónica , Hepatitis C , Humanos , Vacunas contra la COVID-19 , SARS-CoV-2/genética , COVID-19/prevención & control , Vacunación , Hepacivirus , Anticuerpos AntiviralesRESUMEN
Sequencing of the spa gene of methicillin-resistant Staphylococcus aureus (MRSA) is used for assigning spa types to e.g., detect transmission and control outbreaks. Traditionally, spa typing is performed by Sanger sequencing but has in recent years been replaced by whole-genome sequencing (WGS) in some laboratories. Spa typing by WGS involves de novo assembly of millions of short sequencing reads into larger contiguous sequences, from which the spa type is then determined. The choice of assembly program therefore potentially impacts the spa typing result. In this study, WGS of 1,754 MRSA isolates was followed by de novo assembly using the assembly programs SPAdes (with two different sets of parameters) and SKESA. The spa types were assigned and compared to the spa types obtained by Sanger sequencing, regarding the latter as the correct spa types. SPAdes with the two different settings resulted in assembly of the correct spa type for 84.8% and 97.6% of the isolates, respectively, while SKESA assembled the correct spa type in 98.6% of cases. The misassembled spa types were generally two spa repeats shorter than the correct spa type and mainly included spa types with repetition of the same repeats. WGS-based spa typing is thus very accurate compared to Sanger sequencing, when the best assembly program for this purpose is used. IMPORTANCE spa typing of methicillin-resistant Staphylococcus aureus (MRSA) is widely used by clinicians, infection control workers, and researchers both in local outbreak investigations and as an easy way to communicate and compare MRSA types between laboratories and countries. Traditionally, spa types are determined by Sanger sequencing, but in recent years a whole-genome sequencing (WGS)-based approach has become increasingly used. In this study, we compared spa typing by WGS using different methods for assembling the genome from short sequencing reads and compared to Sanger sequencing as the gold standard. We find substantial differences in correct assembly of spa types between the assembly methods. Our findings are therefore important for the quality of WGS based spa typing data being exchanged by clinical microbiology laboratories.
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Staphylococcus aureus Resistente a Meticilina , Infecciones Estafilocócicas , Humanos , Staphylococcus aureus Resistente a Meticilina/genética , Epidemiología Molecular/métodos , Infecciones Estafilocócicas/microbiología , Secuenciación Completa del Genoma , Brotes de EnfermedadesRESUMEN
Synbiotics combine probiotics and prebiotics and are being investigated for potential health benefits. In this single-group-design trial, we analyzed changes in the gut microbiome, stool quality, and gastrointestinal well-being in 15 healthy volunteers after a synbiotic intervention comprising Lacticaseibacillus rhamnosus (LGG), Lactobacillus acidophilus (LA-5), Lacticaseibacillus paracasei subsp. paracasei (L. CASEI 431), and Bifidobacterium animalis subsp. lactis BB-12 and 20 g of chicory-derived inulin powder consumed daily for 4 weeks. Fecal samples were collected at baseline and at completion of the intervention, and all participants completed a fecal diary based on the Bristol Stool Scale and recorded their gastrointestinal well-being. No adverse effects were observed after consumption of the synbiotic product, and stool consistency and frequency remained almost unchanged during the trial. Microbiome analysis of the fecal samples was achieved using shotgun sequencing followed by taxonomic profiling. No changes in alpha and beta diversity were seen after the intervention. Greater relative abundances of Bifidobacteriaceae were observed in 12 subjects, with indigenous bifidobacteria species constituting the main increase. All four probiotic organisms increased in abundance, and L. rhamnosus, B. animalis, and L. acidophilus were differentially abundant, compared to baseline. Comparison of the fecal strains to the B. animalis subsp. lactis BB-12 reference genome and the sequenced symbiotic product revealed only a few single-nucleotide polymorphisms differentiating the probiotic B. animalis subsp. lactis BB-12 from the fecal strains identified, indicating that this probiotic strain was detectable after the intervention. IMPORTANCE The effects of probiotics/synbiotics are seldom investigated in healthy volunteers; therefore, this study is important, especially considering the safety aspects of multiple probiotics together with prebiotic fiber in consumption by humans. The study explores at the potential of a synbiotic intervention with lactobacilli, bifidobacteria, and inulin in healthy volunteers and tracks the ingested probiotic strain B. animalis subsp. lactis.
