Antimicrobial susceptibility testing breakpoints and methods from BSAC to EUCAST.

The BSAC Standing Committee on Antimicrobial Susceptibility Testing is one of several European national breakpoint committees that agreed in 2002 to harmonize clinical MIC breakpoints. The process of harmonization has since been completed for commonly used agents, and breakpoints for new agents have been set by EUCAST in accordance with a procedure defined by the EMA. EUCAST breakpoints have now been adopted by a large majority of laboratories in Europe. BSAC implemented the EUCAST breakpoints in its own disc diffusion susceptibility testing method as harmonized breakpoints were agreed to over the years. Since the development of the EUCAST disc diffusion method, several countries with their own disc diffusion methods have switched to the EUCAST method, and BSAC will replace support of its own disc diffusion method with support for the EUCAST method from January 2016. The EUCAST breakpoints are also available in automated systems. The harmonized breakpoints and methods will help to avoid different reports of susceptibility for the same isolate in different countries and enable more reliable comparison of resistance rates in surveillance studies in different countries.

Non-susceptibility trends among enterococci and non-pneumococcal streptococci from bacteraemias in the UK and Ireland, 2001-06.

OBJECTIVES: To describe the current patterns and trends in antimicrobial susceptibility in enterococci and streptococci (excepting pneumococci) from bacteraemia in the UK and Ireland from 2001 to 2006. METHODS: In each year 2001-06, blood culture isolates were collected by 25 laboratories distributed across the UK and Ireland. In total, there were 1408 isolates of enterococci, 1332 of beta-haemolytic streptococci and 1012 of alpha- and non-haemolytic streptococci. A single central laboratory re-identified the isolates and measured MICs by the BSAC agar dilution method. RESULTS: The prevalence of reduced susceptibility in streptococci and enterococci did not change significantly for most antibiotics, but trends were noted to increased ampicillin, imipenem and vancomycin resistance in Enterococcus faecium. The prevalence of reduced susceptibility to macrolides and tetracycline in streptococci, to tetracycline and gentamicin (high level) in enterococci and to beta-lactams and glycopeptides in E. faecium were all high, with some differences in the prevalence among species or groups. CONCLUSIONS: Reduced susceptibility to some antimicrobial agents among streptococci and enterococci remains common and continued surveillance is warranted.

Real-time TaqMan PCR for rapid detection and typing of genes encoding CTX-M extended-spectrum beta-lactamases.

The prevalence of CTX-M-producing members of the Enterobacteriaceae is increasing worldwide. A novel, multiplex, real-time TaqMan PCR assay to detect and type bla(CTX-M) genes is described which is an improvement on previously described techniques with respect to reduced assay time, elimination of the need for protracted post-PCR processing and the convenience of a single reaction vessel. Based on beta-lactam antibiogram and MIC data, 478 of 1279 Enterobacteriaceae isolates from clinical blood and urine culture specimens were selected and tested for extended-spectrum beta-lactamase (ESBL) production using phenotypic methods. The new TaqMan assay detected and typed bla(CTX-M) genes in 21 of 28 ESBL-producing isolates.

Meticillin-resistant Staphylococcus aureus with a novel mecA homologue in human and bovine populations in the UK and Denmark: a descriptive study.

