• Overview
• Pseudomonas aeruginosa
• Burkholderia
• Burkholderia cepacia
• Burkholderia pseudomallei
• Burkholderia mallei
• Delftia acidovorans
• Brevundimonas diminuta and Brevundimonas vesicularis
• Stenotrophomonas maltophilia
• Sphingomonas paucimobilis

Overview

Pseudomonads are an ubiquitous group of environmental gram negative bacterial organisms. They comprise a number of true Pseudomonas species as well as many species formerly classified in the genus. Pseudomonas aeruginosa is the type strain of the genus. They are natural residents of soil and water and may cause primary skin infections in the healthy. This is most associated with recreational water activities and is characterised by a self-limiting skin rash or folliculitis. In the immunocompromised infections can occur in almost any body system and may be severe and accompanied by high mortality. The genus Pseudomonas once comprised over 100 species but over the last decade many of these have been reclassified into different genera. There are five main groups of pseudomonads of medical interest. These are the fluorescent or 'true' pseudomonas, P. aeruginosa (formerly known as P. pyocyanea), P. fluorescens and P. putida. The second group are contained within the genus Burkholderia and within this genus, three species B. cepacia, B. pseudomallei and B. mallei, are associated with human and animal infection. B. cepacia is a complex of 10 genomovars (genetically distinct but phenotypically similar). B. pseudomallei is the aetiological agent of melioidosis, a life threatening septic infection prevalent in SE Asia and Northern Australia and B. mallei causes glanders in horses and other species. The other three generic groups of pseudomonads are Delftia, Brevundimonas and Stenotrophomonas.

With the exception of the medically important species ( P. aeruginosa, B. cepacia, B. mallei and B. pseudomallei) the organisms within the five groups are best regarded as true opportunists. They are relatively insusceptible to antiseptics, disinfectants and antibiotics compared with the natural surrounding bacterial flora and are usually associated with contaminated water reservoirs in respiratory equipment in hospitals. Instillation, injection or inhalation of these organisms by immunocompromised or post surgical patients may pose a significant threat to health. However their isolation from clinical specimens most often reflects colonization rather than invasive infection owing to their relatively low virulence.

Pseudomonas and related species are aerobically growing bacteria which are gram negative. They are able to grow over a wide range of temperatures (11C to 44C) but P. fluorescens and P. putida are psychrophylic and are able to multiply at 4C. They occasionally contaminate refrigerated blood products which when transfused into a patient may cause endotoxic shock. It is also a recognized food spoilage agent, particularly of refrigerated meat, and it may spoil UHT milk if this is stored above 5 ^oC. They may be recovered from fresh vegetables or plants, and sinks, taps and drains in the hospital environment. Most species are motile and can utilize a range of simple organic compounds as energy sources, and metabolize glucose by an oxidative pathway. Some can reduce nitrate or form ammonia from arginine. Anaerobic growth is possible only in the presence of an alternative electron acceptor such as nitrate or arginine. 

Pseudomonas aeruginosa

P. aeruginosas found almost anywhere in the natural habitat in sites ranging from surface waters to vegetation and soil. It can multiply in distilled water but is rarely isolated from sea water except near sewage outfalls and polluted river estuaries. It is not a fish pathogen. The organism is a resident of the soil and rhizosphere and is frequently recovered from fresh vegetables and plants. It is pathogenic for plants such as tobacco, cucumbers and lettuce and is also a well-established pathogen of grasshoppers and insects.

P. aeruginosa has been isolated from a variety of sources including, among others, aviation fuel, cutting oils, cosmetics, plasticizers, photographic materials etc. Hospital and domestic sink traps, taps and drains are invariably colonised by pseudomonads.

Faecal carriage rates vary from 25 to 15% and is higher in vegetarians. P. aeruginosa dies rapidly on dry human skin but in conditions of superhydration of the skin, such as divers in long term saturation chambers and military personnel in swampy terrain, the frequency of colonization is markedly increased and infections such as otitis externa in divers and toe web rot in soldiers are common.

Despite the ubiquity of the organism, community-acquired infections with P.aeruginosa are relatively rare. In hospitals, however, it may account for about 10% of all infections acquired during the patients' stay. It is a frequent cause of pneumonia and urinary tract, surgical wound and blood stream infections are common. The species is particularly frequent as a cause of chronic respiratory infection in cystic fibrosis (CF) patients. P. aeruginosa has low intrinsic virulence in man and animals. Thermal injury or neutropenia or the introduction of relatively large inocula direct into tissues are often necessary prerequisites for the establishment of infection.

