Clostridium difficile infections in teaching hospital in northern Finland.

BACKGROUND: The aim of this study to compare the incidence of Clostridium difficile (CD) infections in the five university hospital districts in Finland based on national register. The clinical findings of CD cases in the Oulu University Hospital (OUH) in one-year cohort were also analyzed. METHODS: The numbers of the CD cases from the national register were used for the hospital district comparison. A retrospective cohort study was conducted among all adult (> 16 years) patients treated in the OUH in 2013, who had positive CD toxin B gene test in stools. The selection of the cohort was based on the data from the OUH microbiology laboratory and the clinical characteristics were collected from hospital records. RESULTS: The incidence of CD findings in 2013 was higher in the OUH district than in the other four university hospital districts: 159 vs. 70 to 84 per 100,000 inhabitants. In 2013, 261 patients had CD infection treated in the OUH. The yearly number of CD cases treated in the OUH in 2009-2016 varied between 221 and 287, and the corresponding proportion of positive CD findings out of all samples taken varied from 10.0 to 17.8%. A recurrent infection was seen in 58 patients (22%) while the all-cause 30 day mortality was 7.3%. CONCLUSIONS: Diagnostic strategies differed nationally, which may explain the differences in CD incidence between the university hospital districts. In the OUH, no increase in the number of CD infections was seen in 2009-2016. Main characteristics of the patient cohort in the OUH were in harmony with earlier literature.

Clonality behind the increase of multidrug-resistance among non-invasive pneumococci in Southern Finland.

Multidrug-resistance among Streptococcus pneumoniae isolates, especially of serotype 19A, has increased in several countries recently. Even before the introduction of the pneumococcal conjugate vaccine into the Finnish National Vaccination Programme, the proportion of multidrug-resistant (MDR) pneumococci had doubled from 2007 to 2008, when it reached 3.6% in Southern Finland. Our aim was to look for a possible association between antimicrobial susceptibility and clonality among the MDR isolates. Twelve non-invasive isolates non-susceptible to penicillin, erythromycin, clindamycin, trimethoprim/sulfamethoxazole, and doxycycline from 2008 were available for serotyping, genotyping by multilocus sequence typing (MLST), and detection of genes encoding macrolide resistance and adherence-promoting pili. Two isolates were also resistant to ceftriaxone. Five serotypes, 19F, 19A, 6B, 23F, and 14, and six genotypes from three genetic lineages were found, among which CC320 was the largest. All isolates in this study carried the erm(B) macrolide resistance gene, and the CC320 isolates additionally carried the mef(A/E) macrolide resistance gene. Eleven isolates carried pilus islet 1, while the CC320 isolates also carried the pilus islet 2 genes. The findings emphasize the importance of the careful monitoring of antimicrobial susceptibility and serotype distribution among pneumococci, especially now that antimicrobials and pneumococcal vaccines are in widespread use.

Antifungal susceptibility and virulence attributes of bloodstream isolates of Candida from Hong Kong and Finland.

Candida bloodstream infection has dramatically increased in the last decade due to the growing number of immunocompromised populations worldwide. In this study, we evaluated the antifungal susceptibility profiles and virulence attributes of Candida bloodstream isolates (CBIs) derived from Hong Kong and Finland, information which are vital for devising empirical clinical strategies. Susceptibility testing of a wide range of antifungals including fluconazole, itraconazole, voriconazole, ketoconazole, 5-fluorocytosine, amphotericin B and caspofungin was performed. Haemolytic activity and secretion of proteinase of CBIs were also examined. All CBIs derived from Hong Kong were susceptible to all the antifungals tested whilst some CBIs from Finland were resistant to azoles and caspofungin. C. albicans, C. glabrata and C. tropicalis showed higher haemolytic activity whereas C. parapsilosis and C. guilliermondii were non-haemolytic in general. Proteinase activity of the Finland C. albicans isolates was significantly higher than the Hong Kong isolates. Our data provide a glimpse of the possible evolutionary changes in pathogenic potential of Candida that may be occurring in different regions of the world. Therefore, continuous surveillance and availability of local data should be taken into consideration when treating candidemia patients.

