Correction to: Bacteria: back pain, leg pain and Modic sign-a surgical multicenter comparative study.

Unfortunately, the 5th author name was incorrectly published in the original paper. The complete correct name is given below.

Bacteria: back pain, leg pain and Modic sign-a surgical multicentre comparative study.

PURPOSE: To compare bacterial findings in pain-generating degenerated discs in adults operated on for lumbar disc herniation (LDH), and mostly also suffering from low back pain (LBP), with findings in adolescent patients with non-degenerated non-pain-generating discs operated on for scoliosis, and to evaluate associations with Modic signs on magnetic resonance imaging (MRI). Cutibacterium acnes (Propionibacterium acnes) has been found in painful degenerated discs, why it has been suggested treating patients with LDH/LBP with antibiotics. As multidrug-resistant bacteria are a worldwide concern, new indications for using antibiotics should be based on solid scientific evidence. METHODS: Between 2015 and 2017, 40 adults with LDH/LBP (median age 43, IQR 33-49) and 20 control patients with scoliosis (median age 17, IQR 15-20) underwent surgery at seven Swedish hospitals. Samples were cultured from skin, surgical wound, discs and vertebrae. Genetic relatedness of C. acnes isolates was investigated using single-nucleotide polymorphism analysis. DNA samples collected from discs/vertebrae were analysed using 16S rRNA-based PCR sequencing. MRI findings were assessed for Modic changes. RESULTS: No bacterial growth was found in 6/40 (15%) LDH patients, compared with 3/20 (15%) scoliosis patients. Most positive samples in both groups were isolated from the skin and then from subcutis or deep within the wound. Of the four disc and vertebral samples from each of the 60 patients, 235/240 (98%) were DNA negative by bacterial PCR. A single species, C. acnes, was found exclusively in the disc/vertebra from one patient in each group. In the LDH group, 29/40 (72%) patients had at least one sample with growth of C. acnes, compared to 14/20 (70%) in the scoliosis group. Bacterial findings and Modic changes were not associated. CONCLUSIONS: Cutibacterium acnes found in discs and vertebrae during surgery for disc herniation in adults with degenerated discs may be caused by contamination, as findings in this group were similar to findings in a control group of young patients with scoliosis and non-degenerated discs. Furthermore, such findings were almost always combined with bacterial findings on the skin and/or in the wound. There was no association between preoperative Modic changes and bacterial findings. Antibiotic treatment of lumbar disc herniation with sciatica and/or low back pain, without signs of clinical discitis/spondylitis, should be seriously questioned. These slides can be retrieved under Electronic Supplementary Material.

Evaluation of MLVA for epidemiological typing and outbreak detection of ESBL-producing Escherichia coli in Sweden.

BACKGROUND: To identify the spread of nosocomial infections and halt outbreak development caused by Escherichia coli that carry multiple antibiotic resistance factors, such as extended-spectrum beta-lactamases (ESBLs) and carbapenemases, is becoming demanding challenges due to the rapid global increase and constant and increasing influx of these bacteria from the community to the hospital setting. Our aim was to assess a reliable and rapid typing protocol for ESBL-E. coli, with the primary focus to screen for possible clonal relatedness between isolates. All clinical ESBL-E. coli isolates, collected from hospitals (n = 63) and the community (n = 41), within a single geographical region over a 6 months period, were included, as well as clinical isolates from a polyclonal outbreak (ST131, n = 9, and ST1444, n = 3). The sporadic cases represented 36 STs, of which eight STs dominated i.e. ST131 (n = 33 isolates), ST648 (n = 10), ST38 (n = 9), ST12 and 69 (each n = 4), ST 167, 405 and 372 (each n = 3). The efficacy of multiple-locus variable number tandem repeat analysis (MLVA) was evaluated using three, seven or ten loci, in comparison with that of pulsed-field gel electrophoresis (PFGE) and multi locus sequence typing (MLST). RESULTS: MLVA detected 39, 55 and 60 distinct types, respectively, using three (GECM-3), seven (GECM-7) or ten (GECM-10) loci. For GECM-7 and -10, 26 STs included one type and eleven STs each included several types, the corresponding numbers for GECM-3 were 29 and 8. The highest numbers were seen for ST131 (7,7 and 8 types, respectively), ST38 (5,5,8) and ST648 (4,5,5). Good concordance was observed with PFGE and GECM-7 and -10, despite fewer types being identified with MLVA; 78 as compared to 55 and 60 types. The lower discriminatory power of MLVA was primarily seen within the O25b-ST131 lineage (n = 34) and its H30-Rx subclone (n = 21). Epidemiologically unrelated O25b-ST131 isolates were clustered with O25b-ST131 outbreak isolates by MLVA, whereas the ST1444 outbreak isolates were accurately distinguished from unrelated isolates. CONCLUSION: MLVA, even when using only three loci, represents an easy initial typing tool for epidemiological screening of ESBL-E. coli. For the ST131-O25b linage, complementary methods may be needed to obtain sufficient resolution.

