Long-term oral antibiotic use in people with acne vulgaris in UK primary care: a drug utilization study.

BACKGROUND: The inappropriate use of antibiotics is understood to contribute to antimicrobial resistance. Oral antibiotics are regularly used to treat moderate-to-severe acne vulgaris. In practice, we do not know the typical length of oral antibiotic treatment courses for acne in routine primary care and what proportion of people receive more than one course of treatment following a new acne diagnosis. OBJECTIVES: To describe how oral antibiotics are prescribed for acne over time in UK primary care. METHODS: We conducted a descriptive longitudinal drug utilization study using routinely collected primary care data from the Clinical Practice Research Datalink GOLD (2004-2019). We included individuals (8-50 years) with a new acne diagnosis recorded between 1 January 2004 and 31 July 2019. RESULTS: We identified 217 410 people with a new acne diagnosis. The median age was 17 years [interquartile range (IQR) 15-25] and median follow-up was 4.3 years (IQR 1.9-7.6). Among people with a new acne diagnosis, 96 703 (44.5%) received 248 560 prescriptions for long-term oral antibiotics during a median follow-up of 5.3 years (IQR 2.8-8.5). The median number of continuous courses of antibiotic therapy (≥ 28 days) per person was four (IQR 2-6). The majority (n = 59 010, 61.0%) of first oral antibiotic prescriptions in those with a recorded acne diagnosis were between the ages of 12 and 18. Most (n = 71 544, 74.0%) first courses for oral antibiotics were for between 28 and 90 days. The median duration of the first course of treatment was 56 days (IQR 50-93 days) and 18 127 (18.7%) of prescriptions of ≥ 28 days were for < 6 weeks. Among people who received a first course of oral antibiotic for ≥ 28 days, 56 261 (58.2%) received a second course after a treatment gap of ≥ 28 days. The median time between first and second courses was 135 days (IQR 67-302). The cumulative duration of exposure to oral antibiotics during follow-up was 255 days (8.5 months). CONCLUSIONS: Further work is needed to understand the consequences of using antibiotics for shorter periods than recommended. Suboptimal treatment duration may result in reduced clinical effectiveness or repeated exposures, potentially contributing to antimicrobial resistance.

MdaB and NfrA, Two Novel Reductases Important in the Survival and Persistence of the Major Enteropathogen Campylobacter jejuni.

The paralogues RrpA and RrpB, which are members of the MarR family of DNA binding proteins, are important for the survival of the global bacterial foodborne pathogen Campylobacter jejuni under redox stress. We report that RrpA is a positive regulator of mdaB, encoding a flavin-dependent quinone reductase that contributes to the protection from redox stress mediated by structurally diverse quinones, while RrpB negatively regulates the expression of cj1555c (renamed nfrA for NADPH-flavin reductase A), encoding a flavin reductase. NfrA reduces riboflavin at a greater rate than its derivatives, suggesting that exogenous free flavins are the natural substrate. MdaB and NfrA both prefer NADPH as an electron donor. Cysteine substitution and posttranslational modification analyses indicated that RrpA and RrpB employ a cysteine-based redox switch. Complete genome sequence analyses revealed that mdaB is frequently found in Campylobacter and related Helicobacter spp., while nfrA is predominant in C. jejuni strains. Quinones and flavins are redox cycling agents secreted by a wide range of cell types that can form damaging superoxide by one-electron reactions. We propose a model for stress adaptation where MdaB and NfrA facilitate a two-electron reduction mechanism to the less toxic hydroquinones, thus aiding survival and persistence of this major pathogen. IMPORTANCE Changes in cellular redox potential result in alteration in the oxidation state of intracellular metabolites and enzymes; consequently, cells make adjustments that favor growth and survival. The work we present here answers some of the many questions that have remained elusive over the years of investigation into the enigmatic microaerophile bacterium Campylobacter jejuni. We employed molecular approaches to understand the regulation mechanisms and functional analyses to reveal the roles of two novel quinone and flavin reductases; both serve as major pools of cellular redox-active molecules. This work extends our knowledge on bacterial redox sensing mechanisms and the significance of hemostasis.

