The distribution limit of the common tick, Ixodes ricinus, and some associated pathogens in north-western Europe.

In north-western Europe, the common tick, Ixodes ricinus, is widely established, its distribution appears to be increasing and the spread of tick-borne diseases is of increasing concern. The project 'Flått i Nord' (Ticks in northern Norway) commenced in spring 2009 with the intention of studying the tick's distribution and that of its pathogens in northern Norway. Several methods were used: cloth-dragging, collecting from trapped small mammals, and collecting from pets. Since 2010, the occurrence of ticks in the region of northern Norway was determined directly by cloth-dragging 167 times in 109 separate locations between the latitudes of 64 °N and 70 °N (included seven locations in the northern part of Trøndelag County). The northernmost location of a permanent I. ricinus population was found to be Nordøyvågen (66.2204 °N, 12.59 °E) on the Island of Dønna. In a sample of 518 nymphal and adult ticks, the Borrelia prevalence collected close to this distribution limit varied but was low (1-15 %) compared with the locations in Trøndelag, south of the study area (15-27 %). Five specimens (1 %) were positive for Rickettsia helvetica. The length of the vegetation growing season (GSL) can be used as an approximate index for the presence of established populations of I. ricinus. The present study suggests that the threshold GSL for tick establishment is about 170 days, because the median GSL from 1991 to 2015 was 174-184 days at sites with permanent tick populations, showing a clear increase compared with the period 1961-1990. This apparent manifestation of climate change could explain the northward extension of the range of I. ricinus.

Emergence of clonally related multidrug resistant Haemophilus influenzae with penicillin-binding protein 3-mediated resistance to extended-spectrum cephalosporins, Norway, 2006 to 2013.

Resistance to cephalosporins in Haemophilus influenzae is usually caused by characteristic alterations in penicillin-binding protein 3 (PBP3), encoded by the ftsI gene. Resistance to extended-spectrum cephalosporins is associated with high-level PBP3-mediated resistance (high-rPBP3), defined by the second stage S385T substitution in addition to a first stage substitution (R517H or N526K). The third stage L389F substitution is present in some high-rPBP3 strains. High-rPBP3 H. influenzae are considered rare outside Japan and Korea. In this study, 30 high-rPBP3 isolates from Norway, collected between 2006 and 2013, were examined by serotyping, multilocus sequence typing (MLST), ftsI sequencing, detection of beta-lactamase genes and minimum inhibitory concentration (MIC) determination. MICs were interpreted according to clinical breakpoints from the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Respiratory isolates predominated (proportion: 24/30). The 30 isolates included one serotype f isolate, while the remaining 29 lacked polysaccharide capsule genes. Resistance to extended-spectrum cephalosporins (cefixime, 29 isolates/30 isolates; cefepime, 28/30; cefotaxime, 26 /30; ceftaroline, 26/30; ceftriaxone, 14/30), beta-lactamase production (11/30) and co-resistance to non-beta-lactams (trimethoprim-sulfamethoxazole, 13/30; tetracycline, 4/30; chloramphenicol, 4/30; ciprofloxacin, 3/30) was frequent. The N526K substitution in PBP3 was present in 23 of 30 isolates; these included a blood isolate which represents the first invasive S385T + N526K isolate reported from Europe. The L389F substitution, present in 16 of 30 isolates, coincided with higher beta-lactam MICs. Non-susceptibility to meropenem was frequent in S385T + L389F + N526K isolates (8/12). All 11 beta-lactamase positive isolates were TEM-1. Five clonal groups of two to 10 isolates with identical MLST-ftsI allelic profiles were observed, including the first reported high-rPBP3 clone with TEM-1 beta-lactamase and co-resistance to ciprofloxacin, tetracycline, chloramphenicol and trimethoprim-sulfamethoxazole. Prior to this study, no multidrug resistant high-rPBP3 H. influenzae had been reported in Norway. Intensified surveillance of antimicrobial resistance is needed to guide empiric therapy.

Mutant ftsI genes in the emergence of penicillin-binding protein-mediated beta-lactam resistance in Haemophilus influenzae in Norway.

