Clinical features of gastroenteritis during a large waterborne Campylobacter outbreak in Askøy, Norway.

PURPOSE: Outbreaks of Campylobacter infection are common, but studies exploring the clinical features of acute illness in the outbreak setting are scarce in existing literature. The main purpose of the present study was to investigate the clinical features of self-reported acute illness in gastroenteritis cases during a large waterborne Campylobacter outbreak in Askøy municipality, Norway, in 2019. METHODS: A web-based self-administered questionnaire, and invitation to participate was sent by the municipality of Askøy as text message to mobile phones using the municipality's warning system to the inhabitants during the ongoing outbreak. RESULTS: Out of 3624 participants, 749 (20.7%) were defined as cases, of which 177 (23.6%) reported severe gastroenteritis. The most common symptoms were loose stools (90.7%), abdominal pain (89.3%) and diarrhea (88.9%), whereas 63.8% reported fever, 50.2% joint pain and 14.2% bloody stools. Tiredness, a symptom non-specific to gastroenteritis, was the overall most common symptom (91.2%). CONCLUSION: About one in four of the cases reported symptoms consistent with severe gastroenteritis. We found more joint pain and less bloody stools than reported in published studies of laboratory confirmed campylobacteriosis cases. Tiredness was common in the current study, although rarely described in previous literature of acute illness in the outbreak setting.

An observational study of community-acquired severe sepsis comparing intensive care and non-intensive care patients.

BACKGROUND: Most studies of sepsis are from intensive care units (ICUs). We aimed to investigate community-acquired severe sepsis in a broader population, in order to compare patients treated in or outside an ICU . METHODS: We performed a 1-year prospective observational study with enrollment of patients from three units; a general ICU, a combined ICU/non-ICU and a medical ward with limited surveillance facilities. Hospital survivors were followed up for 5 years. RESULTS: Overall, 220 patients were included, of which 107 received ICU treatment. The majority of abdominal (77%, P = 0.003) and genitourinary (81%, P < 0.001) infections were found in ICU and non-ICU patients, respectively. Time to first antibiotic administration was longer in ICU-patients (median 3.5 vs. 2.0 h in non-ICU patients, P = 0.011). ICU developed more organ dysfunctions than non-ICU patients (P < 0.001), nevertheless supportive therapy with vasoactive drugs and non-invasive ventilation was documented in 22% and 27% of the latter. Median hospital length of stay was 15 vs. 9 days (P = 0.001), and hospital and 5-year mortality rates 35% vs. 16% (P = 0.002) and 57% vs. 58% (P = 0.892) among ICU and non-ICU patients, respectively. Increasing age (HR 1.06 (1.04, 1.07) per year, P < 0.001), not care level during hospitalization (HR 1.19 (0.70, 2.02), P = 0.514), influenced long-term survival. CONCLUSION: Half of the subjects with community-acquired severe sepsis never received ICU treatment. Still, use of organ supportive therapy outside the ICU was considerable. Hospital mortality was higher, whereas 5-year survival was similar when comparing ICU with non-ICU patients.

Sequence diversity of sicG among group C and G Streptococcus dysgalactiae subspecies equisimilis isolates associated with human infections in western Norway.

Streptococcal inhibitor of complement (SIC) and distantly related to SIC (DRS) are well-characterized extracellular virulence factors produced by only a few emm types among group A streptococci. The prevalence and sequence variations of the sic-like gene (sicG) in clinical samples of group C and G Streptococcus dysgalactiae subspecies equisimilis (SDSE), however, have not been widely investigated. We constructed primers targeting sicG and screened 129 geographically matched and previously emm-typed non-invasive (n = 64) and invasive (n = 65) SDSE isolates for the presence of this gene. sicG was detected in seven non-invasive and eight invasive isolates belonging to eight different emm types. Within five of these emm types, sicG-negative isolates were also detected. All three isolates of stG2078.0 possessed sicG and were associated with severe soft tissue infections. Altogether, six sicG alleles (sicG1-6) were identified, and sequence variations were mainly caused by single nucleotide polymorphisms and deletion/insertion mutations. sicG1-6 were predicted to encode SICG proteins of varying length, composition, and homology with SIC and DRS proteins of group A streptococci. Our findings indicate an unpredictable association between sicG and emm type, a limited distribution and substantial sequence diversity of sicG, and no obvious relation between its presence and disease severity.

Early quantification of HCV core antigen may help to determine the duration of therapy for chronic genotype 2 or 3 HCV infection.

