Burkholderia cepacia

Burkholderia cepacia
Pseudomonas cepacia
burkholderia cepacia complex
B. cepacia

1. Burkholderia cepacia Prof . Abdelraouf Elmanama by Ahmed A. Abdallah

2. Intro Burkholderia cepacia , formerly  Pseudomonas cepacia  is a motile gram negative bacillus which is aerobic and glucose non-fermenting. It proliferates under conditions of minimal nutrition and can survive in the presence of certain disinfectants. It has a wide distribution in the natural environment  via  water, soil, fruits, and vegetables. In the last two decades,  B. cepacia  has emerged as a serious human pathogen causing fatal necrotizing pneumonia and bacteremia, especially in patients with cystic fibrosis (CF) or chronic granulomatous disease. Cross-transmission , frequent pulmonary procedures, and central venous access facilitates the nosocomial spread of this organism. High transmissibility in the hospital setting, intrinsic resistance to many antibiotics and association with a poor prognosis highlights the need for early detection and treatment of  B. cepacia   infections These bacteria also have contaminated many drug products and can create public health concerns. Pharmaceutical products that are contaminated with B. cepacia may pose serious consequences to vulnerable patients (e.g., compromised immune system).

3. Burkholderia cepacia Burkholderia cepacia was known as Pseudomonas cepacia prior to 1992. B. cepacia was discovered by Walter Burkholder in 1949 as the cause of onion skin rot, and first described as a human pathogen in the 1950s. It was first isolated in patients with cystic fibrosis (CF) in 1977, when it was known as Pseudomonas cepacia. In the 1980s, outbreaks of B. cepacia in individuals with CF were associated with a 35% death rate.

4. Burkholderia cepacia Similar to other opportunistic pathogens such as Pseudomonas aeruginosa , Bcc strains do not normally infect healthy individuals but only those that are immunocompromised. Specific populations susceptible to infection include elderly people, young children, cancer patients, pregnant women, and people with chronic illness. However, B. cepacia occasionally causes illness in non-immunocompromised patients.

5. Burkholderia cepacia The most serious conditions caused by B. cepacia are pneumonia or patients with impaired immune systems or chronic lung disease, particularly cystic fibrosis (CF ). CF is the most common lethal inherited disease of Caucasian populations, with pulmonary infections being the major cause of morbidity and mortality. The severity of infection or colonization by B. cepacia may be different for individual patients. However , overall, pulmonary colonization reduces survival by 50%; about one third to one-half of patients succumb to cepacia syndrome , a rapidly fatal necrotizing pneumonia.

6. Burkholderia cepacia Burkholderia  is a genus composed of more than 60 organisms. Many of which were formerly classed as  Pseudomonas  species (family: Burkholderiaceae; phylum: Proteobacteria ). Within the Burkholderiaceae is the  Burkholderia cepacia  complex, a group of some 20 organisms (the actual number is a matter of taxonomic debate, falling between 17 and 20 species) subgrouped into nine genomovars (that is, phylogenetically differentiable but phenotypically indistinguishable bacteria ). The BCC is a group of species that are so closely related that they can, for the most part, only be differentiated using a combination of multiple molecular diagnostic procedures. 

7. overview of the  Burkholderia cepacia  complex.

8. Burkholderia cepacia Bcc are widely distributed in natural and man-made habitats. Burkholderia cepacia   is a motile aerobic oxidase positive Gram-negative bacillus commonly found in liquid reservoirs and moist environments. The cells are 0.5 to 1.0mm wide and 5mm in length. They are common in nature and present in soils, plant rhizospheres, water, and agriculture products. Infections caused by Bcc have occurred worldwide , and accumulating evidence implicates contaminated pharmaceuticals, cosmetics, disinfectants, and preservative products as major sources of Bcc.

