Genetic and Phenotypic Characteristics of Staphylococcus aureus Isolates from Cystic Fibrosis Patients in Austria.

BACKGROUND: Cystic fibrosis (CF) is the most common life-limiting inherited disease in Caucasian populations. While pathological changes can be seen in various organs, morbidity and mortality are mainly related to the respiratory tract, with patients suffering from chronic bronchopulmonary infections with characteristic pathogens including Staphylococcus aureus. OBJECTIVES: To date, there is only very limited data on the genetic and phenotypic characteristics of S. aureus in CF patients. Therefore, in our study, we characterized 58 S. aureus isolates collected from CF patients in Austria by spa typing, DNA microarray profiling, as well as antimicrobial susceptibility testing in order to determine common genomic and antimicrobial resistance features. The tested strain collection exhibited high genomic diversity. RESULTS: The 58 isolates were assigned to 16 clonal complexes and 48 spa types and differed greatly regarding their virulence and resistance gene profiles. The predominant clonal complexes were MLST CC30 (22%), CC15 (16%), CC45 (14%), and CC5 (12%), complexes that are highly prevalent worldwide among S. aureus strains isolated from humans colonized or infected with S. aureus. DNA microarray profiles showed a wide variety of genes encoding antimicrobial resistance and virulence factors such as various leukocidins, haemolysins, enterotoxins, exfoliative toxins, toxic shock syndrome toxin, as well as genes involved in adhesion and immune evasion. CONCLUSIONS: While a large number of strains exhibited resistance to one or several antimicrobial agents, methicillin-resistant S. aureus was found at a low prevalence of 3% (n = 2) only. The two methicillin-resistant S. aureus isolates were assigned to CC152/t355 (SCCmecV) and CC5/t001 (SCCmecI). This is the first study to genetically characterize S. aureus isolates in CF patients in Austria.

Modified culture method detects a high diversity of fungal species in cystic fibrosis patients.

Cystic fibrosis (CF) is one of the most common genetic lung diseases worldwide. The production of sticky viscous mucus leads to enhanced bacterial colonization and infection, but yeasts and filamentous fungi are also found abundantly in the mucus of patients suffering from CF. The role of fungi in the airways of CF patients is still not understood completely. Furthermore, recent investigations have shown that the spectrum of fungi isolated from the airways of CF patients depends strongly on the methods used. In this study, different mycological culture methods were compared: culture with a native inoculum, culture with homogenization of CF sputum, and culture after homogenization and serial dilutions of CF sputum. Altogether, 934 sputum samples from 113 patients were examined from July 2009 through December 2011. A total of 1,744 fungal isolates was recovered; 20 different yeasts and 14 filamentous fungal species were identified. Candida albicans, C. dubliniensis, and C. parapsilosis were the most common species of yeast. For the filamentous fungi, Aspergillus fumigatus was the most common, followed by Scedosporium apiospermum/Pseudallescheria boydii group and A. terreus. Many fungal, species such as Exophiala dermatitidis, Rasamsonia (Geosmithia) argillacea, and others, were isolated only from homogenized sputum samples. The longitudinal data also show that fungal colonization of CF patients is quite stable, even when treated with itraconazole. In conclusion, we recommend homogenizing CF sputa with a mucolyticum, to prepare serial dilutions, and to use appropriate fungal culture media with added antibiotics.

Susceptibility of Austrian Clinical Klebsiella and Enterobacter Isolates Linked to Patient-Related Data.

The aim of the study was to analyze the antimicrobial susceptibility of Austrian clinical Klebsiella sp. and Enterobacter sp. isolates linked to patient-related data over a time period from 1998 to 2014. The main findings of this study were (i) a marked difference of antibiotic susceptibility rates between different infection sites for both Klebsiella sp. and Enterobacter sp., (ii) significantly greater percentages of resistant isolates among both Klebsiella sp. and Enterobacter sp. in male patients compared to female patients and (iii) significantly greater percentages of resistant isolates among both Klebsiella sp. and Enterobacter sp. from hospital-derived samples compared to samples from the community. In conclusion, our statistical data analysis clearly indicated a strong association of patient-related data and Klebsiella sp. and Enterobacter sp. susceptibility profiles.