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Bifidobacterium animalis , Probióticos , Simbióticos , Humanos , Bifidobacterium , Heces/microbiología , Voluntarios Sanos , Inulina , Lactobacillus , Lactobacillus acidophilus , Prebióticos , Probióticos/farmacologíaRESUMEN
Daptomycin is a cyclic lipopeptide used in the treatment of vancomycin-resistant Enterococcus faecium (VREfm). However, the development of daptomycin-resistant VREfm challenges the treatment of nosocomial VREfm infections. Resistance mechanisms of daptomycin are not fully understood. Here, we analyzed the genomic changes leading to a daptomycin-susceptible VREfm isolate becoming resistant after 50 days of daptomycin and linezolid combination therapy. A total of seven isogenic VREfm isolates from the same patient (daptomycin-susceptible and daptomycin-resistant) were analyzed using Illumina whole genome sequencing, and two isolates were further characterized with Nanopore sequencing. One nonsynonymous SNP in the rpoC gene previously shown to harbor mutations in daptomycin-resistant VREfm was identified in the daptomycin-resistant isolates. Whole genome comparative analysis identified the loss of a 46.5 kb fragment, duplication of a 29.7 kb fragment, and integration of two plasmids upon acquisition of daptomycin resistance. Transmission electron microscopy showed similar alterations in cell morphology and cell wall structure as have previously been described in daptomycin-resistant E. faecalis.
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Infección Hospitalaria , Daptomicina , Enterococcus faecium , Infecciones por Bacterias Grampositivas , Enterococos Resistentes a la Vancomicina , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Daptomicina/farmacología , Enterococcus faecium/genética , Genómica , Infecciones por Bacterias Grampositivas/tratamiento farmacológico , Humanos , Pruebas de Sensibilidad Microbiana , Vancomicina/farmacología , Enterococos Resistentes a la Vancomicina/genéticaRESUMEN
The purpose of this trial was to evaluate the efficacy of a 4-week supplementation of Lactobacillus rhamnosus GG (LGG) in eliminating the gastrointestinal carrier state of vancomycin-resistant Enterococcus faecium (VREfm) in hospitalized adults. The primary outcome of the study was the number of patients with cleared VREfm colonization after the 4-week intervention. Secondary outcomes were clearance of VREfm colonization at weeks 8, 16, and 24, number of VREfm infections (isolated from nonintestinal foci), and changes in fecal microbiome diversity after the intervention. The trial was a multicenter, randomized, double-blind, placebo-controlled trial in hospitalized adult VREfm carriers. Patients were enrolled and randomized to receive 60 billion CFU of LGG daily or placebo for 4 weeks. For a subgroup of patients, rectal swabs for VREfm were collected also at 8, 16, and 24 weeks and analyzed using shotgun metagenomics. Patients ingesting a minimum of 50% of the probiotic during the 4-week intervention were included in subsequent outcome analyses (48 of 81 patients). Twelve of 21 patients in the LGG group (57%) compared to 15 of 27 patients in the placebo group (56%) cleared their VREfm carriage. Eighteen patients completed the entire 24-week intervention with the same minimum compliancy. Of these, almost 90% in both groups cleared their VREfm carriage. We found a statistically significant difference between VREfm clearers and nonclearers regarding metronidazole and vancomycin usage as well as length of hospitalization after inclusion. The microbiome analyses revealed no significant difference in alpha diversity between the LGG and the placebo group. Beta diversity differed between the groups and the different time points. This study did not show an effect of LGG in eradication of VREfm after a 4-week intervention. IMPORTANCE Whereas other studies exploring the effect of L. rhamnosus in clearing VREfm from the intestine included children and adults, with a wider age range, our study consisted of a geriatric patient cohort. The natural clearance of VREfm in this study was almost 60% after 4 weeks, thus much higher than described previously. Also, this study characterizes the microbiome of VREfm patients in detail. This article showed no effect of the probiotic L. rhamnosus in clearing VREfm from the intestine of patients.
