UK circulating strains of human parainfluenza 3: an amplicon based next generation sequencing method and phylogenetic analysis.

Background: Human parainfluenza viruses type 3 (HPIV3) are a prominent cause of respiratory infection with a significant impact in both pediatric and transplant patient cohorts.  Currently there is a paucity of whole genome sequence data that would allow for detailed epidemiological and phylogenetic analysis of circulating strains in the UK. Although it is known that HPIV3 peaks annually in the UK, to date there are no whole genome sequences of HPIV3 UK strains available.  Methods: Clinical strains were obtained from HPIV3 positive respiratory patient samples collected between 2011 and 2015.  These were then amplified using an amplicon based method, sequenced on the Illumina platform and assembled using a new robust bioinformatics pipeline. Phylogenetic analysis was carried out in the context of other epidemiological studies and whole genome sequence data currently available with stringent exclusion of significantly culture-adapted strains of HPIV3. Results: In the current paper we have presented twenty full genome sequences of UK circulating strains of HPIV3 and a detailed phylogenetic analysis thereof.  We have analysed the variability along the HPIV3 genome and identified a short hypervariable region in the non-coding segment between the M (matrix) and F (fusion) genes. The epidemiological classifications obtained by using this region and whole genome data were then compared and found to be identical. Conclusions: The majority of HPIV3 strains were observed at different geographical locations and with a wide temporal spread, reflecting the global distribution of HPIV3. Consistent with previous data, a particular subcluster or strain was not identified as specific to the UK, suggesting that a number of genetically diverse strains circulate at any one time. A small hypervariable region in the HPIV3 genome was identified and it was shown that, in the absence of full genome data, this region could be used for epidemiological surveillance of HPIV3.Wellcome Open Grant PHE PhD studentshi

Adenovirus pseudo-outbreak in a large UK neonatal intensive care unit.

An unusually high number of positive adenovirus stool antigen tests were observed in neonatal samples from a large tertiary referral unit over a 10-week period, prompting the declaration of an outbreak and escalation of infection control precautions. Subsequent testing of original samples by alternative methods revealed a series of false-positive results. This is the first adenovirus pseudo-outbreak to be reported in the United Kingdom and the first to involve the Proflow Rotavirus-Adenovirus Combi test

Complete Genome Sequences of BK Polyomavirus Strains from Two Patients with Urinary Tract Infection, Sequenced Using the Ion Torrent Platform.

BK polyomavirus is an important pathogen in kidney transplant patients. We report here two complete genome sequences, those of isolates CAMB-1035 and CAMB-1055, identified in two urine samples tested for urinary tract infection at a hospital in eastern England, United Kingdom. Variation and phylogenetic analyses indicate that both isolates belong to subtype Ib-1

UK circulating strains of human parainfluenza 3: an amplicon based next generation sequencing method and phylogenetic analysis [version 2; referees: 2 approved]

Background: Human parainfluenza viruses type 3 (HPIV3) are a prominent cause of respiratory infection with a significant impact in both pediatric and transplant patient cohorts.  Currently there is a paucity of whole genome sequence data that would allow for detailed epidemiological and phylogenetic analysis of circulating strains in the UK. Although it is known that HPIV3 peaks annually in the UK, to date there are no whole genome sequences of HPIV3 UK strains available.  Methods: Clinical strains were obtained from HPIV3 positive respiratory patient samples collected between 2011 and 2015.  These were then amplified using an amplicon based method, sequenced on the Illumina platform and assembled using a new robust bioinformatics pipeline. Phylogenetic analysis was carried out in the context of other epidemiological studies and whole genome sequence data currently available with stringent exclusion of significantly culture-adapted strains of HPIV3. Results: In the current paper we have presented twenty full genome sequences of UK circulating strains of HPIV3 and a detailed phylogenetic analysis thereof.  We have analysed the variability along the HPIV3 genome and identified a short hypervariable region in the non-coding segment between the M (matrix) and F (fusion) genes. The epidemiological classifications obtained by using this region and whole genome data were then compared and found to be identical. Conclusions: The majority of HPIV3 strains were observed at different geographical locations and with a wide temporal spread, reflecting the global distribution of HPIV3. Consistent with previous data, a particular subcluster or strain was not identified as specific to the UK, suggesting that a number of genetically diverse strains circulate at any one time. A small hypervariable region in the HPIV3 genome was identified and it was shown that, in the absence of full genome data, this region could be used for epidemiological surveillance of HPIV3

Influenza-associated mortality in hospital care: a retrospective cohort study of risk factors and impact of oseltamivir in an English teaching hospital, 2016 to 2017.