Compared to enterobacteria, P. aeruginosa is relatively resistant to many antibiotics but there are a number of antimicrobial agents with good to excellent activity against most isolates of the species. These include ceftazidime, ticarcillin, piperacillin, imipenem, meropenem, gentamicin, tobramycin, amikacin, ciprofloxacin and aztreonam.

As many as 80% of CF patients may be colonised in the lung with P. aeruginosa and once established it is particularly refractory to antibiotic treatment. Many isolates grow as mucoid colonies but mixtures of different colonial forms are frequently found on primary plates. These variants invariably prove to be genetically identical. P. aeruginosa from CF patients are often atypical in growth requirements and may be auxotrophic for specific amino acids, be non motile and a minority may exhibit extreme susceptibility to semi synthetic penicillins.

For epidemiological studies, isolates of P. aeruginosa can be serotyped by slide agglutination of live cultures. There are 21 internationally accepted O-serotypes but four types account for approximately 50% of clinical and environmental isolates. Further discrimination between serotypes can be achieved by DNA fingerprinting using pulsed-field gel electrophoresis of XbaI or SpeI restriction endonuclease digests. 

Burkholderia

The genus Burkholderia was defined in 1992 by Yabuuchi for Pseudomonas species formerly of rRNA group II. There are at least 30 validly named species in the genus but the medically important species are B. cepacia, B. pseudomallei and B. mallei. The organism formerly known as Pseudomonas pickettii was reassigned by Yabuuchi to Burkholderia but has subsequently been reclassified as Ralstonia pickettii. Occasional clinically significant isolates of B. pickettii are recovered from hospital patients but they are most often isolated from the ward environment or as a contaminant of antiseptic and disinfectant solutions. It is oxidase and nitrate positive and arginine negative.

Burkholderia cepacia

The B. cepacia complex currently comprises 10 genomic species or genomovars (see B. cepacia fact sheet ). CF patients appear to be susceptible to lung infection with these organisms which can be particularly severe and lead to the death of a minority of patients from a fulminant necrotizing pneumonia. Patients with chronic granulomatous disease may also succumb to B. cepacia infection due to its resistance to opsonophagocytes of patients with this disease. Apart from these conditions, B. cepacia may be acquired by patients in hospitals from contaminated equipment water reservoirs such as nebulizers and contaminated antiseptic irrigation fluids.

They grow moderately well on nutrient agar and a variety of non-fluorescent pigments may be produced by some strains. They grow slowly at 37 ^oC and extended incubation for 48 hours is recommended to optimise their recovery from sputum. Cultures often die rapidly on storage on nutrient agar slopes but survive remarkably well suspended in sterile distilled water. Selective media based on their constitutive resistance to colistin and bile salts have been described but other colistin resistant gram negative rods may also be recovered on these media. Members of the complex are not differentiated well by phenotypic tests but PCR assays specific for individual genomovars have been reported (see, Coeyne et al. 2001).

B. cepacia has high intrinsic resistance to antimicrobials and is generally resistant to the antibiotics active against P. aeruginosa. It is resistant to aminoglycosides, colistin, ticarcillin, azlocillin and imipenem. Variable susceptibility is shown to temocillin, aztreonam, ciprofloxacin and tetracycline and about two-thirds of strains from CF patients are susceptible to ceftazidime, piperacillin/tazobactam and meropenem.

Burkholderia pseudomallei

This is an important pathogen of humans (melioidosis) and farm animals in tropical and subtropical areas of SE Asia and Northern Australia, where it is endemic in rodents and is found in moist soil, on vegetables and on fruit. A closely related but non-pathogenic species, B. thailandensis, has been described from environmental samples. Cultures should be sent to a reference laboratory for species confirmation. For further information on these organisms and melioidosis, see Dance (1999).

Cultures on blood agar and nutrient agar at 37°C give mucoid or corrugated, wrinkled, dry colonies in 1-2 days, and an orange pigment may develop on prolonged incubation. Variation between rough and smooth colonies is frequent. Cells may also exhibit bipolar staining in gram stains. B. pseudomallei is a strict aerobe, it is motile, oxidizes glucose and breaks down arginine. Most isolates are reliably identified by API 20NE microgalleries but must be distinguished from non-pigmented strains of P. aeruginosa, P. stutzeri and B. mallei. It is resistant to colistin and gentamicin but isolates are generally susceptible to imipenem, piperacillin, amoxycillin-clavulanic acid, doxycycline, ceftazidime, aztreonam and chloramphenicol.

