Staphylococcus aureus is a pathogen and a skin commensal that is today also common in the infant gut flora. We examine the role of S. aureus virulence factors for gut colonization. S. aureus isolated from quantitative stool cultures of 49 Swedish infants followed from birth to 12 months of age were assessed for 30 virulence-associated genes, spa type, and agr allele by serial polymerase chain reaction (PCR) assays. Strains carrying genes encoding collagen-binding protein, and the superantigens S. aureus enterotoxin O/M (SEO/SEM) had higher stool counts than strains lacking these genes, whereas genes for S. aureus enterotoxin A (SEA) were associated with low counts. A cluster of strains belonging to agr allele I and the spa clonal cluster 630 (spa-CC 630) that carried genes encoding SEO/SEM, SEC, collagen-binding protein, and elastin-binding protein were all long-time colonizers. Thus, certain S. aureus virulence factors might promote gut colonization.
Laboratory of Medical Microbiology, St. Olavs Hospital, University Hospital, Department of Laboratory Medicine, Children's and Women's Health, Trondheim, Norway. jan.afset@stolav.no
The aim of the present case control study was to investigate the prevalence of atypical enteropathogenic Escherichia coli (EPEC) and its possible role in causing diarrhoea among children or = 14 days had atypical EPEC. The association between atypical EPEC and prolonged diarrhoea (OR = 2.1, P = 0.04) was caused by a high prevalence among female patients (40.6 %). In conclusion, atypical EPEC was found to be slightly more prevalent in patients than controls, without any overall significant association with diarrhoea. However, a significant association was observed with diarrhoea lasting 14 days or more, a finding that may indicate a role for atypical EPEC in prolonged disease.
Campylobacter spp., Enterococcus spp., Escherichia coli, Salmonella spp., Yersinia spp., and Cryptosporidium oocysts in semi-domesticated reindeer (Rangifer tarandus tarandus) in Northern Finland and Norway.
The specific aim of this study was to assess the faecal shedding of zoonotic enteropathogens by semi-domesticated reindeer (Rangifer tarandus tarandus) to deduce the potential risk to human health through modern reindeer herding. In total, 2,243 faecal samples of reindeer from northern regions of Finland and Norway were examined for potentially enteropathogenic bacteria (Campylobacter species, Enterococcus species, Escherichia coli, Salmonella species and Yersinia species) and parasites (Cryptosporidium species) in accordance with standard procedures. Escherichia coli were isolated in 94.7%, Enterococcus species in 92.9%, Yersinia species in 4.8% of the samples and Campylobacter species in one sample only (0.04%). Analysis for virulence factors in E. coli and Yersinia species revealed no pathogenic strains. Neither Salmonella species nor Cryptosporidium oocysts were detected. The public health risk due to reindeer husbandry concerning zoonotic diseases included in this study has to be considered as very low at present but a putative epidemiological threat may arise when herding conditions are changed with respect to intensification and crowding.
Fecal samples collected from 470 slaughtered reindeer 6 to 7 months of age were screened by real-time PCR (after enrichment) for Shiga toxin genes (stx) and then for Escherichia coli serogroup O157. Shiga toxin genes were found frequently (>30% of samples), and serogroup O157 was detected in 20% of the stx-positive samples. From these samples, a total of 25 E. coli O157:H(-) isolates (nonmotile but PCR positive for fliCH7) were obtained. Twenty-four of these E. coli O157:H(-) isolates did not ferment sorbitol and originated from one geographic area. These 24 isolates belonged to the multilocus sequence type 11, typical for Shiga toxin-producing E. coli (STEC) O157:H7 and O157:H(-), and harbored genes stx1a, stx2c, eae, and hlyA; the stx2c subtype has been associated with high virulence. In contrast, one E. coli O157:H(-) isolate (multilocus sequence type 11) did ferment sorbitol, lacked Shiga toxin genes, but was positive for eae, hlyA, and sfpA. This isolate closely resembled an STEC that has lost its Shiga toxin genes. Additional examination revealed that reindeer can be colonized by various other STEC isolates; 21 non-O157 STEC isolates belonged to four multilocus sequence types, harbored stx1a (8 isolates) or stx2b (13 isolates), and in the stx2b-positive isolates the recently described new allelic variants (subAB2-2 and subAB2-3) for subtilase cytotoxin were identified. Hence, slaughtered semidomesticated Finnish reindeer might constitute a little known reservoir for STEC O157:H7/H(-) and other serogroups, and the risk of direct or indirect transmission of these pathogens from reindeer to humans and domestic livestock must not be overlooked.
