Genome-wide association study of hemolytic uremic syndrome causing Shiga toxin-producing Escherichia coli from Sweden, 1994-2018.

Shiga toxin-producing Escherichia coli (STEC) infection can cause clinical manifestations ranging from diarrhea to potentially fatal hemolytic uremic syndrome (HUS). This study is aimed at identifying STEC genetic factors associated with the development of HUS in Sweden. A total of 238 STEC genomes from STEC-infected patients with and without HUS between 1994 and 2018 in Sweden were included in this study. Serotypes, Shiga toxin gene (stx) subtypes, and virulence genes were characterized in correlation to clinical symptoms (HUS and non-HUS), and pan-genome wide association study was performed. Sixty-five strains belonged to O157:H7, and 173 belonged to non-O157 serotypes. Our study revealed that strains of O157:H7 serotype especially clade 8 were most commonly found in patients with HUS in Sweden. stx2a and stx2a + stx2c subtypes were significantly associated with HUS. Other virulence factors associated with HUS mainly included intimin (eae) and its receptor (tir), adhesion factors, toxins, and secretion system proteins. Pangenome wide-association study identified numbers of accessory genes significantly overrepresented in HUS-STEC strains, including genes encoding outer membrane proteins, transcriptional regulators, phage-related proteins, and numerous genes related to hypothetical proteins. Whole-genome phylogeny and multiple correspondence analysis of pangenomes could not differentiate HUS-STEC from non-HUS-STEC strains. In O157:H7 cluster, strains from HUS patients clustered closely; however, no significant difference in virulence genes was found in O157 strains from patients with and without HUS. These results suggest that STEC strains from different phylogenetic backgrounds may independently acquire genes determining their pathogenicity and confirm that other non-bacterial factors and/or bacteria-host interaction may affect STEC pathogenesis.

Escherichia coli strains producing a novel Shiga toxin 2 subtype circulate in China.

Shiga toxin (Stx) is the key virulence factor in Shiga toxin producing Escherichia coli (STEC), which can cause diarrhea and hemorrhagic colitis with life-threatening complications. Stx comprises two toxin types, Stx1 and Stx2. Several Stx1/Stx2 subtypes have been identified in E. coli, which are variable in sequences, toxicity and host specificity. Here, we report the identification of a novel Stx2 subtype, designated Stx2k, in E. coli strains widely detected from diarrheal patients, animals, and raw meats in China over time. Stx2k exhibits varied cytotoxicity in vitro among individual strains. The Stx2k converting prophages displayed considerable heterogeneity in terms of insertion site, genetic content and structure. Whole genome analysis revealed that the stx2k-containing strains were genetically heterogeneous with diverse serotypes, sequence types, and virulence gene profiles. The nine stx2k-containing strains formed two major phylogenetic clusters closely with strains belonging to STEC, enterotoxigenic E. coli (ETEC), and STEC/ETEC hybrid. One stx2k-containing strain harbored one plasmid-encoded heat-stable enterotoxin sta gene and two identical copies of chromosome-encoded stb gene, exhibiting STEC/ETEC hybrid pathotype. Our finding enlarges the pool of Stx2 subtypes and highlights the extraordinary genomic plasticity of STEC strains. Given the wide distribution of the Stx2k-producing strains in diverse sources and their pathogenic potential, Stx2k should be taken into account in epidemiological surveillance of STEC infections and clinical diagnosis.

Protozoan infections are under-recognized in Swedish patients with gastrointestinal symptoms.

