Differences in molecular epidemiology of Staphylococcus aureus and Escherichia coli in nursing home residents and people in unassisted living situations.

BACKGROUND: The usefulness of colonization pressure as a working model and proxy for infection transmission is limited due to the inability to grade or quantify the specific risk within environments that are subject to change. AIM: To elaborate on the colonization pressure model by comparing the molecular epidemiology of two bacteria, Staphylococcus aureus and Escherichia coli, among residents in a nursing home and people in unassisted living situations. METHODS: A cross-sectional study of 73 elderly residents from a village in south-central Sweden was conducted. Of these, 35 were residents of a nursing home, and 34 lived in an own place of residence in the same geographical area. Samples of two representative bacterial species were collected from multiple body sites and analysed for molecular diversity. FINDINGS: Combining all body sites, 47% of the participants were colonized with S. aureus and 93% with E. coli. The nursing home group, the group in unassisted living situations, and both units combined, held 16, 17, and 29 different S. aureus spa types, respectively. The corresponding numbers of different E. coli serogenotypes were 34, 28, and 48. Diabetes mellitus was associated with more frequent colonization with S. aureus. CONCLUSION: The molecular diversity of bacteria found within different forms of accommodation was within the same range. Hospital quality hygiene might have contributed to the absence of homogenization of the molecular diversity within the nursing home group. Diabetes mellitus might have played a role in a patient selection characterized by advanced age.

Rapid diagnosis of acute norovirus-associated gastroenteritis: evaluation of the Xpert Norovirus assay and its implementation as a 24/7 service in three hospitals in Jönköping County, Sweden.

Noroviruses are a leading cause of epidemic and sporadic cases of acute gastroenteritis worldwide. The rapid diagnosis of norovirus infection is important for prompt infection control measures and may reduce the need for additional diagnostic testing. Here we evaluated the performance of the rapid Xpert Norovirus assay, and assessed the turn-around time (TAT) before and after the implementation of the analysis as a 24/7 service at all the three hospitals in Jönköping County, Sweden. We describe the implementation process which was performed in two steps during 2014. A total number of 276 clinical samples (stool and vomitus) from patients with symptoms of acute gastroenteritis were included in 2014-2015. The samples were analysed with the Xpert Norovirus assay and the already existing routine method: an in-house reverse transcription real-time PCR. Samples showing discrepant results with the two assays were further analysed by a third PCR method. The Xpert Norovirus assay performed well with a sensitivity of 100% and a specificity of 93% compared to the gold standard (defined as the result obtained by at least two of the three PCR methods). The median TAT decreased from 22 hours in 2013 to 2.4 hours in 2015 (p<0.001). We conclude that the performance of the Xpert Norovirus assay was excellent, and that the implementation of the analysis as a 24/7 service at all three hospitals in the county has greatly reduced the time to diagnosis which is beneficial for both patients and healthcare providers.

Genetic makeup of Shiga toxin-producing Escherichia coli in relation to clinical symptoms and duration of shedding: a microarray analysis of isolates from Swedish children.

Shiga toxin (Stx)-producing Escherichia coli (STECs) cause non-bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome, and are the primary cause of acute renal failure in children worldwide. This study investigated the correlation of genetic makeup of STEC strains as revealed by DNA microarray to clinical symptoms and the duration of STEC shedding. All STEC isolated (n = 96) from patients <10 years of age in Jönköping County, Sweden from 2003 to 2015 were included. Isolates were characterized by DNA microarray, including almost 280 genes. Clinical data were collected through a questionnaire and by reviewing medical records. Of the 96 virulence genes (including stx) in the microarray, 62 genes were present in at least one isolate. Statistically significant differences in prevalence were observed for 21 genes when comparing patients with bloody diarrhea (BD) and with non-bloody stool (18 of 21 associated with BD). Most genes encode toxins (e.g., stx2 alleles, astA, toxB), adhesion factors (i.e. espB_O157, tir, eae), or secretion factors (e.g., espA, espF, espJ, etpD, nleA, nleB, nleC, tccP). Seven genes were associated with prolonged stx shedding; the presence of three genes (lpfA, senB, and stx1) and the absence of four genes (espB_O157, espF, astA, and intI1). We found STEC genes that might predict severe disease outcome already at diagnosis. This can be used to develop diagnostic tools for risk assessment of disease outcome. Furthermore, genes associated with the duration of stx shedding were detected, enabling a possible better prediction of length of STEC carriage after infection.

