Evaluation of detection of extended-spectrum beta-lactamases among Escherichia coli and Klebsiella spp. isolates by VITEK 2 AST-N029 compared to the agar dilution and disk diffusion methods.

A total of 123 clinical Escherichia coli and Klebsiella spp. isolates were included in the study in order to evaluate VITEK 2 AST-NO29 (Nordic) card for detection of extended-spectrum beta-lactamases (ESBL) and to compare the results with genotypic ESBL verification. The results were also compared to alternative phenotypic methods, i.e. agar dilution and disk diffusion. The strains that were ESBL-positive according to AST-N029 were further analysed with the ESBL test card, VITEK 2 AST-N041. Using genotype as reference, Vitek 2 AES had the highest accuracy of the tested methods in classifying the strains as ESBL-positive or -negative (91.1%). When VITEK 2 gave ESBL as the only option for E. coli or K. pneumoniae, 44 of 45 (97.8%) strains had an ESBL gene. VITEK 2 achieved an accuracy of 94.9% and disk diffusion 95.9% compared to the agar dilution method as the phenotypic reference method for the E. coli and K. pneumoniae strains. For the K. oxytoca strains VITEK 2 achieved the highest accuracy (84.0%) of the methods used in this work.

Aetiological diagnosis of community acquired pneumonia: utility of rapid microbiological methods with respect to disease severity.

The present study investigated the utility of rapid microbiological methods in the aetiological diagnosis of community acquired pneumonia (CAP) according to the severity of CAP. Between 1999 and 2004, 384 adult patients with CAP were studied prospectively. In addition to standard microbiological methods, PCR and antigen detection techniques were used to identify pathogens. A total of 230 microbial agents in 209 patients were identified, with 134 (58.2%) identified by antigen detection or PCR tests. Of these 134 microbial agents, 95 (70.9%) were identified only by these rapid methods. Streptococcus pneumoniae urinary antigen detection was positive in 24.3% (81/333) of the patients with a diagnostic yield of 38.7% in those with severe pneumonia. Respiratory viral antigen detection was positive in 11.1% (35/314) of the patients with the highest diagnostic yield (20.3%) in patients with severe pneumonia. Mycoplasma pneumoniae PCR was positive in 7.5% (13/174) of the patients, all of whom were low-risk patients. Only 1 case of Chlamydia pneumoniae was identified by PCR. In conclusion, besides yielding the aetiological diagnosis rapidly, new methods add to the total diagnostic yield in CAP. The diagnostic yield of rapid methods differs according to the severity of the pneumonia.

The dynamics of bacteria in the middle ear during the course of acute otitis media with tympanostomy tube otorrhea.

BACKGROUND: Dynamics of bacteria during acute otitis media (AOM) has not been thoroughly studied because it requires repeated tympanocentesis. AOM with tympanostomy tube otorrhea provides a unique opportunity to study the appearance and disappearance of pathogens during the course of the disease without stressing the child. METHODS: Middle ear fluid (MEF) samples were taken before treatment (amoxicillin clavulanate or placebo) and then daily during follow-up from 75 children having AOM with otorrhea through a tympanostomy tube. Bacteria were identified by culture, and typical AOM pathogens also by polymerase chain reaction. RESULTS: Bacteria were initially shown in 67 (89%) children. New bacteria appeared in MEF more often in placebo than in amoxicillin clavulanate recipients [9 of 38 (24%) versus 2 of 37 (5%); P = 0.032]. During the follow-up, new occurrences of Moraxella catarrhalis were detected in MEF more frequently than those of Streptococcus pneumoniae or Haemophilus influenzae. Of the 28 patients with bilateral otorrhea, 11 (39%) had disparate bacteria at study entry and/or during the follow-up. CONCLUSIONS: Changes in bacterial findings during the course of AOM are common in patients not receiving treatment, and even possible despite adequate treatment. In bilateral otorrhea, disparate bacterial findings are common.

Bacteria and viruses in maxillary sinuses of patients with primary hypogammaglobulinemia.