In acute gastroenteritis (GE), identification of the infectious agent is important for patient management and surveillance. The prevalence of GE caused by protozoa may be underestimated in Swedish patients. The purpose was to compare the prevalence of E. histolytica, Cryptosporidium spp., G. intestinalis, and C. cayetanensis in samples from patients where the clinician had requested testing for gastrointestinal parasites only (n = 758) to where testing for bacterial GE only (n = 803) or where both parasite and bacterial testing (n = 1259) was requested and a healthy control group (n = 197). This prospective cohort study was conducted in Region Jönköping County, Sweden (October 2018-March 2019). Fecal samples were analyzed with microscopy and real-time PCR. Cryptosporidium spp. was detected in 16 patients in the bacterial GE group and in 13 in the both bacterial and parasite group; no cases were detected in the group were only parasite infection was suspected. C. cayetanensis was detected in two patients in the bacterial GE group. One case of E. histolytica was detected in the bacterial group and one in the both bacterial and parasite group. G. intestinalis was detected in 14 patients in the parasite only group, 12 in the both parasite and bacterial group, three in the bacterial GE group, and one in the control group. Diarrhea caused by protozoa, especially Cryptosporidium was under-recognized by clinicians and is likely more common than hitherto estimated in Sweden. A more symptom-based diagnostic algorithm may increase detection and knowledge about protozoan infections.

Impact of ribotype on Clostridioides difficile diagnostics.

This study investigates the performance of diagnostic methods for detection of Clostridioides difficile infection in Sweden, including impact of PCR ribotype on diagnostic performance. Between 2011 and 2016, a total of 17,878 stool samples from 26 laboratories were tested by either well-type enzyme immunoassays (EIAs), membrane bound EIAs, cell cytotoxicity neutralization assay (CTA), or nucleic acid amplification tests (NAATs) and subsequently cultured for C. difficile. Roughly half of the samples (9454/17878) were subjected to diagnostic testing both on the fecal sample and on the 1323 isolated C. difficile strains. All C. difficile isolates were typed by PCR ribotyping, and the isolates were classified as toxigenic or non-toxigenic based on the empirical knowledge of the association between toxin-positivity and ribotype. The overall sensitivity, specificity, and positive and negative predictive values were highest for NAATs and membrane EIAs. Ribotype-specific sensitivity varied greatly between methods and ribotypes. All methods had 100% sensitivity against ribotype 027 and 013. For other types, the sensitivity ranged from 33 to 85% in fecal samples and from 78 to 100% on isolates. For the most prevalent ribotypes (014, 020, and 001), the sensitivity varied between 38 and 100% in the fecal samples, with the lowest sensitivity observed for well-type EIAs and CTA. The large variation in diagnostic sensitivity implies that type distribution significantly affects the outcome when evaluating diagnostic performance. Furthermore, performing comparative studies of diagnostic tests in settings with high prevalence of ribotype 027 will overestimate the general performance of diagnostic tests.

Hemolytic uremic syndrome caused by Shiga toxin-producing Escherichia coli in children: incidence, risk factors, and clinical outcome.

BACKGROUND: Hemolytic uremic syndrome (HUS) is a multisystemic disease. In a nationwide study, we characterized the incidence, clinical course, and prognosis of HUS caused by Shiga toxin (Stx)-producing Escherichia coli (STEC) strains with emphasis on risk factors, disease severity, and long-term outcome. METHODS: The data on pediatric HUS patients from 2000 to 2016 were collected from the medical records. STEC isolates from fecal cultures of HUS and non-HUS patients were collected from the same time period and characterized by whole genome sequencing analysis. RESULTS: Fifty-eight out of 262 culture-positive cases developed verified (n = 58, 22%) STEC-HUS. Another 29 cases had probable STEC-HUS, the annual incidence of STEC-HUS being 0.5 per 100,000 children. Eleven different serogroups were detected, O157 being the most common (n = 37, 66%). Age under 3 years (OR 2.4), stx2 (OR 9.7), and stx2a (OR 16.6) were found to be risk factors for HUS. Fifty-five patients (63%) needed dialysis. Twenty-nine patients (33%) developed major neurological symptoms. Complete renal recovery was observed in 57 patients after a median 4.0 years of follow-up. Age under 3 years, leukocyte count over 20 × 109/L, and need for dialysis were predictive factors for poor renal outcome. CONCLUSIONS: Age under 3 years, stx2, and stx2a were risk factors for HUS in STEC-positive children. However, serogroup or stx types did not predict the renal outcome or major CNS symptoms.