Staphylococcus aureus colonization related to severity of hand eczema.

Knowledge on Staphylococcus aureus colonization rates and epidemiology in hand eczema is limited. The aim of this study was to clarify some of these issues. Samples were collected by the "glove juice" method from the hands of 59 patients with chronic hand eczema and 24 healthy individuals. Swab samples were taken from anterior nares and throat from 43 of the 59 patients and all healthy individuals. S. aureus were spa typed and analysed by DNA-microarray-based genotyping. The extent of the eczema was evaluated by the hand eczema extent score (HEES). The colonization rate was higher on the hands of hand eczema patients (69 %) compared to healthy individuals (21 %, p < 0.001). This was also seen for bacterial density (p = 0.002). Patients with severe hand eczema (HEES ≥ 13) had a significantly higher S. aureus density on their hands compared to those with milder eczema (HEES = 1 to 12, p = 0.004). There was no difference between patients and healthy individuals regarding colonization rates in anterior nares or throat. spa typing and DNA-microarray-based genotyping indicated certain types more prone to colonize eczematous skin. Simultaneous colonization, in one individual, with S. aureus of different types, was identified in 60-85 % of the study subjects. The colonization rate and density indicate a need for effective treatment of eczema and may have an impact on infection control in healthcare.

Dientamoeba fragilis prevalence coincides with gastrointestinal symptoms in children less than 11 years old in Sweden.

Dientamoeba fragilis is a protozoan with a debated role in gastrointestinal (GI) disease. Although correlated to GI symptoms, no virulence factors have been described. In this study, we evaluated the cause of GI symptoms in children at two schools, with children aged 1 to 10 years, in the county of Jönköping, Sweden. D. fragilis infection correlated to GI symptoms in children and Enterobius vermicularis correlated to D. fragilis infection.

Staphylococcus aureus spa type t437: identification of the most dominant community-associated clone from Asia across Europe.

Methicillin-resistant Staphylococcus aureus (MRSA) belonging to the multilocus sequence type clonal complex 59 (MLST CC59) is the predominant community-associated MRSA clone in Asia. This clone, which is primarily linked with the spa type t437, has so far only been reported in low numbers among large epidemiological studies in Europe. Nevertheless, the overall numbers identified in some Northern European reference laboratories have increased during the past decade. To determine whether the S. aureus t437 clone is present in other European countries, and to assess its genetic diversity across Europe, we analysed 147 S. aureus t437 isolates from 11 European countries collected over a period of 11 years using multiple locus variable number tandem repeat fingerprinting/analysis (MLVF/MLVA) and MLST. Additionally 16 S. aureus t437 isolates from healthy carriers and patients from China were included. Most isolates were shown to be monophyletic with 98% of the isolates belonging to the single MLVA complex 621, to which nearly all included isolates from China also belonged. More importantly, all MLST-typed isolates belonged to CC59. Our study implies that the European S. aureus t437 population represents a genetically tight cluster, irrespective of the year, country and site of isolation. This underpins the view that S. aureus CC59 has been introduced into several European countries, not being restricted to particular geographical regions or specific host environments. The European S. aureus t437 isolates thus bear the general hallmarks of a high-risk clone.

Laboratory diagnosis of Lyme neuroborreliosis: a comparison of three CSF anti-Borrelia antibody assays.