ß-Lactam-Resistant Streptococcus pneumoniae Dynamics Following Treatment: A Dose-Response Meta-analysis.

BACKGROUND: Patient exposure to antibiotics promotes the emergence of drug-resistant pathogens. The aim of this study was to identify whether the temporal dynamics of resistance emergence at the individual-patient level were predictable for specific pathogen-drug classes. METHODS: Following a systematic review, a novel robust error meta-regression method for dose-response meta-analysis was used to estimate the odds ratio (OR) for carrying resistant bacteria during and following treatment compared to baseline. Probability density functions fitted to the resulting dose-response curves were then used to optimize the period during and/or after treatment when resistant pathogens were most likely to be identified. RESULTS: Studies of Streptococcus pneumoniae treatment with ß-lactam antibiotics demonstrated a peak in resistance prevalence among patients 4 days after completing treatment with a 3.32-fold increase in odds (95% confidence interval [CI], 1.71-6.46). Resistance waned more gradually than it emerged, returning to preexposure levels 1 month after treatment (OR, 0.98 [95% CI, .55-1.75]). Patient isolation during the peak dose-response period would be expected to reduce the risk that a transmitted pathogen is resistant equivalently to a 50% longer isolation window timed from the first day of treatment. CONCLUSIONS: Predictable temporal dynamics of resistance levels have implications both for surveillance and control.

Ethnically diverse urban transmission networks of Neisseria gonorrhoeae without evidence of HIV serosorting.

OBJECTIVE: We aimed to characterise gonorrhoea transmission patterns in a diverse urban population by linking genomic, epidemiological and antimicrobial susceptibility data. METHODS: Neisseria gonorrhoeae isolates from patients attending sexual health clinics at Barts Health NHS Trust, London, UK, during an 11-month period underwent whole-genome sequencing and antimicrobial susceptibility testing. We combined laboratory and patient data to investigate the transmission network structure. RESULTS: One hundred and fifty-eight isolates from 158 patients were available with associated descriptive data. One hundred and twenty-nine (82%) patients identified as male and 25 (16%) as female; four (3%) records lacked gender information. Self-described ethnicities were: 51 (32%) English/Welsh/Scottish; 33 (21%) white, other; 23 (15%) black British/black African/black, other; 12 (8%) Caribbean; 9 (6%) South Asian; 6 (4%) mixed ethnicity; and 10 (6%) other; data were missing for 14 (9%). Self-reported sexual orientations were 82 (52%) men who have sex with men (MSM); 49 (31%) heterosexual; 2 (1%) bisexual; data were missing for 25 individuals. Twenty-two (14%) patients were HIV positive. Whole-genome sequence data were generated for 151 isolates, which linked 75 (50%) patients to at least one other case. Using sequencing data, we found no evidence of transmission networks related to specific ethnic groups (p=0.64) or of HIV serosorting (p=0.35). Of 82 MSM/bisexual patients with sequencing data, 45 (55%) belonged to clusters of ≥2 cases, compared with 16/44 (36%) heterosexuals with sequencing data (p=0.06). CONCLUSION: We demonstrate links between 50% of patients in transmission networks using a relatively small sample in a large cosmopolitan city. We found no evidence of HIV serosorting. Our results do not support assortative selectivity as an explanation for differences in gonorrhoea incidence between ethnic groups.

Impact of Eimeria tenella Coinfection on Campylobacter jejuni Colonization of the Chicken.

Eimeria tenella can cause the disease coccidiosis in chickens. The direct and often detrimental impact of this parasite on chicken health, welfare, and productivity is well recognized; however, less is known about the secondary effects that infection may have on other gut pathogens. Campylobacter jejuni is the leading cause of human bacterial foodborne disease in many countries and has been demonstrated to exert negative effects on poultry welfare and production in some broiler lines. Previous studies have shown that concurrent Eimeria infection can influence the colonization and replication of bacteria, such as Clostridium perfringens and Salmonella enterica serovar Typhimurium. Through a series of in vivo coinfection experiments, this study evaluated the impact that E. tenella infection had on C. jejuni colonization of chickens, including the influence of variations in parasite dose and sampling time after bacterial challenge. Coinfection with E. tenella resulted in a significant increase in C. jejuni colonization in the cecum in a parasite dose-dependent manner but a significant decrease in C. jejuni colonization in the spleen and liver of chickens. The results were reproducible at 3 and 10 days after bacterial infection. This work highlights that E. tenella not only has a direct impact on the health and well-being of chickens but can have secondary effects on important zoonotic pathogens.