Serological diagnostics of Lyme borreliosis: comparison of assays in twelve clinical laboratories in Northern Europe.

Lyme borreliosis (LB), caused by spirochetes belonging to the Borrelia burgdorferi sensu lato complex, is the most common tick-borne infection in Europe. Laboratory diagnosis of LB is mainly based on the patients' medical history, clinical signs and symptoms in combination with detection of Borrelia-specific antibodies where indirect enzyme-linked-immunosorbent assay (ELISA) is the most widely used technique. The objective of the study was to evaluate and compare the diagnostic accuracy (sensitivities and specificities) of serological tests that are currently in use for diagnosis of LB in clinical laboratories in Northern Europe, by use of a large serum panel. The panel consisted of 195 serum samples from well-characterized and classified patients under investigation for clinically suspected LB (n = 59) including patients with Lyme neuroborreliosis, Lyme arthritis, acrodermatitis chronica atrophicans, erythema migrans or other diseases (n = 112). A total of 201 serum samples from healthy blood donors were also included. The panel (396 serum samples altogether) was sent to 12 clinical laboratories (using five different ELISA methods) as blinded for group affiliation and the laboratories were asked to perform serological analysis according to their routine procedure. The results from the study demonstrated high diagnostic concordance between the laboratories using the same diagnostic assay and lower diagnostic concordance between laboratories using different diagnostic assays. For IgG, the results were in general rather homogenous and showed an average sensitivity of 88% (range 85-91%) compared to IgM which showed lower average sensitivity of 59% (range 50-67%) and more heterogeneous results between assays and laboratories.

PDGF-D gene polymorphism is associated with increased cardiovascular mortality in elderly men.

BACKGROUND: Platelet-derived growth factor (PDGF) D has been reported to be active in fibroblasts, and in areas of myocardial infarction. In this longitudinal study we evaluated the association between PDGF-D polymorphism and cardiovascular mortality, and attempted to discover whether specific genotype differences regarding risk could be observed, and if gender differences could be seen. METHODS: Four hundred seventy-six elderly community participants were included in this study. All participants underwent a clinical examination, echocardiography, and blood sampling including PDGF-D single nucleotide polymorphism (SNP) analyses of the rs974819 A/A, G/A and G/G SNP. The follow-up time was 6.7 years. RESULTS: No specific genotype of rs974819 demonstrated increased cardiovascular mortality in the total population, however, the male group with genotypes A/A and G/A demonstrated an increased risk that persisted in a multivariate evaluation where adjustments were made for well-known cardiovascular risk factors (2.7 fold compared with the G/G genotype). No corresponding finding was observed in the female group. CONCLUSION: We report here for the first time that the genotypes G/A or A/A of the SNP rs974819 near PDGF-D exhibited a 2.7 fold increased cardiovascular mortality risk in males. Corresponding increased risk could not be observed in either the total population and thus not in the female group. However, the sample size is was small and the results should be regarded as hypothesis-generating, and thus more research in the field is recommended.

Altered PPARγ Coactivator-1 Alpha Expression in Abdominal Aortic Aneurysm: Possible Effects on Mitochondrial Biogenesis.

