The nasopharyngeal commensal bacteria Streptococcus pneumoniae is also a frequent cause of serious infections. Nasopharyngeal colonisation with S. pneumoniae inhibits subsequent re-colonisation by inducing Th17-cell adaptive responses, whereas vaccination prevents invasive infections by inducing antibodies to S. pneumoniae capsular polysaccharides. In contrast, protection against invasive infection after nasopharyngeal colonisation with mutant S. pneumoniae strains was associated with antibody responses to protein antigens. The role of colonisation-induced Th17-cell responses during subsequent invasive infections is unknown. Using mouse models, we show that previous colonisation with S. pneumoniae protects against subsequent lethal pneumonia mainly by preventing bacteraemia with a more modest effect on local control of infection within the lung. Previous colonisation resulted in CD4-dependent increased levels of Th17-cell cytokines during subsequent infectious challenge. However, mice depleted of CD4 cells prior to challenge remained protected against bacteraemia, whereas no protection was seen in antibody deficient mice and similar protection could be achieved through passive transfer of serum. Serum from colonised mice but not antibody deficient mice promoted phagocytosis of S. pneumoniae, and previously colonised mice were able to rapidly clear S. pneumoniae from the blood after intravenous inoculation. Thus, despite priming for a Th17-cell response during subsequent infection, the protective effects of prior colonisation in this model was not dependent on CD4 cells but on rapid clearance of bacteria from the blood by antibody-mediated phagocytosis. These data suggest that whilst nasopharyngeal colonisation induces a range of immune responses, the effective protective responses depend upon the site of subsequent infection.
Despite the sparsity of Pseudomonas aeruginosa in the environment colonisation and infection with this organism was found at several sites by selective culture in 20 out of 46 patients in an intensive therapy unit. Three patients developed Ps aeruginosa pneumonia. Serial serogrouping and phage typing identified multiple strains in the unit and in the same patient. Rectal carriage occurred in 16 patients but rectal strains did not subsequently appear in tracheal aspirates; strains varied in their affinity for the upper respiratory tract. Colonisation was not directly related to length of stay and was detected in 16 of those colonised within 24 hours of admission. In intubated patients, who were colonised more frequently than those not intubated, upper respiratory tract colonisation correlated strongly with low initial arterial pH values. Personnel were probably responsible for cross infection among patients when the unit was busy. Strain differences and the susceptibility of patients also influenced colonisation and infection. Elimination of major reservoirs of Ps aeruginosa and compliance with procedures to control cross infection remain essential if patients in hospital are to escape colonisation by the organism.
Candida albicans systemic dissemination in immunocompromised patients is thought to develop from initial gastrointestinal (GI) colonisation. It is unclear what components of the innate immune system are necessary for preventing C. albicans dissemination from the GI tract, but studies in mice have indicated that both neutropenia and GI mucosal damage are crucial for allowing widespread invasive C. albicans disease. Mouse models, however, provide limited applicability to genome-wide screens for pathogen or host factors – factors that might influence systemic dissemination following GI colonisation. For this reason we developed a Drosophila model to study intestinal infection by Candida. We found that commensal flora aided host survival following GI infection. Candida provoked extensive JNK-mediated death of gut cells and induced antimicrobial peptide expression in the fat body. From the side of the host, nitric oxide and blood cells influenced systemic antimicrobial responses. The secretion of SAP4 and SAP6 (secreted aspartyl proteases) from Candida was also essential for activating systemic Toll-dependent immunity.
A biochemical typing method is described for Klebsiella pneumoniae (sensu lato) and Enterobacter aerogenes. The technique depends on differences in metabolism of five carbon substrates--glycerol, inositol, lactose, glucose, and xylose--at two concentrations. Reproducibility is satisfactory and is monitored by the incorporation of control klebsiellae of known biotype. The method has been used for 12 months in the surveillance of urinary tract colonisation in this hospital. Gut carriage of klebsiellae, implicated by several workers as a source of infection, was common among staff and new admissions. Many biotypes were represented which were sensitive to most antibiotic except ampicillin. Klebsiellae, all multiply resistant, were isolated most frequently from urine specimens in two orthopaedic wards. In a longitudinal study in these wards, a sequential dominance in urinary tract colonisation by two klebsiella biotypes was shown, which suggested the presence of cross infection or an environmental reservoir. Confirmatory evidence was obtained from capsular serotypes and R-factor studies.