BackgroundEvidence of an oseltamivir treatment effect on influenza A(H3N2) virus infections in hospitalised patients is incomplete.AimsThis cohort study aimed to evaluate risk factors for death among PCR-confirmed hospitalised cases of seasonal influenza A(H3N2) of all ages and the impact of oseltamivir.MethodsParticipants included all 332 PCR-confirmed influenza A(H3N2) cases diagnosed between 30 August 2016 and 17 March 2017 in an English university teaching Hospital. Oseltamivir treatment effect on odds of inpatient death was assessed by backward stepwise multivariable logistic regression analysis.ResultsThe odds of death were reduced by two thirds (odds ratio (OR): 0.32; 95% confidence interval (CI): 0.11-0.93), in inpatients treated with a standard course of oseltamivir 75 mg two times daily for 5 days - compared with those untreated with oseltamivir, after adjustment for age, sex, current excess alcohol intake, receipt of 2016/17 seasonal influenza vaccine, serum haemoglobin and hospital vs community attribution of acquisition of influenza.ConclusionsOseltamivir treatment given according to National Institutes of Clinical Excellence (NICE); United States Centres for Disease Control and Prevention (CDC); Infectious Diseases Society of America (IDSA) and World Health Organization (WHO) guidelines was shown to be effective in reducing the odds of mortality in inpatients with PCR-confirmed seasonal influenza A(H3N2) after adjustment in a busy routine English hospital setting. Our results highlight the importance of hospitals complying with relevant guidelines for prompt seasonal influenza PCR testing and ensuring standard oseltamivir treatment to all PCR-confirmed cases of seasonal influenza

Screening for thrombophilia by clotting based and molecular (polymerase chain reaction - PCR) methods in patients from a tertiary hospital

