Wednesday, August 24, 2011

Pseudomonas aeruginosa (Part 1)

Ps. aeruginosa is a classic opportunist pathogen with innate resistance to many antibiotics and disinfectants. It flourishes as a saprophyte in warm moist situations in the human environment, including sinks, drains, respirators, humidifiers and disinfectant solutions. Infections due to Ps. aeruginosa are seldom encountered in healthy adults but in the last two decades the organism has become increasingly recognized as the aetiological agent in a variety of serious infections in hospitalized patients with impaired immune defenses (Neu, 1983).
Identification and Nomenclature

Ps. aeruginosa is member of the Gamma Proteobacteria class of bacteria. It is gram-negative, aerobic rod belonging to the bacterial family Pseudomonaceae. According to taxonomy based on conserved macromolecules (e.g. 16S ribosomal RNA) the family includes only the members of the genus Pseudomonas which are cleaved into eight groups. Ps. aeruginosa is the type species of its group, which contains 12 other members (Todar, 2008). The word Pseudomonas means false unit, from the Greek pseudo- (Greek: ψευδο, false) and monas (Latin: monas, from Greek: μονος, a single unit). The stem word mon was used early in the history of microbiology to refer to germs, e.g., Kingdom Monera. The species name aeruginosa is a Latin word meaning copper rust, as seen with the oxidized copper patina on the Statue of Liberty. This also describes the blue-green bacterial pigment seen in laboratory cultures of the species. This blue-green pigment is a combination of two metabolites of Ps. aeruginosa, pyocyanin (blue) and pyoverdine (green), which impart the blue-green characteristic color of cultures. Pyocyanin biosynthesis is regulated by quorum sensing as in the biofilms associated with colonization of the lungs in cystic fibrosis patients. Another assertion is that the word may be derived from the Greek prefix ae- meaning"old or aged, and the suffix ruginosa means wrinkled or bumpy. The derivations of pyocyanin and pyoverdine are of the Greek, with pyo-, meaning pus, cyanin, meaning blue, and verdine, meaning green. Pyoverdine in the absence of pyocyanin is a fluorescent-yellow color (Brown, 1956).
Morphology and staining
Gram-negative, non-sporing rods; motile; usually with a single polar flagellum. Fimbriae may be present and are usually polar and non-haemagglutinating. Some Ps. aeruginosa, notably from respiratory tract infections in patients with cystic fibrosis, produce large amount of alginate, an exopolysaccharide consisting of mannuronic and guluronic acids which gives rise to strikingly mucoid colonies. The alginate produced by these mucoid colonial forms of Ps. aeruginosa is distinct from pseudomonas slime, a heterogeneous mixture of hexoses which is produced by all strains of the species on prolonged incubation in media with high carbon, low nitrogen content (Govan, 1996).
Cultural characters
Ps. aeruginosa, like other members of the genus, is extremely adaptable in nutritional terms and can utilize a very wide range of organic substrates as sources of C and N. cultures produce a characteristic sweet, musty smell of aminoacetophenone. Strict aerobe, although NO3 can be used as an electron acceptor to permit anaerobic growth. Colonies are usually very small or extremely flat. Temperature range 5-42°C, optimum 37°C; Ps. aeruginosa, unlike most other pseudomonas species, will grow in serial subculture at 42°C. optimum pH 7.4-7.6 (Govan, 1996).
After aerobic incubation on nutrient agar for 24 h at 37°C six distinct colonial types of Ps. aeruginosa are encountered (Phillips, 1969). Type 1 is the most common and easily recognized; the colonies are large, low convex, rough in appearance and often oval with the long axis in the line of the inoculum streak. Type 2 colonies are small, domed and smooth in appearance and described as coliform-like. Colony type 3 and 4 are also small and appear rough and rugose respectively. The mucoid alginate-producing type 5 colony is very striking. The dwarf colony type 6 is the smallest colony form of Ps. aeruginosa and may appear slightly mucoid (Govan, 1996).
Many strains exhibit the phenomenon known as iridescence, which is observed as a moth-eaten type of colonial lysis with a metallic sheen. Colonies on blood agar may be surrounded by a zone of haemolysis. On MacConkey agar, Ps. aeruginosa colonies are pale, i.e. non-lactose fermenters, and, as on blood agar, the characteristic pigments are often poorly observed (Govan, 1996).
Production of pigments
Demonstration of the presence of the blue pigment pyocyanin is absolute confirmation of a strain as Ps. aeruginosa and thus the major diagnostic test. The yellow pigment fluorescein is also produced by many strains, giving the characteristic blue-green appearance of infected pus or cultures. Pyocyanin, fluorescein and the more rarely observed pyomelanin and pyorubrin are easily identified on nutrient or sensitivity test agars, particularly after prolonged incubation (Govan, 1996).
Biochemical reactions
In general, Ps. aeruginosa appears inert in the usual tests used for the fermentative gram-negative bacilli, e.g. indole and H2S are not produced, Voges-proskauer and methyl red reactions are negative. Cetrimide agar has been traditionally used as selective media. Tests that may be used to characterize Ps. aeruginosa, particularly when pyocyanin production is absent or doubtful, are oxidase, gelatinase positive and can reduce Nitrate (Collee et al., 1996).

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