The diagnosis of Lyme neuroborreliosis (LNB) requires the detection of intrathecal synthesis of Borrelia-specific antibodies, but in very early disease, the sensitivity may be low. We compared the performance of the second-generation IDEIA Lyme Neuroborreliosis test (Oxoid), based on purified native flagellum antigen, with two newly developed tests based on several recombinant antigens for the diagnosis of LNB. Patients investigated for LNB during 2003 through 2007 were included (n = 175); 52 with definite LNB, four with possible LNB and 119 non-LNB patients. Serum and cerebrospinal fluid (CSF) were analysed with the IDEIA Lyme Neuroborreliosis (Oxoid), VIDAS Lyme IgG (bioMérieux) and recomBead Borrelia IgM and IgG (Mikrogen) assays. Intrathecal antibody indices (AIs) were calculated according to the manufacturers' protocols. The IDEIA test performed with an overall sensitivity (IgM and IgG AIs taken together) of 88 % and a specificity of 99 %. The VIDAS test showed a sensitivity of 86 % and a specificity of 97 %. An overall sensitivity of 100 % and a specificity of 97 % were achieved by the recomBead test. We conclude that the three assays performed equally well regarding specificity, but our data suggest an improved diagnostic sensitivity with the recomBead Borrelia test.

Prevalence and molecular epidemiology of Staphylococcus aureus in Swedish nursing homes - as revealed in the SHADES study.

SUMMARY: Knowledge of carriage and population dynamics of Staphylococcus aureus is crucial for infection risk assessment and to reveal transmission patterns of strains. We report the prevalence and molecular epidemiology of S. aureus in elderly people (n = 290) living in nursing homes in three cities in the south of Sweden. The overall carriage prevalence rate was 48% when results from nares (31%) and throat (34%) samples were combined. Common spa types were equally distributed but a frequent type, t160, was found only in one of the regions. Carriage of different spa types was detected in 23% of individuals and antimicrobial resistance rates were higher in S. aureus isolates from those carrying more than one spa type. Five of the 21 individuals who carried different spa types were colonized simultaneously with resistant and non-resistant strains. Seventeen per cent of the individuals carried S. aureus of the same spa type on all occasions. Methicillin resistance was not detected. In conclusion we found a high prevalence of S. aureus in this elderly population with a high rate of dual colonization with different spa types. We also found signs of institutional spread of one strain.

A recommendation to perform a blood culture before the administration of intravenous antibiotics increased the detection of Staphylococcus aureus bacteremia.

In 2004, the Surviving Sepsis Campaign was launched to increase awareness and improve the outcome of severe sepsis. Accordingly, in Jönköping County, Sweden, a strong recommendation to perform a blood culture before the start of intravenous antibiotic treatment was introduced in 2007. Moreover, a reminder was included in the laboratory report to consult an infectious disease specialist when Staphylococcus aureus was isolated from a blood culture. Retrospectively, patients with at least one blood culture growing S. aureus during 2002 through 2003 (pre intervention n = 58) or during 2008 through 2009 (post intervention n = 100) were included. Medical records were evaluated regarding clinical data and outcome. Blood culture isolates were characterized by antibiotic susceptibility testing (AST) and S. aureus protein A (spa) gene typing. The annual incidence of S. aureus bacteremia (SAB) increased from 28 per 100,000 inhabitants at the pre intervention period to 45 per 100,000 at the post intervention period (p = 0.046). During post intervention, the SAB incidence was significantly higher in men (p = 0.009). The mortality rate during hospital stay was 14 % during pre intervention and 18 % during post intervention (p = 0.47). The most common spa types were t012 and t084. The Surviving Sepsis Campaign resulted in an increased number of detected cases of SAB. The mortality rate was the same before and after the intervention, and no spa type correlated to certain clinical manifestations or mortality.

Possible transmission of Candida albicans on an intensive care unit: genotype and temporal cluster analyses.

