Candidaemia in Central Slovenia: A 12-year retrospective survey.

BACKGROUND: Candida bloodstream infections (BSI) became an important invasive disease in the late 20th century, in particular among immunocompromised patients. Although considerable progress has been made in the management of patients with invasive mycoses, Candida BSI are still widespread among hospitalised patients and are associated with relatively high mortality. OBJECTIVES: We conducted a retrospective study to evaluate patient characteristics, incidence, species distribution and antifungal susceptibility of BSI isolates of Candida spp. as well as outcomes of Candida BSI from 2001 to 2012, before the widespread use of echinocandins. This is the first epidemiological study of Candida BSI in Slovenia so far. METHODS: All documented candidaemia cases from 2001 to 2012 in two major hospitals-University Medical Centre and Institute of Oncology in Ljubljana, Slovenia-were taken into consideration. Candida BSI were identified in 422 patients (250 male, 172 female). Laboratory and clinical data of these patients were retrospectively analysed. Mann-Whitney U test was used to compare continuous variables and Fisher's exact test or chi-squared test for categorical variables. RESULTS AND CONCLUSIONS: The average incidence of Candida BSI was 0.524/10.000 patient-days (0,317/1000 admissions); 16/422 were younger than 1 year and 251/422 patients were over 60 years old. The most commonly isolated species were Candida albicans and Candida glabrata, followed by Candida parapsilosis. Majority of the patients had a single episode of Candida BSI, multiple episodes of Candida BSI occurred in 18/434 patients (4.1%); in 25/434 patients (5.8%) mixed Candida BSI were observed. Crude 30-day case-fatality rate was 55.4%.

Diagnostic utility of broad range bacterial 16S rRNA gene PCR with degradation of human and free bacterial DNA in bloodstream infection is more sensitive than an in-house developed PCR without degradation of human and free bacterial DNA.

We compared a commercial broad range 16S rRNA gene PCR assay (SepsiTest) to an in-house developed assay (IHP). We assessed whether CD64 index, a biomarker of bacterial infection, can be used to exclude patients with a low probability of systemic bacterial infection. From January to March 2010, 23 patients with suspected sepsis were enrolled. CD64 index, procalcitonin, and C-reactive protein were measured on admission. Broad range 16S rRNA gene PCR was performed from whole blood (SepsiTest) or blood plasma (IHP) and compared to blood culture results. Blood samples spiked with Staphylococcus aureus were used to assess sensitivity of the molecular assays in vitro. CD64 index was lower in patients where possible sepsis was excluded than in patients with microbiologically confirmed sepsis (P = 0.004). SepsiTest identified more relevant pathogens than blood cultures (P = 0.008); in three patients (13%) results from blood culture and SepsiTest were congruent, whereas in four cases (17.4%) relevant pathogens were detected by SepsiTest only. In vitro spiking experiments suggested equal sensitivity of SepsiTest and IHP. A diagnostic algorithm using CD64 index as a decision maker to perform SepsiTest shows improved detection of pathogens in patients with suspected blood stream infection and may enable earlier targeted antibiotic therapy.

Multi locus sequence typing (MLST) used as a tool to confirm the ability of susceptible Helicobacter pylori strains to gain resistance to clarithromycin during eradication therapy.

BACKGROUND/AIMS: Primary resistance of H. pylori to clarithromycin is the most common reason for eradication failure, followed by mixed susceptible/ resistant H. pylori strain infection. To distinguish between mixed infections and H. pylori switch to resistance phenotype during eradication therapy, we proceeded with multi locus sequence typing (MLST) of H. pylori strains isolated from gastric biopsy samples of patients before and after eradication therapy. METHODOLOGY: We collected H. pylori isolates from gastric biopsies from 133 patients who were never treated for H. pylori. Five patients had eradication failure with the first isolate susceptible and second isolate resistant to clarithromycin. To analyse genotypes of first and second H. pylori isolates, we compared H. pylori strain sequences of 7 housekeeping genes with MLST. RESULTS: Five patients had clarithromycin-sensitive H. pylori before eradication therapy and gained H. pylori-resistant to clarithromycin after eradication therapy. The sensitive and resistant colonies of each of the H. pylori populations, taken from patients before/after antibiotic therapy, had identical sequence types (ST) obtained with MLST. CONCLUSIONS: The factors favouring H. pylori survival and switch to antibiotic-resistance during eradication therapy probably enable milder environmental conditions for H. pylori persistence during therapy. One of such factor is the ineffective destruction of mucosa-adhered H. pylori by immune cells during therapy which may be due to locally induced immune deficit by H. pylori molecules like strain specific H. pylori lipopolysaccharides.

