Consideration of within-patient diversity highlights transmission pathways and antimicrobial resistance gene variability in vancomycin-resistant Enterococcus faecium.

BACKGROUND: WGS is increasingly being applied to healthcare-associated vancomycin-resistant Enterococcus faecium (VREfm) outbreaks. Within-patient diversity could complicate transmission resolution if single colonies are sequenced from identified cases. OBJECTIVES: Determine the impact of within-patient diversity on transmission resolution of VREfm. MATERIALS AND METHODS: Fourteen colonies were collected from VREfm positive rectal screens, single colonies were collected from clinical samples and Illumina WGS was performed. Two isolates were selected for Oxford Nanopore sequencing and hybrid genome assembly to generate lineage-specific reference genomes. Mapping to closely related references was used to identify genetic variations and closely related genomes. A transmission network was inferred for the entire genome set using Phyloscanner. RESULTS AND DISCUSSION: In total, 229 isolates from 11 patients were sequenced. Carriage of two or three sequence types was detected in 27% of patients. Presence of antimicrobial resistance genes and plasmids was variable within genomes from the same patient and sequence type. We identified two dominant sequence types (ST80 and ST1424), with two putative transmission clusters of two patients within ST80, and a single cluster of six patients within ST1424. We found transmission resolution was impaired using fewer than 14 colonies. CONCLUSIONS: Patients can carry multiple sequence types of VREfm, and even within related lineages the presence of mobile genetic elements and antimicrobial resistance genes can vary. VREfm within-patient diversity could be considered in future to aid accurate resolution of transmission networks.

E. coli bacteraemia and antimicrobial resistance following antimicrobial prescribing for urinary tract infection in the community.

BACKGROUND: Urinary tract infections are one of the most common infections in primary and secondary care, with the majority of antimicrobial therapy initiated empirically before culture results are available. In some cases, however, over 40% of the bacteria that cause UTIs are resistant to some of the antimicrobials used, yet we do not know how the patient outcome is affected in terms of relapse, treatment failure, progression to more serious illness (bacteraemia) requiring hospitalization, and ultimately death. This study analyzed the current patterns of antimicrobial use for UTI in the community in Scotland, and factors for poor outcomes. OBJECTIVES: To explore antimicrobial use for UTI in the community in Scotland, and the relationship with patient characteristics and antimicrobial resistance in E. coli bloodstream infections and subsequent mortality. METHODS: We included all adult patients in Scotland with a positive blood culture with E. coli growth, receiving at least one UTI-related antimicrobial (amoxicillin, amoxicillin/clavulanic acid, ciprofloxacin, trimethoprim, and nitrofurantoin) between 1st January 2009 and 31st December 2012. Univariate and multivariate logistic regression analysis was performed to understand the impact of age, gender, socioeconomic status, previous community antimicrobial exposure (including long-term use), prior treatment failure, and multi-morbidity, on the occurrence of E. coli bacteraemia, trimethoprim and nitrofurantoin resistance, and mortality. RESULTS: There were 1,093,227 patients aged 16 to 100 years old identified as receiving at least one prescription for the 5 UTI-related antimicrobials during the study period. Antimicrobial use was particularly prevalent in the female elderly population, and 10% study population was on long-term antimicrobials. The greatest predictor for trimethoprim resistance in E. coli bacteraemia was increasing age (OR 7.18, 95% CI 5.70 to 9.04 for the 65 years old and over group), followed by multi-morbidity (OR 5.42, 95% CI 4.82 to 6.09 for Charlson Index 3+). Prior antimicrobial use, along with prior treatment failure, male gender, and higher deprivation were also associated with a greater likelihood of a resistant E. coli bacteraemia. Mortality was significantly associated with both having an E. coli bloodstream infection, and those with resistant growth. CONCLUSION: Increasing age, increasing co-morbidity, lower socioeconomic status, and prior community antibiotic exposure were significantly associated with a resistant E. coli bacteraemia, which leads to increased mortality.