The most important mechanism for beta-lactam resistance in beta-lactamase-negative ampicillin-resistant (BLNAR) isolates of Haemophilus influenzae is the alteration of penicillin-binding protein 3 (PBP3) as a result of ftsI gene mutations. The present study aimed to map PBP3 alterations and to determine the correlation to beta-lactam resistance in respiratory tract isolates of H. influenzae in Norway, as well as assess the contribution of clonal spread to the emergence of PBP3-mediated resistance. Twenty-three beta-lactamase negative respiratory tract isolates with resistance to penicillins and 23 susceptible control isolates were examined by determination of beta-lactam MICs, ftsI sequencing and molecular typing by pulsed-field gel electrophoresis (PFGE). Ampicillin MIC ranges in the resistant group and the control group were 1-2 mg/L and 0.125-0.5 mg/L, respectively. All isolates in the resistant group had the PBP3 substitution Asn526-->Lys and were thus categorized as group II low-BLNAR. No control isolate met the genetic BLNAR (gBLNAR) criteria. The PBP3 substitution patterns corresponded well to those observed in previous European studies. Eighty-three percent (19/23) of the resistant isolates belonged to two clones, demonstrating the capability of low-BLNAR strains of clonal dissemination. Combined analysis of ftsI DNA sequences and PFGE patterns revealed distinctly different ftsI alleles in genetically indistinguishable isolates and identical copies of the same ftsI allele in unrelated isolates. A possible explanation of this observation is the recombinational exchange of ftsI alleles. This phenomenon, as well as the possibility of endemic European gBLNAR strains, should be further investigated.

Fluoroquinolone-resistant uropathogenic Escherichia coli in Norway: evidence of clonal spread.

Prevalence, resistance profiles, virulence gene complements, and phylogenetic and clonal affinities of fluoroquinolone-resistant Escherichia coli from urinary tract infections (UTIs) in Norway were investigated. Of 7302 E. coli UTI isolates from 2003, 1.2% were fluoroquinolone-resistant; 35 of these fluoroquinolone-resistant isolates were included in the present study. The isolates were predominantly multiresistant, carried few virulence factors, and tended to belong to the less-virulent phylogroups A and B1. Although the isolates were genetically heterogeneous, there was evidence of a limited degree of clonal dissemination.

A comparison of phylogenetic group, virulence factors and antibiotic resistance in Russian and Norwegian isolates of Escherichia coli from urinary tract infection.

Isolates of Escherichia coli from 31 Norwegian and 31 Russian females with significant bacteruria who presented with clinical signs of urinary tract infection (UTI) were tested for antimicrobial sensitivity, the presence of virulence genes, phylogroup distribution and clonal affinity. Twenty isolates, representing the full clonal diversity of a collection of 138 intestinal isolates of E. coli from healthy Norwegian females, served as a reference group. Russian UTI isolates belonged more often to phylogroup A and possessed fewer virulence genes than did Norwegian isolates. UTI isolates of E. coli were genetically heterogeneous and had a high degree of antimicrobial sensitivity.

Comparison of PCR detection of mecA with agar dilution and Etest for oxacillin susceptibility testing in clinical isolates of coagulase-negative staphylococci.

Oxacillin-resistant staphylococci are heterogeneous in their expression of resistance to beta-lactam antibiotics. Different recommendations regarding screening methods for routine use have been published. In this study, the susceptibility to oxacillin of 232 coagulase-negative staphylococci (CoNS) was determined by agar dilution, Etest and presence of the mecA gene. When an oxacillin resistance breakpoint of > or = 0.5 mg/L was used, the sensitivity and specificity for agar dilution were 97.6% and 100%, and those for Etest were 100% and 95.4%. The current National Committee for Clinical Laboratory Standards oxacillin breakpoint recommendation will categorise accurately the CoNS species encountered commonly.

Identification of Aerococcus urinae in urine samples.

To evaluate procedures for the identification of Aerococcus urinae, we examined 24 alpha-hemolytic non-enterococcal bacterial isolates from 4373 urine samples. Published procedures were compared with 16s rRNA sequencing and biochemical profiling (BBL-Crystal-GP). 16s rRNA sequencing and BBL-Crystal-GP identified the same 13 isolates as A. urinae. Published tests failed to distinguish the 13 A. urinae isolates from eight non-A. urinae isolates; several tests exhibited no discrimination. Ciprofloxacin and trimethoprim susceptibility and growth at 45 degrees C improved discrimination. For urinary isolates, standard procedures for identification of A. urinae are redundant and insufficiently discriminatory, and may need revision. BBL-Crystal-GP is an accurate alternative.

Heterogeneity of methicillin-resistant Staphylococcus aureus isolated in Norway.

Our objective was to look for differences in susceptibility patterns between Norwegian and imported methicillin-resistant Staphylococcus aureus (MRSA) strains. All MRSA isolates from the participating hospitals (87 isolates from 81 patients) throughout the period 1994-98 were examined, to study the clonal distribution of MRSA isolated in Norway and to identify any epidemic clones among the isolates. We found that imported isolates were resistant to an average of 5.6 antibiotics, while Norwegian isolates were resistant to an average of 2.6 antibiotics. MRSA isolates imported to Norway are more often multiresistant than domestic isolates. MRSA isolates in Norway show a striking diversity. Epidemic clones are present, but no single clone is predominant.