INTRODUCTION: Abdominal aortic aneurysm (AAA) is a complex and deadly vascular disorder. The pathogenesis of AAA includes destruction and phenotypic alterations of the vascular smooth muscle cells (VSMCs) and aortic tissues. PPARγ coactivator-1 alpha (PGC1α) regulates VSMC migration and matrix formation and is a major inducer of mitochondrial biogenesis and function, including oxidative metabolism. METHODS: Protein and gene expression of PGC1α and markers for mitochondria biogenesis and cell type-specificity were analysed in AAA aortas from humans and mice and compared against control aortas. RESULTS: Gene expression of PPARGC1A was decreased in human AAA and angiotensin (Ang) II-induced AAA in mice when compared to control vessels. However, high expression of PGC1α was detected in regions of neovascularisation in the adventitia layer. In contrast, the intima/media layer of AAA vessel exhibited defective mitochondrial biogenesis as indicated by low expression of PPARGC1A, VDAC, ATP synthase and citrate synthase. CONCLUSION: Our results suggest that mitochondrial biogenesis is impaired in AAA in synthetic SMCs in the media, with the exception of newly formed supporting vessels in the adventitia where the mitochondrial markers seem to be intact. To our knowledge, this is the first study investigating PGC1α and mitochondria biogenesis in AAA.

Prognostic significance of PLA2G4C gene polymorphism in patients with stage II colorectal cancer.

BACKGROUND: Phospholipase A2 Group IV C (PLA2G4C) catalyzes the release of certain fatty acids from phospholipids and plays a role in a range of physiological functions, such as remodeling of cell membranes and the production of prostaglandins. Furthermore, it has been proposed that PLA2G4C plays an important role in breast cancer cell chemotaxis. This study aimed to investigate the effect of a single nucleotide polymorphism (SNP) rs1549637 (T>A) of the PLA2G4C gene on the prognosis of colorectal cancer (CRC). MATERIAL AND METHODS: Whole blood DNA was extracted from 381 patients with CRC and 618 controls, and a TaqMan SNP genotyping assay was used to determine the distribution of the genotypes. Cancer-specific and disease-free survival was analyzed by Kaplan-Meier graphs and by uni- and multivariable Cox regression. RESULTS: The cancer-specific survival differed between the genotypes (p = 0.019) and the carriers of the A allele were associated with the highest risk of CRC death, with a hazard ratio (HR) of 1.72 [95% confidence interval (CI) 1.17-2.53, p = 0.006] compared with homozygous carriers of the T allele. This increased mortality in the carriers with the allele A was especially marked in stage II with an HR of 3.84 (95% CI 1.51-9.78, p = 0.005). CONCLUSION: The A allele in PLA2G4C SNP (rs1549637) is associated with a worse prognosis in patients with CRC, especially in stage II disease, and it could be a potential prognostic biomarker in the planning of individual adjuvant therapy in stage II patients.

Norovirus Dynamics in Wastewater Discharges and in the Recipient Drinking Water Source: Long-Term Monitoring and Hydrodynamic Modeling.

Norovirus (NoV) that enters drinking water sources with wastewater discharges is a common cause of waterborne outbreaks. The impact of wastewater treatment plants (WWTPs) on the river Göta älv (Sweden) was studied using monitoring and hydrodynamic modeling. The concentrations of NoV genogroups (GG) I and II in samples collected at WWTPs and drinking water intakes (source water) during one year were quantified using duplex real-time reverse-transcription polymerase chain reaction. The mean (standard deviation) NoV GGI and GGII genome concentrations were 6.2 (1.4) and 6.8 (1.8) in incoming wastewater and 5.3 (1.4) and 5.9 (1.4) log10 genome equivalents (g.e.) L-1 in treated wastewater, respectively. The reduction at the WWTPs varied between 0.4 and 1.1 log10 units. In source water, the concentration ranged from below the detection limit to 3.8 log10 g.e. L-1. NoV GGII was detected in both wastewater and source water more frequently during the cold than the warm period of the year. The spread of NoV in the river was simulated using a three-dimensional hydrodynamic model. The modeling results indicated that the NoV GGI and GGII genome concentrations in source water may occasionally be up to 2.8 and 1.9 log10 units higher, respectively, than the concentrations measured during the monitoring project.

Detection of Anaplasma phagocytophilum in Ixodes ricinus ticks from Norway using a realtime PCR assay targeting the Anaplasma citrate synthase gene gltA.