Genomic Analysis of Global Staphylococcus argenteus Strains Reveals Distinct Lineages With Differing Virulence and Antibiotic Resistance Gene Content.

Infections due to Staphylococcus argenteus have been increasingly reported worldwide and the microbe cannot be distinguished from Staphylococcus aureus by standard methods. Its complement of virulence determinants and antibiotic resistance genes remain unclear, and how far these are distinct from those produced by S. aureus remains undetermined. In order to address these uncertainties, we have collected 132 publicly available sequences from fourteen different countries, including the United Kingdom, between 2005 and 2018 to study the global genetic structure of the population. We have compared the genomes for antibiotic resistance genes, virulence determinants and mobile genetic elements such as phages, pathogenicity islands and presence of plasmid groups between different clades. 20% (n = 26) isolates were methicillin resistant harboring a mecA gene and 88% were penicillin resistant, harboring the blaZ gene. ST2250 was identified as the most frequent strain, but ST1223, which was the second largest group, contained a marginally larger number of virulence genes compared to the other STs. Novel S. argenteus pathogenicity islands were identified in our isolates harboring tsst-1, seb, sec3, ear, selk, selq toxin genes, as well as chromosomal clusters of enterotoxin and superantigen-like genes. Strain-specific type I modification systems were widespread which would limit interstrain transfer of genetic material. In addition, ST2250 possessed a CRISPR/Cas system, lacking in most other STs. S. argenteus possesses important genetic differences from S. aureus, as well as between different STs, with the potential to produce distinct clinical manifestations.

Origin, maintenance and spread of antibiotic resistance genes within plasmids and chromosomes of bloodstream isolates of Escherichia coli.

Blood stream invasion by Escherichia coli is the commonest cause of bacteremia in the UK and elsewhere with an attributable mortality of about 15-20 %; antibiotic resistance to multiple agents is common in this microbe and is associated with worse outcomes. Genes conferring antimicrobial resistance, and their frequent location on horizontally transferred genetic elements is well-recognised, but the origin of these determinants, and their ability to be maintained and spread within clinically-relevant bacterial populations is unclear. Here, we set out to examine the distribution of antimicrobial resistance genes in chromosomes and plasmids of 16 bloodstream isolates of E. coli from patients within Scotland, and how these genes are maintained and spread. Using a combination of short and long-read whole genome sequencing methods, we were able to assemble complete sequences of 44 plasmids, with 16 Inc group F and 20 col plasmids; antibiotic resistance genes located almost exclusively within the F group. blaCTX-M15 genes had re-arranged in some strains into the chromosome alone (five strains), while others contained plasmid copies alone (two strains). Integrons containing multiple antibiotic genes were widespread in plasmids, notably many with a dfrA7 gene encoding resistance to trimethoprim, thus linking trimethoprim resistance to the other antibiotic resistance genes within the plasmids. This will allow even narrow spectrum antibiotics such as trimethoprim to act as a selective agent for plasmids containing antibiotic resistance genes mediating much broader resistance, including blaCTX-M15. To our knowledge, this is the first analysis to provide complete sequence data of chromosomes and plasmids in a collection of pathogenic human bloodstream isolates of E. coli. Our findings reveal the interplay between plasmids and integrative and conjugative elements in the maintenance and spread of antibiotic resistance genes within pathogenic E. coli.

A highly conserved complete accessory Escherichia coli type III secretion system 2 is widespread in bloodstream isolates of the ST69 lineage.

