[{"data":1,"prerenderedAt":-1},["ShallowReactive",2],{"$fxLN3MUwXCdr5RPjwZYIDpOj8CHyjOmngWTgoKXPtZbg":3,"$fQigZlmxiKiuBUDK-P2QxZXMXtj4qTlWmd6E4inQkXXQ":32,"$f3Ft0rKFJHppdzE-vuveecxx1BUcg9iOlMLtyzf_MJDg":63},[4,8,12,16,20,24,28],{"title":5,"slug":6,"path":7},"About Microbeonline.com","about-microbeonline-com","\u002Fabout-microbeonline-com\u002F",{"title":9,"slug":10,"path":11},"About Me","about-me","\u002Fabout-microbeonline-com\u002Fabout-me\u002F",{"title":13,"slug":14,"path":15},"Advertise with Us","advertise-us","\u002Fadvertise-us\u002F",{"title":17,"slug":18,"path":19},"Privacy Policy","privacy-policy","\u002Fprivacy-policy\u002F",{"title":21,"slug":22,"path":23},"Abbreviations","abbreviations","\u002Fabbreviations\u002F",{"title":25,"slug":26,"path":27},"Microbes","microbes","\u002Fmicrobes\u002F",{"title":29,"slug":30,"path":31},"Books","recommended-books","\u002Frecommended-books\u002F",{"type":33,"data":34},"blog",{"slug":35,"title":36,"description":37,"seoTitle":38,"seoDescription":38,"author":39,"createdDate":40,"lastUpdatedDate":41,"draft":42,"category":43,"image":38,"body":44,"faq":45,"tags":61,"related":62},"cetrimide-agar-composition-principle-preparation-uses","Cetrimide Agar: Composition, Principle, and the Confirmatory Test Non-Fermenters Fail","Why an unpigmented, unremarkable-looking colony from a CF sputum sample can still turn out to be Pseudomonas aeruginosa, and the one temperature test that separates it from its fluorescent look-alikes.",null,"Nisha Rijal","2015-05-24","2026-07-05",false,"culture-media","**The colony that didn't look like much, until it was given the right stage**\n\nA sputum sample from a cystic fibrosis patient grows a non-fermenting, Gram-negative rod on the primary plate. No strong odor, no obvious pigment, nothing that immediately says \"*Pseudomonas aeruginosa*,\" the single most consequential chronic colonizer in CF lung disease, one whose presence changes antibiotic strategy, prognosis, and infection control practice.\n\nSubcultured onto cetrimide agar, the same organism looks different. Cetrimide, a cationic detergent, doesn't just suppress most competing organisms, it specifically enhances *P. aeruginosa*'s own production of pyocyanin and fluorescein, the pigments responsible for its signature blue-green color. A strain that looked unremarkable on the primary plate can reveal its true colors once it's given the right medium to do so.\n\nThat's the entire reason this medium exists: not every *P. aeruginosa* strain announces itself, and cetrimide agar is built specifically to coax out the signature even when a strain doesn't show it readily elsewhere.\n\n## Principle\n\nCetrimide agar, also called pseudosel agar, is used for the selective isolation and presumptive identification of *Pseudomonas aeruginosa*. Cetrimide is a quaternary ammonium salt acting as a cationic detergent; on contact with a bacterial cell, it releases nitrogen and phosphorus, denaturing membrane proteins and inhibiting a wide range of organisms, including *Pseudomonas* species other than *P. aeruginosa*. Cetrimide also enhances *P. aeruginosa*'s production of pyocyanin (blue pigment) and fluorescein\u002Fpyoverdin (yellow-green fluorescent pigment); the combination produces the organism's characteristic bright green appearance.\n\n## Composition of Cetrimide Agar\n\nFinal pH: 7.2 ± 0.2 at 25°C\n\n| Ingredients | Gram\u002Fliter |\n| --- | --- |\n| Enzymatic digest of gelatin | 20 g |\n| Magnesium chloride | 1.4 g |\n| Potassium chloride | 10 g |\n| Cetrimide (cetyltrimethylammonium bromide) | 0.3 g |\n| Glycerol | 10 mL |\n| Agar | 13.6 g |\n\n Cetrimide media also contains gelatin pancreatic digest which provides nitrogen, vitamins, minerals, and amino acids essential for growth.\n\n- Glycerol acts as the carbon source.\n- Magnesium chloride and potassium chloride enhance the production of pyocyanin and fluorescein.