The aim of this study was to compare conventional 16S rRNA gene PCR, real-time 16S rRNA gene PCR and real-time Mycoplasma genitalium adhesin protein (MgPa) gene PCR as detection methods for M. genitalium infection. The study also determined the prevalence of M. genitalium in male and female patients attending a sexually transmitted infections clinic in a rural area in the west of Sweden. First void urine (FVU) and/or urethral swabs were collected from 381 men, and FVU and/or cervical swabs and/or urethral swabs were collected from 298 women. A total of 213 specimens were used in the PCR comparative study: 98 consecutively sampled specimens from patients enrolled in the prevalence study, 36 consecutively sampled specimens from patients with symptoms of urethritis and 79 specimens from patients positive for M. genitalium by real-time MgPa gene PCR in the prevalence study. A true-positive M. genitalium DNA specimen was defined as either a specimen positive in any two PCR assays or a specimen whose PCR product was verified by DNA sequencing. The prevalence of M. genitalium infection in men and women was 27/381 (7.1 %) and 23/298 (7.7 %), respectively. In the PCR comparative study, M. genitalium DNA was detected in 61/76 (80.3 %) of true-positive specimens by conventional 16S rRNA gene PCR, in 52/76 (68.4 %) by real-time 16S rRNA gene PCR and in 74/76 (97.4 %) by real-time MgPa gene PCR. Real-time MgPa gene PCR thus had higher sensitivity compared with conventional 16S rRNA gene PCR and had considerably increased sensitivity compared with real-time 16S rRNA gene PCR for detection of M. genitalium DNA. Real-time MgPa gene PCR is well suited for the clinical diagnosis of M. genitalium.
Investigation of an Escherichia coli O145 outbreak in a child day-care centre--extensive sampling and characterization of eae- and stx1-positive E. coli yields epidemiological and socioeconomic insight.
On October 29th 2009 the health authorities in the city of Trondheim, Norway were alerted about a case of Shiga toxin-positive E. coli (STEC) O145 in a child with bloody diarrhoea attending a day-care centre. Symptomatic children in this day-care centre were sampled, thereby identifying three more cases. This initiated an outbreak investigation.
A case was defined as a child attending the day-care centre, in whom eae- and stx1- but not stx2-positive E. coli O145:H28 was diagnosed from a faecal sample, with multilocus variable number of tandem repeat analysis (MLVA) profile identical to the index isolate. All 61 children, a staff of 14 in the day-care centre, and 74 close contacts submitted faecal samples. Staff and parents were interviewed about cases' exposure to foods and animals. Faecal samples from 31 ewes from a sheep herd to which the children were exposed were analyzed for E. coli O145.
Sixteen cases were identified, from which nine presented diarrhoea but not haemolytic uremic syndrome (HUS). The attack rate was 0.26, and varied between age groups (0.13-0.40) and between the three day-care centre departments (0.20-0.50), and was significantly higher amongst the youngest children. Median duration of shedding was 20 days (0-71 days). Children were excluded from the day-care centre during shedding, requiring parents to take compassionate leave, estimated to be a minimum total of 406 days for all cases. Atypical enteropathogenic E. coli (aEPEC) were detected among 14 children other than cases. These isolates were genotypically different from the outbreak strain. Children in the day-care centre were exposed to faecal pollution from a sheep herd, but E. coli O145 was not detected in the sheep.
We report an outbreak of stx1- and eae-positive STEC O145:H28 infection with mild symptoms among children in a day-care centre. Extensive sampling showed occurrence of the outbreak strain as well as other STEC and aEPEC strains in the outbreak population. MLVA-typing of the STEC-isolates strongly indicates a common source of infection. The study describes epidemiological aspects and socioeconomic consequences of a non-O157 STEC outbreak, which are less commonly reported than O157 outbreaks.
Notes
Cites: Appl Environ Microbiol. 1996 Jul;62(7):2567-708779595
Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), ?6 (n = 4), ?25 (n = 5), ?26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and sulphonamides dfrA17-aadA5 and dfrA12-orfF-aadA2. One isolate ETEC_Ef-6 was found to be a multidrug-resistant (MDR) pathogen that carried both the beta-lactamase gene and class 1 integron. These data suggest the circulation of ETEC in Russia. Further investigations are necessary to study the spread of the revealed ETEC sequence types (STs) and serotypes. Their role in the etiology of diarrhea should be also estimated.