Urinary tract infections in Norway: bacterial etiology and susceptibility. A retrospective study of clinical isolates.

OBJECTIVES: The objective of the present study was to determine bacterial etiology and susceptibility in urinary tract infections. The study was designed as a retrospective study of urine samples from patients both inside and outside hospitals and nursing homes that were received at our laboratory between 1 January 1997 and 31 October 1999. The Telemark Biomedical Center receives all the medical microbiology specimens from hospitals, nursing homes and general practitioners in the County of Telemark (165,000 inhabitants), Norway. All urines fulfilling the criteria for significant bacteriuria [> or =10,000 colony-forming units/mL urine] were included in the study. METHODS: Bacterial susceptibility testing was performed using breakpoint methodology. During the study period, we received 52 350 urine samples, of which 28,066 (53.6%) fulfilled the criteria for significant bacteriuria (pure growth of > or =10,000 CFU/mL urine). RESULTS: Escherichia coli was the most predominant bacterium in the urine from both inpatients (56.7%) and outpatients (68.3%). Coagulase-negative staphylococci and enterococci occurred significantly more often (P < 0.001) in urine samples from inpatients (12.5% and 7.9%) than in urine samples from outpatients (7.5% and 4.7%). Escherichia coli from both outpatients and hospitalised patients was highly susceptible (>93%) to cefalothin, mecillinam and nitrofurantoin, and more than 75% of E. coli isolates were also sensitive to ampicillin. Overall, the susceptibility to nitrofurantoin in bacteria from outpatients was 90% and from hospitalised patients was 85%. The corresponding figures for cefalothin were 92% and 90%, and for trimethoprim were 81% and 76%, respectively. CONCLUSIONS: Bacteria causing urinary tract infections in Norway are less resistant to antibacterial medication than in other western countries and the reason for this may be the low consumption of antibacterials by the Norwegian population. During the period from 1990 to 1999 the mean total annual consumption of antibacterial drugs in Norway was 15.3 defined daily doses per 1000 inhabitants per year.

Borrelia burgdorferi sensu lato and Ehrlichia spp. in Ixodes ticks from southern Norway.

We report the results of a study of the prevalence of Ehrlichia and Borrelia species in 341 questing Ixodes ricinus ticks from two locations in southern Norway. The prevalences of Borrelia burgdorferi sensu lato and Ehrlichia spp. were, respectively, 16 and 11.5% at site 1 and 17 and 6% at site 2. Prevalence and species composition of Borrelia and Ehrlichia varied with location and date of collection. The dominant Borrelia species at both sites was Borrelia afzelii, followed by Borrelia burgdorferi sensu stricto. Borrelia garinii was found in only a single tick. The dominant member of the Ehrlichia group was a recently described Ehrlichia-like organism related to the monocytic ehrlichiae. Variants of Ehrlichia phagocytophila and the agent of human granulocytic ehrlichiosis were also found. The highest prevalences for B. afzelii, B. burgdorferi sensu stricto, and the Ehrlichia-like organism were observed in May. B. afzelii was most prevalent in females, less prevalent in nymphs, and least prevalent in males, while the prevalence of Ehrlichia was highest in nymphs, lower in females, and least in males. Double infections with B. afzelii and B. burgdorferi sensu stricto and with B. afzelii and the Ehrlichia-like organism were significantly overrepresented. Tick densities were highest in May, when densities of more than 200 ticks/100 m2 were observed, and declined during the summer months to densities as low as 20 ticks/100 m2. We conclude that estimates of the prevalence of tick-borne bacteria are sensitive to the choice of date and site for collection of ticks. This is the first study of tick-borne Borrelia and Ehrlichia in Norway and the lowest reported B. garinii prevalence in Northern Europe. The prevalence of the Ehrlichia-like organism is described for the first time in questing ticks.

[Human granulocytic ehrlichiosis in Norway]. / Human granulocytaer ehrlichiose i Norge.

BACKGROUND: The bacterium that causes human granulocytic ehrlichiosis may be transmitted by ticks. MATERIAL AND METHODS: We describe two patients with human granulocytic ehrlichiosis. During the summer of 1998, both patients were bitten by ticks. Four to 7 days later they developed influenza-like symptoms with fever, headache and myalgia. After 4 and 21 days, respectively, both patients were given doxycycline for suspected bacterial respiratory diseases, and recovered. RESULTS: Blood samples for human granulocytic ehrlichiosis antibodies showed a fourfold increase in titer in one patient and a remaining high titer in the other. Both patients had a positive polymerase chain reaction with primers specific for the Ehrlichia phagocytophilae genogroup. INTERPRETATION: The two patients fulfill the human granulocytic ehrlichiosis diagnostic criteria set by Centers for Disease Controls and Prevention, and are the first two human granulocytic ehrlichiosis cases described in Norway.