Prevalence, antibiotic resistance patterns and molecular characterization of Escherichia coli from Austrian sandpits.

The aim was to determine the prevalence of E. coli and coliform bacteria in playground sand of all public children's sandpits in Graz (n = 45), Austria, and to assess the frequency of antimicrobial resistance in E. coli. Molecular characterization included the discrimination of O-serotypes and H-antigens and the determination of virulence and resistance genes, using a microarray technology. E. coli isolates were tested for susceptibility to a set of antibiotics by VITEK2 system and disk diffusion method. In total, 22 (49%) and 44 (98%) sandpits were positive for E. coli and coliform bacteria. Median concentrations of E. coli and coliform bacteria in the sand samples were: 2.6 × 10(4) CFU/100 g and 3.0 × 10(5) CFU/100 g. Resistance rates were: ampicillin, 12.5%; piperacillin, 10.4%; amoxicillin/clavulanic acid, 9.4%; cotrimoxazole, 6.3%; tetracycline, 6.3%; piperacillin/tazobactam, 5.2%. No ESBL- or carbapenemase-producing isolates were found. The most prevalent serogroups were O15, O6 and O4. Isolates harbored 0 up to 16 different virulence genes.

Molecular epidemiology of Pseudomonas aeruginosa in cystic fibrosis patients from Southeast Austria.

Pseudomonas aeruginosa is the major pathogen in the cystic fibrosis (CF) lung, the predominant source of its acquisition, however, is under discussion. In order to study the molecular epidemiology, we evaluated 86 P. aeruginosa isolates from 43 CF patients from southeast Austria. The DiversiLab system was used to identify genetic relationships among the isolates. Antibiotic susceptibilities were tested with a broth microdilution method (Micronaut Merlin). A total of 39 unrelated P. aeruginosa genotypes were found of which 34 were unique to a single patient and one was unique to a sibling pair. We found low rates of resistance for ß-lactams with resistance to piperacillin/tazobactam and ceftazidime ranging from 4 to 6%. Resistance rates for meropenem and ciprofloxacin were 11% and 15%, respectively. The prevalence of multidrug-resistant isolates was 2%. We conclude that the majority of P. aeruginosa isolates from CF patients originate from environmental sources and patient-to-patient spread is very uncommon in our centre.

Occurrence and genotyping using automated repetitive-sequence-based PCR of methicillin-resistant Staphylococcus aureus ST398 in Southeast Austria.

In this retrospective study, the occurrence and genetic relatedness of methicillin-resistant Staphylococcus aureus (MRSA) ST398 in Austrian MRSA patients was investigated. From 2002 to 2008, 14 MRSA ST398 were detected. First occurrence of MRSA ST398 was already found in 2004. Spa ribotyping assigned 12 isolates to spa type t011 and 1 each to spa type t034 and spa type t1451. Isolated MRSA ST398 was nontypeable by pulsed-field gel electrophoresis (NT-MRSA) using restriction enzyme SmaI; therefore, genotyping was performed using automated repetitive-sequence-based PCR (rep-PCR) on the DiversiLab system. Rep-PCR results assigned 10 (71%) of the 14 MRSA ST398 into 1 cluster with a similarity >95%; there was 1 cluster consisting of 2 isolates with a similarity >99% and 2 unique MRSA ST398 isolates. In conclusion, MRSA ST398 was continuously detected in Southeast Austria; first in 2004 with up to 5 MRSA ST398 isolates in 2008. Automated rep-PCR proved as a reliable technique in determining genetic relatedness of NT-MRSA ST398 and demonstrates clonal spread of MRSA ST398 in the investigated region.