Cepacia Colonization Research Articles

Humidification Solution as a Source for Spreading Burkholderia cepacia in a Neonatal Intensive Care Unit

Abstract Burkholderia cepacia is an important opportunistic organism in hospitalized and immunocompromised patients especially in newborns. The natural ecology of these bacteria associated with plants is also a cause of infectious potential. The disease-causing potential of bacteria as a nosocomial pathogen may be due to its ability to survive in antiseptic solutions, contamination equipment. The patient was hospitalized for prematurity and respiratory distress syndrome. He was treated with surfactant intratracheally for the respiratory distress syndrome. Umbilical catheter was inserted. Ampicillin and gentamicin treatments were initiated. The patient who received respiratory support for a long time was given a steroid protocol because of bronchopulmonary dysplasia. Burkholderia cepacia was detected in the blood and tracheal aspirate cultures of the patient, whose infection markers increased and a new area of infection was detected on the chest radiograph. Colistin and ciprofloxacin treatments were given according to the culture antibiogram. Screening tests revealed B. cepacia colonization in incubator moistening solutions. All incubator humidification solutions in the hospital were changed. Burkholderia cepacia is a rare cause of nosocomial infection in intensive care units but resistant to many treatments. With its capability to colonize water and grow on microbicides, the presence of B. cepacia in a patient's blood warrants further investigation in institutions providing care.

A National Guard outbreak of Burkholderia Cepacia infection and colonization secondary to intrinsic contamination of Albuterol nebulization solution

Burkholderia cepacia, a gram-negative pathogen, has been a known cause of hospital outbreaks because of a contaminated common source such as multidose medications. We describe an outbreak with Burkholderia cepacia infection in 2 major hospitals affiliated to the National Guard, related to an intrinsic contamination of a locally manufactured, multidose Albuterol nebulization solution (Tabouk Pharmaceutical Company, Tabouk, Saudi Arabia) and we report the interventions taken to interrupt this outbreak.During the outbreak period between May 2003 and March 2004, a combined prospective surveillance and a retrospective chart and microbiologic data review were conducted in 4 major hospitals affiliated to the National Guard. Microbiologic cultures were also performed on environmental objects of concern, as well as certain medications. In addition, a questionnaire was distributed to the respiratory therapy staff to evaluate the process of administering respiratory medications and their adherence to sound infection control practices.An intrinsic contamination of a locally manufactured brand of multidose Albuterol nebulization with B cepacia was identified. Two of the 4 hospitals were found to be involved: hospital A a 700-bed tertiary care center and Hospital B a 150-bed hospital. A total of 2121 patients were exposed to Albuterol nebulization as inpatients at hospital A and 318 as outpatients. For hospital B, a total of 283 inpatients and 34 outpatients were exposed to the Albuterol nebulization. Forty and 12 patients, from hospital A and hospital B, respectively, were found to have at least 1 positive culture for B cepacia. From hospital A, most samples were respiratory, and, from hospital B, most were from blood. Molecular typing of 34 available isolates showed that 23 cases were of a single strain of B cepacia that matched the strain isolated from the 3 different batches of multidose Albuterol nebulization. Three culture-positive patients never received Albuterol nebulization of that brand but were in the same room of a patient who had been receiving the medication.We identified a large outbreak of B cepacia in 2 major hospitals affiliated with the National Guard, linked to an intrinsic contamination of a multidose Albuterol nebulization solution. During the period of prospective surveillance, only a few cases were identified as a result of nosocomial transmission. Immediate notification of the Ministry of Health and withdrawal of the medication and revisiting the respiratory therapy practices were necessary to halt this outbreak.

Immigration and emigration of Burkholderia cepacia and Pseudomonas aeruginosa between and within mixed biofilm communities.