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Enterococcus faecium , Infecciones por Bacterias Grampositivas , Lacticaseibacillus rhamnosus , Microbiota , Probióticos , Enterococos Resistentes a la Vancomicina , Adulto , Anciano , Niño , Infecciones por Bacterias Grampositivas/tratamiento farmacológico , Humanos , Probióticos/uso terapéutico , Vancomicina/farmacología , Vancomicina/uso terapéuticoRESUMEN
The prevalence of vancomycin-resistant Enterococcus faecium has increased rapidly, and in Denmark, we are facing an endemic outbreak situation in hospitals. The aim of this study was to use whole-genome sequencing (WGS) and core genome multilocus sequencing typing (cgMLST) to determine the duration of VREfm outbreaks and thereby evaluate the effect of our infection control strategies. We included all VREfm isolates from six hospitals in the Capital Region of Denmark that were sequenced between 2012 and 2020. Ward data were collected from our laboratory information system. A ward outbreak was defined as two patient samples from the same ward within a period of 30 days belonging to the same cgMLST cluster. cgMLST complex types were determined using Ridom SeqSphere v7.2.3, where a maximum of 20 allelic differences between isolates defines a cluster. We included 1690 patient isolates between 2012 and 2020. Our collection consisted of 45 unique clusters and 227 ward outbreaks. The median duration of outbreaks was 20 days. We reported a median outbreak duration of VREfm outbreaks based on WGS data to be 20 days, and thus concluded that our infection control precautions are adequate.
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Infección Hospitalaria , Enterococcus faecium , Infecciones por Bacterias Grampositivas , Enterococos Resistentes a la Vancomicina , Infección Hospitalaria/epidemiología , Brotes de Enfermedades , Enterococcus faecium/genética , Infecciones por Bacterias Grampositivas/epidemiología , Humanos , Tipificación de Secuencias Multilocus , Vancomicina/farmacología , Enterococos Resistentes a la Vancomicina/genéticaRESUMEN
INTRODUCTION: Lactobacilli, especially Lactobacillus (L.) rhamnosus, are common and well-documented components of commercial probiotics [1]. Whole genome sequencing (WGS) is often used to compare bacterial genomes and their relatedness. In outbreak situations, it is used to investigate the transmission of pathogenic bacteria. WGS has also been used to determine safety in probiotics, by looking at potential virulence factors and resistance genes. CASE PRESENTATION: This case report describes a 56-year old multi-traumatised, immunocompetent woman who was given L. rhamnosus GG as a probiotic, and later developed a blood stream infection with L. rhamnosus GG.The patient was fed by a nasogastric tube, and she also had a central venous catheter for parenteral feeding. When the patient developed diarrhoea after long-term hospitalisation, she was given L. rhamnosus GG, as a probiotic, which was standard care on the ward where she was hospitalised. In this case report we describe the use of WGS to demonstrate that a patient fed with L. rhamnosus GG as a probiotic, developed a blood stream infection with the same strain. CONCLUSION: In this case WGS was applied to show the relatedness of a probiotic and a pathogenic strain of L. rhamnosus GG. This case emphasises the need for caution when administering probiotics to patients with indwelling catheters. The patient was immunocompetent and she cleared the infection without the need for antibiotics.
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BACKGROUND: Vancomycin-resistant Enterococcus faecium (VREfm) cause a wide range of hospital infections. Ireland has had one of the highest invasive VREfm infection rates in Europe over the last decade, yet little is known about Irish VREfm. OBJECTIVES: To investigate the population structure of Irish VREfm, explore diversity by analysing the vanA transposon region and compare Irish, Danish and global isolates. METHODS: E. faecium (nâ=â648) from five Irish hospitals were investigated, including VREfm [547 rectal screening and 53 bloodstream infection (BSI)] isolates and 48 vancomycin-susceptible (VSEfm) BSI isolates recovered between June 2017 and December 2019. WGS and core-genome MLST (cgMLST) were used to assess population structure. Genetic environments surrounding vanA were resolved by hybrid assembly of short-read (Illumina) and long-read (Oxford Nanopore Technologies) sequences. RESULTS: All isolates belonged to hospital-adapted clade A1 and the majority (435/648) belonged to MLST ST80. The population structure was highly polyclonal; cgMLST segregated 603/648 isolates into 51 clusters containing mixtures of screening and BSI isolates, isolates from different hospitals, and VREfm and VSEfm. Isolates within clusters were closely related (mean average ≤16 allelic differences). The majority (96.5%) of VREfm harboured highly similar vanA regions located on circular or linear plasmids with multiple IS1216E insertions, variable organization of vanA operon genes and 78.6% harboured a truncated tnpA transposase. Comparison of 648 Irish isolates with 846 global E. faecium from 30 countries using cgMLST revealed little overlap. CONCLUSIONS: Irish VREfm are polyclonal, yet harbour a characteristic plasmid-located vanA region with multiple IS1216E insertions that may facilitate spread.