\n- [Agar is the solidifying agent.](\u002Fagar-properties-uses\u002F)\n- Cetrimide is the selective agent. It is a toxic substance that inhibits the growth of many bacteria.\n\n## Quality Control\n\nPerform QC on each new lot or shipment of media prior to using them. Inspect cetrimide agar for evidence of contamination, cracks, dehydration, and bubbles prior to storage and before use. Perform performance testing of the media with the following organisms;\n\n1. *P. aeruginosa* ATCC 27853—growth; yellow-green to blue pigment\n2. *Escherichia coli* ATCC 25922—inhibited\n\n## Media Preparation\n\n![Cetrimide Agar A. Positive, B. Negative - Cetrimide AgarA. Positive, B. Negative](https:\u002F\u002Fassets.microbeonline.com\u002Fblogs\u002FCetrimide-agar-positive-and-negative-test-result-300x220.jpg)Figure: Cetrimide Agar A. Positive, B. Negative\n\n1. Suspend 45.3 g of the medium and 10 ml of glycerol in one liter of purified water.\n2. Heat with frequent agitation and boil for one minute to completely dissolve the medium.\n3. Autoclave at 121°C for 15 minutes.\n4. Mix well and pour into sterile Petri plates.\n\n## Colony Characteristics\n\nAfter 18 – 48 hr of incubation, the plates should show isolated colonies in streaked areas and confluent growth in areas of heavy inoculation. Colonies may be presumptively identified as *Pseudomonas aeruginosa* when it exhibits a blue-green to green pigment and fluoresces under short wavelength (254 nm) ultraviolet light.\n\n> Note:  Certain strains of P. aeruginosa may not produce pyocyanin. Other species of Pseudomonas do not produce pyocyanin, but fluoresce under UV light. Some non-fermenters and some aerobic spore formers may exhibit a water-soluble tan to brown pigmentation on this medium. Serratia may exhibit pink pigmentation\n\n## Limitations\n\n- Growth on cetrimide agar alone is not sufficient for [identification of P. aeruginosa ](\u002Fpseudomonas-aeruginosa-infection-mortality-pathogenesis-and-diagnosis\u002F)to the species level, since other non-glucose-fermenting species such as *Achromobacter xylosoxidans subsp. xylosoxidans* and *Alcaligenes faecalis* may grow. Pigment must also be present.\n- Lack of growth on cetrimide agar does not rule out the identification of *P. aeruginosa*.\n\n## How to Remember?\n\n**Anchor for the two pigments: \"one glows, one just is.\"** Fluorescein needs short-wavelength UV light to reveal its yellow-green glow clearly. Pyocyanin doesn't need any special light at all, it's visible as a blue-green color under ordinary conditions. A strain can show either, both, or, occasionally, neither.\n\n**Anchor for the definitive confirmatory step: \"same glow, different comfort zone.\"** *P. fluorescens* and *P. putida* can mimic *P. aeruginosa*'s fluorescent glow on cetrimide agar, but only true *P. aeruginosa* tolerates growth at 42°C. When the pigment picture is ambiguous, this temperature test, not another look at the color, is what actually settles it.\n\n### Pulling the Real Bench Traps Together\n\n- **A non-pigmented colony doesn't rule out *P. aeruginosa*.** Some strains genuinely don't produce pyocyanin. The absence of visible pigment is a limitation of this medium, not proof the organism isn't there.\n- **Growth alone, without pigment, isn't sufficient for species-level identification.** Other non-glucose-fermenting genera, *Achromobacter xylosoxidans* and *Alcaligenes faecalis* among them, can also grow on cetrimide agar. Pigment has to be present for a presumptive call based on this medium alone.\n- **Fluorescence under UV light doesn't confirm *P. aeruginosa* on its own.** *Pseudomonas fluorescens* and *Pseudomonas putida* can also grow and fluoresce on cetrimide agar. The feature that actually separates them is growth at 42°C: true *P. aeruginosa* tolerates it, its fluorescent look-alikes don't.\n- **A pink-pigmented colony on cetrimide agar isn't automatically ruled out as contamination.