To investigate the dynamics of binary culture biofilm formation through use of both the Sorbarod model of biofilm growth and the constant depth film fermenter (CDFF). Pseudo steady-state biofilm cultures of laboratory and clinical strains of Pseudomonas aeruginosa, selected on the basis of their ability to produce a Burkholderia cepacia growth-inhibitory substance, were established on Sorbarod filters and challenged with corresponding planktonic grown cultures of B. cepacia. Reverse challenges were also conducted. Both B. cepacia and P. aeruginosa were able to form steady-state monoculture biofilms after 48 h growth. When steady-state biofilms of B. cepacia NTCT 10661 were challenged with planktonically grown P. aeruginosa PAO1 known to produce a B. cepacia growth-inhibitory substance, the immigrant population was rapidly and almost completely bound to the biofilm, displacing B. cepacia. By contrast, established biofilms of P. aeruginosa PAO1 resisted immigration of B. cepacia 10661. Similar experiments conducted with a nongrowth inhibitory substance producing clinical pairing of P. aeruginosa 313113 and B. cepacia 313113 led to the formation of stable, mixed biofilm populations in both instances. Moreover, co-inoculation with these clinical isolates resulted in a stable, mixed steady-state biofilm. Similar observations were made for biofilms generated in CDFFs. In such instances following pan-swapping between two monoculture CDFFs, B. cepacia 313113 was able to integrate into an established P. aeruginosa 313113 biofilm to form a stable binary biofilm. Establishment of a mixed species community follows a specific sequence of inoculation that may either be due to some degree of match between co-colonizers or that P. aeruginosa predisposes uncolonized sections of the surface to permit B. cepacia colonization. Colonization of a surface with one bacterial species confers colonization resistance towards other species. Disinfection of a surface might well increase the probability of pathogen harbourage.

Epidemiologic investigation of Burkholderia cepacia acquisition in two pediatric intensive care units.

To investigate and describe an outbreak of Burkholderia cepacia in a neonatal intensive care unit (NICU) and a pediatric intensive care unit (PICU), and to report the interventions leading to the cessation of the outbreak. We conducted an epidemiologic investigation of an outbreak of B. cepacia colonization or infection in two clinical wards during a 35-month period (December 1998 to October 2001). A 500-bed, university hospital-affiliated, tertiary-care pediatric institution in Paris, France, with a 22-bed PICU and 31-bed NICU. Ribotyping was used to determine the genotypes of B. cepacia isolates. Procedures for the maintenance and disinfection of respiratory therapy devices were reviewed. Thirty-two children were colonized (n = 14) or infected (n = 18) by B. cepacia in 2 wards (28 in the PICU and 4 in the NICU). In the PICU, a single ribotype was found among the isolates obtained from all of the patients except 1, and from the 6 isolates obtained from respiratory therapy devices (ie, heated humidifier water). In the NICU, the isolates obtained from the patients harbored a single ribotype unrelated to that of the epidemic strain isolated in the PICU; no environmental source of infection was found. Two different outbreaks appeared to be associated with 2 ribotypes, 1 of which was linked to patient-to-patient transmission via respiratory therapy devices. Complete elimination of the outbreak was achieved only when disposable, sterilizable, or easy-to-disinfect materials were used in the PICU. The source of infection in the NICU was not found.

Molecular typing of the bacterial flora in sputum of cystic fibrosis patients

AbstractDespite recent advances in therapy, lower airway infections remain the major cause of morbidity and mortality in cystic fibrosis (CF) patients. Bacterial colonisation of the lower airways in CF is limited to a few bacterial species, commonly Staphylococcus aureus, Pseudomonas aeruginosa and Haemophilus influenzae. Burkholderia cepacia colonisation is much rarer, but it has been thought to be associated with more advanced lung disease and increased mortality. A rapid characterisation of the bacterial flora in sputum of CF patients is of great importance for proper treatment. The aim of this study was to establish bacterial profiles and to identify pathogenic bacteria in respiratory specimens by means of molecular methods including temporal temperature gradient gel electrophoresis (TTGE) and DNA sequencing of PCR amplicons derived from 16S rDNA variable V3 and V6 regions. Sputa of 13 CF patients (7 males/6 females, age 19 – 59 years) collected at the Stockholm CF centre were analysed. TTGE revealed the presence of complex bacterial profiles in all samples. The V3 and V6 PCR amplicons were cloned and sequenced by real-time DNA PyrosequencingTM. DNA from Staphylococcus aureus, Haemophilus influenzae and Pseudomonas aeruginosa respectively, was identified together with sequences from normal oral cavity flora. The results were in reasonable agreement with those obtained by conventional bacterial culture, considering that only known CF pathogens are included in routine reports. However, the methodology seems too elaborate to be introduced into daily routine

Effect of Burkholderia cepacia infection in the clinical course of patients with cystic fibrosis: a pilot study in a Sydney clinic.