The aim of the present study was to evaluate the utility of hepatitis C virus (HCV) core antigen (coreAg) assessment for the identification of candidates for short-term therapy. Plasma samples from HCV genotype 2 or 3-infected patients participating in the NORDynamIC trial (n=382) comparing 12 and 24 weeks of combination treatment with pegylated interferon-α2a and a fixed dose of 800 mg ribavirin daily were analyzed for coreAg. Among the 126 patients (33% of the intention-to-treat population) achieving HCV coreAg levels in plasma below 0.2 pg/mL when assayed on treatment day 3, sustained viral response (SVR) rates of 86% and 84% were achieved in the 12- and 24-week arms, respectively. Similarly, among patients having received at least 80% of the target dose of both pegylated interferon α-2a and of ribavirin for at least 80% of the target treatment duration (per-protocol analysis), the SVR rates were 89% and 95%, respectively. Twelve weeks of combination treatment may be sufficient for genotype 2 or 3-infected patients achieving HCV coreAg levels below 0.2 pg/mL by day 3, signaling a rapid clearance of HCV viremia.

Hepatitis C treatment response kinetics and impact of baseline predictors.

The optimal duration of treatment for hepatitis C virus (HCV) infections is highly variable but critical for achieving cure (sustained virological response, SVR). We prospectively investigated the impact of age, fibrosis, baseline viraemia and genotype on the early viral kinetics and treatment outcome. Patients treated with peginterferon alfa-2a and ribavirin in standard dosing were included: 49 with genotype 1 treated for 48weeks and 139 with genotype 2 or 3 treated for 24weeks. The reduced SVR rates in patients older than 45years, with severe liver fibrosis or pretreatment viraemia above 400,000IU/mL were strongly associated with slower second phase declines of HCV RNA. Genotype 2/3 infections responded more rapidly than genotype 1, reaching week 4 negativity (RVR) in 59%vs 22%. We conclude that baseline response predictors such as age, fibrosis and viral load were well reflected by the early viral kinetics as assessed by repeated HCV RNA quantifications. The kinetic patterns and the high relapse rate in genotype 2/3 patients without RVR suggest that this group might benefit from treatment durations longer than 24weeks.

Ribavirin plasma concentration is a predictor of sustained virological response in patients treated for chronic hepatitis C virus genotype 2/3 infection.

In hepatitis C virus (HCV) genotype 1 infection, the likelihood of obtaining sustained virological response (SVR) is associated with higher ribavirin exposure. Such an association has not been demonstrated for HCV genotype 2/3 infection, where a fixed 800 mg daily dosing of ribavirin is generally recommended. The primary aim of this study was to investigate the correlation between ribavirin concentration at day 29 and therapeutic response in patients with HCV genotype 2/3 infection. A total of 382 patients were randomized to 12 or 24 weeks of treatment with pegylated interferon-alfa 2a 180 µg weekly and 800 mg ribavirin daily. Trough plasma concentration of ribavirin was measured at day 29 and week 12 and the primary outcome was SVR (HCV-RNA undetectable 24 weeks after treatment). Of the 382 patients, 355 had a ribavirin concentration available at day 29. SVR was 84% among patients with a ribavirin concentration ≥2 mg/L at day 29 compared to 66% in those with concentrations <2 mg/L (P = 0.002). The corresponding figures in the 12-week treatment group were 74% and 57% (P = 0.12), and in the 24-week treatment group 91% and 75% (P = 0.02), respectively. In a multivariate analysis, ribavirin concentration at day 29 was an independent predictor of SVR (P = 0.002). In conclusion, a higher plasma ribavirin concentration is associated with an increased likelihood of achieving SVR in HCV genotype 2/3 infection. Individualization of ribavirin dosing may be helpful in improving outcome, especially in the presence of unfavourable baseline characteristics. This, however, requires evaluation in a prospective trial.

emm gene diversity, superantigen gene profiles and presence of SlaA among clinical isolates of group A, C and G streptococci from western Norway.

In order to investigate molecular characteristics of beta-hemolytic streptococcal isolates from western Norway, we analysed the entire emm gene sequences, obtained superantigen gene profiles and determined the prevalence of the gene encoding streptococcal phospholipase A2 (SlaA) of 165 non-invasive and 34 contemporary invasive group A, C and G streptococci (GAS, GCS and GGS). Among the 25 GAS and 26 GCS/GGS emm subtypes identified, only emm3.1 was significantly associated with invasive disease. M protein size variation within GAS and GCS/GGS emm types was frequently identified. Two non-invasive and one invasive GGS possessed emm genes that translated to truncated M proteins as a result of frameshift mutations. Results suggestive of recombinations between emm or emm-like gene segments were found in isolates of emm4 and stG485 types. One non-invasive GGS possessed speC, speG, speH, speI and smeZ, and another non-invasive GGS harboured SlaA. speA and SlaA were over-represented among invasive GAS, probably because they were associated with emm3. speG(dys) was identified in 83% of invasive and 63% of non-invasive GCS/GGS and correlated with certain emm subtypes. Our results indicate the invasive potential of isolates belonging to emm3, and show substantial emm gene diversity and possible lateral gene transfers in our streptococcal population.