New 16-plex PCR method for rapid detection of diarrheagenic Escherichia coli directly from stool samples.

A rapid 16-plex polymerase chain reaction (PCR) suitable for routine diagnostics of diarrheagenic Escherichia coli (EHEC, EIEC, EAEC, ETEC, and EPEC) was developed, validated with control strains, and tested with 250 diarrhoeal stool samples. The specificity was 100% when tested with 289 control bacterial strains, and the analytical sensitivity of automated DNA extraction directly from stool samples was made by boiling the bacterial culture (10(4)-10(5) colony forming units/ml). The assay design starting directly from extraction of stool DNA allowed same day analysis without compromising sensitivity and specificity, which makes it superior compared to PCR after culturing the bacteria. The 16-plex PCR method demonstrated high prevalence of diarrheagenic E. coli in stool samples of patients returning from abroad (39.0%) in contrast to the patients with no travel history (8.7%; p < 0.001). The high prevalence of diarrheagenic E. coli suggests that their screening should be part of normal diarrhoea diagnostics, at least in the leading diagnostic laboratories.

Evaluation of a commercial MRSA assay when multiple MRSA strains are causing epidemics.

Rapid and reliable diagnostic methods are needed to control methicillin-resistant Staphylococcus aureus (MRSA) transmission. We studied the BD GeneOhm MRSA Assay which is based on one specific amplification product at the junction of the right extremity sequence of the staphylococcal cassette chromosome mec (SCCmec) and the chromosomal sequence of orfX of S. aureus. The test was applied on 95 clinical isolates in Finland: 83% were positive. The isolates giving negative results represented several pulsed-field gel electrophoresis (PFGE) types and harboured SCCmec types IV, V, VI or were new types with different combinations of ccr genes.

First isolations of KPC-2-carrying ST258 Klebsiella pneumoniae strains in Finland, June and August 2009.

The first two Klebsiella pneumoniae carbapenemase-producing (KPC) type 2 strains carrying ST258 were detected in Finland in June and early August 2009. They were found colonising two patients transferred from the Mediterranean; one patient referred from a hospital in Greece where isolates were first found in 2007 and another from Italy where the first isolates have been described only very recently.

Non-tuberculous Mycobacteria can Cause Disseminated Mycobacteriosis in Cats.

Mycobacteriosis caused by non-tuberculous mycobacteria (NTM) is a rising concern in human medicine both in immunocompromised and immunocompetent patients. In cats, mycobacteriosis caused by NTM is considered mostly to be a focal or dermal infection, with disseminated disease mostly caused by Mycobacterium avium. We describe three cases of disseminated mycobacteriosis in cats, caused by Mycobacterium malmoense, Mycobacterium branderi/shimoidei and M. avium, with no identified underlying immunosuppression. In all cases, extracellular mycobacteria were seen in the pulmonary epithelium, intestinal lumen and glomerular tufts, which could affect the shedding of the organism. The present study highlights the importance of mycobacteriosis as a differential even in immunocompetent animals. Considering the close relationship of owners and pets and the potential presence of free mycobacteria in secretions, cats should be considered as a possible environmental reservoir for mycobacteria.

Costs of an outbreak of meticillin-resistant Staphylococcus aureus.

An outbreak of meticillin-resistant Staphylococcus aureus (MRSA) occurred in surgical and internal medicine units of a 1752-bed Finnish tertiary care hospital during 2003-2004. In order to analyse the costs of this 14-month outbreak, patients were categorized as follows: patients with MRSA infections; patients with MRSA colonization; patients exposed to MRSA but whose MRSA status remained inconclusive; and exposed patients who were negative for MRSA. We reviewed a sample of patients' charts to determine the types of clinical infections and interviewed staff about the practical implementation of control measures. The number of patients and patient-days involved in the outbreak were identified from the hospital's databases, with the administrative database supplying unit costs of work and materials. Loss of income due to closed beds was analysed. A total of 266 MRSA-positive patients (114 with infections and 152 colonized) and 797 patients exposed to MRSA were identified (11,744 contact isolation days). There were 1240 patients negative after screening (9880 contact isolation days). Total additional costs of MRSA were 386,062 euro (70% for screening and 25% for contact isolation). Costs due to meticillin resistance in treatment of MRSA infections were 16,000 euro. The income loss for this hospital due to closed beds was 1,183,808 euro. The high cost of MRSA screening underlines the importance of appropriate screening methods. Our model of analysing costs might be useful for other hospitals after adapting variables such as local control measures.