Burkholderia mallei

B.mallei is the causative agent of glanders, a rare disease of horses and no isolates of the organism have been recovered in the UK since the last World War.

Both B. pseudomallei and B. mallei must be handled in category 3 containment facilities and their exchange between laboratories is restricted. 

Delftia acidovorans

This organism was reclassified from the genus Comamonas. It is found on occasion in clinical specimens and the hospital environment. Isolates grow as non-pigmented colonies overnight at 37 ^oC but incubation should be extended to 48 hours for slow growing strains. Some isolates exhibit resistance to colistin and gentamicin and may grow on B. cepacia selective media. Antimicrobial susceptibility is variable but most isolates are susceptible to ureidopenicillins, tetracycline, the quinolones and trimethoprim-sulphamethoxazole.

Brevundimonas diminuta and Brevundimonas vesicularis

These are closely related species previously of rRNA homology group IV of Pseudomonas, are rare in clinical specimens and of doubtful clinical significance. They grow slowly on nutrient agar and require 48 hours incubation at 37 ^oC. B. vesicularis grows as orange pigmented colonies on nutrient agar and gives a weak oxidase reaction. B. diminuta is not pigmented.

Stenotrophomonas maltophilia

This species was once a member of the genus Xanthomonas. It has been isolated from a variety of hospital environmental sources and may be clinically significant in severely immunocompromised patients. The extensive use of imipenem, to which S. maltophilia is resistant, appears to be associated with nosocomial outbreaks. S. maltophilia is increasingly isolated from CF sputum and is often misidentified as B. cepacia as it grows reasonably well on colistin-containing media. Most strains are susceptible to co-trimoxazole, doxycycline and minocycline and third-generation cephalosporins but are resistant to aminoglycosides.

Colonies resemble those of P. aeruginosa but a yellow or brown diffusible pigment may be produced; on blood agar they can appear as faint lavender. It is usually oxidase negative, does not hydrolyse arginine and does not grow on cetrimide agar. It is the only pseudomonad that gives a positive lysine decarboxylase reaction.

Sphingomonas paucimobilis

S. paucimobilis produces a non-diffusible yellow pigment and is most likely to be confused with flavobacteria. Motility is poor and best seen in cultures incubated at room temperature. It has been found in clinical material and recovered from hospital equipment. Most strains are susceptible to erythromycin, tetracycline, chloramphenicol and aminoglycosides.

References and further reading

Coeyne T, Vandamme P, Govan JRW, LiPuma JJ. Taxonomy and identification of the Burkholderia cepacia complex. Journal of Clinical Microbiology 2001; 39:3427-3436.

Chiarini L, Bevivino A, Dalmastri C, Tabacchioni S, Visca P. Burkholderia cepacia complex species: health hazards and biotechnological potential. Trends in Microbiology 2006; 14:277-286.

Pitt TL, Dance DAB. Burkholderia spp. and related genera. In: Topley and Wilson's Microbiology and Microbial Infections. Tenth Edition, Vol 2. (Eds. SP Borriello, PR Murray, G Funke) Hodder Arnold, London. (2005):1607-1648.

Pitt TL, Simpson AJ. Pseudomonas and Burkholderia spp. In: Principles and Practice of Clinical Bacteriology. Second Edition. (Eds. SH Gillespie, PM Hawkey). John Wiley & Sons, London. (2006): 427-443.

Yabuuchi E, Kosaka Y, Oyaizu H, Yano I, Hotta H, Hashimoto Y, Ezaki T, Arakawa M. Proposal of Burkholderia gen. nov; and transfer of seven species of the Pseudomonas homology group II to the new genus, with the type species Burkholderia cepacia (Palleroni and Holmes 1981) comb.nov. Microbiology and Immunology 1992; 36: 1251-1275.

Yamamoto S, Kasai H, Arnold DL, Jackson RW, Vivian A, Harayama S. Phylogeny of the genus Pseudomonas: intrageneric structure reconstructed from the nucleotide sequences of gyrB and rpoD genes. Microbiology 2000; 146:2385-2394.
 

TL Pitt
August 2007