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Infección Hospitalaria , Enterococcus faecium , Infecciones por Bacterias Grampositivas , Enterococos Resistentes a la Vancomicina , Infección Hospitalaria/epidemiología , Enterococcus faecium/genética , Genómica , Infecciones por Bacterias Grampositivas/epidemiología , Humanos , Irlanda/epidemiología , Tipificación de Secuencias Multilocus , Plásmidos/genética , Prevalencia , Vancomicina , Enterococos Resistentes a la Vancomicina/genéticaRESUMEN
Vancomycin-resistant Enterococcus faecium (VREfm) is a globally significant nosocomial pathogen with a rapidly increasing prevalence. The objectives were to investigate VREfm outbreak duration and study the additional impact that infection control bundle strategies (ICBSs) set up to curb coronavirus disease 2019 (COVID-19) spreading had on VREfm outbreaks. Outbreak data set were collected prospectively from April 2, 2014 to August 13, 2020 at Copenhagen University Hospital Bispebjerg, Denmark. All VREfm samples had polymerase chain reaction performed for vanA/vanB genes before whole genome sequencing using the Illumina MiSeq platform. The relatedness of isolates was studied by core genome multilocus sequence typing (cgMLST) using Ridom SeqSphere. Eighty-one outbreaks had a median outbreak duration of 32.5 days (range 5-204 days) and 1,161 VREfm isolates were sequenced. The same cgMLST cluster types reappeared after outbreaks were terminated. When comparing the first 5 months of the COVID-19 pandemic with the corresponding period in 2019, we found a 10-fold decrease in VREfm outbreak patients and median outbreak duration decreased from 56 to 7 days (88%). Several COVID-19 ICBSs were implemented from March 13 through summer 2020. VREfm outbreaks lasted up to 204 days, but our findings suggest that outbreaks might last longer since the same cgMLST persisted in the same wards for years implying an endemic situation with recurrent outbreaks caused by hospital reservoirs or readmittance of unknown VREfm carriers. The sharp decline in VREfm outbreaks during the COVID-19 pandemic was most likely due to the ICBSs, resulting in a decrease in VREfm transmission.
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COVID-19 , Enterococcus faecium/genética , Pandemias , Cuarentena , Infecciones Estreptocócicas/epidemiología , Resistencia a la Vancomicina/genética , Anciano , Portador Sano/microbiología , Dinamarca/epidemiología , Enterococcus faecium/efectos de los fármacos , Femenino , Hospitalización , Humanos , Masculino , Infecciones Estreptocócicas/microbiología , Secuenciación Completa del GenomaRESUMEN
We investigated the prevalence of coronaviruses in 44 bats from four families in northeastern Eswatini using high-throughput sequencing of fecal samples. We found evidence of coronaviruses in 18% of the bats. We recovered full or near-full-length genomes from two bat species: Chaerephon pumilus and Afronycteris nana, as well as additional coronavirus genome fragments from C. pumilus, Epomophorus wahlbergi, Mops condylurus, and Scotophilus dinganii. All bats from which we detected coronaviruses were captured leaving buildings or near human settlements, demonstrating the importance of continued surveillance of coronaviruses in bats to better understand the prevalence, diversity, and potential risks for spillover.