Increase of prostate biopsy-related bacteremic complications in southern Finland, 2005-2013: a population-based analysis.

BACKGROUND: The most severe manifestations of prostate biopsy complications are bacteremic infections. These complications are increasing alarmingly. METHODS: A retrospective cohort study of 17 183 transrectal prostate biopsies performed at the Helsinki and Uusimaa hospital district in southern Finland during 2005-2013. Biopsies were linked to a database of positive blood cultures, yielding 111 bacteremic cases, and yearly bacteremia rates were determined. By multiple regression analysis, demographic risk factors of the whole biopsy cohort for developing bacteremia or fluoroquinolone (FQ)-resistant bacteremia were studied. Clinical risk factors for bacteremia caused by an FQ-resistant organism and for serious bacteremic outcomes were studied by univariate and multivariate analyzes. RESULTS: The average bacteremia rate was 0.7% (111 of 17 183 biopsies) and an increase was observed from 0.5% in 2005 (95% confidence interval (CI): 0.3-0.9) to 1.2% in 2012 (95% CI 0.8-1.8); 53.2% were caused by an FQ-resistant organism. In univariate regression analysis, previous biopsy sessions and increasing calendar year of biopsy associated with the risk of developing bacteremia (odds ratio (OR) 1.232, 95% CI: 1.020-1.488, P=0.030 and OR 1.164, 95% CI: 1.079-1.255, P<0.001, respectively), but only increasing calendar year of biopsy remained statistically significant (OR 1.155, 95% CI: 1.070-1.247, P<0.001) in multivariate analysis. Foreign travel within 3 months was associated with FQ resistance in multivariate analysis (OR 7.158, 95% CI: 1.042 to infinite, P=0.045). The study failed to show any significant clinical risk factors for serious bacteremic outcomes (requiring intensive care, developing deep infection foci or death). CONCLUSIONS: The postbiopsy bacteremia rate doubled during the study period and half of the cases were caused by FQ-resistant organisms. Recent foreign travel increased the risk for FQ resistance. Future research efforts should be aimed at better identifying risk factors, targeted prophylaxis and reducing the need for biopsies.

Partitioning of hydrophobic probes into lipopolysaccharide bilayers.

Lipophilic solutes permeate rapidly through lipid bilayer membranes. However, the outer membrane of enteric bacteria, which is composed of a lipopolysaccharide monolayer outer leaflet and the glycerophospholipid inner leaflet, shows extremely low permeability to hydrophobic solutes. In order to examine the cause of this exceptionally low permeability, the lipid/water partition behavior of various lipophilic probes was determined by using lipopolysaccharides of various chemotypes and glycerophospholipids. With all probes, under many different conditions, the lipopolysaccharide/water partition coefficients were generally about an order of magnitude smaller than the phospholipid/water partition coefficients, and this result is consistent with the low permeability of the lipopolysaccharide monolayer, and hence the asymmetric bilayer found in the outer membrane. Furthermore, organic polycations significantly increased the partition of N-phenylnaphthylamine into lipopolysaccharides, a result again consistent with the permeability-increasing effect of such cations on intact outer membrane. Very defective, 'deep rough' lipopolysaccharides of chemotypes Rd2, Rd1 and Re, had only slightly (20-75%) higher partition coefficients in comparison with the more complete lipopolysaccharides, and this difference is probably not enough to explain the approximately 100-fold increase in lipophile permeability seen in deep rough strains.

The nucleotide and deduced amino acid sequence of the cationic 19 kDa outer membrane protein OmpH of Yersinia pseudotuberculosis.