BACKGROUND: Nosocomial transmission of Candida spp. has not been fully explored and previous studies have shown conflicting results. AIM: To evaluate the possible nosocomial transmission of Candida spp. on an intensive care unit (ICU). METHODS: A prospective study was conducted for a period of 19 months, including all patients on our ICU with growth of Candida spp. from surveillance and directed cultures. Molecular typing with repetitive sequence-based polymerase chain reaction was used to define genotype relationships between the Candida albicans and Candida glabrata isolates. Candida isolates obtained from blood cultures taken from patients in our county outside the ICU were used as a reference. Temporal cluster analysis was performed to evaluate genotype distribution over time. FINDINGS: Seventy-seven patients with 78 ICU stays, representing 12% of all ICU stays, were found to harbour 180 isolates of Candida spp. Molecular typing revealed 27 C. albicans genotypes and 10 of C. glabrata. Possible clustering, indicated by overlapping stays of patients with indistinguishable candida genotypes, was observed on seven occasions with C. albicans and on two occasions with C. glabrata. Two C. albicans genotypes were found significantly more often in the ICU group compared with the reference group. Moreover, C. albicans genotypes isolated from more than one patient were significantly more often found in the ICU group. Temporal cluster analysis revealed a significantly increased number of pairs with indistinguishable genotypes at a 21-day interval, indicating clustering. CONCLUSION: This study indicates possible transmission of C. albicans between ICU patients based on genotyping and temporal cluster analysis.

The role of broth enrichment in Staphylococcus aureus cultivation and transmission from the throat to newborn infants: results from the Swedish hygiene intervention and transmission of S. aureus study.

Staphylococcus aureus is detected by direct plating, whereas incubation in enrichment broth prior to plating to increase the proportion of positive samples has not been fully evaluated. S. aureus throat colonization has been suggested to be more common than colonization of the anterior nares, but no data are available on the transmission of S. aureus from the throat. Swab samples were collected from the anterior nares and umbilicus from newborn infants (n = 168), anterior nares, throat, skin lesions, and vagina from parents (n = 332), and anterior nares, throat, and skin lesions from healthcare workers (n = 231) at three maternity wards. spa typing was used to elucidate the transmission routes of S. aureus. The use of enrichment broth prior to plating increased the proportion of positive samples by 46%. The prevalence of S. aureus colonization in adults was 58%. Throat colonization (47%) was significantly more common than colonization in any of the other screened sites (p < 0.001). In total, 103 out of 168 (61%) newborn infants were colonized during their hospital stay. Overall, 124 S. aureus transmissions to newborn infants were detected. Although we detected an increased risk of transmission from the nares as compared to the throat, with an odds ratio of 4.8 [95% confidence interval (CI) 1.8-12.7], we detected a transmission rate of 7 % from the throat. We show that S. aureus throat colonization is more common than colonization in any of the other sites among the parents and staff. We also show evidence of transmission from the throat.

The inverted CD4/CD8 ratio and associated parameters in 66-year-old individuals: the Swedish HEXA immune study.

The Swedish OCTO and NONA immune longitudinal studies were able to identify and confirm an immune risk profile (IRP) predictive of an increased 2-year mortality in very old individuals, 86-94 years of age. The IRP, was associated with persistent cytomegalovirus infection and characterized by inverted CD4/CD8 ratio and related to expansion of terminally differentiated effector memory T cells (TEMRA phenotype). In the present HEXA immune longitudinal study, we have examined a younger group of elderly individuals (n = 424, 66 years of age) in a population-based sample in the community of Jönköping, Sweden, to examine the relevance of findings previously demonstrated in the very old. Immunological monitoring that was conducted included T cell subsets and CMV-IgG and CMV-IgM serology. The result showed a prevalence of 15 % of individuals with an inverted CD4/CD8 ratio, which was associated with seropositivity to cytomegalovirus and increases in the level of TEMRA cells. The proportion of individuals with an inverted CD4/CD8 ratio was significantly higher in men whereas the numbers of CD3+CD4+ cells were significantly higher in women. In conclusion, these findings are very similar to those previously found by us in the Swedish longitudinal studies, suggesting that an immune profile previously identified in the very old also exists in the present sample of hexagenerians. Therefore, it will be important to examine clinical parameters, including morbidity and mortality, to assess whether the immune profile also is a risk profile associated with higher mortality in this sample of hexagenerians.