Klebsiella pneumoniae is an opportunistic pathogen responsible for nosocomial infections that initially colonize the intestinal tract of patients. Signature-tagged mutagenesis was used to identify genes required for this function. A library of 2,200 mutants was analyzed for the inability of the mutants to survive in a murine model of intestinal colonization and to adhere to human intestinal cells (Int-407) in vitro. Twenty-nine attenuated mutants were selected for further analyses after competition assays against the wild-type strain. Whatever the screening model, most of the transposon insertions occurred in genes involved in metabolic pathways, membrane transport, DNA metabolism, transcriptional regulation, and unknown functions. Only one mutant was attenuated in both the murine colonization and the in vitro adhesion models, and the sequence disrupted by the transposon had homology to adhesin-encoding genes of Haemophilus sp.
Group B streptococci (GBS) are a major cause of pneumonia, sepsis, and meningitis in newborns and infants. GBS initiate infection of the lung by colonizing mucosal surfaces of the respiratory tract; adherence of the bacteria to host cells is presumed to be the initial step in and prerequisite for successful colonization (G. S. Tamura, J. M. Kuypers, S. Smith, H. Raff, and C. E. Rubens, Infect. Immun. 62:2450-2458, 1994). We have performed a genome-wide screen to identify novel genes of GBS that mediate adherence to fibronectin. A shotgun phage display library was constructed from chromosomal DNA of a serotype Ia GBS strain and affinity selected on immobilized fibronectin. DNA sequence analysis of different clones identified 19 genes with homology to known bacterial adhesin genes, virulence genes, genes involved in transport or metabolic processes, and genes with yet-unknown function. One of the isolated phagemid clones showed significant homology to the gene (scpB) for the GBS C5a peptidase, a surface-associated serine protease that specifically cleaves the complement component C5a, a chemotaxin for polymorphonuclear leukocytes. In this work we have demonstrated that affinity-purified recombinant ScpB and a peptide ScpB fragment (ScpB-PDF), similar to the peptide identified in the phagemid, bound fibronectin in a concentration-dependent manner. Adherence assays to fibronectin were performed, comparing an isogenic scpB mutant to the wild-type strain. Approximately 50% less binding was observed with the mutant than with the wild-type strain. The mutant phenotype could be fully restored by in trans complementation of the mutant with the cloned wild-type scpB gene, providing further evidence for the role of ScpB in fibronectin adherence. Our results suggest that C5a peptidase is a bifunctional protein, which enzymatically cleaves C5a and mediates adherence to fibronectin. Since binding of fibronectin has been implicated in attachment and invasion of eukaryotic cells by streptococci, our results may imply a second important role for this surface protein in the pathogenesis of GBS infections.
A novel in vivo expression technology (IVET) was performed to identify Klebsiella pneumoniae CG43 genes that are specifically expressed during infection of BALB/c mice. The IVET employed a UDP glucose pyrophosphorylase (galU)-deficient mutant of K. pneumoniae which is incapable of utilizing galactose and synthesizing capsular polysaccharide, as demonstrated by its low virulence to BALB/c mice and a white nonmucoid colony morphology on MacConkey-galactose agar. By using a functional galU gene as the reporter, an IVE promoter could render the galU mutant virulent while maintaining the white nonmucoid colony phenotype. A total of 20 distinct sequences were obtained through the in vivo selection. Five of them have been identified previously as virulence-associated genes in other pathogens, while another five with characterized functions are involved in regulation and transportation of nutrient uptake, biosynthesis of isoprenoids, and protein folding. No known functions have been attributed to the other 10 sequences. We have also demonstrated that 2 of the 20 IVE genes turn on under iron deprivation, whereas the expression of another five genes was found to be activated in the presence of paraquat, a superoxide generator.