OBJECTIVE: To study bacteria and viruses in maxillary sinuses of patients with primary hypogammaglobulinemia receiving immunoglobulin therapy. DESIGN: Prospective cross-sectional study during 6 months. SETTING: Tertiary care university hospital. PATIENTS: Seventeen patients with primary hypogammaglobulinemia (10 males and 7 females; mean age, 39 years [age range, 11-71 years]). Sixteen patients had common variable immunodeficiency, and 1 patient had X-linked agammaglobulinemia. MAIN OUTCOME MEASURES: Magnetic resonance imaging and x-ray imaging of paranasal sinuses when patients did not have signs of acute infection and reevaluation 6 months later. Maxillary sinus aspiration and lavage were performed at a follow-up visit. Sinus fluid analysis for bacteria and viruses was performed by culture and by polymerase chain reaction. A questionnaire on symptoms related to sinusitis was administered during the follow-up period. RESULTS: Among 17 patients, 9 (53%) had radiologically defined sinusitis without subjective symptoms at study enrollment. At reevaluation 6 months later, radiological findings remained unchanged in two thirds of the patients. Among 15 patients, bacteria were found in sinus lavage samples from 13 patients, and viruses were found in samples from 7 patients. Eight patients had 2 pathogens or more on bacterial culture. Rhinovirus was identified from sinus lavage samples in 5 patients (33%), enterovirus in 3 patients (20%), and respiratory syncytial virus in 1 patient (7%). Pathogenic bacteria were found in maxillary sinuses of all patients who tested positive for rhinovirus and enterovirus. No fungi were found. During the follow-up period, 6 patients reported mucopurulent drainage. CONCLUSIONS: Bacteria and viruses were commonly found in maxillary sinuses of patients with primary hypogammaglobulinemia. Yearly evaluation by an ear, nose, and throat surgeon is recommended.

In vitro susceptibility of viridans group streptococci isolated from blood in southwest Finland in 1993-2004.

We studied in vitro activity against invasive viridans group streptococci (VGS) of penicillin and 4 newer antibiotics, i.e. telithromycin, linezolid, levofloxacin and quinupristin-dalfopristin. Also 7 other antibiotics were tested. Antibiotic susceptibility of 263 VGS blood isolates, collected from southwest Finland during a 12-y period was determined. We wished to discover whether there is an increasing trend of antimicrobial resistance among VGS in Finland. Our results showed that penicillin is still a good choice for treating VGS infections based on the considerably low resistance percentage, 2.3%. Also newer antibiotics showed good in vitro activity: susceptibilities for telithromycin, linezolid and levofloxacin were 100%, 98.9% and 94.6%, respectively. However, quinupristin-dalfopristin was not as effective as described in previous studies, with only 57% susceptibility.

Occurrence of Scopulariopsis and Scedosporium in nails and keratinous skin. A 5-year retrospective multi-center study.

A 5-year retrospective multicenter study was performed for microascaceous moulds (Microascaceae, Ascomycetes) in Finnish clinical specimens. The files from 1993-1997 of six clinical mycology laboratories in Finland were searched for reports of these fungi, mainly Scopulariopsis and Scedosporium anamorphs in keratinous specimens. From the 521 primary findings, 165 cases were selected for further study based on direct microscopy, colony numbers and accompanying fungi. The clinical records of 148 cases (141 Scopulariopsis, 7 Scedosporium) were studied. Of the nail infections from which Scopulariopsis was recovered, 39 cases were further separated which showed clinical or laboratory-based evidence of dermatophytosis. In the remaining 90 'non-dermatophyte' nail cases, Scopulariopsis spp. were the only documented fungal agents (c. 6 cases/million/year). The patients were mainly elderly, 66% of whom had problems involving their big toe nails. For 74% of them, the nail problem was mentioned as their reason for visiting the physician. However, only 18% had documented benefit from treatment. The Scopulariopsis nail infections seem to be treatment-resistant and the pathogenesis and etiological role of Scopulariopsis remain poorly understood.

Identification of alpha-hemolytic streptococci by pyrosequencing the 16S rRNA gene and by use of VITEK 2.

