A study of Clostridium difficile in biofilm mode of growth using the Sorbarod filter system

In all ecosystems, bacteria are likely to grow in biofilms, and the organisms making up the natural flora of the colon are considered to be no exception to this. One of the most important hospital-acquired infections is antibiotic associated diarrhoea, with the causative agent being the Gram-positive anaerobe, Clostridium difficile. This organism elaborates two exotoxins, A and B, which cause disease by interaction with the enterocytes of the colon. Treatment of the condition can be difficult, with treatment failures not uncommon. As the organism produces a heat stable spore, it can survive with ease in the environment. There are no previous reports concerned with the study of anaerobes in an in vitro biofilm system. The work described here is a study of C. docile, using the Sorbarod filter system. This was to determine its suitability in establishing continuous culture biofilms of the organism. It was shown that C. difficile readily established itself on the filters, maintained a titre in excess of 109 cfu/filter for at least 72h. The lack of sporulation in the system confirmed that vegetative growth was being maintained. In the filters, the organism exhibited elongated forms, sometimes in excess of 10 microns in length. Electron microscopy showed that this was probably due to lack of cell separation at cell division. Toxin production appeared to be higher when C. difficile was grown in biofilm in comparison with growth in broth and biofilm effluent. The susceptibility of the organism to benzylpenicillin, vancomycin, metronidazole and rifampicin was determined in broth culture, biofilm and biofilm effluent. Broth minimum inhibitory and minimum bacteriocidal (MIC/MBC) experiments showed, that apart from rifampicin, the organism was tolerant to the antibiotics. This was essentially repeated in the filter system. Rifampicin had a significantly better bactericidal activity against the organism, and synergistic killing was achieved when rifampicin was combined with vancomycin. The growth of C. difficile in biofilm with other common members of the bowel flora was investigated in combination experiments with Bacteroidesfragilis, Enterococcus faecalis and Escherichia coli. No antagonistic effect was demonstrated and antimicrobial susceptibility experiments using C. difficile and E. coli in combination showed resistance of both organisms to broad-spectrum antibiotics. Screening for bacteriophages and other biological agents with activity against C. difficile identified an isolate of Lactobacillus paracasei, which had marked activity against the clostridium as shown by the "sloppy agar" method, but there was a very variable effect in broth and the Sorbarod biofilms. Interestingly, the bacteriocin-like agent had activity against not only a number of other clostridial species, but also against Streptococcus pneumoniae. The work here is thus a novel investigation of an important infection control problem, and is the basis for further detailed work examining the growth and pathogenic properties of C. difficile in biofilm

A study of Clostridium difficile in biofilm mode of growth using the Sorbarod filter system

In all ecosystems, bacteria are likely to grow in biofilms, and the organisms making up the natural flora of the colon are considered to be no exception to this. One of the most important hospital-acquired infections is antibiotic associated diarrhoea, with the causative agent being the Gram-positive anaerobe, Clostridium difficile. This organism elaborates two exotoxins, A and B, which cause disease by interaction with the enterocytes of the colon. Treatment of the condition can be difficult, with treatment failures not uncommon. As the organism produces a heat stable spore, it can survive with ease in the environment. There are no previous reports concerned with the study of anaerobes in an in vitro biofilm system. The work described here is a study of C. docile, using the Sorbarod filter system. This was to determine its suitability in establishing continuous culture biofilms of the organism. It was shown that C. difficile readily established itself on the filters, maintained a titre in excess of 109 cfu/filter for at least 72h. The lack of sporulation in the system confirmed that vegetative growth was being maintained. In the filters, the organism exhibited elongated forms, sometimes in excess of 10 microns in length. Electron microscopy showed that this was probably due to lack of cell separation at cell division. Toxin production appeared to be higher when C. difficile was grown in biofilm in comparison with growth in broth and biofilm effluent. The susceptibility of the organism to benzylpenicillin, vancomycin, metronidazole and rifampicin was determined in broth culture, biofilm and biofilm effluent. Broth minimum inhibitory and minimum bacteriocidal (MIC/MBC) experiments showed, that apart from rifampicin, the organism was tolerant to the antibiotics. This was essentially repeated in the filter system. Rifampicin had a significantly better bactericidal activity against the organism, and synergistic killing was achieved when rifampicin was combined with vancomycin. The growth of C. difficile in biofilm with other common members of the bowel flora was investigated in combination experiments with Bacteroidesfragilis, Enterococcus faecalis and Escherichia coli. No antagonistic effect was demonstrated and antimicrobial susceptibility experiments using C. difficile and E. coli in combination showed resistance of both organisms to broad-spectrum antibiotics. Screening for bacteriophages and other biological agents with activity against C. difficile identified an isolate of Lactobacillus paracasei, which had marked activity against the clostridium as shown by the "sloppy agar" method, but there was a very variable effect in broth and the Sorbarod biofilms. Interestingly, the bacteriocin-like agent had activity against not only a number of other clostridial species, but also against Streptococcus pneumoniae. The work here is thus a novel investigation of an important infection control problem, and is the basis for further detailed work examining the growth and pathogenic properties of C. difficile in biofilm.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Incidence, management and outcomes of the first cfr-mediated linezolid-resistant Staphylococcus epidermidis outbreak in a tertiary referral centre in the Republic of Ireland.

