Violet Red Bile Agar (VRBA) 50 FAQs and 30 MCQs:
Violet Red Bile Agar (VRBA) is a selective and differential medium widely used in microbiology for the detection and enumeration of gram-negative bacteria, particularly Enterobacteriaceae like E. coli and Salmonella. Its unique formulation—containing crystal violet, bile salts, lactose, and neutral red dye—inhibits gram-positive bacteria while allowing lactose-fermenting gram-negative organisms to form distinctive red-purple colonies. VRBA is indispensable in food safety, water quality testing, and clinical diagnostics. To deepen your understanding, this article consolidates 50 frequently asked questions (FAQs) and 30 multiple-choice questions (MCQs) covering VRBA’s principles, applications, protocols, and limitations. Whether you’re a student, researcher, or lab professional, these resources will reinforce your knowledge and practical proficiency.
Violet Red Bile Agar (VRBA) 50 FAQs
What is Violet Red Bile Agar (VRBA)?
VRBA is a selective and differential medium used to detect and enumerate lactose-fermenting coliform bacteria in food, dairy, and water samples.
What type of bacteria does VRBA detect?
It detects gram-negative, lactose-fermenting coliforms (e.g., E. coli, Enterobacter aerogenes).
Why is VRBA used in food and dairy testing?
It helps assess microbial contamination, particularly by coliforms, which indicate poor hygiene or fecal contamination.
Can VRBA be used for clinical diagnosis?
No, VRBA is not intended for human disease diagnosis; it is for food, water, and dairy testing.
What are the key components of VRBA?
Peptic digest, yeast extract, lactose, bile salts, neutral red, crystal violet, and agar.
How does VRBA inhibit gram-positive bacteria?
Bile salts and crystal violet suppress gram-positive bacterial growth.
What indicates lactose fermentation on VRBA?
Red/pink colonies with bile precipitation (due to acid production detected by neutral red).
What do colorless colonies on VRBA indicate?
Non-lactose fermenters (e.g., Salmonella).
Can gram-negative non-coliforms grow on VRBA?
Yes, but they typically appear as colorless or pale colonies.
Do enterococci grow on VRBA?
Some may grow as tiny pink pinpoint colonies but are not target organisms
How is VRBA prepared?
Suspend 41.53g in 1L distilled water, boil to dissolve (do not autoclave), cool to 45°C, and pour plates.
Why should VRBA not be autoclaved?
Autoclaving degrades bile salts and crystal violet, reducing selectivity.
What is the recommended incubation temperature?
35°C (18–24 hrs); 32°C for dairy products.
How is VRBA used for water testing?
A 1 mL sample is mixed with VRBA, solidified, and incubated.
Can VRBA be stored after preparation?
Yes, but it should be kept refrigerated and used within a few days.
What do red colonies with bile precipitation mean?
Presumptive coliforms (e.g., E. coli).
What does mucoid pink growth indicate?
Likely Enterobacter aerogenes.
What if colonies are colorless?
Non-lactose fermenters (e.g., Salmonella).
How are E. coli colonies identified on VRBA?
Pink-red colonies with bile precipitation zones.
Can Staphylococcus aureus grow on VRBA?
No, it is mostly inhibited due to bile salts and crystal violet.
Is VRBA 100% specific for coliforms?
No, some non-coliform gram-negatives may grow.
What are VRBA’s limitations?
Some strains may grow poorly, and gram-positive inhibition is not absolute.
How are VRBA results confirmed?
Further biochemical, molecular, or mass spectrometry tests are needed.
Why might some coliforms not grow on VRBA?
Nutritional variations or sensitivity to bile salts/crystal violet.
Can VRBA detect Salmonella?
Yes, but they appear as colorless colonies (non-lactose fermenters).
Which organizations recommend VRBA?
FDA, APHA (American Public Health Association), and ISO standards.
Is VRBA used in pasteurization testing?
Yes, for checking coli-aerogenes content in milk and equipment.
What is the pH of VRBA?
7.4 ± 0.2 at 25°C.
Why is neutral red used in VRBA?
It acts as a pH indicator, turning red in acidic conditions (lactose fermentation).
What is the role of bile salts in VRBA?
They inhibit gram-positive bacteria and precipitate around lactose fermenters.
What if VRBA doesn’t solidify properly?
Ensure correct agar concentration (15g/L) and avoid overheating.
Why might colonies appear too small?
Insufficient incubation time or low bacterial load.
What if no colonies grow despite contamination?
Check for overheating during preparation or expired medium.
Can VRBA be used for high-fat dairy products?
Yes, but sample pre-treatment (e.g., dilution) may be needed.
How long should VRBA plates be incubated?
18–24 hours; extended incubation may allow non-target growth.
How does VRBA compare to MacConkey Agar?
Both detect lactose fermenters, but VRBA has bile salts for stronger gram-positive inhibition.
Is VRBA better than EMB Agar for coliforms?
VRBA is more selective, while EMB is more differential for E. coli.
Can VRBA replace chromogenic media for E. coli?
No, chromogenic media provide higher specificity.
Does VRBA detect Klebsiella?
Yes, as pink pinpoint colonies.
Is VRBA suitable for environmental swabs?
Yes, but confirmatory testing is recommended.
Why does Salmonella grow on VRBA but not ferment lactose?
VRBA supports growth of non-fermenters, which appear colorless.
What happens if VRBA is overheated?
Bile salts and crystal violet degrade, reducing selectivity.
Can VRBA detect gas production?
No, gas production requires Durham tubes in broth media.
Does VRBA work for frozen food testing?
Yes, but thawing and proper homogenization are crucial.
How does VRBA compare to Colilert® for water testing?
Colilert® is faster (24 hrs) and more specific, but VRBA is cost-effective for enumeration.