The aim of the study was to determine inter- and intrapatient variation of Helicobacter pylori strains based on genomic fingerprinting and cagA (cytotoxin-associated gene A) status. Ten bacterial colonies from each of 10 patients with gastric cancer (GC), 10 with duodenal ulcer (DU), and 10 with gastritis (GI) were used. The presence of the putative adhesin gene, the cagA gene, and the strain specific banding pattern obtained by arbitrary primed (AP-) PCR was analyzed. Genomic fingerprinting showed extensive interpatient variation, but the banding patterns obtained from colonies from the same patient were always identical (intrapatient variation). In five patients, the cagA status varied between the colonies despite identical banding patterns. Among patients in a developed country such as Sweden, the proportion with multiple-strain infection of H. pylori is low, but subclones with differing cagA status exist within the strain.
[PHENOTYPICAL CHARACTERISTICS AND GENETICAL DETERMINANTS OF PATHOGENICITY OF STAPHYLOCOCCUS AUREUS, ISOLATED FROM BACTERIAL CARRIERS, RESIDING ON THE TERRITORIES WITH VARIOUS LEVELS OF ANTHROPOGENIC POLLUTION OF AIR ENVIRONMENT].
Comparative phenotypical and genetical evaluation of pathogenic potential of Staphylococcus aureus strains, isolated from resident bacterial carriers, residing on the territories with anthropogenic pollution of air environment of varying intensity.
S. aureus, isolated 3 times from mucous membrane of the anterior of nose from 210 children, were the object of the study. Anti-carnosine activity and biofilm formation was determined by a photometric method, antibiotics resistance--by a disc diffusion method. lukS, lukF, sec 3, clfA, clfB, agr and mecA gene detection, that are associated with S. aureus, was carried out by PCR.
S. aureus strains, isolated from children, residing on the territories with a high level of anthropogenic pollution of air environment, were characterized by antibiotics resistance, higher values of anti-carnosine activity, 2 times more frequently formed biofilms with higher values of the parameter. clfA and clfB genes, that determine colonization of mucous membranes, and agr gene were detected in all the studied S. aureus strains, lukF and sec 3 genes were detected in 20-40% of the strains, isolated from children, residing on both territories. mecA and lukS genetical determinants were not detected.
S. aureus, isolated from children, residing on the territories with high levels of anthropogenic pollution of air environment; were characterized by higher values of the studied factors of persistence and stability against antibiotics. Genetical determinants of pathogenicity were not detected in S. aureus, isolated from individuals, residing on both territories.
Phylogenetic backgrounds and virulence profiles of atypical enteropathogenic Escherichia coli strains from a case-control study using multilocus sequence typing and DNA microarray analysis.
Atypical enteropathogenetic Escherichia coli (EPEC) strains are frequently detected in children with diarrhea but are also a common finding in healthy children. The aim of this study was to compare the phylogenetic ancestry and virulence characteristics of atypical (eae positive, stx and bfpA negative) EPEC strains from Norwegian children with (n = 37) or without (n = 19) diarrhea and to search for an association between phylogenetic ancestry and diarrhea. The strains were classified in phylogenetic groups by phylogenetic marker genes and in sequence types (STs) by multilocus sequence typing. Phylogenetic ancestry was compared to virulence characteristics based on DNA microarray analysis. Serotyping and pulsed-field gel electrophoresis (PFGE) were also performed. All four phylogenetic groups, 26 different STs, and 20 different clonal groups were represented among the 56 atypical EPEC strains. The strains were separated into three clusters by overall virulence gene profile; one large cluster with A, B1, and D strains and two clusters with group B2 strains. There was considerable heterogeneity in the PFGE profiles and serotypes, and almost half of the strains were O nontypeable. The efa1/lifA gene, previously shown to be statistically linked with diarrhea in this strain collection (J. E. Afset et al., J. Clin. Microbiol. 44:3703-3711, 2006), was present in 8 of 26 STs. The two phylogenetic groups B1 and D were weakly associated with diarrhea (P = 0.06 and P = 0.09, respectively). In contrast, group B2 was isolated most frequently from healthy controls (P = 0.05). In conclusion, the atypical EPEC strains were heterogeneous both phylogenetically and by virulence profile. Phylogenetic ancestry was less useful as a predictor of diarrhea than were specific virulence genes.