Multilocus Sequence Analysis of Clinical "Candidatus Neoehrlichia mikurensis" Strains from Europe.

"Candidatus Neoehrlichia mikurensis" is the tick-borne agent of neoehrlichiosis, an infectious disease that primarily affects immunocompromised patients. So far, the genetic variability of "Ca. Neoehrlichia" has been studied only by comparing 16S rRNA genes and groEL operon sequences. We describe the development and use of a multilocus sequence analysis (MLSA) protocol to characterize the genetic diversity of clinical "Ca. Neoehrlichia" strains in Europe and their relatedness to other species within the Anaplasmataceae family. Six genes were selected: ftsZ, clpB, gatB, lipA, groEL, and 16S rRNA. Each MLSA locus was amplified by real-time PCR, and the PCR products were sequenced. Phylogenetic trees of MLSA locus relatedness were constructed from aligned sequences. Blood samples from 12 patients with confirmed "Ca. Neoehrlichia" infection from Sweden (n = 9), the Czech Republic (n = 2), and Germany (n = 1) were analyzed with the MLSA protocol. Three of the Swedish strains exhibited identical lipA sequences, while the lipA sequences of the strains from the other nine patients were identical to each other. One of the Czech strains had one differing nucleotide in the clpB sequence from the sequences of the other 11 strains. All 12 strains had identical sequences for the genes 16S rRNA, ftsZ, gatB, and groEL. According to the MLSA, among the Anaplasmataceae, "Ca. Neoehrlichia" is most closely related to Ehrlichia ruminantium, less so to Anaplasma phagocytophilum, and least to Wolbachia endosymbionts. To conclude, three sequence types of infectious "Ca. Neoehrlichia" were identified: one in the west of Sweden, one in the Czech Republic, and one spread throughout Europe.

Real-time PCR threshold cycle cutoffs help to identify agents causing acute childhood diarrhea in Zanzibar.

Molecular assays might improve the identification of causes of acute diarrheal disease but might lead to more frequent detection of asymptomatic infections. In the present study, real-time PCR targeting 14 pathogens was applied to rectal swabs from 330 children aged 2 to 59 months in Zanzibar, including 165 patients with acute diarrhea and 165 asymptomatic control subjects. At least one pathogen was detected for 94% of the patients and 84% of the controls, with higher rates among patients for norovirus genogroup II (20% versus 2.4%; P<0.0001), rotavirus (10% versus 1.8%; P=0.003), and Cryptosporidium (30% versus 11%; P<0.0001). Detection rates did not differ significantly for enterotoxigenic Escherichia coli (ETEC)-estA (33% versus 24%), ETEC-eltB (44% versus 46%), Shigella (35% versus 33%), and Campylobacter (35% versus 33%), but for these agents threshold cycle (CT) values were lower (pathogen loads were higher) in sick children than in controls. In a multivariate analysis, CT values for norovirus genogroup II, rotavirus, Cryptosporidium, ETEC-estA, and Shigella were independently associated with diarrhea. We conclude that this real-time PCR allows convenient detection of essentially all diarrheagenic agents and provides CT values that may be critical for the interpretation of results for pathogens with similar detection rates in patients and controls. The results indicate that the assessment of pathogen loads may improve the identification of agents causing gastroenteritis in children.