9. Transmission In epidemiological terms, the mode of interpersonal transmission for this opportunistic pathogen primarily occurs through direct contact with other people (e.g ., a handshake), or through contact with body perspiration . Although B. cepacia does not appear to survive on completely dry surfaces for more than one week, it can survive for many months in water. B . cepacia can use other routes of transmission including contact with hard surfaces. Perhaps most important to note is this microbe’s ability to remain viable under harsh conditions (e.g., organic solvents, antiseptics, low nutrients, etc.) for many months .

10. Resistance Bcc are multi-drug resistant organisms. The multi resistance of Bcc bacteria appears to result from various efflux pumps that efficiently remove antibiotics from the cell, decreased contact of antibiotics with the bacterial cell surface due to Bcc’s ability to form biofilms , and changes in the cell envelope that reduce the permeability of the membrane to the antibiotic. B. cepacia is also resistant to many disinfectant cleansers and is unaffected by many preservatives including Betadine. Bcc are among the most antimicrobial agent–resistant organisms encountered in the clinical laboratory. Due to mucin-binding proteins, this species can form biofilms and contaminate plastics, metals, water systems, hospital equipment, catheters, and living tissue .

11. Resistance Microbial biofilms develop when microorganisms adhere to a surface by producing extracellular polymers that facilitate adhesion and provide a structural matrix. Once these cells attach and produce extracellular polysaccharides in the biofilm, their rate of growth is influenced by flow rate, nutrient composition of the medium , antimicrobial-drug concentration, and ambient temperature. It is well established that microbial biofilms can impart physiological resistance to antimicrobial treatment a thousand fold greater compared to exposure to the same bacteria exposed as individual cells .

12. Resistance Research indicates that B. cepacia strains exhibit random genetic changes that can confer a high frequency of transmissibility, or given the right epidemiologic circumstances , these altered genetic changes can benefit the organism’s communal survival and pathogenicity . Exchange of genes among different species for virulence factors might increase adaptability and diminish the effectiveness of treatment and control.

13. Virulence An enormous amount of research has been undertaken to define those virulence factors, expressed by B. cepacia, which interact with the host, and account for the greater morbidity and mortality. Amongst virulence factors, endotoxin which has been clearly shown to have a role in the pathogenesis of B. cepacia infection. Lipopolysaccharide from clinical isolates has endotoxin activity and the capacity to induce tumor necrosis factor (TNF-α ) levels over nine times more than endotoxin extracted from Pseudomonas aeruginosa. This conforms to the evidence that CF infected B. cepacia patients, when compared with P. aeruginosa colonized patients, have an up-regulated inflammatory response when measuring plasma neutrophil elastase.

14. Virulence Nitric oxide (NO) and hydrogen peroxide are important bactericidal mediators in lung defense against  B. cepacia . Inducible NO synthetase is deficient in the bronchial epithelium of CF patients. This may contribute to their susceptibility to  B. cepacia  as a pulmonary pathogen .

15. Complications Bcc-contaminated products are most harmful to CF patients receiving lung transplantations. The mortality rate is high. Mortality rates in the United Kingdom for the first year of infection were reported at 50 –100 %. Of the 11 patients with cystic fibrosis who were also infected with B. cepacia complex , five died post-transplant because of progressive B. cepacia related sepsis. All five patients were clinically unresponsive to cyclical antibiotics and thoracostomy drainage.

16. Complications - In Toronto, Canada Of the 53 [transplant] recipients, 19 have died ( 15 of 28 [54%] B. cepacia positive and 4 of 25 [16%] B. cepacia –negative ). One-year survival was 67% for B . cepacia –positive patients and 92% for B. cepacia –negative patients. Another study in Liverpool, England Thirty-seven patients had been colonized by epidemic B. cepacia and these patients had four times the mortality of the remainder