Colonisation with Burkholderia cepacia complex in patients with cystic fibrosis (CF) has been associated with adverse outcomes. The aim of the present study was to determine the actuarial survival of CF patients colonized with B. cepacia and to evaluate the efficacy of the Royal Prince Alfred Hospital segregation policy. A secondary aim was to characterize the specific genomovars and strains of B. cepacia isolated in an Australian clinic. Retrospective review of spirometric and microbiological data on all patients colonized with B. cepacia. Each B. cepacia-colonized subject was matched with three case-control subjects. Phenotype and genomovar typing, random amplified polymorphic DNA strain type and B. cepacia epidemic strain marker analyses were performed. The effect of B. cepacia colonization on transplant-free survival was estimated by Cox's proportional hazards regression using the entire clinic population. Fifteen patients were colonized with B. cepacia, of whom six (40%) had died from CF-related disease by August 1998, compared with 30 of 173 (17.3%) of the entire clinic population. Cepacia status had a significant adverse effect on survival, with a hazard ratio of 2.16 (95% confidence interval 1.0-4.69; P = 0.05). The outcome was variable in subgroups of B. cepacia. Colonization with B. cepacia had a significant adverse effect on survival within the study population. Genomovar and strain typing contributed usefully in accessing the effectiveness of the hospital's segregation policy in preventing cross-colonization.

Outcome of Burkholderia cepacia colonisation in an adult cystic fibrosis centre

Background: Colonisation with Burkholderia cepacia is a poor prognostic indicator in subjects with cystic fibrosis (CF), but outcome prediction is impossible since patients are colonised by different strains with differing...

Outbreak of nosocomial Burkholderia cepacia infection and colonization associated with intrinsically contaminated mouthwash.

From August 1996 through June 1998, 69 ventilated, intensive care unit patients at two Arizona hospitals had nosocomial respiratory tract cultures positive for Burkholderia cepacia. Intrinsically contaminated alcohol-free mouthwash was identified by pulsed-field gel electrophoresis as the source of the outbreak.

Differential binding of mannose-binding lectin to respiratory pathogens in cystic fibrosis

We have found that mannose-binding lectin binds to Burkholderia cepacia, an important pathogen in patients with cystic fibrosis, and leads to complement activation, but that this is not the case for Pseudomonas aeruginosa, the more common colonising organism in this disease. We suggest that patients with cystic fibrosis with mannose-binding-lectin deficiency will be at particular risk of B cepacia colonisation.

Transmission and prevalence of Burkholderia cepacia in welsh cystic fibrosis patients

From 1987 to 1994, 16 of 162 cystic fibrosis (CF) patients attending CF clinics at three different hospitals in South Wales, U.K. were found to have respiratory secretions colonized with Burkholderia cepacia (B. cepacia). Bacteriological typing by polymerase chain reaction (PCR) ribotyping demonstrated seven strains of B. cepacia among these 16 CF patients. This typing confirmed that cross-infection was the mechanism of colonization in six of the nine patients who were colonized at the paediatric CF clinic at the University Hospital of Wales in Cardiff, and in three of the six patients who were colonized at the adult CF clinic at Llandough Hospital in Cardiff (cross-infection rate nine of 16 patients or 56%). A search was made for a nosocomial source, with screening of wards and clinics. Swabs from fomites produced four positive cultures for B. cepacia. Two isolates had the same PCR ribotype as that of the previous CF room occupant. To establish prevalence of B. cepacia among CF children living throughout Wales, respiratory secretions were cultured from 151 of 186 CF children (age <16 years). This failed to demonstrate B. cepacia colonization other than in the CF patients already identified.