Simultaneous colonization by Escherichia coli and Klebsiella pneumoniae harboring mcr-1 in Brazil.

CASE PRESENTATION: We present a case report of a woman, concurrently colonized by polymyxin-resistant E. coli and K. pneumoniae. A Brazilian female patient, in her mid-fifties, was hospitalized with schistosomiasis. During hospitalization, polymyxin-resistant E. coli and K. pneumoniae were isolated from surveillance cultures. METHODS: Identification, antimicrobial susceptibility testings, PCR for mcr-1, plasmid transfer by conjugation and whole genome sequencing were performed. RESULTS: E. coli ST744 and K. pneumoniae ST101 carrying mcr-1 gene were described. Transconjugant E. coli was positive for mcr-1 and IncX4 by PCR. The plasmid is a 33,304-base pair plasmid, and the mcr-1 gene was the only antimicrobial resistance gene present in the plasmid. CONCLUSIONS: This study presents a case report of a hospitalized woman, concurrently colonized by mcr-1-harboring E. coli ST744, a different ST from previously described in Brazil, and a K. pneumoniae ST101.

Genome-Wide Identification by Transposon Insertion Sequencing of Escherichia coli K1 Genes Essential for In Vitro Growth, Gastrointestinal Colonizing Capacity, and Survival in Serum.

Escherichia coli K1 strains are major causative agents of invasive disease of newborn infants. The age dependency of infection can be reproduced in neonatal rats. Colonization of the small intestine following oral administration of K1 bacteria leads rapidly to invasion of the blood circulation; bacteria that avoid capture by the mesenteric lymphatic system and evade antibacterial mechanisms in the blood may disseminate to cause organ-specific infections such as meningitis. Some E. coli K1 surface constituents, in particular the polysialic acid capsule, are known to contribute to invasive potential, but a comprehensive picture of the factors that determine the fully virulent phenotype has not emerged so far. We constructed a library and constituent sublibraries of ∼775,000 Tn5 transposon mutants of E. coli K1 strain A192PP and employed transposon-directed insertion site sequencing (TraDIS) to identify genes required for fitness for infection of 2-day-old rats. Transposon insertions were lacking in 357 genes following recovery on selective agar; these genes were considered essential for growth in nutrient-replete medium. Colonization of the midsection of the small intestine was facilitated by 167 E. coli K1 gene products. Restricted bacterial translocation across epithelial barriers precluded TraDIS analysis of gut-to-blood and blood-to-brain transits; 97 genes were required for survival in human serum. This study revealed that a large number of bacterial genes, many of which were not previously associated with systemic E. coli K1 infection, are required to realize full invasive potential.IMPORTANCEEscherichia coli K1 strains cause life-threatening infections in newborn infants. They are acquired from the mother at birth and colonize the small intestine, from where they invade the blood and central nervous system. It is difficult to obtain information from acutely ill patients that sheds light on physiological and bacterial factors determining invasive disease. Key aspects of naturally occurring age-dependent human infection can be reproduced in neonatal rats. Here, we employ transposon-directed insertion site sequencing to identify genes essential for the in vitro growth of E. coli K1 and genes that contribute to the colonization of susceptible rats. The presence of bottlenecks to invasion of the blood and cerebrospinal compartments precluded insertion site sequencing analysis, but we identified genes for survival in serum.

Frequent Undetected Ward-Based Methicillin-Resistant Staphylococcus aureus Transmission Linked to Patient Sharing Between Hospitals.