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Quirópteros , Infecciones por Coronavirus , Coronavirus , Metagenómica , Animales , Quirópteros/virología , Coronavirus/genética , Infecciones por Coronavirus/veterinaria , Esuatini , Variación Genética , Genoma Viral , FilogeniaRESUMEN
BACKGROUND: During 2018-19, an increase of vanB vancomycin-resistant Enterococcus faecium (VREfm) was observed in the Capital Region of Denmark. vanA/vanB PCR performed directly on rectal swabs is accurate in detection of vanA; however, the positive predictive value for vanB-positive samples is low because of the presence of vanB in non-enterococcal gut commensals. OBJECTIVES: We investigated the epidemiology and clonal relatedness of vanB VREfm from the period 2015-19 and describe the application of a clone-specific vanB VREfm PCR assay for rapid and accurate detection of vanB VREfm in rectal screening samples. METHODS: vanB VREfm were investigated using epidemiological data and WGS data. The SeqSphere+ software was used to analyse MLST and cgMLST, and de novo assemblies were annotated to determine insertion sites for the vanB transposon (Tn1549). A clone-specific vanB VREfm PCR assay was designed to detect the sequence bridging Tn1549 and the E. faecium chromosome (araA2) in the dominant cluster. RESULTS: Two hundred and seventy-five vanB VREfm isolates were identified, of which 76% were identified in 2019. A dominant cluster (Cluster 1, nâ=â204, 74%), six minor clusters and 15 singletons were identified. All Cluster 1 isolates and six non-Cluster 1 isolates had Tn1549 integrated into araA2. In 2019, the PCR assay would have detected 92% of all rectal screening samples containing vanB VREfm. CONCLUSIONS: vanB VREfm increased due to the introduction and nosocomial transmission of the successful Cluster 1. The clone-specific PCR assay detected vanB VREfm outbreak isolates in rectal screening samples rapidly and accurately.
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Infección Hospitalaria , Enterococcus faecium , Infecciones por Bacterias Grampositivas , Enterococos Resistentes a la Vancomicina , Proteínas Bacterianas/genética , Células Clonales , Dinamarca/epidemiología , Enterococcus faecium/genética , Infecciones por Bacterias Grampositivas/epidemiología , Humanos , Tipificación de Secuencias Multilocus , Reacción en Cadena de la Polimerasa , Enterococos Resistentes a la Vancomicina/genéticaRESUMEN
BACKGROUND: Since 2012, the incidence of vancomycin-resistant Enterococcus faecium (VREfm) has increased dramatically in Copenhagen and vanA E. faecium has become endemic and polyclonal. OBJECTIVES: To examine whether a patient with a positive VRE clinical sample had the same VREfm in a preceding screening sample (within 60 days). METHODS: We performed a 30 month retrospective study. From our laboratory information system (LIS), we identified all patients with an invasive VREfm isolate and a VREfm rectal screening isolate within 60 days before infection. VREfm pairs (screening isolate and invasive isolate) were whole-genome sequenced. All isolates were analysed using SeqSphere and core-genome MLST (cgMLST) types were determined. We examined all isolates for the presence of the three most dominant vanA plasmids in the Capital Region of Denmark. Two novel vanA plasmids were closed by Nanopore/Illumina sequencing. RESULTS: We found a total of 19 VREfm pairs. Of these, 13 patients had pairs with matching cgMLST types and vanA plasmids and a median number of 6 days from identification of carriage to clinical infection. One patient had a pair with non-matching cgMLST types but matching vanA plasmids and 24 days between identification of carriage to clinical infection. Five patients had pairs with non-matching cgMLST types and non-matching vanA plasmids and a median number of 18 days from identification of carriage to clinical infection. CONCLUSIONS: Of our 19 pairs, 13 were a match regarding cgMLST types (68%) and 1 more (5%) had matching vanA plasmids. Infection was thus preceded by colonization with the same isolates in 13 out of 19 patients. The five mismatches (26%) could be explained by the longer interval between colonization and infection.
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Infección Hospitalaria , Enterococcus faecium , Infecciones por Bacterias Grampositivas , Enterococos Resistentes a la Vancomicina , Proteínas Bacterianas/genética , Enterococcus faecium/genética , Infecciones por Bacterias Grampositivas/epidemiología , Humanos , Tipificación de Secuencias Multilocus , Estudios Retrospectivos , Vancomicina , Enterococos Resistentes a la Vancomicina/genéticaRESUMEN
The surveillance and correct subtyping of hepatitis C virus strains require available and up-to-date publicly available reference genomes. Here, we present the complete open reading frame sequence of a hepatitis C virus genotype 6 strain of an unknown subtype that was discovered during routine subtyping of patients in the clinic.