Exercise testing and prognosis in adult cystic fibrosis.

BACKGROUND: The assessment of prognosis is an important issue in cystic fibrosis. The prognostic value of exercise testing in comparison with other predictors of mortality was examined. METHODS: Ninety two...

Burkholderia (Pseudomonas) cepacia epidemiology in a cystic fibrosis population: a genome finger-printing study.

In patients with cystic fibrosis, infection with Pseudomonas cepacia is associated with poor outcomes. However, the epidemiology of Burkholderia cepacia is still unclear. The aim of this study was to investigate the epidemiology of Burkholderia (Pseudomonas) cepacia colonization among cystic fibrosis patients attending the Verona CF Centre, a large specialized unit to which patients from different parts of Italy are admitted. We used a genome finger-printing system to analyse the nucleotidic structure of B. cepacia isolates from 60 colonized cystic fibrosis patients. Forty-two different finger-printing patterns were identified. Thirty-two patients were colonized by individual B. cepacia strains (53.3%). The remaining 28 subjects were divided into 10 different subgroups, each exhibiting a distinct strain of B. cepacia (46.7%). Nevertheless, direct, person-to-person transmission was evident in only 10 cases (16.7%). The stability up to 12 months, of the B. cepacia colonizing strain was documented in 36 individuals. Consistent with other reports, risk of B. cepacia transmission between cystic fibrosis patients through intimacy or nosocomial contact was found in our study. However, besides low contagiousness, our data suggest that the environmental reservoir of B. cepacia outside the hospital seems to play an important role in B. cepacia infection of our cystic fibrosis population.

Epidemiology of Pseudomonas cepacia colonization among patients with cystic fibrosis.

Colonization with Pseudmonas cepacia in patients with cystic fibrosis (CF) has been associated with increased morbidity and early death, compared with colonization by P. aeruginosa. The mode of acquisition of P. cepacia is not fully understood, although person-to-person spread appears likely. Recent epidemiologic studies support the importance of social contact in the spread of P. cepacia among patients with CF. This study was undertaken to investigate the epidemiology of P. cepacia colonization among patients with CF attending the CF clinic at our center. Isolates of P. cepacia were collected from patients at two CF treatment centers, including ours. Additional isolates were collected from patients without CF in the hospital ICU, from other teaching hospitals, and from the environment. Profiles of enzymes were obtained by ultrathin polyacrylamide gel electrophoresis of P. cepacia extracts. A predominant electromorphic type (ET) was found among P. cepacia isolates from patients at both centers, suggesting a common source or person-to-person transmission. The majority of hospital isolates fell into a single, different ET. Surveillance swabs of respiratory equipment at our CF clinic did not grow P. cepacia. Attendance of patients at CF summer camp correlated strongly with P. cepacia colonization (P < 0.0001).

Inapparent transmission of Pseudomonas (Burkholderia) cepacia among patients with cystic fibrosis.

Pseudomonas cepacia is a significant pathogen in children and young adults with cystic fibrosis, and prevention of its acquisition has become an important goal in patient management. Although it is now clear that this bacterium can be transmitted from person to person, the frequency of this mode of acquisition and the measures required to prevent it are controversial. In this report we describe the use of a novel genotyping method to extend our previous investigation of person to person transmission of P. cepacia among patients with cystic fibrosis attending an educational program. Three (20%) of 15 individuals acquired P. cepacia after contact with a chronically colonized patient. Analysis revealed that the isolates recovered from the three newly colonized patients were the same as that from the index patient. We also demonstrated that pulmonary colonization with P. cepacia may not be detected by currently recommended culture methods for as long as 2 years after acquisition. These data indicate a need to develop more sensitive means of detecting P. cepacia colonization in order better to understand host-pathogen interaction and to optimize preventive strategies.

Serum IgG and sputum IgA antibody to core lipopolysaccharide antigen from Pseudomonas cepacia in patients with cystic fibrosis.