peer-reviewedAim: To report the first Irish outbreak of cfr-mediated linezolid-resistant Staphylococcus epidermidis. Methods: Linezolid-resistant S. epidermidis isolated at University Hospital Limerick from four blood cultures, one wound and four screening swabs (from nine patients) between April and June 2013 were characterized by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and staphylococcal cassette chromosome (SCCmec) typing. Antibiotic susceptibilities were determined according to the guidelines of the British Society for Antimicrobial Chemotherapy. The outbreak was controlled through prohibiting prescription and use of linezolid, adherence to infection prevention and control practices, enhanced environmental cleaning, isolation of affected patients, and hospital-wide education programmes. Findings: PFGE showed that all nine isolates represented a single clonal strain. MLST showed that they belonged to ST2, and SCCmec typing showed that they encoded a variant of SCCmecIII. All nine isolates were cfr positive, and eight isolates were positive for the G2576T 23S rRNA mutation commonly associated with linezolid resistance. Isolates exhibited multiple antibiotic resistances (i.e. linezolid, gentamicin, methicillin, clindamycin, ciprofloxacin, fusidic acid and rifampicin). The adopted infection prevention intervention was effective, and the outbreak was limited to the affected intensive care unit.PUBLISHEDpeer-reviewe

A comparative study of hand hygiene, including alcohol-based hand rub use, among Irish medical and nursing students.

Background: In Ireland, the setting for this study, the national prevalence rate of health careassociated infection (HCAI) in acute-care facilities is 5.2%. Hand hygiene and in particular hand rubbing using alcohol-based hand rub (ABHR) is highly efficacious in preventing HCAI transmission. Yet, compliance among healthcare professionals is sub-optimal. Less is known about the practices of nursing and medical students and no study comparing practices among these groups in Ireland was found. Hence, the aim of this study was to provide insight into the current hand hygiene and hand rubbing practices of nursing and medical students in Ireland and, by doing so, contribute to the broader understanding of this topic. Methods: This observational study employed a cross-sectional, self-reported design. An electronically administered questionnaire was sent to all nursing and medical students from one university. Data were analysed using appropriate software. Results: The response rate was 37% (323/872). Higher compliance with the World Health Organisation ‘my five moments for hand hygiene’ model was reported among nursing students (NS) than medical students (MS), with scope for improvement in both disciplines identified. Hand hygiene compliance was highest after body fluid exposure (99.5% NS, 91% MS) and lowest after touching a patient’s surroundings (61.5 % NS, 57.5% MS). Attitudes towards hand rubbing were largely positive in both disciplines. 16% of NS were not aware of the clinical contraindications to ABHR use, compared to 45% of MS. 9% of NS did not know when to use soap and water and when to use ABHR, compared to 36% of MS. In contrast, more medical students (46%) than nursing students (22%) were routinely using alcohol-based hand rub for decontamination of hands as recommended. Conclusions: Results suggest scope to review current hand hygiene curricula focusing on the knowledge gaps, the practice deficits and the barriers to ABHR usage identified

A pyo-hydropneumothorax with sepsis, secondary to Gardnerella vaginalis infection in a post-partum female

A 20 year old female, 14 days post partum, presented to the Emergency Department in severe respiratory distress. Imaging of her chest revealed a left tension hydropneumothorax with significant mediastinal displacement. A chest drain was inserted and over 4L of cloudy-brown malodourous fluid was drained. Gardnerella Vaginalis was isolated on culture of the fluid. The pyohydropneumothorax and associated sepsis, was refractory to management with a chest drain and antimicrobial therapy. She required a video-assisted thoracoscopic surgery and decortication of her unexpanded lung. She ultimately made a full recovery. Gardnerella vaginalis is considered a dysbiosis of the genitourinary tract, rather than an overtly virulent pathogen. Although extremely rare, there are occasional reports of Gardnerella vaginalis causing infection, even at sites distant from the genitourinary tract. To our knowledge, this is the first documented case of Gardnerella vaginalis causing respiratory sepsis and a pyohydropneumothorax in a healthy, immunocompetent female during the post-partum period. Although it is a unique case, nevertheless, it highlights the need for physicians to be congnisant of G. vaginalis as a potential pathogen when treating post-partum sepsis and indeed, even as a potential pathogen when treating pulmonary infections in obstetric patients. This will lend to prompt initiation of appropriate antimicrobial treatment. Keywords: Pneumothorax, Post-partum, Sepsis, Gardnerella vaginali

Opportunities lost may be the greatest cost of CPE outbreaks

no abstract availabl

Timely publishing of experiences and sharing of knowledge.