Bacterial type III secretion systems (T3SSs) play an important role in pathogenesis of Gram-negative infections. Enteropathogenic and enterohemorrhagic Escherichia coli contain a well-defined T3SS but in addition a second T3SS termed E. coli T3SS 2 (ETT2) has been described in a number of strains of E. coli. The majority of pathogenic E. coli contain elements of a genetic locus encoding ETT2, but which has undergone significant mutational attrition rendering it without predicted function. Only a very few strains have been reported to contain an intact ETT2 locus. To investigate the occurrence of the ETT2 locus in strains of human pathogenic E. coli, we carried out genomic sequencing of 162 isolates obtained from patient blood cultures in Scotland. We found that 22 of 26 sequence type (ST) 69 isolates from this collection contained an intact ETT2 together with an associated eip locus which encodes putative secreted ETT2 effectors as well as eilA, a gene encoding a putative transcriptional regulator of ETT2 associated genes. Using a reporter gene for eilA activation, we defined conditions under which this gene was differentially activated. Analysis of published E. coli genomes with worldwide representation showed that ST69 contained an intact ETT2 in these strains as well. The conservation of the genes encoding ETT2 in human pathogenic ST69 strains strongly suggests it has importance in infection, although its exact functional role remains obscure.

Genetic analysis of invasive Escherichia coli in Scotland reveals determinants of healthcare-associated versus community-acquired infections.

Bacteraemia caused by Escherichia coli is a growing problem with a significant mortality. The factors that influence the acquisition and outcome of these infections are not clear. Here, we have linked detailed genetic data from the whole-genome sequencing of 162 bacteraemic isolates collected in Scotland, UK, in 2013-2015, with clinical data in order to delineate bacterial and host factors that influence the acquisition in hospital or the community, outcome and antibiotic resistance. We identified four major sequence types (STs) in these isolates: ST131, ST69, ST73 and ST95. Nearly 50 % of the bacteraemic isolates had a urinary origin. ST69 was genetically distinct from the other STs, with significantly less sharing of accessory genes and with a distinct plasmid population. Virulence genes were widespread and diversely distributed between the dominant STs. ST131 was significantly associated with hospital-associated infections (HAIs), and ST69 with those from the community. However, there was no association of ST with outcome, although patients with HAI had a higher immediate mortality compared to those with community-associated infections (CAIs). Genome-wide association studies revealed genes involved in antibiotic persistence as significantly associated with HAIs and those encoding elements of a type VI secretion system with CAIs. Antibiotic resistance was common, and there were networks of correlated resistance genes and phenotypic antibiotic resistance. This study has revealed the complex interactions between the genotype of E. coli and its ability to cause bacteraemia, and some of the determinants influencing hospital or community acquisition. In part, these are shaped by antibiotic usage, but strain-specific factors are also important.

The Where is Norovirus Control Lost (WINCL) Study: an enhanced surveillance project to identify norovirus index cases in care settings in the UK and Ireland.

BACKGROUND: Norovirus outbreaks have a significant impact on all care settings; little is known about the index cases from whom these outbreaks initiate. AIM: To identify and categorise norovirus outbreak index cases in care settings. METHODS: A mixed-methods, multi-centre, prospective, enhanced surveillance study identified and categorised index cases in acute and non-acute care settings. RESULTS: From 54 participating centres, 537 outbreaks were reported (November 2013 to April 2014): 383 (71.3%) in acute care facilities (ACF); 115 (21.4%) in residential or care homes (RCH) and 39 (7.3%) in other care settings (OCS). Index cases were identified in 424 (79%) outbreaks. Of the 245 index cases who were asymptomatic on admission and not transferred within/into the care setting, 123 (50%) had been an inpatient/resident for 4 days. Four themes emerged: missing the diagnosis, care service under pressure, delay in outbreak control measures and patient/resident location and proximity. CONCLUSION: The true index case is commonly not identified as the cause of a norovirus outbreak with at least 50% of index cases being misclassified. Unrecognised norovirus cross-transmission occurs frequently suggesting that either Standard Infection Control Precautions (SICPs) are being insufficiently well applied, and or SICPs are themselves are insufficient to prevent outbreaks.