Postinfectious and sporadic functional gastrointestinal disorders have different prevalences and rates of overlap: results from a controlled cohort study 3 years after acute giardiasis.

BACKGROUND: Irritable bowel syndrome (IBS) is a common complication following gastroenteritis, and a high prevalence of postgiardiasis IBS has previously been reported. This study aims to investigate the prevalence, adjusted relative risk (RRadj), and overlap of different functional gastrointestinal disorders (FGID) according to Rome III criteria following infection with Giardia lamblia. METHODS: All patients ≥18 years of age with verified giardiasis during an outbreak in 2004, and a control group matched by age and gender, were mailed a questionnaire 3 years later. KEY RESULTS: The prevalence of functional dyspepsia (FD) was 25.9% in the exposed and 6.9% in the control group, RRadj: 3.9 (95% confidence intervals [CI]: 3.1-4.8). The prevalence of IBS was 47.9% and 14.3%, respectively, with RRadj: 3.4 (95% CI: 3.0-3.8). Prevalence of other gastrointestinal symptoms ranged from 70.0% vs 39.7% for bloating (RRadj: 1.8) to 8.3% vs 2.9% for nausea (RRadj: 3.0) in the Giardia and the control group, respectively. Among individuals fulfilling criteria for IBS 44% in the exposed group and 29% in the control group also fulfilled criteria for FD. IBS subtypes based on Rome III criteria (stool consistency) showed poor agreement with subtypes based on frequency of bowel movements (Kappa-values: 0.17 and 0.27). CONCLUSIONS & INFERENCES: There were high prevalences and RRs of IBS, FD and other gastrointestinal symptoms following acute giardiasis, and a high degree of overlap between the disorders. The agreement between different IBS subtype criteria varied, and there were also differences between the exposed and control group.

Salivary IgA from the sublingual compartment as a novel noninvasive proxy for intestinal immune induction.

Whole-saliva IgA appears like an attractive noninvasive readout for intestinal immune induction after enteric infection or vaccination, but has failed to show consistent correlation with established invasive markers and IgA in feces or intestinal lavage. For reference, we measured antibodies in samples from 30 healthy volunteers who were orally infected with wild-type enterotoxigenic Escherichia coli. The response against these bacteria in serum, lavage, and lymphocyte supernatants (antibody-in-lymphocyte-supernatant, ALS) was compared with that in targeted parotid and sublingual/submandibular secretions. Strong correlation occurred between IgA antibody levels against the challenge bacteria in sublingual/submandibular secretions and in lavage (r=0.69, P<0.0001) and ALS (r=0.70, P<0.0001). In sublingual/submandibular secretions, 93% responded with more than a twofold increase in IgA antibodies against the challenge strain, whereas the corresponding response in parotid secretions was only 67% (P=0.039). With >twofold ALS as a reference, the sensitivity of a >twofold response for IgA in sublingual/submandibular secretion was 96%, whereas it was only 67% in the parotid fluid. To exclude that flow rate variations influenced the results, we used albumin as a marker. Our data suggested that IgA in sublingual/submandibular secretions, rather than whole saliva with its variable content of parotid fluid, is a preferential noninvasive proxy for intestinal immune induction.

Neomycin inhibits PDGF-induced IP3 formation and DNA synthesis but not PDGF-stimulated uptake of inorganic phosphate in C3H/10T1/2 fibroblasts.

Porcine PDGF was found to increase [3H]inositol trisphosphate, [3H]thymidine incorporation and 32P-labelling of polyphosphoinositides in C3H/10T1/2 Cl 8 fibroblasts. These responses to PDGF stimulation were all inhibited by 5 mM neomycin, a polycationic aminoglycoside formerly known to inhibit polyphosphoinositide turnover. PDGF also markedly increased the cellular uptake of inorganic [32P]Pi. This response of PDGF was not inhibited by neomycin (5 mM). Thus, neomycin inhibited PDGF-induced IP3 formation, 32P-labelling of polyphosphoinositides and DNA synthesis, but not cellular uptake of inorganic phosphate. These effects of neomycin suggest a bifurcation of the initial part of the PDGF-induced signal transduction, separating at the receptor level or before phospholipase C activation.