Lactobacillus reuteri strain 100-23 together with a Lactobacillus-free mouse model, provides a system with which the molecular traits underpinning bacterial commensalism in vertebrates can be studied. A polysaccharide was extracted from sucrose-containing liquid cultures of strain 100-23. Chemical analysis showed that this exopolysaccharide was a levan (β-2, 6-linked fructan). Mutation of the fructosyl transferase (ftf) gene resulted in loss of exopolysaccharide production. The ftf mutant was able to colonise the murine gastrointestinal tract in the absence of competition, but colonisation was impaired in competition with the wild type. Biofilm formation by the mutant on the forestomach epithelial surface was not impaired and the matrix between cells was indistinguishable from that of the wild type in electron micrographs. Colonisation of the mouse gut by the wild-type strain led to increased proportions of regulatory T cells (Foxp3+) in the spleen, whereas colonisation by the ftf mutant did not. Survival of the mutant in sucrose-containing medium was markedly reduced relative to the wild type. Comparison of the genomic ftf loci of strain 100-23 with other L. reuteri strains suggested that the ftf gene was acquired by lateral gene transfer early in the evolution of the species and subsequently diversified at accelerated rates. Levan production by L. reuteri 100-23 may represent a function acquired by the bacterial species for life in moderate to high-sucrose extra-gastrointestinal environments that has subsequently been diverted to novel uses, including immunomodulation, that aid in colonisation of the murine gut.
Lactobacillus reuteri; exopolysaccharide; survival; regulatory T cells; evolution
Klebsiella pneumoniae is an important gram-negative opportunistic pathogen causing primarily urinary tract infections, respiratory infections, and bacteraemia. The ability of bacteria to form biofilms on medical devices, e.g. catheters, has a major role in development of many nosocomial infections. Most clinical K. pneumoniae isolates express two types of fimbrial adhesins, type 1 fimbriae and type 3 fimbriae. In this study, we characterized the role of type 1 and type 3 fimbriae in K. pneumoniae biofilm formation.
Isogenic fimbriae mutants of the clinical K. pneumoniae isolate C3091 were constructed, and their ability to form biofilm was investigated in a flow cell system by confocal scanning laser microscopy. The wild type strain was found to form characteristic biofilm and development of K. pneumoniae biofilm occurred primarily by clonal growth, not by recruitment of planktonic cells. Type 1 fimbriae did not influence biofilm formation and the expression of type 1 fimbriae was found to be down-regulated in biofilm forming cells. In contrast, expression of type 3 fimbriae was found to strongly promote biofilm formation.
By use of well defined isogenic mutants we found that type 3 fimbriae, but not type 1 fimbriae, strongly promote biofilm formation in K. pneumoniae C3091. As the vast majority of clinical K. pneumoniae isolates express type 3 fimbriae, this fimbrial adhesin may play a significant role in development of catheter associated K. pneumoniae infections.
Neural crest cells (NCCs) form at the dorsal margin of the neural tube and migrate along distinct pathways throughout the vertebrate embryo to generate multiple cell types. A subpopulation of vagal NCCs invades the foregut and colonises the entire gastrointestinal tract to form the enteric nervous system (ENS). The colonisation of embryonic gut by NCCs has been studied extensively in chick embryos, and genetic studies in mice have identified genes crucial for ENS development, including Ret. Here, we have combined mouse embryo and organotypic gut culture to monitor and experimentally manipulate the progenitors of the ENS. Using this system, we demonstrate that lineally marked intestinal ENS progenitors from E11.5 mouse embryos grafted into the early vagal NCC pathway of E8.5 embryos colonise the entire length of the gastrointestinal tract. By contrast, similar progenitors transplanted into Ret-deficient host embryos are restricted to the proximal foregut. Our findings establish an experimental system that can be used to explore the interactions of NCCs with their cellular environment and reveal a previously unrecognised non-cell-autonomous effect of Ret deletion on ENS development.
Enteric nervous system; Neural crest; Ret
HIV infected adults have increased susceptibility to bacterial pneumonia but the underlying immune defect is poorly understood. We tested the hypothesis that HIV infection might be associated with increased bacterial colonisation of distal airways by nasal flora, which would then predispose patients to bacterial pneumonia.
Healthy volunteer adults with normal chest radiographs were recruited. Bronchoscopy was carried out and uncontaminated mucosal samples were collected from proximal and distal sites in the large airways using a protected specimen brush. Samples were cultured to detect typical respiratory tract colonising organisms, and the proportion of samples found to contain colonising bacteria compared between HIV infected and uninfected subjects using non-parametric tests.