Can VRBA detect Shigella?
Rarely, as Shigella is a non-lactose fermenter (colorless colonies).
Is VRBA suitable for probiotic testing?
No, as it inhibits many beneficial bacteria.
What is the shelf life of VRBA powder?
Typically 2–3 years if stored properly.
Can VRBA be used for antibiotic susceptibility testing?
No, it is not designed for AST.
Why is sodium chloride included in VRBA?
To maintain osmotic balance for bacterial growth.
Violet Red Bile Agar (VRBA) 30 MCQs:
- What is the primary purpose of Violet Red Bile Agar (VRBA)?
a) To culture gram-positive bacteria
b) To isolate and enumerate lactose-fermenting coliforms✔
c) To diagnose human infections
d) To grow fungi in dairy products - Which component in VRBA inhibits gram-positive bacteria?
a) Lactose
b) Neutral red
c) Bile salts and crystal violet✔
d) Yeast extract - What indicates lactose fermentation on VRBA?
a) Colorless colonies
b) Pink/red colonies with bile precipitation✔
c) Black colonies
d) Greenish colonies - VRBA is NOT recommended for:
a) Water testing
b) Dairy product analysis
c) Clinical diagnosis of human disease✔
d) Food contamination studies - What is the pH of VRBA?
a) 5.4 ± 0.2
b) 7.4 ± 0.2✔
c) 8.0 ± 0.2
d) 6.2 ± 0.2
- Which sugar in VRBA helps differentiate lactose fermenters?
a) Glucose
b) Sucrose
c) Lactose✔
d) Fructose - What is the role of neutral red in VRBA?
a) Inhibits gram-positive bacteria
b) Acts as a pH indicator for acid production✔
c) Provides essential vitamins
d) Enhances gram-negative growth - Which ingredient maintains osmotic balance in VRBA?
a) Sodium chloride✔
b) Bile salts
c) Crystal violet
d) Yeast extract - Why is crystal violet added to VRBA?
a) To enhance lactose fermentation
b) To inhibit gram-positive bacteria✔
c) To provide nitrogen
d) To solidify the medium - What happens if VRBA is autoclaved?
a) Improves selectivity
b) Degrades bile salts and crystal violet✔
c) Enhances lactose fermentation
d) Kills all bacteria
- Red colonies with bile precipitation on VRBA indicate:
a) Non-lactose fermenters
b) Gram-positive bacteria
c) Lactose-fermenting coliforms✔
d) Fungi - Colorless colonies on VRBA suggest:
a) E. coli
b) Salmonella✔
c) Enterobacter
d) Staphylococcus - Mucoid pink colonies on VRBA are typical of:
a) E. coli
b) Enterobacter aerogenes✔
c) Staphylococcus aureus
d) Pseudomonas - Pinpoint pink colonies may indicate:
a) Klebsiella✔
b) Salmonella
c) Shigella
d) Yeast - Which organism is inhibited on VRBA?
a) E. coli
b) Enterobacter
c) Staphylococcus aureus✔
d) Salmonella
- How should VRBA be prepared?
a) Autoclave at 121°C
b) Boil but do not autoclave✔
c) Microwave until dissolved
d) Filter-sterilize - The correct incubation temperature for dairy products is:
a) 35°C
b) 37°C
c) 32°C✔
d) 42°C - VRBA is used for:
a) Antibiotic testing
b) Coliform enumeration✔
c Viral culture
d) Spore germination - A 1 mL sample is mixed with how much VRBA?
a) 5 mL
b) 10 mL✔
c) 15 mL
d) 20 mL - What is the incubation period for VRBA?
a) 6–8 hours
b) 12 hours
c) 18–24 hours✔
d) 48 hours
- VRBA may not inhibit:
a) All gram-positive bacteria✔
b) All gram-negative bacteria
c) Fungi
d) Viruses - Confirmatory tests for VRBA isolates include:
a) Gram stain and biochemical tests✔
b) PCR only
c) Microscopy only
d) No further testing needed - Enterococci on VRBA appear as:
a) Large red colonies
b) Pinpoint pink colonies✔
c) Black colonies
d) Green colonies - Overheating VRBA during preparation affects:
a) Only the agar
b) Bile salts and crystal violet✔
c) Lactose fermentation
d) Neutral red stability - VRBA is recommended by:
a) FDA and APHA✔
b) WHO only
c) CDC only
d) USDA only
- Which organism grows as colorless colonies on VRBA?
a) E. coli
b) Enterobacter
c) Salmonella✔
d) Klebsiella - VRBA is unsuitable for detecting:
a) Coliforms in milk
b) Gram-negative pathogens
c) Probiotic bacteria✔
d) Water contaminants - The bile precipitation zone is due to:
a) Acid production✔
b) Alkaline byproducts
c) Gas formation
d) Protein breakdown - Which is a limitation of VRBA?
a) Detects only gram-positive bacteria
b) Cannot differentiate coliform species✔
c) Kills all bacteria
d) Requires anaerobic incubation - Sodium chloride in VRBA functions to:
a) Inhibit gram-negatives
b) Maintain osmotic balance✔
c) Enhance lactose fermentation
d) Act as a pH indicator
Conclusion
Mastering Violet Red Bile Agar (VRBA) is crucial for accurate microbiological analysis in both academic and industrial settings. The 50 FAQs address conceptual nuances—from the role of selective agents to troubleshooting common protocol errors—while the 30 MCQs provide an interactive tool to test comprehension and application skills. Together, these resources demystify VRBA’s methodology, enhance diagnostic confidence, and underscore its significance in safeguarding public health. For further study, refer to standards like the FDA’s Bacteriological Analytical Manual (BAM) or ISO 6579. Keep exploring, stay curious, and leverage these FAQs and MCQs to excel in your microbiological endeavors!