Comparison of rectal swabs and faeces for real-time PCR detection of enteric agents in Rwandan children with gastroenteritis.

BACKGROUND: Molecular diagnostics have emerged as an efficient and feasible alternative for broad detection of pathogens in faeces. However, collection of stool samples is often impractical in both clinical work and in epidemiology studies. The aim of this study was to investigate the diagnostic performance of rectal swabs as compared with traditional faeces samples for detection of enteric agents by PCR. METHOD: Three hundred twenty-six pairs of rectal swab and stool samples, obtained from Rwandan children aged 0.5-4.99 years, with or without diarrhoea, were analysed by multiple real-time PCR amplifying 3 viral, 6 bacterial and one protozoan target. RESULTS: For all agents there was a significant correlation (R2 0.31-0.85) between Ct values in faeces and rectal swabs. For most agents the Ct values, a marker for target concentration, were significantly lower (by 1-3 cycles) in faeces, indicating pathogen content up to ten times higher than in rectal swabs. Despite this, there was no significant difference in detection rate between faeces and rectal swabs for any agent, reflecting that pathogen concentration was far above the limit of detection in the majority of cases. CONCLUSION: The similar detection rates and the Ct value correlations as compared with traditional faeces samples indicate that rectal swabs are accurate for real-time PCR-based identification of enteric agents and may be used also for quantitative estimation of pathogen load.

First case of human "Candidatus Neoehrlichia mikurensis" infection in a febrile patient with chronic lymphocytic leukemia.

An immunocompromised patient presented with febrile episodes, an erysipelas-like rash, and thromboembolic complications. Amplification of 16S rRNA gene sequences from blood and sequence analysis revealed "Candidatus Neoehrlichia mikurensis." We report the first case of human disease caused by "Ca. Neoehrlichia mikurensis."

Multiplex quantitative PCR for detection of lower respiratory tract infection and meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis.

BACKGROUND: Streptococcus pneumoniae and Haemophilus influenzae cause pneumonia and as Neisseria meningitidis they are important agents of meningitis. Although several PCR methods have been described for these bacteria the specificity is an underestimated problem. Here we present a quantitative multiplex real-time PCR (qmPCR) for detection of S. pneumoniae (9802 gene fragment), H. influenzae (omp P6 gene) and N. meningitidis (ctrA gene). The method was evaluated on bronchoalveolar lavage (BAL) samples from 156 adults with lower respiratory tract infection (LRTI) and 31 controls, and on 87 cerebrospinal fluid (CSF) samples from meningitis patients. RESULTS: The analytical sensitivity was not affected by using a combined mixture of reagents and a combined DNA standard (S. pneumoniae/H. influenzae/N. meningitidis) in single tubes. By blood- and BAL-culture and S. pneumoniae urinary antigen test, S. pneumoniae and H. influenzae were aetiological agents in 21 and 31 of the LTRI patients, respectively. These pathogens were identified by qmPCR in 52 and 72 of the cases, respectively, yielding sensitivities and specificities of 95% and 75% for S. pneumoniae, and 90% and 65% for H. influenzae, respectively. When using a cut-off of 105 genome copies/mL for clinical positivity the sensitivities and specificities were 90% and 80% for S. pneumoniae, and 81% and 85% for H. influenzae, respectively. Of 44 culture negative but qmPCR positive for H. influenzae, 41 were confirmed by fucK PCR as H. influenzae. Of the 103 patients who had taken antibiotics prior to sampling, S. pneumoniae and H. influenzae were identified by culture in 6% and 20% of the cases, respectively, and by the qmPCR in 36% and 53% of the cases, respectively.In 87 CSF samples S. pneumoniae and N. meningitidis were identified by culture and/or 16 S rRNA in 14 and 10 samples and by qmPCR in 14 and 10 samples, respectively, giving a sensitivity of 100% and a specificity of 100% for both bacteria. CONCLUSIONS: The PCR provides increased sensitivity and the multiplex format facilitates diagnosis of S. pneumoniae, H. influenzae and N. meningitidis and the assay enable detection after antibiotic treatment has been installed. Quantification increases the specificity of the etiology for pneumonia.