Background: Recent evidence suggests that hospital transmission of methicillin-resistant Staphylococcus aureus (MRSA) is uncommon in UK centers that have implemented sustained infection control programs. We investigated whether a healthcare-network analysis could shed light on transmission paths currently sustaining MRSA levels in UK hospitals. Methods: A cross-sectional observational study was performed in 2 National Health Service hospital groups and a general district hospital in Southeast London. All MRSA patients identified at inpatient, outpatient, and community settings between 1 November 2011 and 29 February 2012 were included. We identified genetically defined MRSA transmission clusters in individual hospitals and across the healthcare network, and examined genetic differentiation of sequence type (ST) 22 MRSA isolates within and between hospitals and inpatient or outpatient and community settings, as informed by average and median pairwise single-nucleotide polymorphisms (SNPs) and SNP-based proportions of nearly identical isolates. Results: Two hundred forty-eight of 610 (40.7%) MRSA patients were linked in 90 transmission clusters, of which 27 spanned multiple hospitals. Analysis of a large 32 patient ST22-MRSA cluster showed that 26 of 32 patients (81.3%) had multiple contacts with one another during ward stays at any hospital. No residential, outpatient, or significant community healthcare contacts were identified. Genetic differentiation between ST22 MRSA inpatient isolates from different hospitals was less than between inpatient isolates from the same hospitals (P ≤ .01). Conclusions: There is evidence of frequent ward-based transmission of MRSA brought about by frequent patient admissions to multiple hospitals. Limiting in-ward transmission requires sharing of MRSA status data between hospitals.

High-throughput analysis of Yersinia pseudotuberculosis gene essentiality in optimised in vitro conditions, and implications for the speciation of Yersinia pestis.

BACKGROUND: Yersinia pseudotuberculosis is a zoonotic pathogen, causing mild gastrointestinal infection in humans. From this comparatively benign pathogenic species emerged the highly virulent plague bacillus, Yersinia pestis, which has experienced significant genetic divergence in a relatively short time span. Much of our knowledge of Yersinia spp. evolution stems from genomic comparison and gene expression studies. Here we apply transposon-directed insertion site sequencing (TraDIS) to describe the essential gene set of Y. pseudotuberculosis IP32953 in optimised in vitro growth conditions, and contrast these with the published essential genes of Y. pestis. RESULTS: The essential genes of an organism are the core genetic elements required for basic survival processes in a given growth condition, and are therefore attractive targets for antimicrobials. One such gene we identified is yptb3665, which encodes a peptide deformylase, and here we report for the first time, the sensitivity of Y. pseudotuberculosis to actinonin, a deformylase inhibitor. Comparison of the essential genes of Y. pseudotuberculosis with those of Y. pestis revealed the genes whose importance are shared by both species, as well as genes that were differentially required for growth. In particular, we find that the two species uniquely rely upon different iron acquisition and respiratory metabolic pathways under similar in vitro conditions. CONCLUSIONS: The discovery of uniquely essential genes between the closely related Yersinia spp. represent some of the fundamental, species-defining points of divergence that arose during the evolution of Y. pestis from its ancestor. Furthermore, the shared essential genes represent ideal candidates for the development of novel antimicrobials against both species.

Role of Glycosyltransferases Modifying Type B Flagellin of Emerging Hypervirulent Clostridium difficile Lineages and Their Impact on Motility and Biofilm Formation.

Clostridium difficile is the principal cause of nosocomial infectious diarrhea worldwide. The pathogen modifies its flagellin with either a type A or type B O-linked glycosylation system, which has a contributory role in pathogenesis. We study the functional role of glycosyltransferases modifying type B flagellin in the 023 and 027 hypervirulent C. difficile lineages by mutagenesis of five putative glycosyltransferases and biosynthetic genes. We reveal their roles in the biosynthesis of the flagellin glycan chain and demonstrate that flagellar post-translational modification affects motility and adhesion-related bacterial properties of these strains. We show that the glycosyltransferases 1 and 2 (GT1 and GT2) are responsible for the sequential addition of a GlcNAc and two rhamnoses, respectively, and that GT3 is associated with the incorporation of a novel sulfonated peptidyl-amido sugar moiety whose structure is reported in our accompanying paper (Bouché, L., Panico, M., Hitchen, P., Binet, D., Sastre, F., Faulds-Pain, A., Valiente, E., Vinogradov, E., Aubry, A., Fulton, K., Twine, S., Logan, S. M., Wren, B. W., Dell, A., and Morris, H. R. (2016) J. Biol. Chem. 291, 25439-25449). GT2 is also responsible for methylation of the rhamnoses. Whereas type B modification is not required for flagellar assembly, some mutations that result in truncation or abolition of the glycan reduce bacterial motility and promote autoaggregation and biofilm formation. The complete lack of flagellin modification also significantly reduces adhesion of C. difficile to Caco-2 intestinal epithelial cells but does not affect activation of human TLR5. Our study advances our understanding of the genes involved in flagellar glycosylation and their biological roles in emerging hypervirulent C. difficile strains.

