Overview of glucose nonfermenters

History

• 1949: B. cepacia discovered by Walter Burkholder as the cause of onion skin rot
• 1950’s: described as a human pathogen.
• 1977: First isolated in patients with cystic fibrosis (CF), (then called Pseudomonas cepacia).
• 1980’s: Outbreaks of B. cepacia in CF patients have 35% mortality.
  • B. cenocepacia (particularly strain ET12) assc. with outbreaks of ‘cepacia syndrome’ in CF units

Taxonomy

• 79 Burkholderia spp.
• Recent research has resulted in a number of changes to the taxonomy of Burkholderia cepacia complex (Bcc). The Bcc currently comprises 21 species which exhibit a high degree of 16S rRNA (98–100%) and recA (94–95%) gene sequence similarity, and moderate levels of DNA–DNA hybridization (30–50%).

Genomovars listed in the table (Genomovar = closely-related but distinct genetic spp):

| Genomovar | Species (Bold = Oxidase variable) | Commonly in CF Lung | | --- | --- | --- | | I | Burkholderia cepacia | X | | II | Burkholderia multivorans | X | | III (a, b etc) | Burkholderia cenocepacia | X | | IV | Burkholderia stabilis | | | V | Burkholderia vietnamiensis | X | | VI | Burkholderia dolosa | X | | VII | Burkholderia ambifaria | | | VIII | Burkholderia anthinia | | | IX | Burkholderia pyrrocinia | | | X | B. ubonensis | | | Other (genomovar) | B. arboris (BCC3) B. contaminans (group K, BBC AT) B. diffusa (BCC2) B. lata (group K) B. latens (BCC1), B. metallica (BCC8) B. paludis B. pseudomultivorans B. seminalis (BCC7) B. stagnalis (BCC B) B. territorii (BCC L) | |

| Site | - Soil

• Plant rhizosphere (the region of soil near plant roots)
• Freshwater environments such as river sediments
• “Marine environments (marine sponges) … some strains can tolerate high salinity” | | --- | --- | | Risk Factors | - CF (NB: person-person spread documented)
• Prior colonisation with Pseudomonas
• Chronic granulomatous disease (resist neutrophil-mediated non-oxidative killing & induces neutrophil necrosis)
• Sickle cell | | Pathogenic mechanisms | Low virulence
• Adhere to plastics
• Long survival in moist environments (can survive on skin for <30 minutes, on sputum- contaminated surfaces for weeks & in distilled water for many years)
• Survival in air (After Bcc+ Pt leaves room): ~45m
• Produce Elastase/Gelatinase
• Relatively resistant to Neutrophils (see above)
• Invade & survive inside airway epithelial cells & macrophages

Transmissibility markers:

• cblA gene encoding a major structural subunit of cable-like and mucin binding pili.
• The DNA transcription regulator BCESM (Burkholderia cepacia epidemic strain marker) BCESM & cblA are not found in all transmissible Bcc strains, but B.cenocepacia strain ET12 uniquely possesses both markers. | | Clinical syndromes | CF Lung infection
• Isolated from 3% CF Patient’s sputum
• 90% are B.cenocepacia or multivorans
• 1/3 CF patients: slight decline in lung Fn noted (i.e. not pathogenic in all colonised)
• More harmful: B.cenocepacia, dolosa

Transient infection:

• Less common with B. cenocepacia
• Determines if they can be cohorted with Bcc- Pts
• Definition: 3 neg sputum Cx over 1yr period

Cepacia syndrome:

• Fulminant LRTI
• Sx: High fever, leucocytosis, sepsis & severe progressive respiratory failure
• High-risk strains: B.cenocepacia, multivorans
• Mean time to onset (from colonisation): 5 years

Effects on Lung transplant outcomes:

• Excess mortality for CFs infected pre-op with Bcc vs uninfected (33% v 12%).
• 1yr post-transplant survival: 67% (Bcc-infected) vs 92% uninfected | | Transmission risk (Between CF Patients) | Low: Brief encounters indoors or outdoors

High:

• Closer social contact – evenings in the pub or restaurant
• Hand shaking
• Contacts involving siblings with cystic fibrosis
• Sharing bedrooms
• Social kissing
• Travelling together in closed conditions e.g. car or lift
• Sports or exercise classes
• Sharing eating or drinking utensils
• Intimate contact - kissing, sexual relationships | | IPC (CF Trust recommendations) | - Segregate Bcc+ CF patients (even Bcc patients from each other)
• Regular microbiological surveillance in CF centres
• See Bcc+ patients on separate days in clinic, see high-risk strain Pts separately etc. Avoid communal CF camps, spas/aerated baths, segregate CF schoolchildren from each other. etc.
• See here (Section 12) for full recommendations |

Lab diagnostics

Microbiology

• Bug:
  • Gm-neg bacilli
  • Slender motile rods.
• Agar:
  • BCSA (Burkholderia cepacia selective agar)
    • Containse crystal violet and bile salts (inhibit GPC), & ticarcillin/polymyxin B (inhibit GNB).
    • Contains phenol red pH indicator (turns pink reacting to alkaline byproducts of Burkholderia)
    • DDx of a colony: Candida spp, Steno, R.pickettii, P.aeruginosa, some Pseudomonas spp & other colistin-R GNB
  • Alternative: OFPBL (Oxidation-Fermentation Polymyxin Bacitracin Lactose agar)
    • Polymyxin: Kills GNB
    • Bacitracin: Kills GPC + Neisseria
    • Yellow colonies = non-lactose fermenting org (i.e. Burkholderia)
    • LF colonies = Candida, Steno, P.fluorescens
  • Aerobic
  • Temp: Prefer 25-35; can grow at 41°C; No growth at 42 or 4 °C.
  • Time: 2 days @ 35-37.
• Colony:
  • 1-2mm Circular, with the medium turning pink
  • Cat +
  • Ox variable
  • Nitrate neg
  • ONPG pos
• Species ID
  • CF patient: Commercial ID system 1st
    • e.g. MALDI
  • 1st time isolate: RefLab (Colindale or Edinburgh)
    • Conduct specific phenotypic tests & recA-based PCR.
    • PCR identification of epidemic markers (BCESM and cblA) is also available
    • Genomic fingerprinting (RAPD, PFGE, MLRT) to investigate clonality of individual isolates for infection control surveillance & other studies
    • MLST: Allows strain & species ID simultaneously