Comparison of BD Phoenix, Vitek 2, and MicroScan automated systems for detection and inference of mechanisms responsible for carbapenem resistance in Enterobacteriaceae.

We assessed the ability of three commercial systems to infer carbapenem resistance mechanisms in 39 carbapenemase-producing and 16 other carbapenem-resistant Enterobacteriaceae. The sensitivity/specificity values for "flagging" a likely carbapenemase were 100%/0% (BD Phoenix), 82 to 85%/6 to 19% (MicroScan), and 74%/38% (Vitek 2), respectively. OXA-48 producers were poorly detected, but all systems reliably detected isolates with KPC and most with metallo-carbapenemases.

Microbiology of odontogenic infections in deep neck spaces: a retrospective study.

The primary treatment of deep neck spaces odontogenic infection (DNSOI) with suppuration is surgery. Systemic antimicrobial therapy is an important adjunct. The initial prescription of antimicrobial therapy is empirical. Over the last decade we have observed a change in practice with the use of second-generation cephalosporins, in conjunction with metronidazole, replacing benzylpencillin and metronidazole. More recently evidence has emerged suggesting that antimicrobial resistance in nosocomial infections could be related to the widespread use of second and third-generation cephalosporins. This study was therefore initiated to determine whether this change in prescribing was justified. A total of 75 cases were retrospectively identified by scrutiny of the operating theatre data. These patients presented with significant DNSOI that required surgical drainage. Streptococcus milleri and mixed anaerobes were predominant. Only in three cases (4%) there were penicillin-resistant microorganisms. The substitution of benzylpenicillin for cefuroxime as an initial empiric therapy for DNSOI seems likely to have been equally efficacious in the large majority of cases. On the other hand, studies in preference of cephalosporins are based on in vitro trials. A multi-centre randomized controlled clinical trial directly comparing initial empiric second-generation cephalosporin therapy with benzylpenicillin in non-allergic patients is justified.

Quantitative analysis of bacteria in forefoot surgery: a comparison of skin preparation techniques.

BACKGROUND: Currently a lack of consensus exists on the optimum solution and preparation methods needed to decrease bacteria present during forefoot surgery. We therefore compared the effect of povidine-iodine and chlorhexidine gluconate on lowering bacterial load and to study any additional benefits gained by pre-treatment with the use of a bristled brush. MATERIALS AND METHODS: Fifty consecutive patients undergoing forefoot surgery were recruited into the study and randomized to receive one of two surgical skin preparations (Povidine-iodine 1% with isopropyl alcohol 23% or Chlorhexidine gluconate 0.5% with isopropyl alcohol 70%). In addition to the skin preparation of the foot with the randomized solution, the subjects other foot was also scrubbed with a sterile surgical bristled brush for three minutes and then painted with the same solution. Swabs were taken from three sites and analyzed via qualitative and quantitative analysis before and after prepping. RESULTS: All four preparation methods significantly decreased (p < 0.001), in all three sites, the number of colony forming units. Using two-way analysis of variance, no significant interaction was observed between preparation method and number of colony-forming units, suggesting that no difference in bacterial inhibition between preparation methods. CONCLUSION: We suggest that either povidone-iodine with no more that 23% isopropyl alcohol or chlorhexidine gluconate with 70% isopropyl alcohol be used for surgical preparation in forefoot surgery. No additional benefit in reduction in bacterial load was gained by scrubbing the foot with bristles prior to painting.

Haemophilus influenzae in acute exacerbations of chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a common progressive respiratory disease that is associated with infective exacerbations that lead to worsening of symptoms. Many organisms are thought to trigger infective exacerbations, but Haemophilus influenzae is the most commonly isolated bacterium. The role of H. influenzae in infective exacerbations remains uncertain, mainly because the organism chronically colonises patients whose clinical condition is stable. H. influenzae may also comprise part of the normal nasopharyngeal flora in man, making the interpretation of positive cultures difficult in some cases.