** *Serratia* species can produce a pink pigment on this medium, worth remembering before dismissing an unexpected color as an artifact.\n\n## Cetrimide test\n\nTo perform cetrimide test, take isolated colonies of non-glucose-fermentative, Gram-negative rods that are suggestive of *P. aeruginosa and inoculate on cetrimide agar slant.*\n\n**Procedure**\n\n1. Streak the cetrimide agar slant back and forth with inoculum picked from the center of a well-isolated colony.\n2. Place cap loosely on tube.\n3. Incubate aerobically at 35 to 37°C for up to 7 days.\n4. Observe for growth and pigment.\n5. If no pigment is visible, examine growth under UV light for the presence of fluorescein.\n\n**Expected Results**\n\n1. Positive: Growth of organism is seen in the slant. Optionally a yellow-green (fluorescein) to dark blue-green (pyocyanin) color may be observed.\n2. Negative: No growth\n\n**Reporting**\n\n- Gram-negative oxidase-positive bacilli that grow on cetrimide agar and produce a blue-green (pyocyanin) pigment can be definitively identified as *P. aeruginosa.*\n- *Pseudomonas fluorescens* and *Pseudomonas putida* may also grow on cetrimide agar and may produce a fluorescent pigment but are separated from *P. aeruginosa* because they do not grow at 42°C.\n\n## Key exam facts in one table\n\n| Fact | Detail |\n| --- | --- |\n| Purpose | Selective isolation and presumptive identification of *Pseudomonas aeruginosa* |\n| Selective mechanism | Cetrimide, a cationic detergent, inhibits most other organisms while enhancing pyocyanin\u002Ffluorescein production in *P. aeruginosa* |\n| Two pigments | Pyocyanin (blue, visible in ordinary light) and fluorescein\u002Fpyoverdin (yellow-green, fluoresces under 254 nm UV) |\n| QC organisms | *P. aeruginosa* ATCC 27853 (growth, pigment); *E. coli* ATCC 25922 (inhibited) |\n| Key limitation 1 | Some *P. aeruginosa* strains don't produce pyocyanin; absence of pigment doesn't rule out the organism |\n| Key limitation 2 | Other genera (*Achromobacter*, *Alcaligenes*) can also grow; growth alone isn't sufficient for species ID |\n| Fluorescent look-alikes | *P. fluorescens*, *P. putida* |\n| Definitive differentiator | Growth at 42°C — true *P. aeruginosa* grows; its fluorescent look-alikes don't |\n| Unexpected pigment to watch for | *Serratia* species can produce a pink color on this medium |\n\n**References and further readings**\n\n1. Forbes, B. A., Sahm, D. F., & Weissfeld, A. S. *Bailey & Scott's Diagnostic Microbiology* (11th ed.).\n2. Linscott, A. J. (2016). *Clinical Microbiology Procedures Handbook* (4th ed.). ASM Press. \u003Chttps:\u002F\u002Fdoi.org\u002F10.1128\u002F9781683670438>\n\n3) Lowbury, E. J., & Collins, A. G. (1955). The use of a new cetrimide product in a selective medium for *Pseudomonas pyocyanea*. *Journal of Clinical Pathology*.",[46,49,52,55,58],{"question":47,"answer":48},"What is cetrimide agar used for?","Selective isolation and presumptive identification of Pseudomonas aeruginosa, based on its characteristic pigment production.",{"question":50,"answer":51},"Why does cetrimide agar enhance pigment production in P. aeruginosa?","Cetrimide, a cationic detergent, selectively inhibits most competing organisms while specifically enhancing P. aeruginosa's production of pyocyanin and fluorescein, making a strain's signature pigment more apparent than it might be on routine media.",{"question":53,"answer":54},"Does a lack of pigment mean an organism isn't Pseudomonas aeruginosa?","No. Some P. aeruginosa strains genuinely don't produce pyocyanin. Absence of pigment is a known limitation of this medium, not proof the organism is absent.",{"question":56,"answer":57},"How do you distinguish P. aeruginosa from P. fluorescens or P. putida on cetrimide agar?","By growth at 42°C. True P. aeruginosa tolerates that temperature; its fluorescent look-alikes, P. fluorescens and P. putida, do