Clinical severity of Mycoplasma pneumoniae (MP) infection is associated with bacterial load in oropharyngeal secretions but not with MP genotype.

BACKGROUND: Disease severity in Mycoplasma pneumoniae (MP) infection could potentially be related to bacterial factors such as MP genotype (MP1 or MP2; distinguished by different adhesions proteins) or bacterial load in airway secretions. We have compared these parameters in patients who were hospitalized for MP pneumonia, with outpatients with mild MP disease. METHODS: MP bacterial load was measured by real-time PCR in 45 in- and outpatients ("clinical study group") in whom MP DNA had been detected in oropharyngeal secretions by PCR. In addition, genotype and phylogenetic relationships were determined. The phylogenetical assessment was done by partial DNA sequencing of the P1 gene on isolates from 33 patients in the clinical study-group where sufficient DNA was available. The assessment was further extended to isolates from 13 MP-positive family members and 37 unselected MP positive patients from the two subsequent years and two different geographical locations. In total 83 strains were molecular characterized. RESULTS: Mean MP loads were significantly higher in 24 hospitalized patients than in 21 outpatients (1600 vs. 170 genomic equivalents/microL, p = 0.009). This difference remained significant after adjustment for age and days between disease onset and sampling. Hospitalized patients also had higher C-reactive protein levels. Mean levels were 188 vs 20 mg/L (p = 0,001). The genotype assessment showed MP genotype 1 in 17 of the 33 sequenced strains from the clinical study-group, and type 2 in 16 of these patients. Within each genotype, sequence differences were minimal. No association between disease severity and MP genotype was observed. In the extended genotype assessment, MP1 was found in similar proportions. In family contacts it was found in 53% and among patients from the two subsequent years 53% and 40%. CONCLUSIONS: A higher MP bacterial load in throat secretions at diagnosis was associated with more advanced respiratory disease in patients, but MP genotype did not influence disease severity. Both MP genotypes co-circulated during recent outbreaks in Sweden.

Transmission dynamics study of tuberculosis isolates with whole genome sequencing in southern Sweden.

Epidemiological contact tracing complemented with genotyping of clinical Mycobacterium tuberculosis isolates is important for understanding disease transmission. In Sweden, tuberculosis (TB) is mostly reported in migrant and homeless where epidemiologic contact tracing could pose a problem. This study compared epidemiologic linking with genotyping in a low burden country. Mycobacterium tuberculosis isolates (n = 93) collected at Scania University Hospital in Southern Sweden were analysed with the standard genotyping method mycobacterial interspersed repetitive units-variable number tandem repeats (MIRU-VNTR) and the results were compared with whole genome sequencing (WGS). Using a maximum of twelve single nucleotide polymorphisms (SNPs) as the upper threshold of genomic relatedness noted among hosts, we identified 18 clusters with WGS comprising 52 patients with overall pairwise genetic maximum distances ranging from zero to nine SNPs. MIRU-VNTR and WGS clustered the same isolates, although the distribution differed depending on MIRU-VNTR limitations. Both genotyping techniques identified clusters where epidemiologic linking was insufficient, although WGS had higher correlation with epidemiologic data. To summarize, WGS provided better resolution of transmission than MIRU-VNTR in a setting with low TB incidence. WGS predicted epidemiologic links better which could consolidate and correct the epidemiologically linked cases, avoiding thus false clustering.

Infection with Panton-Valentine leukocidin-positive methicillin-resistant Staphylococcus aureus t034.

Panton-Valentine leukocidin (PVL)-positive methicillin-resistant Staphylococcus aureus (MRSA), sequence type 398 is believed to be of animal origin. We report 2 cases of infection due to PVL-positive MRSA, spa type t034, in patients in Sweden who had had no animal contact.

Time window for positive cerebrospinal fluid broad-range bacterial PCR and Streptococcus pneumoniae immunochromatographic test in acute bacterial meningitis.

