Pseudomonas Media and Tests

By: Jvo Siegrist, Product Manager Microbiology, ivo.siegrist@sial.com, Analytix Volume 2007 Article 5

Detection, identification, differentiation and cultivation of Pseudomonas species


Figure 1. HiFluoro Pseudomonas Agar under UV lightFigure 1.HiFluoro Pseudomonas Agar under UV light


Pseudomonas are motile (one or more polar flagella), rod shaped and aerobe Gram-negative bacteria. They are found almost everywhere, in soil, water, plants and animals. In most cases it is not pathogenic and in fact can be beneficial. For example, P. putida is used as a bio-scrubber to aid in the biodegradation of diverse organic compounds in polluted air and waste water. However, P. aeruginosa is an infamous opportunistic human pathogen most commonly affecting immuno-compromised patients. Along with P. maltophilia, it accounts for the majority of human infections. Pathogenic Pseudomonas are found throughout the body, most commonly in the urinary tract, respiratory tract, blood and wounds [1].

Rugged and opportunistic, Pseudomonas use a wide range of nutritional sources, even very simple nutritional environments without any organic compounds. They can remain viable for long periods of time in many different habitats and under very adverse conditions. They are also widespread, being found in water, saline solutions, utensils and even in cosmetics, pharmaceuticals and disinfectants, and many natural and manufactured foods. Psychrotrophic (cold-tolerant) Pseudomonas species are a significant food spoilage problem in refrigerated meat, fish, shell fish and dairy products. Because Pseudomonas thrive in water systems, they can be the source of contamination in the food and beverage industry [2].

Pseudomonas are not generally fastidious microorganisms. They can grow on very simple media like Kind Agar, for example, which contains a protein hydrolysate, magnesium chloride, potassium sulphate and agar. Analytical microbiology leverages a microbe’s unique biochemistry to aid in its identification. For example, selective Pseudomonas media use cetrimide, nalidixic acid, cephaloridine, penicillin G, pimaricin, malachite green and other inhibitory agents. The proteolytic activity, lipolytic activity, fluorescent pigment formation, nitrate utilisation, glutamate utilisation, hemolytic reaction and other biochemical reactions are used in the media for the identification and differentiation of Pseudomonas species.

Pseudomonas gives negative Voges Proskauer, indole and methyl red tests, but a positive catalase test. While some species show a negative reaction in the oxidase test, most species, including P. fluorescens, give a positive result (see Figure 2). Another feature associated with Pseudomonas is the secretion of pyoverdin (fluorescein, a siderophore), a fluorescent yellow-green pigment under iron-limiting conditions [3]. Certain Pseudomonas species may also produce additional pigments, such as pyocyanin (blue pigment, a siderophore) by P. aeruginosa [4], quinolobactin (yellow, dark green in presence of iron, a siderophore) by P. fluorescens [5], a reddish pigment called pyorubrin and pyomelanin (brown pigment). On blood agar a hemolytic reaction can be observed.


Figure 2. Oxidase test

Figure 2.Oxidase test


Scientific classification of Pseudomonas:

Kingdom: Bacteria Order: Pseudomonadales
Phylum: Proteobacteria Family: Pseudomonadaceae
Class: Gamma Proteobacteria Genus: Pseudomonas






Pseudomonas
utilizes sugars as an energy source by using the Entner-Doudoroff pathway with pyruvate as the end product (dissimilation). The reaction utilizes a different set of enzymes from those used in glycolysis and the pentose phosphate pathway. Fermentation catabolism is not observed in Pseudomonas, but some species, like P. aeruginosa, P. stutzeri and P. denitrificans, are able to use nitrate as an electron acceptor instead of oxygen. Growth can also occur under anaerobic conditions when the denitrification pathway is used. Sigma-Aldrich supplies a wide array of products for the detection, identification, differentiation, enumeration and cultivation of Pseudomonas, using its biochemical characteristics, including Gram staining kit, and many types of selective growth media (Table 1) and diagnostic tests (Table 2). Additional information on media and tests for Pseudomonas and a wide range of other microbes can be found on our web site: www.sigmaaldrich.com/microbiology.