Ischemic stroke (IS) is a leading cause of death in developed countries; its prevalenceamong young adults (aged 18 to 50 years) ranges from 3% to 5%.The potential reduction in quality of life, as well as the obvious long-term significantsocioeconomic consequences urges for efficient primary and secondary prevention so as tofacilitate the likelihood of an increased number of productive years with good quality of life.IS is a heterogenous, multifactorial disease caused by conventional vascular risk factors andgenetic factors. Over the last years several studies have been performed to elucidate the mechanisms ofthis ischemic event. The well-established risk factors of hypertension, obesity and smoking,account for only a small percentage of cerebrovascular events.Common variations in the genes encoding factor V, prothrombin, 5,10methylenetetrahydrofolate reductase (MTHFR), factor XIII and plasminogen activatorinhibitor-1 are clotting factor mutations or polymorphisms that are associated with anincreased tendency towards thrombosis. The evidence of a role for these variants in the risk ofischemic stroke is controversial.In young adults with a history of ischemic stroke, we evaluated the role of theseinherited prothrombotic factors.The aim of the study was to investigate and compare the prevalence of theaforementioned classic vascular risk factors and polymorphisms, that is FVLeiden R506Q(FVL), the prothrombin variant (allele 20210A, FIIG20210A), C677T MTHFR(C677TMTHFR) polymorphism, the insertion/deletion (4G/5G) variation (-6754G/5GPAI-1)as well as the single nucleotide polymorphism -844G/A of in the plasminogen activatorinhibitor-1 (-844G/APAI-1) gene promoter and factor XIIIA Val34Leu polymorphism(FXIIIAVal34leu)and the impact of these gene mutations and classic vascular risk factors on theoverall stroke risk in individuals aged 50 years or younger. We also evaluated the role ofProtein C (PrC), Protein S (PrS) and Antithrombin (AT) deficiencies.In this respect, we conducted a case control study of 51 hospital cases of first-everischemic stroke in patients and 70 community-based controls.Patients and controls did not differ regarding age (p=0.876), sex ratios (p=0.567),smoking (p=0.559), hypercholesterolemia (p=0.112) or family history of stroke (p=0.20).Hypertension was more frequent in patients (8/51, 15.7%) (p=0.0001) than in the controls(0/70, 0%) (p=0.0001). All the patients and all the control subjects were of Caucasian origin(same ethnic background population).The prevalence of FVL, FIIG20210A, C677T MTHFR gene mutations, -6754G/5GPAI, -844G/A PAI-1 and FXIII-AVal34leu polymorphisms, as well as the plasma levelsof PrC, PrS and AT were assessed (after being tested by Polymerase ChainReaction/molecular techniques and clotting assays) and compared with controls.As data regarding strokes in young adults from Greece is scarce, the present study hasbeen carried out with the aim to test the possible association between -675 4G/5G PAI-1polymorphism, FVL, FIIG20210A, MTHFR C677T, the -844G/A PAI-1 single nucleotide polymorphism (SNP) and polymorphism FXIIIA Val34leu with ischemic stroke in the youngage in this specific South European population. We further evaluated the role of plasma levelsof PrC, PrS and AT.As this is the first study of this kind in Greek population, the additional goal was to evaluatethe importance of these mutations as risk factors for development of IS in the young age aswell as the prevalence of these mutations to the general population.In our setting of patients with young adult ischemic stroke, we came to the followingconclusions:No significant difference was found between patients and controls regarding thedistribution of FVL (p=0.199 ), FIIG20210A (p= 0.410), MTHFR (p= 0.788), -844G/A PAI-1(p=0.493) and FXIII-AVal34leu (p= 0.160) mutations/polymorphisms.These data are compatible with several reports showing that the presence of theabovementioned variants is not associated with an increased risk of ischemic stroke in theyoung age. However, they are at variance with other studies which showed some association. The 4G/5G genotype of the -675 4G/5G was significantly more frequent amongpatients as compared to controls. Moreover, we found a significantly lower frequency of the5G/5G genotype in patients as compared to control subjects. (p=0.02)Hence, this study suggests that the polymorphism of the PAI-1 gene promoter is agenetic risk factor for IS in the young age.We failed to demonstrate a significant association between patients and controlsregarding the rest of the inherited thrombophilia risk factors screened. Thus, those factors werenot found to increase the risk of IS among the group of young patients of Caucasian origin westudied.Further studies including all incoming young patients with IS are necessary to evaluatethe impact of those variants as risk factors in IS in the young age. Moreover, other novelgenetic defects and/or environmental factors could play a role and need to be investigatedΤα ισχαιμικά αγγειακά εγκεφαλικά επεισόδια (ΙΑΕΕ) αποτελούν μια από τις πιο σημαντικές αιτίες θανάτου στις ανεπτυγμένες χώρες. Η συχνότητα τους, σε νέους ενήλικες κυμαίνεται μεταξύ 3% και 5%.