BACKGROUND: Reliable microbiological tests are essential for the diagnosis of acute bacterial meningitis (ABM). In this study we investigated the time period after the start of antibiotic therapy during which culture, polymerase chain reaction (PCR) and the immunochromatographic test (ICT) are able to detect bacteria in cerebrospinal fluid (CSF). METHODS: The study was performed on CSF samples from adults with ABM admitted to the Department of Infectious Diseases, Sahlgrenska University Hospital, Gothenburg, Sweden, from January 2007 to April 2014. In addition to the initial lumbar puncture (LP), the participants underwent one or two more LPs during 10 days following the start of antibiotics. The analyses performed on the CSF samples were culture, PCR and ICT. RESULTS: The study comprised 70 CSF samples from 25 patients with ABM. A bacterium could be identified by CSF culture in 44%, by blood culture in 58% and by PCR in 100% of the patients. There were no positive CSF cultures in samples taken later than the day of starting antibiotics. PCR was positive in 89% on days 1-3, 70% on days 4-6 and 33% on days 7-10. For cases of pneumococcal meningitis, the ICT was positive in 88% on days 1-3, 90% on days 4-6 and 75% on days 7-10. CONCLUSIONS: This study shows that PCR is highly sensitive for bacterial detection in CSF samples taken up to 1 week into antibiotic therapy. The ICT is highly sensitive for the detection of pneumococci in CSF samples taken during the first week of antibiotic treatment.

Analyses of black fungi by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS): species-level identification of clinical isolates of Exophiala dermatitidis.

Conventional mycological identifications based on the recognition of morphological characteristics can be problematic. A relatively new methodology applicable for the identification of microorganisms is based on the exploitation of taxon- specific mass patterns recorded from abundant cell proteins directly from whole-cell preparations, using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). This study reports the application of MALDI-TOF MS for the differentiation and identifications of black yeasts, isolated from the respiratory tracts of patients with cystic fibrosis (CF). Initial phenotypic and DNA sequence-based analyses identified these isolates to be Exophiala dermatitidis. The type strains of E. dermatitidis (CBS 207.35(T)) and other species of Exophiala were included in the MALDI-TOF MS analyses to establish the references for comparing the mass spectra of the clinical isolates of Exophiala. MALDI-TOF MS analyses exhibited extremely close relationships among the clinical isolates and with the spectra generated from the type strain of E. dermatitidis. The relationships observed between the E. dermatitidis strains from the MALDI-TOF MS profiling analyses were supported by DNA sequence-based analyses of the rRNA ITS1 and ITS2 regions. These data demonstrated the applicability of MALDI-TOF MS as a reliable, rapid and cost-effective method for the identification of isolates of E. dermatitidis and other clinically relevant fungi and yeasts that typically are difficult to identify by conventional methods.

Streptococcus devriesei sp. nov., from equine teeth.

Phenotypic and phylogenetic studies were performed on four unidentified Gram-positive staining, catalase-negative, alpha-hemolytic Streptococcus-like organisms recovered from the teeth of horses. SDS PAGE analysis of whole-cell proteins and comparative 16S rRNA gene sequencing demonstrated the four strains were highly related to each other but that they did not correspond to any recognised species of the genus Streptococcus. Phylogenetic analysis based on 16S rRNA gene sequences showed the unidentified organisms form a hitherto unknown sub-line within the Streptococcus genus, displaying a close affinity with Streptococcus mutans, Streptococcus ferus and related organisms. Sequence divergence values of > 5% with these and other reference streptococcal species however demonstrated the organisms from equine sources represent a novel species. Based on the phenotypic distinctiveness of the new bacterium and molecular chemical and molecular genetic evidence, it is proposed that the unknown species be classified as Streptococcus devriesei sp. nov. The type strain of Streptococcus devriesei is CCUG 47155T (= CIP 107809T).

[How an outbreak of MRSA in Gothenburg was eliminated: by strict hygienic routines and massive control-culture program]. / Så hävdes MRSA-utbrottet i Göteborg: med strikta hygienrutiner och omfattande kontrollodlingsprogram.