Table 1 Media for Pseudomonas

Cat. No. Brand Nonselective Broths
A0465 Sigma Alternative Thioglycollate Medium
53286 Fluka Brain Heart Broth
B5051 Sigma Bushnell Haas Broth
D3435 Sigma Dey-Engley Neutralizing Broth
63649 Fluka Membrane filter Rinse Fluid (USP)
70149 Fluka Nutrient Broth No 3
03856 Fluka Nutrient Broth No. 4
70179 Fluka Peptone Water
77187 Fluka Peptone Water, phosphate-buffered
40893 Fluka Peptone Water, phosphate-buffered,Vegitone
70157 Fluka Thioglycollate Broth (USP Alternative)
41960 Fluka Vegitone Infusion Broth


Cat. No. Brand Selective Enrichment Broths & Biochemical Identification Broths
00185 Fluka Acetamide Nutrient Broth
17129 Fluka Asparagine Proline Broth
78886 Fluka Cetrimide Broth
63163 Fluka Malachite Green Broth
39484 Fluka Methyl Red Voges Proskauer Broth
14305 Fluka Motility Nitrate Medium
72548 Fluka Nitrate Broth


Cat. No. Brand Nonselective Agars for Cultivation, Enumeration and Isolation
70147 Fluka Milk Agar
70148 Fluka Nutrient Agar
44776 Fluka Nutrient Agar Plates (Diameter 55 mm)
80957 Fluka Plate Count Skim Milk Agar
17209 Fluka R-2A Agar
17175 Fluka Skim Milk Agar, modified
51414 Fluka Skim Milk Agar, modified
70159 Fluka Tryptone Glucose Extract Agar
T2188 Sigma Tryptone Glucose Yeast Extract Agar
01497 Fluka Yeast Extract Agar


Cat. No. Brand Nonselective Agars for Differentiation
70133 Fluka Blood Agar (Base)
21065 Fluka Calcium caseinate Agar
55420 Fluka CLED Agar
16636 Fluka HiCrome(™) UTI Agar, modified
60788 Fluka King Agar A
60786 Fluka King Agar B
75315 Fluka OF Test Nutrient Agar
P1852 Sigma Pseudomonas Agar (for Fluorescein)
91015 Fluka Tributyrin Agar


Cat. No. Brand Selective Agars for Detection and Isolation
11012 Fluka Cetrimide Nalidixic acid Agar
22470 Fluka Cetrimide Agar
70887 Fluka Cetrimide Agar
P2102 Sigma Pseudomonas Agar Base
14521 Fluka Cetrimide Agar Plates (Diameter 55 mm)
17168 Fluka Milk Agar, modified according to Brown & Scott
17208 Fluka Pseudomonas Isolation Agar


Cat. No. Brand Selective Agars with Differential System for Differentiation, Detection and Isolation
50875 Fluka GSP Agar
78996 Fluka HiFluoro (™) Pseudomonas Agar Base


Table 2 Test for identification and differentiation of Pseudomonas

Cat. No. Brand Diagnostic Tests for Pseudomonas
88597 Fluka Catalase Test
05686 Fluka DMACA Indole Disks
49825 Fluka DMACA Reagent
78719 Fluka Kovac‘s Reagent Strips
60983 Fluka Kovac‘s Reagent for indoles
67309 Fluka Kovac‘s Reagent for indoles
08714 Fluka Methyl Red Solution
70439 Fluka Oxidase Test
40560 Fluka Oxidase Strips
07345 Fluka Oxidase Reagent acc. Gaby-Hadley A
07817 Fluka Oxidase Reagent acc. Gaby-Hadley B
18502 Fluka Oxidase Reagent acc. Gordon-McLeod

back to top

Materials

     

References

  1. Collins, F.M. Pasteurella, Yersinia, and Francisella. In Barron‘s Medical Microbiology, 4th ed., S. Barron, et al., eds., University of Texas Medical Branch at Galveston: Galveston, TX, 1996; Chapter 27.
  2. Mead, G.C. et al. Br. Poult. Sci. 1977, 18, 661-670.
  3. Meyer, J.M.; Geoffroy, V.A.; Baida, Appl. Environ. Microbiol. 2002, 68 (6), 2745-2753.
  4. Lau, G.W.; Hassett, D.J.; Ran, H.; Kong, F. Trends in molecular medicine 2004, 10 (12), 599-606.
  5. Matthijs, S.; Tehrani, K.A.; Laus, G.; Jackson, R.W.; Cooper, R.M.; Cornelis, P. Environ. Microbiol. 2007, 9 (2), 425-

back to top

Related Links