A monoclonal antibody against PDGF B-chain inhibits PDGF-induced DNA synthesis in C3H fibroblasts and prevents binding of PDGF to its receptor.

A monoclonal antibody (MAb 6D11) against platelet-derived growth factor (PDGF) was studied. We found that the MAb 6D11 in concentrations equimolar to PDGF blocked the [3H]thymidine incorporation in C3H/10T1/2 C18 fibroblasts stimulated by PDGF B-B and PDGF A-B. This inhibition was overcome by high doses of PDGF. The [3H]thymidine incorporation stimulated by other growth factors (aFGF, bFGF and bombesin) was not inhibited by the antibody. The MAb 6D11 blocked receptor binding of PDGF B-B, but not PDGF A-A. These findings suggest that the MAb 6D11 abolishes PDGF-induced DNA synthesis by blocking PDGF receptor binding. In this communication we demonstrate an isoform-specific monoclonal antibody against PDGF.

Whole genome sequencing of clinical isolates of Giardia lamblia.

Clinical isolates from protozoan parasites such as Giardia lamblia are at present practically impossible to culture. By using simple cyst purification methods, we show that Giardia whole genome sequencing of clinical stool samples is possible. Immunomagnetic separation after sucrose gradient flotation gave superior results compared to sucrose gradient flotation alone. The method enables detailed analysis of a wide range of genes of interest for genotyping, virulence and drug resistance.

Reduction of HSV-1 binding to BHK cells after treatment with phosphatidylinositol-specific phospholipase C.

Baby-hamster kidney cells were treated with unspecific and phosphatidylinositol-specific phospholipase C (PI-PLC) prior to infection with herpes simplex virus type 1 or 2. Subsequent to PI-PLC treatment, a 30% reduction of infectivity and receptor binding was observed for type 1 virus, while type 2 was unaffected. Treating the cells with unspecific phospholipase C did not affect subsequent infection with either virus. Treatment of the virus particles with the unspecific phospholipase reduced its infectivity, probably due to loss of the viral envelope. PI-PLC treatment of virus particles did not have any such effect on virus infectivity.

Human interferon reduces surface expression but not total production of herpes simplex virus type 1 glycoproteins gC and gE in heterologous hamster cells.

The effect of interferon (IFN) on herpes simplex virus type 1 (HSV-1)-induced glycoproteins gC and gE was investigated in a heterologous IFN/cell model. In this model, the effect on surface expression of the glycoproteins could be studied separately from the effect on virus multiplication. Pretreatment of baby hamster kidney cells (BHK) with heterologous human leukocyte IFN suppressed surface expression of HSV-1-encoded gC and gE but had no influence on total production of the glycoproteins. This was in contrast to the effect on human embryonic fibroblast cells (HE) (homologous IFN and cells), where surface expression as well as total production of glycoproteins were reduced. The surface expression was demonstrated by antibody-sensitized monodisperse polystyrene beads, and immunoblotting and two-dimensional electrophoretic analysis of radioisotope-labeled proteins were used to study the total production.

Two-dimensional gel analysis of [35S]methionine labelled and phosphorylated proteins present in virions and light particles of herpes simplex virus type 1, and detection of potentially new structural proteins.

Cells infected with herpes simplex virus (HSV) synthesize both infectious viruses and non-infectious light particles (L-particles). The latter contain the envelope and tegument components of the virions, but lack virus capsid and DNA. Electrophoresis in SDS-polyacrylamide gels (SDS-PAGE) has been used extensively for analysis of structural proteins in virions and L-particles. Two-dimensional (2-D) gel electrophoresis, however has a markedly higher resolution, and in the present work we have used this technique to study both [35S]methionine labelled and phosphorylated structural proteins in virions and L-particles. Proteins were assigned to the tegument or the envelope by the analysis of L-particles. Localization of structural proteins was also determined by stepwise solubilization in the presence of the neutral detergent NP-40 and NaCl, and by isolation of capsids from nuclei of infected cells. Different steps in posttranslational modification can be detected by 2-D gel electrophoresis such that a single polypeptide may appear as several spots. This was most clearly observed for some of the HSV-encoded glycoproteins which were shown to exist in multiple forms in the virion. Some polypeptides apparently not identified previously were either capsid associated, or localized in the tegument or envelope. The degrees of phosphorylation in L-particles and virions are almost identical for some proteins, but markedly different for others. Thus, glycoprotein E of HSV-1 is for the first time shown to be phosphorylated, and most heavily so in virions. The IE VMW)110 protein represents a group of proteins which are more phosphorylated in L-particles than in virions. Attempts are made to correlate the proteins detected by 2-D analysis with those previously separated by SDS-PAGE.