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BACKGROUND: Viruses and other infectious agents cause more than 15% of human cancer cases. High-throughput sequencing-based studies of virus-cancer associations have mainly focused on cancer transcriptome data. METHODS: In this study, we applied a diverse selection of presequencing enrichment methods targeting all major viral groups, to characterize the viruses present in 197 samples from 18 sample types of cancerous origin. Using high-throughput sequencing, we generated 710 datasets constituting 57 billion sequencing reads. RESULTS: Detailed in silico investigation of the viral content, including exclusion of viral artefacts, from de novo assembled contigs and individual sequencing reads yielded a map of the viruses detected. Our data reveal a virome dominated by papillomaviruses, anelloviruses, herpesviruses, and parvoviruses. More than half of the included samples contained 1 or more viruses; however, no link between specific viruses and cancer types were found. CONCLUSIONS: Our study sheds light on viral presence in cancers and provides highly relevant virome data for future reference.
Asunto(s)
Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Metagenoma/genética , Neoplasias/virología , Anelloviridae/genética , Anelloviridae/aislamiento & purificación , Biopsia , Conjuntos de Datos como Asunto , Femenino , Herpesviridae/genética , Herpesviridae/aislamiento & purificación , Humanos , Masculino , Neoplasias/patología , Papillomaviridae/genética , Papillomaviridae/aislamiento & purificación , Parvovirus/genética , Parvovirus/aislamiento & purificaciónRESUMEN
We describe a ceftriaxone-resistant Salmonella Typhi bacteraemia in a pregnant woman returning from a family visit in Pakistan. Whole genome sequencing confirmed similarity to a Pakistani outbreak clone. Pregnancy and unawareness of this outbreak delayed appropriate antibiotic therapy. Concurrently, we detected faecal carriage of a carbapenemase-producing Escherichia coli. Awareness of the ongoing outbreak should affect empiric treatment of typhoid fever and hygiene precautions in travellers returning from Pakistan. Meropenem may be warranted in severe cases.
Asunto(s)
Antibacterianos/uso terapéutico , Azitromicina/uso terapéutico , Ceftriaxona/farmacología , Meropenem/uso terapéutico , Salmonella typhi/genética , Salmonella typhi/aislamiento & purificación , Fiebre Tifoidea/diagnóstico , Fiebre Tifoidea/tratamiento farmacológico , Dolor Abdominal/etiología , Adulto , Pruebas de Aglutinación , Antibacterianos/farmacología , Bacteriemia/tratamiento farmacológico , Enterobacteriaceae Resistentes a los Carbapenémicos , Ceftriaxona/uso terapéutico , Dinamarca , Resistencia a Medicamentos , Escherichia coli/genética , Escherichia coli/aislamiento & purificación , Femenino , Fiebre/etiología , Humanos , Pruebas de Sensibilidad Microbiana , Pakistán , Plásmidos/análisis , Reacción en Cadena de la Polimerasa , Embarazo , Salmonella typhi/efectos de los fármacos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Viaje , Fiebre Tifoidea/microbiología , Secuenciación Completa del GenomaRESUMEN
The purpose of the present study was to perform a next-generation sequencing (NGS) based analysis of viruses in ocular adnexal extranodal marginal zone B-cell lymphoma (EMZL). Eight patients with extraocular EMZL were identified in the archives of Department of Ophthalmology, Copenhagen University Hospital. All cases were validated according to the World Health Organization classification. We subjected samples to enrichment of virion-associated (encapsidated) nucleic acids which included sample homogenization, filtration, and nuclease treatment. Both DNA and RNA were sequenced, and we analyzed the sequencing data for the presence of viral sequences. We detected no pathogenic viruses likely to be associated to development of EMZL. In one case, we detected human polyomavirus 7 and traces of Epstein-Barr virus (EBV) (human herpesvirus 4 (HHV4)) and a human papillomavirus. In conclusion, no viral pathogens were consistently detected in the extraocular EMZL samples when applying NGS-based methods.