Forty-nine subjects were studied of whom 27 were HIV infected. Colonising bacteria were identified in the nasopharynx of all subjects including Streptococcus pneumoniae in 6/49 subjects (5 HIV uninfected). Colonising bacteria were found in the distal airway of 6 subjects (3/27 HIV infected vs 3/22 HIV uninfected ; χ2 = 0.07, p = 0.8). Streptococcus pneumoniae was identified in the trachea of all subjects with nasal colonisation but in the distal airway of only 1 subject.
There was no evidence to support a hypothesis of increased airway bacterial colonisation in healthy HIV infected subjects.
The colonization of the gastrointestinal tract of patients by the opportunistic gram-negative bacillus Klebsiella pneumoniae generally occurs prior to the development of nosocomial infections. Mutant strain C-81 was isolated owing to its reduced capacity to colonize the digestive tract in a murine model following transposon mutagenesis (N. Maroncle, D. Balestrino, C. Rich, and C. Forestier, Infect. Immun. 70:4729-4734, 2002). Nucleotide sequence analysis showed that the transposon had inserted into the first open reading frame, eefA, of a three-gene locus (eefABC) whose homologue encodes a tripartite efflux pump in Enterobacter aerogenes (M. Masi, J. M. Pages, C. Villard, and E. Pradel, J. Bacteriol. 187:3894-3897, 2005), and this operon includes an additional short (183-bp) potential open reading frame, eefX, upstream of eefA. In vivo assays showed that a ΔeefA isogenic mutant strain normally colonized the gastrointestinal tract in single-strain tests but was significantly impaired in competition against wild-type strain LM21. Although the cecum was the compartment with the highest number of CFU, the ΔeefA mutant also was detected in the stomach in numbers smaller than those of the wild-type strain. The expression of this potential efflux pump could not be linked to any antimicrobial drug resistance phenotype, but it conferred on the bacteria an acid tolerance response to inorganic acid. The expression of the eef promoter region, measured via a lacZ reporter construction, was slightly induced by an acidic environment and also by hyperosmolarity but not by the presence of bile salts. These results suggest that an efflux pump can confer measurable ecological benefits on K. pneumoniae in an environment with high competition potential.
Backgrounds and aims
Controversy still exists as to whether gastrointestinal colonisation by Candida albicans contributes to aggravation of atopic dermatitis. We hypothesised that Candida colonisation promotes food allergy, which is known to contribute to a pathogenic response in atopic dermatitis. We tested this using a recently established murine Candida colonisation model.
Candida colonisation in the gastrointestinal tract was established by intragastric inoculation with C albicans in mice fed a synthetic diet. To investigate sensitisation against food antigen, mice were intragastrically administered with ovalbumin every other day for nine weeks, and antiovalbumin antibody titres were measured weekly. To examine gastrointestinal permeation of food antigen, plasma concentrations of ovalbumin were measured following intragastric administration of ovalbumin.
Ovalbumin specific IgG and IgE titres were higher in BALB/c mice with Candida colonisation than in normal mice. Gastrointestinal permeation of ovalbumin was enhanced by colonisation in BALB/c mice. Histological examination showed that colonisation promoted infiltration and degranulation of mast cells. Candida colonisation did not enhance ovalbumin permeation in mast cell deficient W/Wv mice but did in congenic littermate control +/+ mice. Reconstitution of mast cells in W/Wv mice by transplantation of bone marrow derived mast cells restored the ability to increase ovalbumin permeation in response to Candida colonisation.
These results suggest that gastrointestinal Candida colonisation promotes sensitisation against food antigens, at least partly due to mast cell mediated hyperpermeability in the gastrointestinal mucosa of mice.
Candida albicans ; food allergy; atopic dermatitis; mast cells
Invasive community-onset staphylococcal disease has emerged worldwide associated with Panton-Valentine leucocidin (PVL) toxin. Whether PVL is pathogenic or an epidemiological marker is unclear. We investigate the role of PVL in disease, colonisation, and clinical outcome.