Alpha-hemolytic streptococci are very difficult to identify by phenotypic methods. In this study, a pyrosequencing method for the identification of streptococcal species based on two variable regions of the 16S rRNA gene is described. Almost all studied streptococcal species (n = 51) represented by their type strains could be differentiated except for some closely related species of the Streptococcus bovis or S. salivarius group. The pyrosequencing results of alpha-hemolytic streptococci isolated from blood (n = 99) or from the normal pharyngeal microbiota (n = 25) were compared to the results obtained by the VITEK 2 with GP card (bioMérieux, Marcy l'Etoile, France). As expected, the results of the two methods did not completely agree, but 93 (75.0%) of the isolates assigned to the same streptococcal group by both methods and 57 (46.0%) reached consistent results at the species level. However, 10 strains remained unidentified by VITEK 2, and 4 isolates could not be assigned to any streptococcal group by pyrosequencing. Identification of members of the S. mitis and S. sanguinis groups proved difficult for both methods. Furthermore, the pyrosequencing analysis revealed great sequence variation, since only 43 (32.3%) of the 133 isolates analyzed by pyrosequencing had sequences identical to a type strain. The variation was greatest in the pharyngeal isolates, slightly lower in the blood culture isolates, and nonexistent in invasive pneumococcal isolates (n = 17) that all had the S. pneumoniae type strain sequence. The resolution of the results obtained by the two methods is impeded by the lack of a proper gold standard.

Microbiology of acute otitis media in children with tympanostomy tubes: prevalences of bacteria and viruses.

BACKGROUND: Bacteria are found in 50%-90% of cases of acute otitis media (AOM) with or without otorrhea, and viruses are found in 20%-49% of cases. However, for at least 15% of patients with AOM, the microbiological etiology is never determined. Our aim was to specify the full etiology of acute middle ear infection by using modern microbiological methods concomitantly for bacterial and viral detection. METHODS: The subjects were 79 young children having AOM with new onset (<48 h) of otorrhea through a tympanostomy tube. Middle ear fluid samples were suctioned from the middle ear through the tympanostomy tube. Bacteria were sought by culture and polymerase chain reaction; viruses were analyzed by culture, antigen detection, and polymerase chain reaction. RESULTS: At least 1 respiratory tract pathogen was noted in 76 children (96%). Bacteria were found in 73 cases (92%), and viruses were found in 55 (70%). In 52 patients (66%), both bacteria and viruses were found. Bacteria typical of AOM were detected in 86% of patients. Picornaviruses accounted for 60% of all viral findings. CONCLUSIONS: In the great majority of children, AOM is a coinfection with bacteria and viruses. The patent tympanostomy tube does not change the spectrum of causative agents in AOM. A microbiological etiology can be established in practically all cases.

Comparison of workflow and accuracy of identification and antimicrobial susceptibility testing of clinical isolates of Enterobacteriaceae, Pseudomonas aeruginosa and enterococci by Vitek 2 and routine methods.

Three hundred and fifty-three consecutive urine cultures growing Enterobacteriaceae, Pseudomonas aeruginosa or enterococci were subjected to parallel identification (ID) and antimicrobial susceptibility testing (AST) by Vitek 2 and routine methods, including simple screening tests or API 20 E for ID and standardized disc diffusion for AST. Accuracy of results, technician hands-on time required by both methods and time to results were compared. Vitek 2 correctly identified 322 (94.7%) of the 340 gram-negative isolates and 17 (81%) of the 21 Enterococcus faecalis strains. AST by Vitek 2 and disc diffusion gave category agreement for 4,058 (95.5%) of 4,248 organism-antimicrobial agent combinations. With MIC determination by E-test as reference, AST by Vitek 2 and disc diffusion produced 15 and 3 very major errors, respectively. Six (40%) of the fifteen very major errors by Vitek 2 were associated with trimethoprim-sulfamethoxazole. With an average of 22 specimens processed per day, use of Vitek 2 saved 80 min per day of technician hands-on time as compared to routine methods. Regarding the cost of hands-on worktime and consumables, use of Vitek 2 for identification of Escherichia coli-screened Enterobacteriaceae saved 0.70 p per sample in comparison to API 20 E. More than 80% of Enterobacteriaceae introduced to Vitek 2 in the morning could be reported by 16:00.

Diagnostic value of bronchoalveolar lavage in community-acquired pneumonia in a routine setting: a study on patients treated in a Finnish university hospital.