Methicillin-resistant Staphylococcus aureus (MRSA) in municipal wastewater: an uncharted threat?

AIMS: (i) To cultivate methicillin-resistant Staphylococcus aureus (MRSA) from a full-scale wastewater treatment plant (WWTP), (ii) To characterize the indigenous MRSA-flora, (iii) To investigate how the treatment process affects clonal distribution and (iv) To examine the genetic relation between MRSA from wastewater and clinical MRSA. METHODS: Wastewater samples were collected during 2 months at four key sites in the WWTP. MRSA isolates were characterized using spa typing, antibiograms, SSCmec typing and detection of Panton-Valentine leukocidin (PVL). CONCLUSIONS: MRSA could be isolated on all sampling occasions, but only from inlet and activated sludge. The number of isolates and diversity of MRSA were reduced by the treatment process, but there are indications that the process was selected for strains with more extensive antibiotic resistance and PVL+ strains. The wastewater MRSA-flora had a close genetic relationship to clinical isolates, most likely reflecting carriage in the community. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that MRSA survives in wastewater and that the WWTP may be a potential reservoir for MRSA.

Neuroborreliosis--an epidemiological, clinical and healthcare cost study from an endemic area in the south-east of Sweden.

We studied retrospectively the medical records of all patients (n = 150) diagnosed, by cerebrospinal fluid (CSF) analysis, with neuroborreliosis (NB) in Jönköping County, Sweden during 2000-2005. The number of NB cases increased from 5/100,000 to 10/100,000 inhabitants/year. In 17% of the patients, anti-Borrelia antibodies were found in CSF but not in serum at the time of diagnosis. Facial palsy, headache and fever were frequent manifestations in children, whereas unspecific muscle and joint pain were the most commonly reported symptoms in older patients. Post-treatment symptoms persisting for more than 6 months occurred in 13%, and the patients concerned were significantly older, had longer-lasting symptoms prior to treatment, had higher levels of Borrelia-specific IgG in CSF, and more often had radiculitis. The total cost of NB-related healthcare was estimated to be euro500,000 for the entire study group (euro3300 per patient), and the cost of social benefits was estimated to be euro134,000 (euro2000 per patient). CSF analysis is necessary for the diagnosis of NB, because some patients develop antibodies in serum later than in CSF. Early diagnosis of borreliosis would result in reduced human suffering and in economic gain.

Infection of human endothelial cells with Staphylococcus aureus induces transcription of genes encoding an innate immunity response.

Staphylococcus aureus is a gram-positive bacterium frequently isolated from patients with bloodstream infections. Endothelial cells (EC) play an important role in host defence against bacteria, and recent reports have shown that infection of EC with S. aureus induces expression of cytokines and cell surface receptors involved in activating the innate immune response. The ability of S. aureus to invade nonphagocytic cells, including EC, has been documented. However, the knowledge of the role of EC in pathogenesis of S. aureus infection is still limited. In this study, we investigate the gene-expression program in human EC initiated by internalized S. aureus, using microarray analysis. We found 156 genes that were differentially regulated at least threefold, using arrays representing 14,239 genes. Many of the upregulated genes code for proteins involved in innate immunity, such as cytokines, chemokines and cell adhesion proteins. Other upregulated genes encode proteins involved in antigen presentation, cell signalling and metabolism. Furthermore, intracellular bacteria survived for days without inducing EC death.

Rapid detection of enterohemorrhagic Escherichia coli by real-time PCR with fluorescent hybridization probes.

In this report, we present a PCR protocol for rapid identification of enterohemorrhagic Escherichia coli on a LightCycler instrument. In a multiplex assay, the genes encoding Shiga toxin 1 and Shiga toxin 2 are detected in a single reaction capillary. A complete analysis of up to 32 samples takes about 45 min.