Ampicillin susceptibility testing of Haemophilus influenzae in the routine clinical laboratory by the EUCAST methodology compared to broth microdilution and the presence of ftsI gene mutations.

OBJECTIVES: To investigate the prevalence of ampicillin resistance in Haemophilus influenzae and the diagnostic accuracy of the EUCAST recommended disc diffusion method to detect the increasingly prevalent ampicillin resistance due to the presence of PBP3 alterations based on mutations in the ftsI gene. METHODS: During a 6-month period all consecutive non-duplicate H. influenzae isolates were prospectively collected and stored. MICs of ampicillin were determined by broth microdilution (BMD). PCR was performed to detect mutations in the ftsI gene. Results of routine disc diffusion susceptibility testing, including the penicillin screening test in accordance with the current EUCAST methodology, as well as additional Etest results, were compared to the BMD as the reference method. RESULTS: In 102 isolates, the prevalence of ampicillin resistance was 28% (29/102) by BMD. There was a good correlation between MICs of ampicillin and the presence of a ß-lactamase and/or an ftsI gene mutation. The prevalence of ampicillin resistance was overestimated using the EUCAST method (33% (34/102)) and underestimated when an additional Etest was used (24% (24/102)) (not significant). The sensitivity and specificity of the EUCAST methodology for the detection of ampicillin resistance were 97% ((28/29); 95% CI, 82-100%) and 92% ((67/73); 95% CI, 83-97%), respectively. CONCLUSIONS: The prevalence of ampicillin resistance was 28%, as determined by BMD. Although the overall diagnostic accuracy of the EUCAST ampicillin disc diffusion was high, misclassification of ampicillin susceptibility may still occur.

Increased exposure to bacterial antigen RpL7/L12 in early stage colorectal cancer patients.

BACKGROUND: Cancer 2010. (c) 2010 American Cancer Society. : Intestinal bacteria have long been implicated in colorectal cancer pathology, and many reports point to a close linkage between Streptococcus bovis biotype I (recently renamed Streptococcus gallolyticus) infections and tumors of the human colon. This work aims to investigate the humoral immune response to this bacterium during different stages of colorectal cancer. METHODS: The presence of serum antibodies against S. bovis antigen RpL7/L12, previously assigned as a potential diagnostic antigen, was evaluated in Dutch (n = 209) and American (n = 112) populations using a newly developed enzyme-linked immunosorbent assay. RESULTS: The analyses consistently showed that an immune response against this bacterial antigen was increased in polyp patients and stage I/II colorectal cancer patients as compared with asymptomatic individuals. This was not paralleled by increased antibody production to endotoxin, an intrinsic cell wall component of the majority of intestinal bacteria, which implies that the humoral immune response against RpL7/L12 is not a general phenomenon induced by the loss of colonic barrier function. Notably, increased anti-RpL7/L12 levels were not or were only mildly detected in late stage colorectal cancer patients having lymph node or distant metastasis. CONCLUSIONS: Cancer 2010. (c) 2010 American Cancer Society. : These findings are indicative of an increased exposure to antigen RpL7/L12 during early stages of colon carcinogenesis and suggest that intestinal bacteria such as S. bovis constitute a risk factor for the progression of premalignant lesions into early stage carcinomas. Clearly, the current findings emphasize the necessity for further studies on the possible etiologic relationship between intestinal bacteria and human colorectal cancer.

Evaluation of real-time PCR and conventional diagnostic methods for the detection of Clostridium difficile-associated diarrhoea in a prospective multicentre study.

In this prospective multicentre study, an enzyme-linked fluorescent assay (VIDAS CDA2; bioMérieux), an enzyme-linked assay [Premier Toxins A and B (PTAB); Meridian] and an in-house real-time PCR amplifying the tcdB gene were compared with the cell cytotoxicity assay used as the 'gold standard' for diagnosis of Clostridium difficile-associated diarrhoea (CDAD). Faecal samples from patients with a request for C. difficile diagnosis and samples from patients with diarrhoea hospitalized for at least 72 h were collected for 3 consecutive months from four university medical centres in The Netherlands. In total, 547 faecal samples were obtained from 450 patients. Of 540 samples available for all of the assays, 84 (15.6 %) showed a positive result in one or more assays. The cell cytotoxicity assay was positive in 31 samples (5.7 %) from 28 patients. A diagnosis of CDAD was not considered by the physician in 5 (23.8 %) of 21 patients with CDAD who were hospitalized for at least 72 h. Compared with the cell cytotoxicity assay, the sensitivity of VIDAS, PTAB and PCR was 83.9, 96.8 and 87.1 %, respectively. The specificity of VIDAS, PTAB and PCR was 97.1, 94.3 and 96.5 %, respectively. The positive and negative predictive values for VIDAS, PTAB and PCR were 63.4 and 99.0 %, 50.9 and 99.8 %, and 60.0 and 99.2 %, respectively. Of 61 samples that were positive in one, two or three of the assays, 56 were available for discordance analysis. Discordance analysis was performed by culture of toxinogenic strains. The concordance of VIDAS, PTAB and PCR with culture was 53.6 % (30/56), 55.4 % (31/56) and 71.4 % (40/56), respectively. It was concluded that real-time PCR had the highest concordance with toxinogenic culture and is therefore the preferred method for diagnosing CDAD in faecal samples. It was also concluded that diagnosis of patients with diarrhoea who have been hospitalized for more than 72 h should focus mainly on the detection of C. difficile, irrespective of the physician's request.

Endogenous endophthalmitis after otitis media.

PURPOSE: To report a case of bacterial endophthalmitis after otitis media in a healthy adult. DESIGN: Interventional case report. METHODS: A healthy 49-year-old Caucasian woman developed acute otitis media of the right ear. Three weeks after the first onset there was a recurrence of the otitis media, with perforation of the tympanic membrane. Two days after this, the woman presented at our clinic with endophthalmitis of the right eye. RESULTS: A culture of vitreous material grew Streptococcus pyogenes (Streptococcus Lancefield group A). The same strain was found in a smear from the perforated ear. Despite aggressive treatment, the affected eye had to be eviscerated. CONCLUSIONS: Otitis media can result in a bacteremia. This may, even in a healthy adult, lead to a devastating endogenous bacterial endophthalmitis.

In vitro activity of the aerosolized agents colistin and tobramycin and five intravenous agents against Pseudomonas aeruginosa isolated from cystic fibrosis patients in southwestern Germany.

Three hundred and eighty-five mucoid and non-mucoid strains of Pseudomonas aeruginosa isolated from 192 sputa from 57 adult cystic fibrosis patients were studied. Susceptibility testing using an agar dilution technique was performed for ceftazidime, ciprofloxacin, fosfomycin, meropenem and piperacillin, and the aerosolized agents colistin and tobramycin. Meropenem, ceftazidime and piperacillin were the most potent agents (susceptibility 86.2%, 84.2% and 84%, respectively). Tobramycin and colistin susceptibility rates were lower, but in light of higher intrabronchial concentrations of these drugs a greater percentage of strains might still be clinically susceptible. Only 46.2% and 41.8% of isolates were susceptible to ciprofloxacin and fosfomycin, respectively.