BACKGROUND: Animals can act as a reservoir and source for the emergence of novel meticillin-resistant Staphylococcus aureus (MRSA) clones in human beings. Here, we report the discovery of a strain of S aureus (LGA251) isolated from bulk milk that was phenotypically resistant to meticillin but tested negative for the mecA gene and a preliminary investigation of the extent to which such strains are present in bovine and human populations. METHODS: Isolates of bovine MRSA were obtained from the Veterinary Laboratories Agency in the UK, and isolates of human MRSA were obtained from diagnostic or reference laboratories (two in the UK and one in Denmark). From these collections, we searched for mecA PCR-negative bovine and human S aureus isolates showing phenotypic meticillin resistance. We used whole-genome sequencing to establish the genetic basis for the observed antibiotic resistance. FINDINGS: A divergent mecA homologue (mecA(LGA251)) was discovered in the LGA251 genome located in a novel staphylococcal cassette chromosome mec element, designated type-XI SCCmec. The mecA(LGA251) was 70% identical to S aureus mecA homologues and was initially detected in 15 S aureus isolates from dairy cattle in England. These isolates were from three different multilocus sequence type lineages (CC130, CC705, and ST425); spa type t843 (associated with CC130) was identified in 60% of bovine isolates. When human mecA-negative MRSA isolates were tested, the mecA(LGA251) homologue was identified in 12 of 16 isolates from Scotland, 15 of 26 from England, and 24 of 32 from Denmark. As in cows, t843 was the most common spa type detected in human beings. INTERPRETATION: Although routine culture and antimicrobial susceptibility testing will identify S aureus isolates with this novel mecA homologue as meticillin resistant, present confirmatory methods will not identify them as MRSA. New diagnostic guidelines for the detection of MRSA should consider the inclusion of tests for mecA(LGA251). FUNDING: Department for Environment, Food and Rural Affairs, Higher Education Funding Council for England, Isaac Newton Trust (University of Cambridge), and the Wellcome Trust.

Clinical severity of Clostridium difficile PCR ribotype 027: a case-case study.

BACKGROUND: Clostridium difficile is a leading infectious cause of health care associated diarrhoea. Several industrialised countries have reported increased C. difficile infections and outbreaks, which have been attributed to the emergent PCR ribotype 027 strain. METHODS AND FINDINGS: We conducted a case-case study to compare severity of C. difficile disease for patients with 027 versus non-027 ribotypes. We retrospectively collected clinical information about 123/136 patients with C. difficile infections admitted to hospitals in the East of England region in 2006 and from whom stool isolates were cultured and ribotyped as part of an earlier national survey. We defined severe C. difficile disease as having one or more of shock, paralytic ileus, pseudo membranous colitis or toxic megacolon. Patient median age was 83 years old (range 3 to 98, interquartile range 75 to 89), 86% were prescribed antibiotics in the eight weeks before illness onset, 41% had ribotype 027 and 30-day all cause mortality during hospital admission was 21%. Severe disease occurred in 24% (95%CI 13% to 37%) and 17% (95%CI 9% to 27%) of patients with PCR ribotype 027 and non-027 ribotypes respectively. In a multivariable model, ribotype 027 was not associated with severe disease after adjusting for sex, discharge from hospital prior to 60 days of current admission, gastroenteritis on admission, number of initiator antibiotics for C. difficile disease, and hospital where the patient was admitted. CONCLUSIONS: Our study found no evidence to support previous assertions that ribotype 027 is more virulent than other PCR ribotypes. This finding raises questions about the contribution of this strain to the recent increase in C. difficile disease throughout North America and Europe.

Antimicrobial susceptibility of community-acquired respiratory tract pathogens in the UK during 2002/3 determined locally and centrally by BSAC methods.