In Vitro Susceptibility to Closthioamide among Clinical and Reference Strains of Neisseria gonorrhoeae.

Neisseria gonorrhoeae is one of the leading antimicrobial resistance threats worldwide. This study determined the MICs of closthioamide to be 0.008 to 0.5 mg/liter for clinical N. gonorrhoeae strains and related species. Cross-resistance with existing antimicrobial resistance was not detected, indicating that closthioamide could be used to treat drug-resistant N. gonorrhoeae.

The In Vitro and In Vivo Effect of Carvacrol in Preventing Campylobacter Infection, Colonization and in Improving Productivity of Chicken Broilers.

The current trend in reducing the antibiotic usage in animal production imposes urgency in the identification of novel biocides. The essential oil carvacrol, for example, changes the morphology of the cell and acts against a variety of targets within the bacterial membranes and cytoplasm, and our in vitro results show that it reduces adhesion and invasion of chicken intestinal primary cells and also biofilm formation. A trial was conducted to evaluate the effects of dietary supplementation of carvacrol at four concentrations (0, 120, 200, and 300 mg/kg of diet) on the performance of Lactobacillus spp., Escherichia coli, Campylobacter spp., and broilers. Each of the four diets was fed to three replicates/trial of 50 chicks each from day 0 to 35. Our results show that carvacrol linearly decreased feed intake, feed conversion rates and increased body weight at all levels of supplementation. Plate count analysis showed that Campylobacter spp. was only detected at 35 days in the treatment groups compared with the control group where the colonization occurred at 21 days. The absence of Campylobacter spp. at 21 days in the treatment groups was associated with a significant increase in the relative abundance of Lactobacillus spp. Also, carvacrol was demonstrated to have a significant effect on E. coli numbers in the cecum of the treatment groups, at all supplementation levels. In conclusion, this study shows for the first time that at different concentrations, carvacrol can delay Campylobacter spp., colonization of chicken broilers, by inducing changes in gut microflora, and it demonstrates promise as an alternative to the use of antibiotics.

Evidence for Community Transmission of Community-Associated but Not Health-Care-Associated Methicillin-Resistant Staphylococcus Aureus Strains Linked to Social and Material Deprivation: Spatial Analysis of Cross-sectional Data.