BACKGROUND: A TaqMan real-time PCR assay targeting the Anaplasma citrate synthase gene, gltA, was developed and used for detection of Anaplasma phagocytophilum in 765 Ixodes ricinus ticks collected from dogs and cats in northern Norway (n = 669) and Telemark county in southern Norway (n = 96). RESULTS: Among the ticks from northern Norway the prevalence of A. phagocytophilum was 3.0 %, while the prevalence in southern Norway was 2.1 % (p = 0.63). The gltA PCR assay showed a high analytical sensitivity (30 genomic units) and efficiency (98.5 %), and its utility in clinical diagnostics should be evaluated in future studies. CONCLUSION: This is the first report of A. phagocytophilum occurrence in ticks collected north of the Arctic Circle in Norway. The prevalence is comparable to that found in Telemark county in southern Norway.

CD93 gene polymorphism is associated with disseminated colorectal cancer.

PURPOSE: Cluster of differentiation 93 (CD93) is involved in apoptosis and inflammation and has a suggested role in angiogenesis, and all of which are involved in the development and dissemination of cancer. We evaluated the expression of CD93 and the association with two single nucleotide polymorphisms (SNPs), rs2749812 and rs2749817, as possible biomarkers in colorectal cancer (CRC). METHODS: Tissue levels and plasma levels of CD93 were measured using an enzyme-linked immunosorbent assay (ELISA). Expression of CD93 was determined by immunohistochemistry, western blot and gene expression analysis. Genotype frequencies were established for the SNPs by real-time polymerase chain reaction (PCR), and the association with tumour stage and survival was analysed. RESULTS: Total CD93 levels were 82% higher (P < 0.001) in tumours compared to matched normal tissues. Mean levels of soluble CD93 in plasma were 30% lower (P < 0.001) in the patients compared to the controls. The T/T genotype of SNP rs2749817 was more common in stage IV patients, with consequently higher risk of CRC death (T/T vs. C/C and C/T; hazard ratio (HR) = 1.73, 95% confidence interval (CI) = 1.11-2.67, P = 0.014), and was associated with a higher risk of CRC recurrence after radical operation (T/T vs. C/C and C/T; HR = 2.07, CI = 1.22-3.51, P = 0.007). CONCLUSIONS: We showed that the T/T genotype of SNP rs2749817 is associated with disseminated cancer at diagnosis and an increased recurrence rate after radical operation. Patients with this genotype may benefit from early identification.

Increased expression of the Wnt signalling inhibitor Dkk-1 in non-lesional skin and peripheral blood mononuclear cells of patients with plaque psoriasis.

Psoriasis is a common inflammatory skin disease characterised by abnormal keratinocyte proliferation, increased dermal angiogenesis and systemic inflammation. The cell signalling cascades provoked by Wnt proteins and their inhibitors, such as Dickkopf-1, play crucial roles to maintain homeostasis of a variety of tissues, including skin, and are also involved in angiogenesis and innate immunity. This study was designed to investigate the distribution of Dickkopf-1, in lesional and non-lesional skin, in serum and in peripheral blood mononuclear cell (PBMCs) of patients with psoriasis compared with healthy controls. Our results showed significantly increased mRNA and protein expression of Dickkopf-1 in non-lesional compared with lesional skin and healthy control skin. No significant differences of Dickkopf-1 serum levels were observed, but Dickkopf-1 protein expression was significantly increased in patients' PBMC. Increased levels of Dickkopf-1 in PBMC, suggest a possible role of Dickkopf-1 in the chronic systemic inflammation of psoriasis. Increased levels of Dickkopf-1 in non-lesional psoriasis skin offers new insights in the local inflammatory processes in psoriasis skin since Wnt signalling regulates angiogenesis. In conclusion, Dickkopf-1 may be a possible target for future treatment options.

Prevalence and Characteristics of Plasmid-Encoded Serine Protease EspP in Clinical Shiga Toxin-Producing Escherichia coli Strains from Patients in Sweden.