Η πιθανή μείωση της ποιότητας ζωής, όπως επίσης και οι προφανείς μακροχρόνιες κοινωνικοοικονομικές συνέπειες, προτρέπουν σε αποτελεσματική πρωτοβάθμια, αλλά και δευτεροβάθμια πρόληψη, ώστε να οδηγηθούμε στην πιθανότητα αύξησης των παραγωγικών χρόνων, καθώς και της καλής ποιότητας ζωής. Τα ΙΑΕΕ αποτελούν μια ετερογενή, πολυπαραγοντική νόσο, προκαλούμενη τόσο από τυπικούς παράγοντες κινδύνου, όσο και γενετικούς παράγοντες.Τα τελευταία χρόνια έχουν διεξαχθεί διάφορες μελέτες, ώστε να διευκρινιστούν οι μηχανισμοί ενός ισχαιμικού συμβάματος. Αποδεδειγμένοι παράγοντες κινδύνου όπως το κάπνισμα, η παχυσαρκία, η αρτηριακή υπέρταση,ενοχοποιούνται για μικρό μόνον ποσοστό ΙΑΕΕ. Κοινές παραλλαγές στα γονίδια που κωδικοποιούν τον παράγοντα V της πήξης, την προθρομβίνη, το ένζυμο 5,10 methylenotetrahydrofolate reductase(MTHFR), τον παράγοντα ΧΙΙΙ, καθώς και τον αναστολέα του ιστικού ενεργοποιητή του πλασμινογόνου 1 (ΡΑΙ-1) αποτελούν μεταλλάξεις ή πολυμορφισμούς των παραγόντων πήξης που σχετίζονται με αυξημένη τάση προς θρόμβωση. Ο ρόλος τους στον κίνδυνο εμφάνισης ΙΑΕΕ παραμένει υπό αμφισβήτηση/συζήτηση.Ο σκοπός αυτής της εργασίας ήταν να ελέγξουμε και να συγκρίνουμε την επίπτωση των προαναφερθέντων κλασσικών παραγόντων κινδύνου και πολυμορφισμών, δηλαδή του FVLeiden (R506Q ή G1691Α), της μετάλλαξης της προθρομβίνης (FIIG202104), της C677TMTHFR μετάλλαξης στο ένζυμο MTHFR,της παραλλαγής «εισαγωγή-διαγραφή» -6754G/5GPAI-1 στην περιοχή του υποκινητή του γονιδίου του ΡΑΙ-1, της απλής σημειακής μετάλλαξης -844G/APAI-1 στην περιοχή του προωθητή του γονιδίου του ΡΑΙ-1, του πολυμορφισμού FXIII-AVal34leuτης Α-υπομονάδας του παράγοντα ΧΙΙΙ καθώς και τη σημασία τους στο συνολικό κίνδυνο εμφάνισης ΙΑΕΕ σε νεαρούς ενήλικες κάτω των 50 ετών. Επίσης αξιολογήσαμε τη σημασία των επιπέδων πρωτεΐνης C, πρωτεΐνης S και αντιθρομβίνης και των πιθανών ανεπαρκειών αυτών των παραγόντων πήξης. Γι’αυτό το λόγο, διεξήχθη η παρούσα μελέτη , η οποία περιελάμβανε 51 επιζήσαντες ασθενείς με ΙΑΕΕ, ηλικίας κάτω των 50 ετών και 70 μάρτυρες. Σε αυτές τις δύο ομάδες εξετάστηκαν οι προαναφερόμενοι παράγοντες με μοριακές τεχνικές,καθώς και με πηκτικολογικές μεθόδους, και έπειτα μετρήθηκε και συγκρίθηκε η συχνότητά τους. Τα στοιχεία που αφορούν ΙΑΕΕ σε νέους ενήλικες από τον Ελλαδικό χώρο είναι σπάνια. Για το λόγο αυτό, η συγκεκριμένη μελέτη διεξήχθη με σκοπό να ελεγχθεί πιθανή συσχέτιση των προαναφερθέντων παραγόντων με τα ΙΑΕΕ στο συγκεκριμένο αυτό πληθυσμό της Νότιας Ευρώπης. Οι μάρτυρες που επελέγησαν προέρχονταν από την ίδια γεωγραφική περιοχή και ήταν σταθμισμένοι ως προς το φύλο (p=0.567) και την ηλικία (p=0.876).Οι ασθενείς επίσης δε διέφεραν στατιστικά σημαντικά ως προς το κάπνισμα,(p=0.559), την ύπαρξη υπερχοληστερολαιμίας (p=0.112) ή το οικογενειακό ιστορικό ΑΕΕ (p=0.20). Η υπέρταση ήταν πιο συχνή στην ομάδα των ασθενών (8/51, 15.7%)σε σχέση με τους μάρτυρες (0/70, 0%) (p=0.0001). Τόσο οι ασθενείς, όσο και οι μάρτυρες ανήκαν στην Καυκάσια φυλή.Καθώς η συγκεκριμένη μελέτη αποτελεί την πρώτη μελέτη αυτών των παραγόντων στο συγκεκριμένο πληθυσμό στον ελλαδικό χώρο, ένας επιπλέον στόχος ήταν να εκτιμηθεί η σημασία αυτών των μεταβλητών στους ΙΑΕΕ σε νέους ενήλικες,αλλά και η συχνότητά τους στο γενικό πληθυσμό.Τα συμπεράσματα που προέκυψαν για τον εξετασθέντα πληθυσμό, από την παρούσα μελέτη είναι: δεν κατεδείχθη στατιστικά σημαντική διαφορά μεταξύ της ομάδας των ασθενών και αυτής των μαρτύρων σε σχέση με τις μεταβλητές FVL(p=0.199), PT20210 (p= 0.410), MTHFR (p= 0.788), -844G/A PAI-1 (p= 0.493) καιFXIII-AVal34leu (p=0.160). Τα παραπάνω δεδομένα έρχονται σε συμφωνία με διάφορες διεθνείς αναφορές, στις οποίες καταδεικνύεται πως οι παραπάνω μεταβλητές δεν σχετίζονται με αυξημένο κίνδυνο εμφάνισης ΙΑΕΕ σε νεαρούς ενήλικες. Παρ΄ολ’ αυτά υπάρχουν και δημοσιεύσεις που καταδεικνύουν συσχέτιση.Η συχνότητα του αλληλίου 4G/5G του πολυμορφισμού -675 4G/5G του PAI-1 ήταν σημαντικά πιο αυξημένη στην ομάδα των ασθενών σε σχέση με τους μάρτυρες.Επίσης η συχνότητα του αλληλίου 5G/5G ήταν σημαντικά πιο ελαττωμένη στην ομάδα των ασθενών σε σχέση με αυτή των μαρτύρων. (p= 0. 02)Επομένως, αυτή η μελέτη υποδεικνύει πως ο πολυμορφισμός -675 4G/5G της περιοχής του υποκινητή του γονιδίου του παράγοντα ΡΑΙ-1, αποτελεί γενετικό παράγοντα κινδύνου για ΙΑΕΕ σε νεαρή ηλικία, ενώ οι υπόλοιποι από τους κληρονομικούς θρομβοφιλικούς παράγοντες που μελετήθηκαν, δεν φαίνεται να αποτελούν παράγοντες κινδύνου.Επιπλέον μελέτες οι οποίες θα περιλαμβάνουν όλους τους νέους ασθενείς που θα εισάγονται σε νοσοκομείο για ΙΑΕΕ, είναι απαραίτητες, ώστε να εκτιμηθεί η σημασία αυτών των μεταβλητών στον κίνδυνο εμφάνισης ΙΑΕΕ σε νέους ενήλικες.Επιπρόσθετα, άλλοι καινούριοι γενετικοί ή περιβαλλοντικοί παράγοντες που δεν έχουν ακόμη διερευνηθεί, θα μπορούσαν να παίζουν ρόλο στην παθογένεια της νόσου και θα έπρεπε να μελετηθούν στο μέλλον