Only a few previous studies have focused on the use or bronchoalveolar lavage (BAL) in patients with community-acquired pneumonia (CAP). Our aim was to evaluate the diagnostic value of BAL in CAP in a routine clinical setting. 71 disease episodes were retrospectively analysed. The patients had undergone BAL for serious or slowly responding pneumonia. All procedures were performed during antimicrobial treatment of the patient. BAL fluid was cultivated for bacteria, fungi, and viruses. In 68 episodes, 1 or several specific polymerase chain reaction tests were performed. Only 1 (1.3%) quantitative bacterial culture was considered diagnostic for CAP, and indicated a change of antimicrobial treatment. The diagnostic yield increased to 9.8% when other methods were used. A respiratory virus was the only aetiology in 3 (6.0%) patients. In slowly responding pneumonia, also hospital-acquired pathogens and malignancies were identified, resulting in a total diagnostic yield of 20.0%. Thus, even when a large array of diagnostic assays was applied, the value of BAL in pretreated patients with CAP was very small, and its therapeutic implications minimal. In a subgroup of slowly responding pneumonia, the procedure was of some usefulness even after commencement of antimicrobial treatment.

Antibiotic treatment of acute otorrhea through tympanostomy tube: randomized double-blind placebo-controlled study with daily follow-up.

OBJECTIVE: The role of routine antimicrobial treatment of acute middle-ear infections is under debate, because the efficacy of antimicrobials in the resolution of middle-ear fluid has not been unambiguously proven. Acute tube otorrhea is regarded as evidence of acute otitis media, and for methodologic reasons it was chosen to provide objectivity for diagnostics and outcome assessment. The objective of this study was to assess whether amoxicillin-clavulanate accelerates the resolution of acute tube otorrhea. DESIGN AND SETTING: Randomized, double-blind, placebo-controlled study in outpatient setting. PATIENTS: Volunteer sample of basically healthy 6- to 72-month-old children with a tympanostomy tube. Eligibility required having acute tube otorrhea of <48 hours' of duration and no prior treatment within the last 2 weeks. The mean age of the participants was 25 months; they had a history of 3 episodes of acute otitis media (median), and 99% had manifestations of a concomitant respiratory infection. Of 79 randomized patients, 7 were withdrawn because of adverse events; 66 patients completed the study. INTERVENTIONS: Amoxicillin-clavulanate (N = 34; 45 mg/kg/d) or matching placebo (N = 32) for 7 days and daily suction of middle-ear fluid through tympanostomy tube. MAIN OUTCOME MEASURES: Duration of acute tube otorrhea and duration of bacterial growth in middle-ear fluid. RESULTS: The median duration of tube otorrhea was significantly shorter in amoxicillin-clavulanate than in the placebo group (3 vs 8 days). At the end of the 7-day medication period, tube otorrhea was resolved in 28 of 34 children receiving amoxicillin-clavulanate compared with 13 of 32 children on placebo (treatment-control difference 41%; 95% confidence interval, 20%-63%; number needed to treat, 2.4). The median duration of bacterial growth in middle-ear fluid was shorter in amoxicillin-clavulanate than in the placebo group (1 vs 8 days). CONCLUSIONS: Oral antibiotic treatment significantly accelerates the resolution of acute tube otorrhea by reducing bacterial growth in middle-ear fluid.

Elimination of epidemic methicillin-resistant Staphylococcus aureus from a university hospital and district institutions, Finland.

From August 1991 to October 1992, two successive outbreaks of methicillin-resistant Staphylococcus aureus (MRSA) occurred at a hospital in Finland. During and after these outbreaks, MRSA was diagnosed in 202 persons in our medical district; >100 cases involved epidemic MRSA. When control policies failed to stop the epidemic, more aggressive measures were taken, including continuous staff education, contact isolation for MRSA-positive patients, systematic screening for persons exposed to MRSA, cohort nursing of MRSA-positive and MRSA-exposed patients in epidemic situations, and perception of the 30 medical institutions in that district as one epidemiologic entity brought under surveillance and control of the infection control team of Turku University Hospital. Two major epidemic strains, as well as eight additional strains, were eliminated; we were also able to prevent nosocomial spread of other MRSA strains. Our data show that controlling MRSA is possible if strict measures are taken before the organism becomes endemic. Similar control policies may be successful for dealing with new strains of multiresistant bacteria, such as vancomycin-resistant strains of S. aureus.