Nature and lability of northern Adriatic macroaggregates.

The key organic constituents of marine macroaggregates (macrogels) of prevalently phytoplankton origin, periodically occurring in the northern Adriatic Sea, are proteins, lipids and especially polysaccharides. In this article, the reactivity of various macroaggregate fractions in relation to their composition in order to decode the potentially ¼bioavailable« fractions is summarized and discussed. The enzymatic hydrolysis of the macroaggregate matrix, using α-amylase, ß-glucosidase, protease, proteinase and lipase, revealed the simultaneous degradation of polysaccharides and proteins, while lipids seem largely preserved. In the fresh surface macroaggregate samples, a pronounced degradation of the α-glycosidic bond compared to ß-linkages. Degradation of the colloidal fraction proceeded faster in the higher molecular weight (MW) fractions. N-containing polysaccharides can be important constituents of the higher MW fraction while the lower MW constituents can mostly be composed of poly- and oligosaccharides. Since the polysaccharide component in the higher MW fraction is more degradable compared to N-containing polysaccharides, the higher MW fraction represents a possible path of organic nitrogen preservation. Enzymatic hydrolysis, using α-amylase and ß-glucosidase, revealed the presence of α- and ß-glycosidic linkages in all fractions with similar decomposition kinetics. Our results indicate that different fractions of macroaggregates are subjected to compositional selective reactivity with important implications for macroaggregate persistence in the seawater column and deposition.

Molecular characterization of Staphylococcus aureus isolates from skin and soft tissue infections samples and healthy carriers in the Central Slovenia region.

Staphylococcus aureus is among the most important human pathogens. It is associated with different infections and is a major cause of skin and soft tissue infections (SSTIs). The aim of our study was to compare S. aureus isolates associated with SSTIs with isolates obtained from healthy carriers in the Central Slovenia region in terms of antimicrobial susceptibility, genetic diversity by clonal complex (CC)/sequence type, spa type, and by toxin gene profiling. In total, 274 S. aureus isolates were collected prospectively by culturing wound samples from 461 SSTI patients and nasal samples from 451 healthy carriers. We have demonstrated high heterogeneity in terms of CCs and spa type in both groups of isolates. The main clone among SSTI strains was Panton-Valentine leukocidin gene (pvl) positive CC121, whereas the main clone among carrier strains was CC45 carrying a large range of toxin genes. The main spa type in both groups was t091. Pvl was more frequently present in SSTI strains (31.2% SSTI vs 3.6% carrier strains) and staphylococcal enterotoxin C was more frequently present in carrier strains (1.6% SSTI vs 17.0% carrier strains). We have also demonstrated that methicillin-resistant S. aureus was a rare cause (2.8%) of SSTIs in our region.

One-year experience with modified BD GeneOhm MRSA assay for detection of methicillin-resistant Staphylococcus aureus from pooled nasal, skin, and throat samples.

We report our 1-year experience with modified GeneOhm MRSA assay (formerly IDI-MRSA) for pooled surveillance specimens in low methicillin-resistant Staphylococcus aureus (MRSA) prevalence clinical setting. We have successfully modified the GeneOhm MRSA assay protocol during the specimen preparation step by adding an extra washing step followed by pooling of up to 3 samples per patient (nose, skin, with or without throat) at the lysis step. The sensitivity of the modified assay compared with conventional cultivation was 94.3%, specificity 99.2%, negative predictive value 99.2%, and positive predictive value 94.3%. The modified test is reliable and performed well compared with conventional culture methods in our clinical setting with low-level prevalence of MRSA colonization. Our findings support the use of pooling of the patients samples as a cost-effective way of screening for MRSA colonization.