In Vivo Acquisition of a Plasmid-Mediated blaKPC-2 Gene among Clonal Isolates of Serratia marcescens▿

Three patients admitted to a Greek hospital were infected with Serratia marcescens isolates that exhibited reduced susceptibility to carbapenems and harbored Klebsiella pneumoniae carbapenemase (KPC) enzymes. In two of these cases, the patients were initially infected by carbapenem-susceptible S. marcescens isolates. Molecular typing and plasmid analysis suggested that all three patients had clonally indistinguishable isolates of S. marcescens that acquired a plasmid-mediated blaKPC-2 gene during the hospitalization

Influenza-associated mortality in hospital care: a retrospective cohort study of risk factors and impact of oseltamivir in an English teaching hospital, 2016 to 2017.

BackgroundEvidence of an oseltamivir treatment effect on influenza A(H3N2) virus infections in hospitalised patients is incomplete.AimsThis cohort study aimed to evaluate risk factors for death among PCR-confirmed hospitalised cases of seasonal influenza A(H3N2) of all ages and the impact of oseltamivir.MethodsParticipants included all 332 PCR-confirmed influenza A(H3N2) cases diagnosed between 30 August 2016 and 17 March 2017 in an English university teaching Hospital. Oseltamivir treatment effect on odds of inpatient death was assessed by backward stepwise multivariable logistic regression analysis.ResultsThe odds of death were reduced by two thirds (odds ratio (OR): 0.32; 95% confidence interval (CI): 0.11-0.93), in inpatients treated with a standard course of oseltamivir 75 mg two times daily for 5 days - compared with those untreated with oseltamivir, after adjustment for age, sex, current excess alcohol intake, receipt of 2016/17 seasonal influenza vaccine, serum haemoglobin and hospital vs community attribution of acquisition of influenza.ConclusionsOseltamivir treatment given according to National Institutes of Clinical Excellence (NICE); United States Centres for Disease Control and Prevention (CDC); Infectious Diseases Society of America (IDSA) and World Health Organization (WHO) guidelines was shown to be effective in reducing the odds of mortality in inpatients with PCR-confirmed seasonal influenza A(H3N2) after adjustment in a busy routine English hospital setting. Our results highlight the importance of hospitals complying with relevant guidelines for prompt seasonal influenza PCR testing and ensuring standard oseltamivir treatment to all PCR-confirmed cases of seasonal influenza

Linezolid Dependence in Staphylococcus epidermidis Bloodstream Isolates

We document linezolid dependence among 5 highly linezolid-resistant (LRSE) Staphylococcus epidermidis bloodstream isolates that grew substantially faster at 32 mu g/mL linezolid presence. These isolates carried the mutations T2504A and C2534T in multiple 23S rRNA copies and 2 mutations leading to relevant amino acid substitutions in L3 protein. Linezolid dependence could account for increasing LRSE emergence