We searched Medline and Embase for original research reporting the prevalence of PVL genes among Staphylococcus aureus pneumonia, bacteraemia, musculoskeletal infection, skin and soft-tissue infection, or colonisation published before Oct 1, 2011. We calculated odds ratios (ORs) to compare patients with PVL-positive colonisation and each infection relative to the odds of PVL-positive skin and soft-tissue infection. We did meta-analyses to estimate odds of infection or colonisation with a PVL-positive strain with fixed-effects or random-effects models, depending on the results of tests for heterogeneity.
Of 509 articles identified by our search strategy, 76 studies from 31 countries met our inclusion criteria. PVL strains are strongly associated with skin and soft-tissue infections, but are comparatively rare in pneumonia (OR 0·37, 95% CI 0·22–0·63), musculoskeletal infections (0·44, 0·19–0·99), bacteraemias (0·10, 0·06–0·18), and colonising strains (0·07, 0·01–0·31). PVL-positive skin and soft-tissue infections are more likely to be treated surgically than are PVL-negative infections, and children with PVL-positive musculoskeletal disease might have increased morbidity. For other forms of disease we identified no evidence that PVL affects outcome.
PVL genes are consistently associated with skin and soft-tissue infections and are comparatively rare in invasive disease. This finding challenges the view that PVL mainly causes invasive disease with poor prognosis. Population-based studies are needed to define the role of PVL in mild, moderate, and severe disease and to inform control strategies.
Infection with Helicobacter species has been associated with the development of mucosal inflammation and inflammatory bowel disease (IBD) in several mouse models. However, consensus regarding the role of Helicobacter as a model organism to study microbial‐induced IBD is confounded by the presence of a complex colonic microbiota.
To investigate the kinetics and inflammatory effects of immune system activation to commensal bacteria following H bilis colonisation in gnotobiotic mice.
C3H/HeN mice harbouring an altered Schaedler flora (ASF) were selectively colonised with H bilis and host responses were investigated over a 10‐week period. Control mice were colonised only with the defined flora (DF). Tissues were analysed for gross/histopathological lesions, and bacterial antigen‐specific antibody and T‐cell responses.
Gnotobiotic mice colonised with H bilis developed mild macroscopic and microscopic lesions of typhlocolitis beginning 3 weeks postinfection. ASF‐specific IgG responses were demonstrable within 3 weeks, persisted throughout the 10‐week study, and presented as a mixed IgG1:IgG2a profile. Lymphocytes recovered from the mesenteric lymph node of H bilis‐colonised mice produced increased levels of interferon γ, tumour necrosis factor α (TNFα), interleukin 6 (IL6) and IL12 in response to stimulation with commensal‐ or H bilis‐specific bacterial lysates. In contrast, DF mice not colonised with H bilis did not develop immune responses to their resident flora and remained disease free.
Colonisation of gnotobiotic C3H/HeN mice with H bilis perturbs the host's response to its resident flora and induces progressive immune reactivity to commensal bacteria that contributes to the development of immune‐mediated intestinal inflammation.
The mechanisms which enable entry into cultured human epithelial cells by Klebsiella pneumoniae were compared with those of Salmonella typhi Ty2. K. pneumoniae 3091, isolated from a urine sample of a patient with a urinary tract infection, invaded human epithelial cells from the bladder and ileocecum and persisted for days in vitro. Electron microscopic studies demonstrated that K. pneumoniae was always contained in endosomes. The internalization mechanism(s) triggered by K. pneumoniae was studied by invasion assays conducted with different inhibitors that act on prokaryotic and eukaryotic cell structures and processes. Chloramphenicol inhibition of bacterial uptake revealed that bacterial de novo protein synthesis was essential for efficient invasion by K. pneumoniae and S. typhi. Interference with receptor-mediated endocytosis by g-strophanthin or monodansylcadaverine and inhibition of endosome acidification by monensin reduced the number of viable intracellular K. pneumoniae cells, but not S. typhi cells. The depolymerization of microfilaments by cytochalasin D inhibited the uptake of both bacteria. Microtubule depolymerization caused by colchicine, demecolcine, or nocodazole and the stabilization of microtubules with taxol reduced only the invasion ability of K. pneumoniae. S. typhi invasion was unaffected by microtubule depolymerization or stabilization. These data suggest that the internalization mechanism triggered by K. pneumoniae 3091 is strikingly different from the solely microfilament-dependent invasion mechanism exhibited by many of the well-studied enteric bacteria, such as enteroinvasive Escherichia coli, Salmonella, Shigella, and Yersinia strains.