17. Burkholderia Cepacia Selective  Agar (BCSA) The slower growing  Burkholderia cepacia  can be missed on conventional media such as blood or MacConkey Agar due to overgrowth caused by other faster growing organisms found in the respiratory tract of CF patients such as Klebsiella  species, Pseudomonas aeruginosa  and  Staphylococcus  species . This may lead to the infection being missed or wrongly diagnosed . The BCSA medium contains peptones and sugars that supply nutrients for the growth of  Burkholderia cepacia  and other microorganisms. All the bacteria in the BCC described to date are capable of growing on this medium. Crystal violet and bile salts are added to inhibit growth of Gram-positive cocci. Antimicrobials such as ticarcillin and polymyxin B used to inhibit the growth of other Gram-negative bacilli

18. Technique and results of BCSA Take a routine respiratory sample from the patient e.g. sputa, deep pharyngeal swabs or bronchial washings, Specimens may also include blood. Streak onto Burkholderia cepacia Medium and incubate at 37°C for 48 to 72 hours . Examine after 48 hours for sage green colonies and the medium turning from straw-green to bright pink. All colonies should be further identified and confirmed. Re-incubate for a further 24 hours if necessary . Typical colonies of  Burkholderia cepacia  are circular, and entire. Burkholderia cepacia  colonies are typically translucent and rough. Suspect colonies on BCSA medium generally appear greenish-brown with a yellowish halo or white with a yellowish-pink halo (phenol red color indicator ). This method is presumptive and requires confirmation of the identification of the suspect colonies.

19. Burkholderia Cepacia Selective Agar (BCSA)

20. PCR-based identification

21. PCR-based identification Genomovars is a term commonly used within the genera Burkholderia and Agrobacterium to denote strains which are phylogenetically differentiable, but are phenotypically indistinguishable. The group of  B .  cepacia  complex (BCC) organisms consists of nine genomovars associated with different levels of virulence and patient-to-patient transmissibility.  To reduce the probability of BCC spreading among patients with CF, a reliable early test that detects small quantities of the bacteria in clinical samples is needed. Conventional microbiological diagnostics of the BCC based on the results of culture and subsequent biochemical identification is insufficient because of the potential risk of misidentification or false negativity. Also, Conventional methods are not able to reliably distinguish the genomovars of the BCC

22. PCR-based identification Two target genes are commonly used for BCC analysis: the 16S rRNA gene and the  recA   gene. Although sequence variation in the 16S rRNA gene is generally useful for differentiating bacterial species, among Bcc species there is limited sequence diversity in this gene. The variation within the rRNA operon is obviously too small to separate all members of the B. cepacia complex. Due to this discriminatory limitation, a novel PCR-based identification assay based on the recA gene developed. The recA gene sequence variations are a useful means of differentiating genomovars, and have become one of the main Bcc identification method. The recA gene shows 94 to 95% similarity between the different genomovars, and typically 98 to 99 % similarity can be found within the genomovars.

23. Prevention The best way to prevent the spread of B. cepacia, and all infections, is to clean your hands often. This includes washing hands with soap and water or using an alcohol-based hand rub. Health care workers should follow specific infection control precautions. Patients and health care workers should clean their hands often.

24. S ources Mahenthiralingam , E., Urban, T. A., & Goldberg, J. B. (2005).  The multifarious, multireplicon Burkholderia cepacia complex. Nature Reviews Microbiology, 3(2), 144–156.   doi:10.1038/nrmicro1085. Coenye , T., Vandamme , P., Govan , J. R. W., & LiPuma , J. J. (2001).  Taxonomy and Identification of the Burkholderia cepacia Complex. Journal of Clinical Microbiology, 39(10), 3427–3436.   doi:10.1128/jcm.39.10.3427-3436.2001. Torbeck , L., Raccasi , D., Guilfoyle , D. E., Friedman, R. L., & Hussong , D. (2011).  Burkholderia cepacia: This Decision Is Overdue. PDA Journal of Pharmaceutical Science and Technology, 65(5), 535–543.   doi:10.5731/pdajpst.2011.00793. A.M. Jones, M.E. Dodd, A.K.  Webb. (2001) Burkholderia cepacia: current clinical issues, environmental controversies and ethical dilemmas. European Respiratory Journal 2001 17:  295-301 .