The immunological response of cystic fibrosis (CF) patients to lipopolysaccharide (LPS) antigens of Pseudomonas cepacia was investigated. Enzyme-linked immunosorbent assays (ELISA) with either P. cepacia whole cells or extracted core LPS from a clinical isolate of P. cepacia as antigen were used to measure serum IgG and sputum IgA anti-P. cepacia antibodies. The ELISA with core LPS distinguished nine CF patients colonised by P. cepacia from nine age- and sex-matched non-colonised CF patients. The rate of increase of anti-P. cepacia IgG antibodies after bacteriologically proven P. cepacia colonisation varied in individual patients: in some patients the first isolation of P. cepacia was preceded or accompanied by a two-to-four-fold rise in anti-P. cepacia LPS IgG titres. Absorption studies and immunoblot analysis of serum from patients colonised with P. cepacia demonstrated that a significant component of the anti-P. cepacia core LPS antibodies was specific for P. cepacia and did not react with the core LPS of P. aeruginosa. Immunoblotting also illustrated that there may be a degree of core heterogeneity between different isolates of P. cepacia. Detection of P. cepacia LPS specific antibodies in serum (IgG) and sputum (IgA) from CF patients is recommended to assist the identification of P. cepacia colonisation in CF patients.

Binding of nonmucoid Pseudomonas aeruginosa to normal human intestinal mucin and respiratory mucin from patients with cystic fibrosis.

Lung infections due to Pseudomonas aeruginosa and Pseudomonas cepacia are common in patients with cystic fibrosis. Initial colonization is due to nonmucoid P. aeruginosa, while later mucoid variants emerge and are associated with chronic infection. P. cepacia colonization tends to be more prevalent in older patients. The present study was conducted to discover whether highly purified mucins (from cystic fibrosis sputum and control intestinal secretions) exhibited specific binding of nonmucoid P. aeruginosa. In vitro solid phase microtiter binding assays (with or without a blocking agent) as well as solution phase assays were conducted. Bacteria bound to both mucins via bacterial pili, but no differences in binding capacity were noted between the mucins. Unlike P. cepacia (described in the accompanying manuscript) there was also no preferential binding of P. aeruginosa to mucins versus bovine serum albumin, casein, gelatin, or a host of structurally unrelated proteins and glycoproteins. Carbohydrate hapten inhibition studies did not suggest the existence of specific mucin carbohydrate receptors for P. aeruginosa. In solid phase assays a low concentration (0.05 M) of tetramethylurea abolished P. aeruginosa bacterial binding to both mucins as well as to BSA, whereas in solution phase assays mucin binding to bacteria was not completely disrupted by tetramethylurea. Specific monoclonal antipilus antibodies did not inhibit binding to a greater extent than did Fab fragments of normal mouse IgG. Binding of strains PAO1 and PAK (and isolated PAK pili) to buccal epithelial cells was not influenced by the presence of mucin in binding assay mixtures. Our findings do not support the widely held notion that specific mucin receptors are responsible for the attachment of P. aeruginosa pili, nor do they support the idea that there is a competitive interference by mucins of bacterial binding to respiratory cells. In patients with cystic fibrosis, it would seem unlikely therefore that initial colonization of the lungs by P. aeruginosa is due to a 'selective tropism' of these bacteria for respiratory mucin.

Longitudinal serum IgG response to Pseudomonas cepacia surface antigens in cystic fibrosis

In cystic fibrosis (CF), serum antibody against surface antigens of Pseudomonas aeruginosa is detected only after colonization. Since pulmonary acquisition of P. cepacia usually follows colonization with P. aeruginosa and since P. aeruginosa-colonized patients with CF have demonstrable antibody against outer membrane proteins of P. cepacia, it appears that acquisition of the latter organism occurs in the presence of specific serum antibody. To test this hypothesis, serum obtained from six P. aeruginosa-colonized patients 4 and 2 years prior to and 3 months and 2 years after P. cepacia colonization were assayed for total and specific IgG to P. cepacia outer membrane components. Four patients demonstrated 6-fold or greater increases in specific IgG titers to whole outer membranes following colonization. By immunoblot, all patients had demonstrable serum IgG against the 27- and 36-kDa outer membrane proteins of P. cepacia 4 and 2 years prior to colonization. Immunoblots after P. cepacia acquisition demonstrated an intensification of the 28- and 36-kDa bands and the appearance of antibody to a very low molecular weight compound which was not hydrolyzed by proteinase K and was present in purified LPS. These observations suggest that low serum titers of antibody against two P. cepacia outer membrane proteins are present in patients with CF prior to P. cepacia colonization, and that these antibodies fail to protect for intrinsic or extrinsic reasons.