Klebsiella pneumoniae is a common cause of septicemia and urinary tract infections. The PCR-supported genomic subtractive hybridization was employed to identify genes specifically present in a virulent strain of K. pneumoniae. Analysis of 25 subtracted DNA clones has revealed 19 distinct nucleotide sequences. Two of the sequences were found to be the genes encoding the transposase of Tn3926 and a capsule polysaccharide exporting enzyme. Three sequences displayed moderate homology with bvgAS, which encodes a two-component signal transduction system in Bordetella pertussis. The rest of the sequences did not exhibit homology with any known genes. The distribution of these novel sequences varied greatly in K. pneumoniae clinical isolates, reflecting the heterogeneous nature of the K. pneumoniae population.
A major bottleneck in understanding zoonotic pathogens has been the analysis of pathogen co-infection dynamics. We have addressed this challenge using a novel direct sequencing approach for pathogen quantification in mixed infections. The major zoonotic food-borne pathogen Campylobacter jejuni, with an important reservoir in the gastrointestinal (GI) tract of chickens, was used as a model. We investigated the co-colonisation dynamics of seven C. jejuni strains in a chicken GI infection trial. The seven strains were isolated from an epidemiological study showing multiple strain infections at the farm level. We analysed time-series data, following the Campylobacter colonisation, as well as the dominant background flora of chickens. Data were collected from the infection at day 16 until the last sampling point at day 36. Chickens with two different background floras were studied, mature (treated with Broilact, which is a product consisting of bacteria from the intestinal flora of healthy hens) and spontaneous. The two treatments resulted in completely different background floras, yet similar Campylobacter colonisation patterns were detected in both groups. This suggests that it is the chicken host and not the background flora that is important in determining the Campylobacter colonisation pattern. Our results showed that mainly two of the seven C. jejuni strains dominated the Campylobacter flora in the chickens, with a shift of the dominating strain during the infection period. We propose a model in which multiple C. jejuni strains can colonise a single host, with the dominant strains being replaced as a consequence of strain-specific immune responses. This model represents a new understanding of C. jejuni epidemiology, with future implications for the development of novel intervention strategies.
Pathogenic bacteria that can be transferred from animals to humans represent a highly potent human health hazard. Understanding the ecology of these pathogens in the animal host is of fundamental importance. A major analytical challenge, however, is the fact that individual animal hosts can be colonised by multiple strains of a given pathogen. We have addressed this challenge by developing a novel high-throughput approach for analyses of mixed strain infections. We chose Campylobacter jejuni colonisation of the chicken gastrointestinal (GI) tract as a model. C. jejuni is a major cause of food-borne disease in humans, and chickens are considered a main reservoir from which this bacterium may enter the food chain. We analysed the co-colonisation of seven C. jejuni strains in two groups of chickens with very different background GI microfloras. We found that mainly two of the C. jejuni strains colonised the chickens, with a shift in the dominant coloniser during the infection period. The C. jejuni colonisation pattern, however, was little affected by the dominating GI microflora. We propose a model where the chicken immune response is the important determinant for C. jejuni colonisation, and suggest that multiple strain colonisation could be a way of maintaining stable infections in the animal host. This new knowledge is very important for future development of novel intervention strategies to prevent C. jejuni from entering the human food chain.
Of the babies admitted to the Special Care Baby Unit of the Royal Free Hospital over 20 months, 10·2% were infected or colonised by klebsiella. The fluorescent antibody technique was used to identify epidemics caused by three strains: capsular type 8 K. aerogenes, type 68 K. oxytoca, or type 13 K. aerogenes, each of which was predominant at a different time, exhibited a difference in virulence, and showed a predilection for different sites of infection. Intestinal colonisation was frequently followed by the presence of sepsis in other sites by the same capsular type. Antibiotic administration led to a higher incidence of klebsiella infection, while the widespread use of compounds containing hexachlorophane could have contributed to skin colonisation and infection by klebsiella. An environmental survey indicated that 1% Hycolin failed to disinfect the incubators, that the babies were the reservoirs of the organisms, and that transmission was due to inadequate hand-washing of nurses and mothers. The mothers were found to have been uninformed of hygienic techniques. They were observed in various practices which could have contributed to the spread of the organism, including contaminating communal areas and handling babies other than their own. It has been recommended that the mothers of premature infants be instructed in the hygienic measures required in dealing with this susceptible population and that the nursing and medical staff be more strict in their own observance of these procedures.