Shiga toxin-producing Escherichia coli (STEC) infection can cause a broad spectrum of symptoms spanning from asymptomatic shedding to mild and bloody diarrhea (BD) and even life-threatening hemolytic-uremic syndrome (HUS). As a member of the serine protease autotransporters of Enterobacteriaceae (SPATE) family, EspP has the ability to degrade human coagulation factor V, leading to mucosal bleeding, and also plays a role in bacteria adhesion to the surface of host cells. Here, we investigated the prevalence and genetic diversity of espP among clinical STEC isolates from patients with mild diarrhea, BD, and HUS, as well as from asymptomatic individuals, and assessed the presence of espP and its subtypes in correlation to disease severity. We found that 130 out of 239 (54.4%) clinical STEC strains were espP positive, and the presence of espP was significantly associated with BD, HUS, and O157:H7 serotype. Eighteen unique espP genotypes (GTs) were identified and categorized into four espP subtypes, i.e., espPα (119, 91.5%), espPγ (5, 3.8%), espPδ (4, 3.1%), and espPε (2, 1.5%). espPα was widely distributed, especially in strains from patients with BD and HUS, and correlated with serotype O157:H7. Serogroup O26, O145, O121, and O103 strains carried espPα only. Ten GTs were identified in espPα, and espPα/GT2 was significantly associated with severe disease, i.e., BD and HUS. Additionally, espP was strongly linked to the presence of eae gene, and the coexistence of espPα and stx2/stx2a + stx2c was closely related to HUS status. To sum up, our data demonstrated a high prevalence and genetic diversity of the espP gene in clinical STEC strains in Sweden and revealed an association between the presence of espP, espP subtypes, and disease severity. espP, particularly the espPα subtype, was prone to be present in more virulent STEC strains, e.g., "top-six" serotypes strains.

Macrophages at the fetal-maternal interface express markers of alternative activation and are induced by M-CSF and IL-10.

During pregnancy, the maternal immune system is challenged by the presence of the fetus, which must be tolerated despite being semiallogeneic. Uterine mucosal (or decidual) macrophages (M), one of the major leukocyte populations at the fetal-maternal interface, have been implicated in fetal tolerance, but information regarding their regulation is scarce. In this study, we investigated the role of several factors potentially involved in the differentiation and polarization of decidual M with an in vitro M differentiation model. By using flow cytometry, we showed that M-CSF and IL-10 were potent inducers of M2 (immunoregulatory) M markers expressed on human decidual M (CD14, CD163, CD206, CD209). In contrast, proinflammatory stimuli, and unexpectedly also the Th2-associated IL-4 and IL-13, induced different patterns of expression, indicating that a Th2-dominated environment is not required for decidual M polarization. M-CSF/IL-10-stimulated and decidual M also showed similar cytokine secretion patterns, with production of IL-10 as well as IL-6, TNF, and CCL4. Conversely, the proinflammatory, LPS/IFN-γ-stimulated M produced significantly higher levels of TNF and no IL-10. We also used a gene array with 420 M-related genes, of which 100 were previously reported to be regulated in a global gene expression profiling of decidual M, confirming that M-CSF/IL-10-induced M are closely related to decidual M. Taken together, our results consistently point to a central role for M-CSF and in particular IL-10 in the shaping of decidual M with regulatory properties. These cytokines may therefore play an important role in supporting the homeostatic and tolerant immune milieu required for a successful pregnancy.

Low level of antimicrobial resistance in Escherichia coli among Swedish nursing home residents.