The largest single-strain outbreak of methicillin resistant Staphylococcus aureus (MRSA) in Scandinavia so far occurred at Sahlgrenska University Hospital in Western Sweden 1997-2000. The strain identified was identical to the UK EMRSA-16 strain. 147 patients at 36 different wards became colonised or infected. Established routines for infection control had to be revised. The endemic situation necessitated an MRSA screening programme in October 1999 for all former hospital patients on re-admission. Since May 2000 no patient has been found with the outbreak strain at Sahlgrenska University Hospital.

Genotyping of Pseudomonas aeruginosa isolates from lung transplant recipients and aquatic environment-detected in-hospital transmission.

Lung infection with Pseudomonas aeruginosa is common in lung transplant recipients and may lead to severe complications. Bacteriological surveillance aims to detect transmission of microbes between hospital environment and patients. We sought to determine whether genotyping of P. aeruginosa isolates could improve identifications of pathways of infection. From 2004 to 2009, we performed genotyping with multiple-locus variable number of tandem repeats analysis (MLVA) and pulsed-field gel electrophoresis (PFGE) of P. aeruginosa isolates cultured from lung transplant recipients at Sahlgrenska University Hospital, Gothenburg. During a small outbreak in 2008, cultivation and genotyping of isolates from sink and drains samples from the hospital ward were performed. Pseudomona aeruginosa from 11/18 patients were genotyped to unique strains. The remaining seven patients were carriers of a P. aeruginosa strain of cluster A genotype. Pseudomona aeruginosa was isolated in 4/8 water samples, typed by MLVA also as cluster A genotype and confirmed by PFGE to be similar or identical to the isolates from four transplanted patients. In conclusion, genotyping of isolates revealed a clonal relationship between patient and water isolates, indicating in-hospital transmission of P. aeruginosa. We suggest genotyping with MLVA for rapid routine surveillance, with the PFGE method used for extended, confirmatory analyses.

Development of IgG antibodies to Exophiala dermatitidis is associated with inflammatory responses in patients with cystic fibrosis.

BACKGROUND: The clinical importance of airway colonisation by the fungus Exophiala dermatitidis in patients with cystic fibrosis (CF) is unclear. We have previously shown that E. dermatitidis frequently colonises the airways of patients with CF. The aims of the present study were to determine whether patients who are colonised by E. dermatitidis have detectable fungal antigens in the circulation, develop anti-fungal antibodies, and show signs of inflammation and impaired respiratory function. METHODS: We collected sputum and serum samples consecutively from 98 sputum-producing patients with CF aged more than 12 years. The serum samples were subjected to bacterial and fungal culturing and analyses for fungal antigens and inflammatory factors. RESULTS: E. dermatitidis was recovered from 17 (17%) patients, the same isolation rate as for Aspergillus fumigatus. There were no difference regarding the levels of ß-glucan in the sera from E. dermatitidis culture-positive and culture-negative patients with CF. Serological analysis revealed significantly higher levels of IgG antibodies to E. dermatitidis cell wall fragments in the E. dermatitidis culture-positive patients. Patients with higher level of E. dermatitidis IgG antibodies were more often colonised with non-tuberculous Mycobacteria, and less often with Staphylococcus aureus. The increased levels of IgG antibodies directed against E. dermatitidis were positively associated with higher white blood cell counts, increased erythrocyte sedimentation rate, pancreatic insufficiency, intravenous antibiotic treatment, and they were negatively associated with respiratory function (FEV1 % predicted). Overall, 4/17 Exophiala-positive patients were diagnosed as having symptomatic infection with E. dermatitidis and were treated with broad-spectrum azoles. CONCLUSION: E. dermatitidis triggers antibody production and may cause significant airway infection in patients with cystic fibrosis.

Virulence gene typing of methicillin-resistant Staphylococcus aureus as a complement in epidemiological typing.