Sir, Philip Larkin wrote “man hands on misery to man”. The poet was born in 1922 at the birth of the modern “antibiotic era” and, while he was not referring to infection, the phrase reflects the microbiological phenomenon. Early career doctors, nurses and IPC professionals should feel empowered to report their work and not be over-awed by perceived restrictions or hierarchical clinical and management structures. However, their early career focus is likely to be dominated by ambition for professional success. Controversy, “rocking of boats” and association with incidents of emerging drug resistance may, perhaps, be viewed as best avoided. And such was the case, in 2005, when a 15 year old male cystic fibrosis patient presented with a linezolid-resistant methicillin-sensitive Staphylococcus aureus G2576T mutation secondary to the prolonged use of linezolid by his attending physican in the absence of any specialist advice. Although now superceded by reports in 2006 and 2010 1,2 of similar incidences, at that time we did not publish our learning regarding risks associated with prolonged monotherapy in this setting, ineffective dosing and subsequent bacteriostatic activity

Attitudes and practices of Irish hospital-based physicians towards hand hygiene and handrubbing using alcohol-based hand rubs, a comparison between 2007 and 2015.

Background: Hand hygiene is the cornerstone of infection prevention and control practices, and reduces healthcare-associated infections significantly. However, international evidence suggests that medical doctors demonstrate poor compliance. Aim: To explore and compare practices and attitudes towards hand hygiene, particularly hand rubbing using alcohol-based hand rub (ABHR), among hospital-based physicians in Ireland between 2007 and 2015. Methods: In 2007, a random sample of doctors in a large teaching hospital was invited to complete a postal survey using a validated questionnaire. In 2015, the study was replicated among all doctors employed in a university hospital group, including the setting of the original study, using an online survey. Data were analysed using SPSS and Survey Monkey. Findings: Predominately positive and improving attitudes and practices were found, with 86% of doctors compliant with hand hygiene before patient contact in 2015, compared with 58% in 2007. Ninety-one percent of doctors were compliant after patient contact in 2015, compared with 76% in 2007. In 2015, only 39% of respondents reported that they ‘almost always’ used ABHR for hand hygiene. However, this represents 13.5% more than in 2007. Stated barriers to use of ABHR included dermatological issues, poor acceptance, tolerance and poor availability of ABHR products. Conclusion: Greater awareness of hand hygiene guidelines and greater governance appear to have had a positive impact on practice. However, despite this, practice remains suboptimal and there is scope for substantial improvement. Continued and sustained efforts are required in order to build on progress achieved since the World Health Organization hand hygiene guidelines were published in 2009

A retrospective observational study of the impact of 16s and 18s ribosomal RNA PCR on antimicrobial treatment over seven years: A tertiary hospital experience

Although culture-based methods remain a staple element of microbiology analysis advanced molecular methods increasingly supplement the testing repertoire. Since the advent of 16s and 18s ribosomal RNA PCR in the 2000s, there has been interest in its utility for pathogen detection. Nonetheless, studies assessing the impact on antimicrobial prescribing are limited. We report a single-centre experience of the influence of 16s and 18s PCR testing on antimicrobial treatment, including a cost-analysis. Methods Data were collected retrospectively for all samples sent for 16s and 18s PCR testing between January 2014 and December 2020. Results were compared to any culture-based result. Assessment focused on any change of antimicrobial treatment based on PCR result, or use of the result as supportive evidence for microbiological diagnosis. Results 310 samples relevant to 268 patients were referred for 16s/18s rRNA PCR testing during the period. Culture was performed for 234 samples. Enrichment culture was performed for 83 samples. 82 of 300 samples sent for 16s PCR had positive results (20.8%). When culture was performed, enrichment reduced the outcome of 16s PCR only positive results (4/36 [11.1%] versus 14/35 [40.0%], p = 0.030 where a pathogen found). 18s PCR yielded 9 positive results from 67 samples. The 16s PCR result influenced antimicrobial change for 6 patients (2.2%). We estimated the cost for 16s PCR testing to result in one significant change in antimicrobial therapy to be €3,340. 18s PCR did not alter antimicrobial treatment

Against the onslaught of endemic KPC, the war is being lost on the Irish Front.

In the context of the excellent report of successful control of an outbreak of carbapenemase-producing Klebsiella pneumoniae in an Italian neonatal intensive care unit published in this journal (1), we wish to report the consequences of the first outbreak of KPC-producing Kliebsiella in Ireland and how, despite identification of operational factors associated with the incidence and best efforts towards rectifying those, our 410-bed hospital in the West of Ireland is failing to control endemic KPCs. Globally, there is recognition of the significant morbidity and mortality implications associated with emergence of carbapenemase-producing bacteria (2). The resulting vigilance has resulted in enhanced reporting of outbreaks, many being the first of their kind in specific countries (3), and descriptions of molecular studies to determine incidence and transfer of the carbapenemase-encoding blaKPC-harboring IncFIA plasmid between clonal variants (4). With indicative rates of carriage being circa 20%, infection control specialists are reacting with novel techniques for microbiological detection, strategies for prevention of nosocomial transmission, and clinical microbiologists are facing therapeutic challenges related to limited, relatively unproven antimicrobial treatment options