BACKGROUND: Screening for bacterial colonization and antimicrobial resistance (AMR) among a defined population could aid in the identification of at-risk populations and provide targets for antibiotic stewardship and infection control programmes. METHODS: Two hundred and sixty-eight participants at 11 Swedish nursing homes underwent serial screening for colonization with Escherichia coli between March 2008 and September 2010. Seventy-two of the 268 participants (27%) were male. The median age was 85 y. Samples were collected from urine, the rectal mucosa, the groin, and active skin lesions. RESULTS: Two hundred and nine of 268 participants (78%) were colonized with E. coli at any body site/fluid. The specific colonization rates were 81% (rectum), 48% (urine), 30% (groin), 59% (unknown), and 13% (skin lesion). An antibiotic-resistant E. coli isolate was identified in 18% of all participants regardless of colonization status; all together, 87 resistant isolates were detected. Only 1 participant carried isolates with resistance to third-generation cephalosporins (cefotaxime and ceftazidime). CONCLUSIONS: The presence of resistance was generally low, and the greater part of the resistant cases was connected with 3 common antibiotics: ampicillin, trimethoprim/sulfamethoxazole, and ciprofloxacin. In spite of generally increasing resistance against third-generation cephalosporins in E. coli in Sweden, this study does not implicate residence at a Swedish nursing home as a risk factor for the acquisition of expressed cephalosporin resistance.

Genetic Characterization of Intimin Gene (eae) in Clinical Shiga Toxin-Producing Escherichia coli Strains from Pediatric Patients in Finland.

Shiga toxin (Stx)-producing Escherichia coli (STEC) infections cause outbreaks of severe disease in children ranging from bloody diarrhea to hemolytic uremic syndrome (HUS). The adherent factor intimin, encoded by eae, can facilitate the colonization process of strains and is frequently associated with severe disease. The purpose of this study was to examine and analyze the prevalence and polymorphisms of eae in clinical STEC strains from pediatric patients under 17 years old with and without HUS, and to assess the pathogenic risk of different eae subtypes. We studied 240 STEC strains isolated from pediatric patients in Finland with whole genome sequencing. The gene eae was present in 209 (87.1%) strains, among which 49 (23.4%) were from patients with HUS, and 160 (76.6%) were from patients without HUS. O157:H7 (126, 60.3%) was the most predominant serotype among eae-positive STEC strains. Twenty-three different eae genotypes were identified, which were categorized into five eae subtypes, i.e., γ1, ß3, ε1, θ and ζ3. The subtype eae-γ1 was significantly overrepresented in strains from patients aged 5-17 years, while ß3 and ε1 were more commonly found in strains from patients under 5 years. All O157:H7 strains carried eae-γ1; among non-O157 strains, strains of each serotype harbored one eae subtype. No association was observed between the presence of eae/its subtypes and HUS. However, the combination of eae-γ1+stx2a was significantly associated with HUS. In conclusion, this study demonstrated a high occurrence and genetic variety of eae in clinical STEC from pediatric patients under 17 years old in Finland, and that eae is not essential for STEC-associated HUS. However, the combination of certain eae subtypes with stx subtypes, i.e., eae-γ1+stx2a, may be used as risk predictors for the development of severe disease in children.

The Local Tumor Microbiome Is Associated with Survival in Late-Stage Colorectal Cancer Patients.

The gut microbiome is associated with survival in colorectal cancer. Single organisms have been identified as markers of poor prognosis. However, in situ imaging of tumors demonstrate a polymicrobial tumor-associated community. To understand the role of these polymicrobial communities in survival, we conducted a nested case-control study in late-stage cancer patients undergoing resection for primary adenocarcinoma. The microbiome of paired tumor and adjacent normal tissue samples was profiled using 16S rRNA sequencing. We found a consistent difference in the microbiome between paired tumor and adjacent tissue, despite strong individual microbial identities. Furthermore, a larger difference between normal and tumor tissue was associated with prognosis: patients with shorter survival had a larger difference between normal and tumor tissue. Within the tumor tissue, we identified a 39-member community statistic associated with survival; for every log2-fold increase in this value, an individual's odds of survival increased by 20% (odds ratio survival 1.20; 95% confidence interval = 1.04 to 1.33). Our results suggest that a polymicrobial tumor-specific microbiome is associated with survival in late-stage colorectal cancer patients. IMPORTANCE Microbiome studies in colorectal cancer (CRC) have primarily focused on the role of single organisms in cancer progression. Recent work has identified specific organisms throughout the intestinal tract, which may affect survival; however, the results are inconsistent. We found differences between the tumor microbiome and the microbiome of the rest of the intestine in patients, and the magnitude of this difference was associated with survival, or, the more like a healthy gut a tumor looked, the better a patient's prognosis. Our results suggest that future microbiome-based interventions to affect survival in CRC will need to target the tumor community.