pneumonia (VAP) caused by Pseudomonas aeruginosa is
usually preceded by colonisation of the respiratory tract. During
outbreaks, colonisation with P aeruginosa is mainly derived from exogenous sources. The relative importance of different pathways of colonisation of P aeruginosa has rarely been
determined in non-epidemic settings.
METHODS—In order to determine the
importance of exogenous colonisation, all isolates of P
aeruginosa obtained by surveillance and clinical cultures from
two identical intensive care units (ICUs) were genotyped with pulsed
field gel electrophoresis.
RESULTS—A total of 100 patients were
studied, 44 in ICU 1 and 56 in ICU 2. Twenty three patients were
colonised with P aeruginosa, seven at the start of the
study or on admission and 16 of the remaining 93 patients became
colonised during the study. Eight patients developed VAP due to P
aeruginosa. The incidence of respiratory tract colonisation and
VAP with P aeruginosa in our ICU was similar to that
before and after the study period, and therefore represents an endemic
situation. Genotyping of 118 isolates yielded 11 strain types: eight in
one patient each, two in three patients each, and one type in eight
patients. Based on chronological evaluation and genotypical identity of
isolates, eight cases of cross-colonisation were identified. Eight
(50%) of 16 episodes of acquired colonisation and two (25%) of eight
cases of VAP due to P aeruginosa seemed to be the result
CONCLUSIONS—Even in non-epidemic settings
cross-colonisation seems to play an important part in the epidemiology
of colonisation and infection with P aeruginosa.
The hygiene hypothesis postulates that the increase in atopic diseases may in part be due to diminished exposure to microorganisms. But it is unknown which type of infection does render protection. An epidemiological study was conducted in Leipzig, Germany, and its rural county, involving 3347 school starters. Two types of infection were considered: (1) gastrointestinal colonisation (Helicobacter pylori detection using in vivo [13C] urea breath test) and (2) respiratory infections (physician‐diagnosed lower (bronchitis) and upper (common cold) respiratory infections). H pylori colonisation was selected because it is very common and plays an important role in gastrointestinal disorders. Atopic eczema was selected as the (allergic) target variable because of its high frequency in the age of the study participants. The results, adjusted for relevant confounders, showed a significant inverse association between H pylori infection and eczema (adjusted odds ratio (aOR) = 0.31, p = 0.006) in children not predisposed to atopy. In contrast, bronchitis increased the risk of eczema (aOR = 1.98, p<0.001). Bacterial digestive tract colonisation (infection) seems to protect against eczema in comparison with the effect of respiratory tract infections. The hygiene hypothesis may be better explained when gastrointestinal and respiratory infections are subtly differentiated.
An outbreak of extended spectrum β lactamase producing
Klebsiella pneumoniae (ESBLKp) in a neonatal unit was
controlled using simple measures. Normally, the control of such
infections can be time consuming and expensive. Seven cases of
septicaemia resulted in two deaths. ESBLKp isolates were subtyped by
pulsed field gel electrophoresis, and four of the five isolates typed
were identical. Control of the outbreak was achieved by altered empiric
antibiotic treatment for late onset sepsis and prevention of cross
infection by strict attention to hand washing. Widespread colonisation
of babies in the unit was presumed, so initial surveillance cultures were not performed. No further episodes of sepsis occurred.
Salmonella enterica is a facultative intracellular pathogen of worldwide importance. Over 2,500 serovars exist and infections in humans and animals may produce a spectrum of symptoms from enteritis to typhoid depending on serovar- and host-specific factors. S. Enteritidis is the most prevalent non-typhoidal serovar isolated from humans with acute diarrhoeal illness in many countries. Human infections are frequently associated with direct or indirect contact with contaminated poultry meat or eggs owing to the ability of the organism to persist in the avian intestinal and reproductive tract. The molecular mechanisms underlying colonisation of poultry by S. Enteritidis are ill-defined. Targeted and genome-wide mutagenesis of S. Typhimurium has revealed conserved and host-specific roles for selected fimbriae in intestinal colonisation of different hosts. Here we report the first systematic analysis of each chromosomally-encoded major fimbrial subunit of S. Enteritidis in intestinal colonisation of chickens.