The OmpH proteins of enteric bacteria are recently described, small (16 kDa), cationic outer membrane proteins. Because a Yersinia pseudotuberculosis cell envelope protein of this size has been found to cross-react serologically with the human histocompatibility antigen HLA-B27 (B*2701), the sequence of Y. pseudotuberculosis OmpH was determined by sequencing the gene region which encodes mature OmpH. A protein consisting of 143 amino acid residues was found. It was 96% homologous with the OmpH of Y. enterocolitica and 62% homologous with that of Escherichia coli. Two separate OmpH regions had sequence similarity with B*2701; they were identical in both Yersinia species.

Safety of probiotics that contain lactobacilli or bifidobacteria.

Lactobacilli and bifidobacteria are extremely rare causes of infection in humans, as are probiotics based on these organisms. This lack of pathogenicity extends across all age groups and to immunocompromised individuals. Strains used for new probiotics should be chosen from the commensal flora of humans and should not carry intrinsic resistance to antibiotics that would prevent treatment of a rare probiotic infection. Vigilance regarding the detection of possible rare cases of infection due to probiotics should be maintained, and isolates should be sent to reference centers for molecular characterization and confirmation.

Complete sequence of the ompH gene encoding the 16-kDa cationic outer membrane protein of Salmonella typhimurium.

The complete nucleotide sequence of the ompH gene encoding the 16-kDa basic outer membrane protein of Salmonella typhimurium was determined. The OmpH protein is synthesized in a precursor form with additional 20 amino acid residues in the N terminus of the protein. This peptide has common characteristics of signal sequences. The promoter region has strong homology to consensus sequences of Escherichia coli. The expression of ompH was detected in minicells.

The Ssc protein of enteric bacteria has significant homology to the acyltransferase Lpxa of lipid A biosynthesis, and to three acetyltransferases.

The Ssc protein, a novel essential protein affecting the function of the enterobacterial outer membrane, matched in a protein homology search best with LpxA (UDP-N-acetylglucosamine 3-hydroxymyristoyl transferase), the enzyme which catalyzes the first step of lipid A biosynthesis. The corresponding genes, located 0.56 kb apart, were 46.7% identical. The search also revealed homology to the bacterial acetyltransferases LacA and NodL, as well as to a hypothetical protein Yglm. The region of residues 109-149 Ssc displayed the highest homology and was also homologous with another bacterial acetyltransferase, CysE, and three other bacterial proteins, two of which are hypothetical. This region and the corresponding regions of all other proteins were found to have a peculiar repeated hexapeptide pattern. Each hexapeptide unit starts with isoleucine (or its equivalent leucine and valine). In most units, the second residue is glycine and the fifth residue either valine or alanine.

Bacterial 'histone-like protein I' (HLP-I) is an outer membrane constituent?

The nucleoid-associated 'histone-like protein I' (HLP-I) protein of E. coli was found to be homologous with the cationic 16-kDa outer membrane protein OmpH of Salmonella typhimurium. Deduced from the nucleotide sequence, the HLP-I protein has 91% identical residues with the OmpH protein. Both proteins have very similar cleavable signal sequences. The nucleotide sequence similarity between the corresponding genes hlpA and ompH is 87%. The ompH gene is located in a gene cluster resembling the hlpA-ORF17 region of E. coli which is close to the Ipx genes involved in the biosynthesis of lipopolysaccharides. The localization of the OmpH/HLP-I protein in the cell is discussed.

Phosphate groups in lipopolysaccharides of Salmonella typhimurium rfaP mutants.

Lipopolysaccharides (LPS) of Salmonella typhimurium rfaP mutants and of a galE strain as a control were subjected to analysis by 31P-NMR in order to assess the location of phosphate groups. This was done to obtain direct proof for our earlier finding by chemical analysis that phosphate was lacking in the core oligosaccharide part of the mutant LPS, whereas the core oligosaccharide normally contains several phosphate groups. Such phosphate deficiency has been associated with the increased susceptibility of the rfaP mutants to hydrophobic antibiotics and detergents. Analysis of the de-O-acylated LPS derivatives of S. typhimurium rfaP strains SH7770, SH8551, and SH8572 by 31P-NMR revealed an almost total lack of phosphate groups in the core oligosaccharide part, the LPS phosphates being largely accounted for by the two monophosphate monoesters of lipid A, linked to positions C-1 and C-4' of the lipid A backbone. Core oligosaccharide-linked phosphates were detected in minor proportions only, indicating the presence of some normally phosphorylated core oligosaccharide, due to the inherently leaky nature of the mutation.