Labeling of surface proteins of herpes simplex virus type 1 using a modified biotin-streptavidin system.

Methods of labeling surface proteins on herpes simplex virus (HSV) which have minimal effect on the biological activity of the virus are useful for the study of both the localization and function(s) of surface proteins. The present work describes a procedure using a water-soluble biotin compound, sulfo-NHS-biotin, which is unable to penetrate biological membranes and reacts with primary amines in proteins. Labeled proteins were detected by binding of [125I]streptavidin. Specific reaction with surface proteins was shown in Western blots using antibodies against selected proteins in the envelope or in the tegument. Proteins susceptible to iodination were also biotinylated, but the efficiency of labeling varied from one protein to another. As a result of freezing and thawing of the virus, as well as the manipulations involved in Ficoll gradient purification, internal proteins were labeled. The infectivity of the virus was reduced by approximately 40% after biotinylation. Labeled viruses were visualized by fluorescein isothiocyanate-conjugated streptavidin, and seen as distinct spots on the surface of the cells.

Outbreak of ampicillin-resistant Enterococcus faecium--risk factors for faecal colonisation.

Since January 1995 there has been a nosocomial outbreak at Haukeland University Hospital involving more than 330 patients with clinical infections caused by ampicillin-resistant Enterococcus faecium (ARE) (minimum inhibitory concentration > or =32 mg/l). Rectal carriage of ARE was initially observed on two medical wards only. Here the ARE colonisation rate has remained high. To assess risk factors for ARE colonisation we performed a case-control study including 37 rectal carriers of ARE and 83 non-carriers on these wards. Significant differences were found between cases and controls with respect to the mean number of days on antimicrobial treatment (13.3 for carriers, 5.5 for non-carriers, p<0.001), mean number of different antibiotics prescribed (2.8 for carriers, 2.1 for non-carriers, p= 0.008) and mean number of days in hospital (18.4 vs 10.2, p=0.001). Unadjusted statistical analysis showed that several antibiotics were risk factors for ARE carriage. Logistic regression analysis showed that fluoroquinolone prescription (OR=3.5, p=0.01) and more than 10 days of antibiotic use (OR= 3.3, p=0.01) were significant risk factors. An additional follow-up screening of previous carriers revealed no colonisation 8 to 36 (median 9) months after discharge from hospital (n=17). Prolonged antimicrobial therapy and broad-spectrum antibiotics seem to facilitate nosocomial ARE colonisation.

Purification of herpes simplex virus type 1 by density gradient centrifugation and estimation of the sedimentation coefficient of the virion.

Purification of herpes simplex virus type 1 (HSV-1) is often performed by centrifugation in gradients of various materials. A major problem with such procedures is a gradual decrease in the infectivity of the virus, probably due to the influence of the gradient material. In the present work we have compared Nycodenz gradients with Ficoll gradients for HSV-1 purification. Both gradient materials have low osmolarities. The purity of the preparations recovered from the two gradients was similar, as measured by electron microscopy and two-dimensional gel electrophoresis, and the yield of infectious virus was approximately the same. Szilágyi & Cunningham (22) reported previously that Ficoll gradients separate one band of complete virions and another band containing light particles devoid of nucleocapsids. In Nycodenz gradients we observed the same bands, and an additional third band which contained aggregates and disrupted particles. In this respect, the separation in Nycodenz is slightly superior to that in Ficoll. The purified HSV-1 virions were subjected to analytical differential centrifugation and an S-value of 670S was calculated for the virus.

Role of penicillin-binding protein 5 C-terminal amino acid substitutions in conferring ampicillin resistance in Norwegian clinical strains of Enterococcus faecium.

The importance of amino acid sequence differences in the C-terminal part and levels of mRNA expression of penicillin-binding protein 5 (PBP5) for ampicillin resistance in Enterococcus faecium was investigated. Seventeen isolates from Norwegian hospitalized patients (ampicillin MIC 0.064->256 mg/L) with different C-terminal pbp5 DNA sequences encoding 11 different amino acid sequences were analyzed with a 14C-radiolabeled penicillin- binding assay to PBP5 and with real-time PCR quantification of pbp5 mRNA expression. Using multiple logistic regression analysis the amino acid substitution Met 485 was linked to ampicillin MIC and levels of 14C-radiolabeled penicillin bound to PBP5; however, there were isolates with identical PBP5 alleles and different ampicillin MICs. There was no relation between the quantity of pbp5 mRNA transcripts and ampicillin resistance. The results cannot explain ampicillin resistance in Norwegian clinical strains of E. faecium and indicate that other factors besides the properties of the C-terminal part of PBP5 are most likely involved.