BACKGROUND: Identifying and tackling the social determinants of infectious diseases has become a public health priority following the recognition that individuals with lower socioeconomic status are disproportionately affected by infectious diseases. In many parts of the world, epidemiologically and genotypically defined community-associated (CA) methicillin-resistant Staphylococcus aureus (MRSA) strains have emerged to become frequent causes of hospital infection. The aim of this study was to use spatial models with adjustment for area-level hospital attendance to determine the transmission niche of genotypically defined CA- and health-care-associated (HA)-MRSA strains across a diverse region of South East London and to explore a potential link between MRSA carriage and markers of social and material deprivation. METHODS AND FINDINGS: This study involved spatial analysis of cross-sectional data linked with all MRSA isolates identified by three National Health Service (NHS) microbiology laboratories between 1 November 2011 and 29 February 2012. The cohort of hospital-based NHS microbiology diagnostic services serves 867,254 usual residents in the Lambeth, Southwark, and Lewisham boroughs in South East London, United Kingdom (UK). Isolates were classified as HA- or CA-MRSA based on whole genome sequencing. All MRSA cases identified over 4 mo within the three-borough catchment area (n = 471) were mapped to small geographies and linked to area-level aggregated socioeconomic and demographic data. Disease mapping and ecological regression models were used to infer the most likely transmission niches for each MRSA genetic classification and to describe the spatial epidemiology of MRSA in relation to social determinants. Specifically, we aimed to identify demographic and socioeconomic population traits that explain cross-area extra variation in HA- and CA-MRSA relative risks following adjustment for hospital attendance data. We explored the potential for associations with the English Indices of Deprivation 2010 (including the Index of Multiple Deprivation and several deprivation domains and subdomains) and the 2011 England and Wales census demographic and socioeconomic indicators (including numbers of households by deprivation dimension) and indicators of population health. Both CA-and HA-MRSA were associated with household deprivation (CA-MRSA relative risk [RR]: 1.72 [1.03-2.94]; HA-MRSA RR: 1.57 [1.06-2.33]), which was correlated with hospital attendance (Pearson correlation coefficient [PCC] = 0.76). HA-MRSA was also associated with poor health (RR: 1.10 [1.01-1.19]) and residence in communal care homes (RR: 1.24 [1.12-1.37]), whereas CA-MRSA was linked with household overcrowding (RR: 1.58 [1.04-2.41]) and wider barriers, which represent a combined score for household overcrowding, low income, and homelessness (RR: 1.76 [1.16-2.70]). CA-MRSA was also associated with recent immigration to the UK (RR: 1.77 [1.19-2.66]). For the area-level variation in RR for CA-MRSA, 28.67% was attributable to the spatial arrangement of target geographies, compared with only 0.09% for HA-MRSA. An advantage to our study is that it provided a representative sample of usual residents receiving care in the catchment areas. A limitation is that relationships apparent in aggregated data analyses cannot be assumed to operate at the individual level. CONCLUSIONS: There was no evidence of community transmission of HA-MRSA strains, implying that HA-MRSA cases identified in the community originate from the hospital reservoir and are maintained by frequent attendance at health care facilities. In contrast, there was a high risk of CA-MRSA in deprived areas linked with overcrowding, homelessness, low income, and recent immigration to the UK, which was not explainable by health care exposure. Furthermore, areas adjacent to these deprived areas were themselves at greater risk of CA-MRSA, indicating community transmission of CA-MRSA. This ongoing community transmission could lead to CA-MRSA becoming the dominant strain types carried by patients admitted to hospital, particularly if successful hospital-based MRSA infection control programmes are maintained. These results suggest that community infection control programmes targeting transmission of CA-MRSA will be required to control MRSA in both the community and hospital. These epidemiological changes will also have implications for effectiveness of risk-factor-based hospital admission MRSA screening programmes.

The Genotoxin Colibactin Is a Determinant of Virulence in Escherichia coli K1 Experimental Neonatal Systemic Infection.

Escherichia coli strains expressing the K1 capsule are a major cause of sepsis and meningitis in human neonates. The development of these diseases is dependent on the expression of a range of virulence factors, many of which remain uncharacterized. Here, we show that all but 1 of 34 E. coli K1 neonatal isolates carried clbA and clbP, genes contained within the pks pathogenicity island and required for the synthesis of colibactin, a polyketide-peptide genotoxin that causes genomic instability in eukaryotic cells by induction of double-strand breaks in DNA. Inactivation of clbA and clbP in E. coli A192PP, a virulent strain of serotype O18:K1 that colonizes the gastrointestinal tract and translocates to the blood compartment with very high frequency in experimental infection of the neonatal rat, significantly reduced the capacity of A192PP to colonize the gut, engender double-strand breaks in DNA, and cause invasive, lethal disease. Mutation of clbA, which encodes a pleiotropic enzyme also involved in siderophore synthesis, impacted virulence to a greater extent than mutation of clbP, encoding an enzyme specific to colibactin synthesis. Restoration of colibactin gene function by complementation reestablished the fully virulent phenotype. We conclude that colibactin contributes to the capacity of E. coli K1 to colonize the neonatal gastrointestinal tract and to cause invasive disease in the susceptible neonate.

Genome-wide evaluation of the interplay between Caenorhabditis elegans and Yersinia pseudotuberculosis during in vivo biofilm formation.