Ehrlichiosis in a moose calf in Norway.

A case of granulocytic ehrlichiosis in a moose calf (Alces alces) in Norway is described. The animal was heavily infested with ticks (Ixodes ricinus), and died from a Klebsiella pneumoniae septicemia. Examination of blood smears from the calf revealed cytoplasmic inclusions (morulae) typical of infection with Ehrlichia phagocytophila in the granulocytes. Ehrlichia sp. was detected by polymerase chain reaction (PCR) in blood from the calf, and in the ticks. Sequence determination identified it as E. phagocytophila. This is the first report of ehrlichiosis in moose.

The prevalence of antibiotic resistance in bacterial respiratory pathogens from Norway is low.

OBJECTIVES: To determine the degree of bacterial susceptibility to the most commonly used drugs for respiratory infections in Norway, and to find if bacterial resistance is emerging. METHODS: Clinical isolates of Streptococcus pneumoniae, Haemophilus influenzae and group A streptococci from respiratory tract specimens and from the eye were collected from different parts of Norway during two study periods. During the first period (1993-1994), three laboratories, covering 15% of the Norwegian population, participated. During the second study period in 1997, five laboratories, covering 27% of the population, collected respiratory isolates. In total, 494 isolates of S. pneumoniae, 696 isolates of H. influenzae and 694 isolates of group A streptococci were included in the study. The study population comprised children and adults attending hospital and general practice. Bacterial susceptibility was determined by the E test, and breakpoints were according to the National Committee for Clinical Laboratory Standards (NCCLS). RESULTS: The prevalence of bacterial resistance was low, and we observed no significant increase in bacterial resistance between the two study periods. In 1997, only 0.6% of pneumococci had decreased susceptibility to penicillin, 1.6% of group A streptococci were resistant to erythromycin, and 6.7% of all isolates of H. influenzae produced beta-lactamase. CONCLUSIONS: The prevalence of antibiotic resistance in respiratory pathogens in Norway is low.

Vancomycin resistance emerging in a clonal outbreak caused by ampicillin-resistant Enterococcus faecium.

OBJECTIVES: To describe the first nosocomial outbreak of ampicillin-resistant Enterococcus faecium (ARE) in Norway, where a few vancomycin-resistant strains have also been identified. METHODS: All cases of ARE and vancomycin-resistant Enterococcus faecium (VRE) diagnosed by the medical microbiological laboratories in a region inhabited by approximately 1 million people were registered. Isolates obtained during the period 1 January 1995 to 31 December 1996 were characterized by pulsed field-gel electrophoresis and the clinical data were recorded. RESULTS: One hundred and forty-nine patients (64 males, 85 females, mean age 70.5 years) were infected with ARE. Isolates from 115 cases were genomically related to the outbreak strain. Infections included bacteremia (14), wound infections (31), urinary tract infections (97) and other infections (seven). Most had a severe underlying disease and 93% of the patients had received antibiotics for a mean time of 23 days. Twenty-four patients (16.1%) died during hospitalization. Four infections were caused by a vanB-type VRE that was genomically related to the ARE outbreak strain. The prescription rate for vancomycin was low, but an increase in vancomycin use paralleled the appearance of VRE. The highest monthly incidence rate was 2.5 per 1000 patient admissions in July 1996 declining to 0.5 in December 1996. CONCLUSIONS: The first nosocomial outbreak caused by ARE was observed in 1995 in Norway and is still ongoing. One year after the onset, VRE occurred in wards which had a relatively high consumption of vancomycin.

[Varying clinical picture and course of human granulocytic ehrlichiosis. Twelve Scandinavian cases of the new tick-borne zoonosis are presented]. / Varierande klinisk bild och förlopp vid human granulocytär ehrlichios. Tolv skandinaviska fall av den nya fästingburna zoonosen redovisas.

In the twelve clinical cases of human granulocytic ehrlichiosis (HGE) so far identified in Scandinavia (ten in Sweden, two in Norway), clinical presentation varied from a mild febrile illness to a severe septic condition with such systemic complications as acute respiratory distress syndrome (ARDS). Laboratory verification was based on PCR (polymerase chain reaction) in ten cases, and on serology in two cases. Sequence analysis of 16S rDNA showed the infectious agents to belong to the Ehrlichia phagocytophila genogroup. Seroprevalence data indicate widespread human exposure to granulocytic Ehrlichia; mean seroprevalence, 15-20% of 1,000 clinical sera from tick-exposed patients (mainly from Sweden and Norway). Proposals for diagnostic criteria and procedures, and case management are presented in the article.