The repertoire, organisation and sequence of the fimbrial operons within members of S. enterica were compared. No single fimbrial locus could be correlated with the differential virulence and host range of serovars by comparison of available genome sequences. Fimbrial operons were highly conserved among serovars in respect of gene number, order and sequence, with the exception of safA. Thirteen predicted major fimbrial subunit genes were separately inactivated by lambda Red recombinase-mediated linear recombination followed by P22/int transduction. The magnitude and duration of intestinal colonisation by mutant and parent strains was measured after oral inoculation of out-bred chickens. Whilst the majority of S. Enteritidis major fimbrial subunit genes played no significant role in colonisation of the avian intestines, mutations affecting pegA in two different S. Enteritidis strains produced statistically significant attenuation. Plasmid-mediated trans-complementation partially restored the colonisation phenotype.
We describe the fimbrial gene repertoire of the predominant non-typhoidal S. enterica serovar affecting humans and the role played by each predicted major fimbrial subunit in intestinal colonisation of the primary reservoir. Our data support a role for PegA in the colonisation of poultry by S. Enteritidis and aid the design of improved vaccines.
Klebsiella pneumoniae is an opportunistic pathogen which frequently causes hospital-acquired urinary and respiratory tract infections. K. pneumoniae may establish these infections in vivo following adherence, using the type 3 fimbriae, to indwelling devices coated with extracellular matrix components. Using a colony immunoblot screen, we identified transposon insertion mutants which were deficient for type 3 fimbrial surface production. One of these mutants possessed a transposon insertion within a gene, designated mrkI, encoding a putative transcriptional regulator. A site-directed mutant of this gene was constructed and shown to be deficient for fimbrial surface expression under aerobic conditions. MrkI mutants have a significantly decreased ability to form biofilms on both abiotic and extracellular matrix-coated surfaces. This gene was found to be cotranscribed with a gene predicted to encode a PilZ domain-containing protein, designated MrkH. This protein was found to bind cyclic-di-GMP (c-di-GMP) and regulate type 3 fimbrial expression.
Klebsiella pneumoniae is an opportunistic gram-negative pathogen involved in outbreaks of nosocomial infections in intensive care units. Strains are resistant to multiple antibiotics, and 15 to 30% of them are also resistant to the broad-spectrum cephalosporins by the production of R plasmid-encoded extended-spectrum beta-lactamases. Because the gastrointestinal tracts of patients have been shown to be the reservoir for nosocomial strains of K. pneumoniae, we looked for a correlation between antibiotic resistance and adhesion of K. pneumoniae strains to intestinal cells. We investigated adhesion to the human intestinal epithelial Caco-2 cell line of 61 clinical K. pneumoniae strains isolated in hospitals in Clermont-Ferrand, France. None of the strains tested expressed the previously described adhesive factors CF29K and KPF-28. Adhesive properties were found for 42.6% of the strains tested (26 strains). Just 7.7% (2 strains) of the 26 strains producing only the chromosomally encoded SHV-1 beta-lactamase adhered to the Caco-2 cell line, whereas 68.5% (24 strains) of the 35 strains producing a plasmid-encoded beta-lactamase were adherent. All the adherent strains, and even the two strains producing only the SHV-1 enzyme, harbored at least one self-transmissible R plasmid. At variance for CAZ-1/TEM-5 or CAZ-5/SHV-4 beta-lactamase-producing K. pneumoniae strains, curing and mating experiments demonstrated that the self-transmissible R plasmids encoding the TEM-1, CTX-1/TEM-3, CAZ-2/TEM-8, CAZ-6/TEM-24, or CAZ-7/TEM-16 beta-lactamase were not involved in the adhesion of K. pneumoniae strains to intestinal epithelial cells. Nevertheless, there was an association of multiple antibiotic resistance, including resistance to extended-spectrum cephalosporins, and adhesive properties in K. pneumoniae clinical isolates.