Effect of Pseudomonas cepacia colonization on survival and pulmonary function of cystic fibrosis patients

We conducted a historical prospective study of 124 cystic fibrosis (CF) patients colonized with Pseudomonas cepacia (cases) and 124 sex and age matched non-colonized with Pseudomonas cepacia (cases) and 124 sex and age matched first year following P. cepacia colonization compared to 8 of the control patients, a highly significant difference ( p < 0.001). In the second year, there was no significant difference in mortality between the two groups. Cases as a group had poorer pulmonary function and chest X-ray scores than controls up to 2 years before P. cepacia first appeared in their sputum or throat cultures. Regression analysis of pulmonary function tests (percent predicted FEV 1 and RV/TLC) for each subject from 3 years before to 2 years after colonization revealed significant differences between cases and controls in slope for FEV 1 and in slope and intercept for RV/TLC. When compared separately according to gender, the differences between cases and controls are significant in females but not in males. These results suggest that patients with poor pulmonary function are more prone to colonization with P. cepacia, that a subgroup of these patients will be dramatically affected and die within a year, and that the organism continues to exert a less dramatic negative effect on the pulmonary function of those patients who survive the initial acute effects of colonization, particularly in female patients.

Purification and characterization of an extracellular protease from Pseudomonas cepacia.

An extracellular proteinase (PSCP) produced by Pseudomonas cepacia was purified from culture supernatants by ammonium sulfate precipitation, anion exchange chromatography on DEAE-Sephacel, and G200 gel filtration chromatography. The protease has an apparent Mr of 34,000 by electrophoresis. Substrates cleaved by the protease include gelatin, hide powder, and collagen but not human immunoglobulin G (IgG), IgM, secretory IgA, or IgA. The enzyme had the characteristics of a metalloprotease, a pH optimum of 6, and a temperature optimum of 45 degrees C. Intratracheal instillation of purified PSCP into rat lungs produced a bronchopneumonia characterized by polymorphonuclear cell infiltration and proteinaceous exudation into large airways. Rats responded immunologically to active immunization with PSCP, but this response was not protective against subsequent lung infection with P. cepacia. PSCP was shown to have antigenic similarity with Pseudomonas aeruginosa elastase by an immunoblotting technique. Sera from 10 cystic fibrosis patients, with and without a previous history of P. cepacia colonization, were shown to possess antibody reactive against PSCP.

The epidemiology of Pseudomonas cepacia in patients with cystic fibrosis.

Pseudomonas cepacia has emerged as an important nosocomial pathogen colonizing and infecting the respiratory tract of patients with cystic fibrosis (CF). Although assessment of outcomes associated with P. cepacia colonization has been difficult, controlled studies have shown that colonized patients experience more adverse outcomes compared with those not colonized. In the United States, an increasing trend in national incidence and prevalence of P. cepacia colonization has been shown, but cases have been unevenly distributed in a few centers. These estimates, however, may be biased by intercenter differences in laboratory methods for detecting P. cepacia in patient sputum. The source and mode of transmission of P. cepacia have not been adequately demonstrated, and may vary from center to center. Until further studies elucidate the epidemiology of P. cepacia in patients with CF, it may be prudent for CF centers to consider the use of selective media to isolate P. cepacia from sputa of patients with CF, to conduct investigations of clusters of P. cepacia-colonized patients, and to consider adopting infection control precautions recommended for control of multiply resistant gram negative organisms.