Methicillin-resistant Staphylococcus aureus (MRSA) has widely spread to all parts of the world. For surveillance and effective infection control molecular typing is required. We have evaluated the utility of virulence gene determination as a complementary tool for epidemiological typing of MRSA in relation to spa-typing and pulsed-field gel electrophoresis (PFGE). We assessed 63 community-acquired MRSA (CA-MRSA) isolates detected in the West part of Sweden for 30 virulence factor genes (VF) and agr allele variations by serial polymerase chain reaction (PCR) assays. These isolates belonged to sequence types (ST) 8, 80, 45 and 30 as classified by multilocus sequence typing. The isolates in each spa-type and PFGE-type were examined over an extended time-period and constituted a varying number of PFGE-subtypes (5-14) and spa-types (3-11) within four major PFGE types. Each ST had a unique VF profile. For isolates within a major PFGE type showing high diversity both in PFGE subtypes and spa the VF profile varied as well in contrast to those with low diversity where no alterations were seen. Thus, the accuracy of each typing method does not only vary by the method per se but is rather dependent on the genetic repertoire of the typed strains and genes evaluated. For strains demonstrating high diversity VF typing may be a useful complement in the epidemiological investigations, and may highlight the accurate discriminatory power of spa or PFGE typing.

Sub-typing of extended-spectrum-ß-lactamase-producing isolates from a nosocomial outbreak: application of a 10-loci generic Escherichia coli multi-locus variable number tandem repeat analysis.

Extended-spectrum ß-lactamase producing Escherichia coli (ESBL-E. coli) were isolated from infants hospitalized in a neonatal, post-surgery ward during a four-month-long nosocomial outbreak and six-month follow-up period. A multi-locus variable number tandem repeat analysis (MLVA), using 10 loci (GECM-10), for 'generic' (i.e., non-STEC) E. coli was applied for sub-species-level (i.e., sub-typing) delineation and characterization of the bacterial isolates. Ten distinct GECM-10 types were detected among 50 isolates, correlating with the types defined by pulsed-field gel electrophoresis (PFGE), which is recognized to be the 'gold-standard' method for clinical epidemiological analyses. Multi-locus sequence typing (MLST), multiplex PCR genotyping of bla CTX-M, bla TEM, bla OXA and bla SHV genes and antibiotic resistance profiling, as well as a PCR assay specific for detecting isolates of the pandemic O25b-ST131 strain, further characterized the outbreak isolates. Two clusters of isolates with distinct GECM-10 types (G06-04 and G07-02), corresponding to two major PFGE types and the MLST-based sequence types (STs) 131 and 1444, respectively, were confirmed to be responsible for the outbreak. The application of GECM-10 sub-typing provided reliable, rapid and cost-effective epidemiological characterizations of the ESBL-producing isolates from a nosocomial outbreak that correlated with and may be used to replace the laborious PFGE protocol for analyzing generic E. coli.

Pertussis after end of a mass vaccination project--end of the "vaccination honey-moon".

After 16 years of no vaccination against pertussis in Sweden, mass vaccination of infants and catch-up vaccination of children up to 10 years with a monocomponent pertussis toxoid vaccine was performed in the Greater Gothenburg area of Sweden between 1995 and 1999. At the end of the project in February 1999, 56% of all 10 year old children born in the Greater Gothenburg area had received 3 doses of the pertussis toxoid. No booster doses were given. This led to a temporary almost complete elimination of the disease. The aim of the present study was to follow the incidence of pertussis after end of the mass vaccination project (1999-2009) as it is reflected by laboratory verified cases (cultures and/or PCR) and pertussis hospitalizations. A reemergence of pertussis was seen from the end of 1999 with a peak in 2004 followed by a decrease when booster doses to both 6 and 10 year old children were introduced in 2005-2006. From July 1, 1999 through December 31, 2009 a total of 1973 cases were diagnosed with culture or PCR. The disease was prevalent in all age groups. The highest documented incidence was seen in infants younger than 12 months. 450 patients with verified pertussis had received 3 doses of the pertussis toxoid vaccine in the mass vaccination project and some other trials (comprising a total of 69,423 children). The mean time from the last dose to the laboratory verification of pertussis was 5 years in these 450 cases. There were 128 hospitalizations, 106 of which were in infants. In conclusion, pertussis is still not eliminated from the area. Booster doses are needed but the numbers and optimal timing are not known.