Which contacts of patients with meningococcal disease carry the pathogenic strain of Neisseria meningitidis? A population based study.

OBJECTIVES: To determine the prevalence of the pathogenic strain of Neisseria meningitidis in contacts of patients with meningococcal disease, and to determine which contact groups are likely to be carriers and warrant chemoprophylaxis. DESIGN: Population based study. SETTING: Norwegian county of Telemark. SUBJECTS: 1535 primary contacts of 48 patients with meningococcal disease, and 78 secondary contacts. INTERVENTIONS: Carriers of the pathogenic strain were treated with rifampicin. All household members and kissing contacts under 15 years of age were treated with oral penicillin. Contacts were taught to recognise the symptoms of meningococcal disease. RESULTS: In 27 of 48 cases investigated, contacts carrying the pathogenic strain of N meningitidis were found. A total of 42 such contacts were identified. Contacts were stratified into three classes according to the assumed closeness of contact with patients. In class 1 (household members and kissing contacts) the prevalence of the pathogenic strain was 12.4% (95% confidence interval 5.5% to 19.3%). In classes 2 and 3 the prevalence was 1.9% (0.9% to 3.4%) and 1.6% (0.14% to 3.1%). CONCLUSIONS: There is a high rate of carriage of the pathogenic strain of N meningitidis in patients' household members and kissing contacts, and this supports the practice of giving chemoprophylaxis to these contacts. The prevalence of carriage among other contacts is 2-3 times that found in the general population (0.7%); the benefits of chemoprophylaxis to these contacts may be marginal.

Clonal distribution of invasive Neisseria meningitidis isolates from the Norwegian county of Telemark, 1987 to 1995.

Forty-two Neisseria meningitidis isolates were obtained from patients with meningococcal disease in the Norwegian county of Telemark (January 1987 to March 1995), and all were compared by PCR amplicon restriction endonuclease analysis (PCR-AREA) of the dhps gene, chromosomal DNA fingerprinting, and serological analysis. PCR-AREA divided the isolates into 11 classes, of which 4, comprising 15, 8, 6, and 2 isolates, were clonal while the remaining 8 classes were genetically heterogeneous or contained only 1 isolate. Three of the four clonal classes could be tentatively equated with recognized epidemic clones (ET5, ET37, and cluster A4) on the basis of their phenotypic characteristics, while the remaining clone appears to be new. There were significant differences in the geographical distribution of clones, with class 1 (ET5-like) isolates significantly overrepresented in rural parts of Telemark. Class 1 (ET5-like) isolates occurred throughout the study period and were dominant in 1987. Class 2 (ET37-like) isolates occurred from 1988 to 1992, and class 3 isolates (with no recognizable ET affinities) were found only in 1991 and 1992.

Cloning, sequence analysis and expression in E. coli of the DNA polymerase I gene from Chloroflexus aurantiacus, a green nonsulfur eubacterium.

We have cloned and sequenced the polA gene from Chloroflexus aurantiacus, a green nonsulfur eubacterium, and expressed the recombinant protein in Escherichia coli. One open reading frame encodes a protein with 942 amino acids showing 38% identity with DNA polymerase I from E. coli. Sequence alignments with other members of DNA polymerase family A and analysis of the separate domains show that the central 3'-5' exonuclease domain is 30% identical to the corresponding E. coli domain and that three sequence motifs associated with 3'-5' exonuclease activity are conserved. Also, a protein fraction from E. coli expressing the Chloroflexus polymerase contains a thermostable 3'-5' exonucleolytic activity, indicating that this activity is present in the enzyme, in agreement with the sequence analysis. The N-terminal 5'-3' exonuclease domain and the C-terminal polymerase domain show 31 and 46% identity, respectively, with the corresponding E. coli domains and all sequence motifs associated with these two enzymatic activities also are conserved. Since several DNA polymerase I enzymes lack the proofreading activity associated with the central domain it has been suggested that the ancestral polA gene contained only the two more conserved N- and C-terminal domains and that the proofreading 3'-5' exonuclease domain was introduced later in those eubacterial branches that have this activity. Our data indicate a different scenario where the ancestral polA gene contained both the exonucleolytic activities in addition to the polymerase activity and where several eubacterial branches lost the polymerase-associated proofreading activity during evolution.