The formation of an incapacitating biofilm on Caenorhabditis elegans by Yersinia pseudotuberculosis represents a tractable model for investigating the genetic basis for host-pathogen interplay during the biofilm-mediated infection of a living surface. Previously we established a role for quorum sensing (QS) and the master motility regulator, FlhDC, in biofilm formation by Y. pseudotuberculosis on C. elegans. To obtain further genome-wide insights, we used transcriptomic analysis to obtain comparative information on C. elegans in the presence and absence of biofilm and on wild-type Y. pseudotuberculosis and Y. pseudotuberculosis QS mutants. Infection of C. elegans with the wild-type Y. pseudotuberculosis resulted in the differential regulation of numerous genes, including a distinct subset of nematode C-lectin (clec) and fatty acid desaturase (fat) genes. Evaluation of the corresponding C. elegans clec-49 and fat-3 deletion mutants showed delayed biofilm formation and abolished biofilm formation, respectively. Transcriptomic analysis of Y. pseudotuberculosis revealed that genes located in both of the histidine utilization (hut) operons were upregulated in both QS and flhDC mutants. In addition, mutation of the regulatory gene hutC resulted in the loss of biofilm, increased expression of flhDC, and enhanced swimming motility. These data are consistent with the existence of a regulatory cascade in which the Hut pathway links QS and flhDC. This work also indicates that biofilm formation by Y. pseudotuberculosis on C. elegans is an interactive process during which the initial attachment/recognition of Yersinia to/by C. elegans is followed by bacterial growth and biofilm formation.

Clonal variation in high- and low-level phenotypic and genotypic mupirocin resistance of MRSA isolates in south-east London.

OBJECTIVES: Both low-level mupirocin resistance (LMR) and high-level mupirocin resistance (HMR) have been identified. The aim of this study was to determine the epidemiology of LMR and HMR in MRSA isolates at five hospitals that have used mupirocin for targeted decolonization as part of successful institutional control programmes. METHODS: All MRSA identified in three microbiology laboratories serving five central and south-east London hospitals and surrounding communities between November 2011 and February 2012 were included. HMR and LMR were determined by disc diffusion testing. WGS was used to derive multilocus sequence types (MLSTs) and the presence of HMR and LMR resistance determinants. RESULTS: Prevalence of either HMR or LMR amongst first healthcare episode isolates from 795 identified patients was 9.69% (95% CI 7.72-11.96); LMR was 6.29% (95% CI 4.70-8.21) and HMR was 3.40% (95% CI 2.25-4.90). Mupirocin resistance was not significantly different in isolates identified from inpatients at each microbiology laboratory, but was more common in genotypically defined 'hospital' rather than 'community' isolates (OR 3.17, 95% CI 1.36-9.30, P = 0.002). LMR was associated with inpatient stay, previous history of MRSA and age ≥65 years; HMR was associated with age ≥65 years and residential postcode outside London. LMR and HMR varied by clone, with both being low in the dominant UK MRSA clone ST22 compared with ST8, ST36 and ST239/241 for LMR and with ST8 and ST36 for HMR. V588F mutation and mupA carriage had high specificity (>97%) and area under the curve (>83%) to discriminate phenotypic mupirocin resistance, but uncertainty around the sensitivity point estimate was large (95% CI 52.50%-94.44%). Mutations in or near the mupA gene were found in eight isolates that carried mupA but were not HMR. CONCLUSIONS: Mupirocin resistance was identified in <10% of patients and varied significantly by clone, implying that changes in clonal epidemiology may have an important role in determining the prevalence of resistance in conjunction with selection due to mupirocin use.

The multidrug-resistant human pathogen Clostridium difficile has a highly mobile, mosaic genome.

We determined the complete genome sequence of Clostridium difficile strain 630, a virulent and multidrug-resistant strain. Our analysis indicates that a large proportion (11%) of the genome consists of mobile genetic elements, mainly in the form of conjugative transposons. These mobile elements are putatively responsible for the acquisition by C. difficile of an extensive array of genes involved in antimicrobial resistance, virulence, host interaction and the production of surface structures. The metabolic capabilities encoded in the genome show multiple adaptations for survival and growth within the gut environment. The extreme genome variability was confirmed by whole-genome microarray analysis; it may reflect the organism's niche in the gut and should provide information on the evolution of virulence in this organism.