not.",{"question":59,"answer":60},"Can other organisms grow on cetrimide agar besides Pseudomonas species?","Yes. Achromobacter xylosoxidans and Alcaligenes faecalis can also grow, which is why growth alone, without pigment, isn't sufficient for a presumptive identification.",[],[],[64,71,78,83,87,91,96,101,105,109],{"slug":65,"name":66,"description":67,"image":68,"body":69,"postCount":70},"acharya-tankeshwar","Acharya Tankeshwar","Editor-in-chief","https:\u002F\u002Fassets.microbeonline.com\u002Fauthors\u002Ftankeshwar-acharya-author-microbeonline.jpg","***Tankeshwar Acharya, MSc (Medical Microbiology)***\n\n*Tankeshwar Acharya is an Assistant Professor in the Department of Microbiology at Patan Academy of Health Sciences (PAHS), Nepal, where he has been teaching and practicing clinical microbiology for over 14 years. He is the founder of Microbe Online, one of the leading free microbiology education resources on the web, covering bacteriology, mycology, parasitology, immunology, and clinical laboratory diagnostics written from direct experience in both the classroom and the diagnostic laboratory.*",433,{"slug":72,"name":73,"description":74,"image":75,"body":76,"postCount":77},"ashma-shrestha","Ashma Shrestha","SEO Copywriter and Science Communicator\nKathmandu, Nepal","https:\u002F\u002Fassets.microbeonline.com\u002Fauthors\u002Fashma-shrestha.png","Ashma Shrestha holds a Master of Science in Medical Microbiology from the Institute of Science and Technology (IOST), Tribhuvan University, Nepal, where she developed a strong foundation in virology, molecular biology, and diagnostic microbiology.\n\nShe now works as an SEO Copywriter at Resolution Digital, where she combines her scientific training with research-driven content strategy. She is certified in Google Analytics and Google Business Profile (GBP), and brings a data-informed approach to science communication writing content that is not only accurate but structured to reach and serve the students who need it most.\n\nAt microbeonline, Ashma contributes articles primarily in virology and molecular biology, areas she finds most compelling for their mechanistic depth and their growing clinical relevance. Her writing reflects the same standard the site is built on: factual rigor, clear explanation of the *why* behind microbiology concepts, and content that helps students move from memorization to genuine understanding.\n\nShe is passionate about making complex microbiological concepts accessible without sacrificing accuracy; a skill that sits at the intersection of her scientific training and her professional work in content and SEO.",81,{"slug":79,"name":80,"description":81,"image":38,"body":38,"postCount":82},"sushmita-baniya","Sushmita Baniya","Author \u002F Contributor",32,{"slug":84,"name":85,"description":81,"image":38,"body":38,"postCount":86},"samikshya-acharya","Samikshya Acharya",20,{"slug":88,"name":89,"description":81,"image":38,"body":38,"postCount":90},"alisha-tripathi","Alisha Tripathi",6,{"slug":92,"name":93,"description":94,"image":38,"body":38,"postCount":95},"aastha-shrestha","Aastha Shrestha"," Author \u002F Contributor",9,{"slug":97,"name":98,"description":99,"image":38,"body":38,"postCount":100},"guest-author","Guest Author","Guest Author \u002F Contributor",2,{"slug":102,"name":103,"description":81,"image":38,"body":38,"postCount":104},"srijana-khanal","Srijana Khanal",18,{"slug":106,"name":107,"description":99,"image":38,"body":38,"postCount":108},"dr-poonam-acharya","Dr. Poonam Acharya",1,{"slug":110,"name":39,"description":81,"image":38,"body":111,"postCount":112},"nisha-rijal","**Nisha Rijal** is a microbiologist and quality assurance specialist. She served for nearly 12 years as a microbiologist at the National Public Health Laboratory (NPHL), Nepal's national reference laboratory, and continues to work as a consultant microbiologist in international public health organization. ",51]