DNA promoter methylation status and protein expression of interleukin-8 in human colorectal adenocarcinomas.

BACKGROUND: Interleukin-8 (IL-8) also referred to as CXCL8, a member of the CXC chemokine family that attracts neutrophils and other leukocytes, has been associated with cancer. Angiogenesis is a prime regulator of tumour expansion and data support that IL-8 is a potent angiogenic factor. Epigenomic instability has been postulated to play a role for the development of multiple neoplasias including colorectal cancer (CRC). DNA methylation of cytosine residues in CpG dinucleotides leads to transcriptional silencing of associated genes. METHOD: In this study, we comparatively analysed the protein expression of IL-8 in plasma, tumour and paired normal tissue and methylation status of the IL-8 gene to evaluate its impact on CRC. RESULTS: Collectively, by using Luminex technology, we noted a significantly higher IL-8 level in cancer tissue compared to paired normal tissue and that CRC patients exhibit significantly higher plasma levels than healthy controls. Analysed by methylation-specific polymerase chain reaction, we detected IL-8 hypomethylation in 64% of the cancerous tissue cases but no hypomethylation was found in paired normal tissue. We noted that the CRC patients with IL-8 hypomethylation revealed a significant higher level of IL-8 protein in cancerous tissue, which tended to be associated with distant metastasis. We also observed that patients with distant metastasis showed a significantly higher plasma level of IL-8 in relation to patients without distant metastasis. CONCLUSION: Our results suggest that the predominance of high plasma levels of IL-8 in patients with distant metastasis in combination with the hypomethylation of the IL-8 promoter region might be a useful marker of the disease advancement.

Borrelia burgdorferi sensu lato in Ixodes ricinus ticks from Norway: evaluation of a PCR test targeting the chromosomal flaB gene.

A consensus TaqMan real-time PCR test targeting the chromosomal flaB gene of Borrelia burgdorferi sensu lato was constructed. The test was compared with a recently published generic Light Upon eXtension (LUX) 16S rRNA real-time PCR test (Wilhelmsson et al. in J Clin Microbiol 48:4169-4176, 2010) on material consisting of 242 Ixodes ricinus ticks collected from dogs and cats in Northern Norway (n = 139) and Telemark County in Southern Norway (n = 103). Ticks positive in either test were further tested by nested PCR amplification of the 5S-23S rRNA intergenic-spacer region followed by sequencing for species identification. A tick was defined as Borrelia positive if two of three tests were positive. Thirty-four of the 242 (14 %) ticks satisfied this definition of positivity. Of these ticks 32 were positive both in the rRNA and flaB test, while two were positive only in the rRNA test. One tick was positive only in the rRNA test and was considered false positive since PCR for sequencing failed. The sensitivity of the flaB test was 94 % and the specificity 100 %. It was possible to determine the species present using Tm analysis. Among ticks from Northern Norway the prevalence of Borrelia was 13 %, whereas the prevalence in Telemark was 16 %. Among identified species (n = 33) B. afzelii was found in 16 (47 %), B. garinii in 15 (44 %) and B. valaisiana in 2 (6 %) ticks, respectively. The flaB test is a rapid, sensitive and specific test for detection and quantification of Borrelia burgdorferi s.l. in I. ricinus ticks. This is the first report on Borrelia prevalence in I. ricinus in Northern Norway.