Host immunity to Clostridium difficile PCR ribotype 017 strains.

Clostridium difficile is an important nosocomial pathogen and the leading cause of antibiotic-associated diarrhea. Multilocus sequence typing indicates that C. difficile strains belong to five distinct genetic clades encompassing several PCR ribotypes (RT). Since their emergence in 2003, hypervirulent RT027 strains have been a major focus of research; in contrast, our current understanding of RT017-mediated disease pathogenesis lags far behind. In this study, we aimed to characterize host immunity to CF5 and M68, two genetically well-defined RT017 strains. Both strains engaged with host Toll-like receptor 2/6 (TLR2/6), TLR2-CD14, and TLR5 to similar extents in a model cell line. Despite this, CF5 mediated significantly greater dendritic cell (DC) interleukin-12 (IL-12), IL-27, and IL-10 immunity than M68. Both strains elicited similar IL-1ß mRNA levels, and yet only M68 caused a marked increase in secretory IL-1ß. A CF5 cocultured-DC cytokine milieu drove an equipotent Th1 and Th17 response, while M68 promoted greater Th17 immunity. Human gastrointestinal ex vivo cytokine responses to both strains were characterized. Taken together, our data suggest that C. difficile strains mediate overlapping and yet distinct mucosal and DC/T cell immunity. Finally, toxin-driven IL-1ß release supports the hypothesis that this cytokine axis is a likely target for therapeutic intervention for C. difficile infection.

Genome Sequences of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli Recovered from Mid-Stream Urine Samples in Accra, Ghana.

Escherichia coli, a member of the commensal intestinal microbiota, is a significant aetiology of urinary tract infections (UTIs) and has a propensity for acquiring multidrug resistance characteristics, such as extended-spectrum beta-lactamases (ESBLs). Despite the increase in the incidence of ESBL-producing E. coli infections in sub-Saharan Africa, routine ESBL detection in Ghana is often absent, and molecular data on ESBL genotypes is scarce. Eleven ESBL-producing E. coli recovered from mid-stream urine samples were subjected to antimicrobial susceptibility testing and whole-genome sequence analyses. All isolates exhibited multidrug resistance, demonstrating phenotypic resistance to third-generation cephalosporins, such as cefotaxime, ceftazidime, and cefpodoxime. Three isolates demonstrated resistance to norfloxacin (a fluoroquinolone), and one isolate demonstrated intermediate resistance to ertapenem (a carbapenem). Analysis of the draft genomes identified multiple antimicrobial resistance genes including ESBL genotypes blaTEM-1B/TEM-190 (6/11 and 1/11, respectively), blaCTX-M-15/CTX-M-3 (7/11 and 1/11) and blaOXA-1/OXA-181 (3/11 and 1/11). The strains belong to 10 different serotypes and 10 different multilocus sequence types. This study provides information on phenotypic resistance in 11 ESBL E. coli from Ghana and AMR genotypes within their genomes.

Altered innate defenses in the neonatal gastrointestinal tract in response to colonization by neuropathogenic Escherichia coli.

Two-day-old (P2), but not 9-day-old (P9), rat pups are susceptible to systemic infection following gastrointestinal colonization by Escherichia coli K1. Age dependency reflects the capacity of colonizing K1 to translocate from gastrointestinal (GI) tract to blood. A complex GI microbiota developed by P2, showed little variation over P2 to P9, and did not prevent stable K1 colonization. Substantial developmental expression was observed over P2 to P9, including upregulation of genes encoding components of the small intestinal (α-defensins Defa24 and Defa-rs1) and colonic (trefoil factor Tff2) mucus barrier. K1 colonization modulated expression of these peptides: developmental expression of Tff2 was dysregulated in P2 tissues and was accompanied by a decrease in mucin Muc2. Conversely, α-defensin genes were upregulated in P9 tissues. We propose that incomplete development of the mucus barrier during early neonatal life and the capacity of colonizing K1 to interfere with mucus barrier maturation provide opportunities for neuropathogen translocation into the bloodstream.