OBJECTIVES: To determine the antimicrobial susceptibility of Haemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumoniae causing community-acquired lower respiratory tract infection in the UK during 2002/2003 and to compare susceptibilities determined locally by disc diffusion with agar dilution MICs determined at a central laboratory. METHODS: H. influenzae, M. catarrhalis and S. pneumoniae were isolated in 30 laboratories and susceptibility determined locally by the BSAC standardized disc diffusion method. At a central laboratory, isolates were re-identified, tested for beta-lactamase production (H. influenzae and M. catarrhalis only) and MICs determined using the BSAC agar dilution method. RESULTS: Five hundred and eighty-one H. influenzae, 269 M. catarrhalis and 519 S. pneumoniae were collected. Over 93% of M. catarrhalis and nearly 15% of H. influenzae were beta-lactamase positive rendering these sub-populations resistant to aminopenicillins. Overall, the antibacterial susceptibility rates for the isolates were high. However, macrolides showed poor activity against H. influenzae (0.86-1.38% susceptible by disc or MIC methods) and, compared with other antimicrobials, against S. pneumoniae (approximately 88% susceptible). Between 84% and 95% of H. influenzae, M. catarrhalis and S. pneumoniae were susceptible to cefuroxime but all isolates were susceptible to cefotaxime. Eighty-five percent of H. influenzae were susceptible to trimethoprim. The fluoroquinolones were very active against the isolates, with moxifloxacin showing lower MICs than levofloxacin against S. pneumoniae. Susceptibility determined locally by disc diffusion was in general agreement with that determined centrally by agar dilution MIC testing. However, there was one inconsistency with H. influenzae where disc diffusion indicated 22.9% and 46.8% resistance to clarithromycin and erythromycin, respectively but by MIC, only 0.9% and 6.9% were resistant, respectively. CONCLUSIONS: Rates of resistance within community-acquired respiratory tract isolates were relatively low in the UK, in agreement with other studies. Moxifloxacin was the only antibacterial with over 99% isolates susceptible for each of the three pathogens investigated where breakpoints are available. The comparison between disc susceptibility testing and MIC determination using BSAC methods indicated generally good correlation but has highlighted a methodological problem with macrolides against H. influenzae in particular.

Guidelines for the laboratory diagnosis and susceptibility testing of methicillin-resistant Staphylococcus aureus (MRSA).

These evidence-based guidelines have been produced after a literature review of the laboratory diagnosis and susceptibility testing of methicillin-resistant Staphylococcus aureus (MRSA). We have considered the detection of MRSA in screening samples and the detection of reduced susceptibility to glycopeptides in S. aureus. Recommendations are given for the identification of S. aureus and for suitable methods of susceptibility testing and screening for MRSA and for S. aureus with reduced susceptibility to glycopeptides. These guidelines indicate what tests should be used but not when the tests are applicable, as aspects of this are dealt with in guidelines on control of MRSA. There are currently several developments in screening media and molecular methods. It is likely that some of our recommendations will require modification as the new methods become available.

Evaluation of selective and enrichment media for isolation of glycopeptide-resistant enterococci from faecal specimens.

OBJECTIVES: Vancomycin-resistant enterococcus enrichment broth (VEB) and vancomycin-resistant enterococcus selective agar with vancomycin 6 mg/L (VSA) are novel azide-aesculin agar-based media that contain meropenem as an additional selective agent. The media were compared with enterococcosel broth (EB) and enterococcosel agar with vancomycin 6 mg/L (EA) for the isolation of glycopeptide-resistant enterococci (GRE) from routine faecal screening specimens. METHODS: Two hundred and eighteen routine faecal screening specimens from patients at Addenbrooke's Hospital were examined. The majority were from patients on haematology wards (155) or the intensive therapy unit (ITU) (21). Specimens were inoculated on to VSA and EA directly, and after enrichment in VEB and EB, respectively. RESULTS: One hundred and twenty-eight GRE isolates were recovered from 93 (43%) specimens with enterococci carrying vanA or vanB genes. There were no statistically significant differences between media (specimens positive; numbers of GRE isolates) on direct plating on VSA (87; 104) or EA (86; 97) or following 24 h enrichment in VEB (89; 103) or EB (86; 98). There was no significant advantage to enrichment compared with direct plating. Incubation of enrichment broth cultures for only 6 h appeared detrimental. Enterococci with vanC were isolated significantly less frequently from VEB and VSA than from EB and EA. Growth of organisms other than GRE was more common on VSA than on EA. CONCLUSIONS: VEB and VSA were at least as effective as EB and EA for the recovery of GRE from faecal screening specimens, but substantially more non-GRE grew on VSA than on EA. Enrichment culture offered no significant advantages over direct plating.