The novel hexapeptide motif found in the acyltransferases LpxA and LpxD of lipid A biosynthesis is conserved in various bacteria.

Two bacterial acyltransferases (LpxA of Escherichia coli, LpxD of E. coli and Salmonella typhimurium) have previously been shown to consist of a very unusual tandem-repeat structure with tens of repeating hexapeptides (24 hexapeptides in LpxA, 26 in LpxD). By sequencing LpxD of Yersinia enterocolitica (a distant relative of E. coli and S. typhimurium within Enterobacteriaceae) as well as LpxA of S. typhimurium and Y. enterocolitica, and by analyzing the existing data on these enzymes of Ricketsia rickettsii, it was now shown that the hexapeptide repeat pattern is a very conservative property of these enzymes. Even though the overall homology (allowing equivalent amino acids) between the four proteins was only 59% in LpxA and 58% in LpxD, the homology in the first residue of each hexapeptide was 87% in LpxA and 100% in LpxD. Secondary structure prediction by PredictProtein server suggested a very strong beta strand dominance in all the hexad regions. Accordingly, LpxA and LpxD of various bacterial origins can now be regarded as structurally very unusual enzymes, largely consisting of hexad repeats.

Effect of bacterial lipopolysaccharide on serum high density lipoprotein cholesterol in rabbits.

The effect of bacterial lipopolysaccharide (LPS) on serum lipids was examined in rabbits. LPS was prepared from the smooth Salmonella typhimurium LT2 strain and given intravenously at a dose of 100 ng/kg b.wt. There were no significant changes in serum triglyceride or cholesterol levels in 1-3 days after the administration of LPS. There was, however, a decrease in serum high density lipoprotein (HDL) cholesterol, which was greatest after 2 days (P less than 0.001). Simultaneously, the HDL/total cholesterol ratio decreased (P less than 0.005).

Effect of bacterial lipopolysaccharide on serum lipids and on the development of aortic atherosclerosis in rabbits.

The effect of repeated intravenous administration of bacterial lipopolysaccharide (LPS) on serum lipids and on aortic atherosclerosis was studied in rabbits on basal diet and on hypercholesterolemic diets containing 0.15-1.0% cholesterol. LPS (10 or 100 ng/kg body weight) was administered 3 times per week for 3 or 6 weeks. No difference was observed in serum lipid levels or in aortic atherosclerosis between LPS- and saline-treated animals. These observations do not support the hypothesis that LPS has an effect on the progression of atherosclerosis.

Intramucosal pH and endotoxin and cytokine release in severe acute pancreatitis.

It has been postulated that in severely ill patients splanchnic hypoperfusion may cause endotoxin release from the gut, and this leakage of endotoxin into the circulation can trigger the cascade of inflammatory cytokines. We tested this hypothesis in 9 patients with acute severe pancreatitis by monitoring gastric intramucosal pH (pHi) as measure of splanchnic hypoperfusion at 12-h intervals trying to correlate it to endotoxin and cytokine release. Only 3 of 59 samples, obtained from 3 patients contained circulating endotoxin. Thirteen of 15 plasma samples drawn at pHi <7.20 did not contain endotoxin. The pHi was significantly lower in patients who subsequently developed 3 or more organ failures (P = 0.0017, analysis of variance). Although endotoxemia was only occasionally found, most patients had measurable interleukin 1beta (IL-1beta), interleukin 6 (IL-6), interleukin 8 (IL-8), and interleukin 10 (IL-10) in their plasma. Concentrations of IL-6, IL-8, and IL-10 on admission correlated to degree of organ dysfunction as measured by the multiple organ system failure score (P = 0.035, r = 0.74; P = 0.010, r = 0.91; P = 0.021, r = 0.82, respectively). In conclusion, patients with acute, severe pancreatitis often have splanchnic hypoperfusion and produce a wide array of cytokines despite a rare occurrence of endotoxemia.