9.1: Mannitol Salt Agar Media, MSA
- Perform aseptic techniques while streaking or inoculating bacteria onto mannitol salt agar plates to prevent contamination.
- Understand the principle behind the mannitol salt agar test, including its selective and differential properties.
- Identify the key components of mannitol salt agar and their roles in the selective and differential processes.
- Describe the significance of mannitol salt agar in the isolation and differentiation of Staphylococcus species, particularly Staphylococcus aureus .
- Interpret the results of the mannitol salt agar test based on the color changes of bacterial colonies and the fermentation of mannitol.
- Compare and contrast the growth of salt-tolerant bacteria versus non-salt-tolerant bacteria on mannitol salt agar plates.
Mannitol Salt Agar
Mannitol salt agar (MSA) is a microbiological medium that is both selective and differential. Selective means the medium will only allow certain microorganisms to grow. Differential means the medium will show some characteristics of the microorganisms growing on it and can be used to differentiate between different species.
MSA is selective since it has a high salt concentration and will only allow halophilic (salt-loving species) or halotolerant (salt-tolerant) species to grow on it. This medium contains 7.5% NaCl (salt), whereas typical media contains about 0.5% NaCl.
- species that grow on MSA: salt resistance positive
- species that do not grow on MSA: salt resistance negative
MSA is differential since it will detect acid production as a result of the fermentation of mannitol (sugar in the medium). The medium contains a pH indicator, phenol red, that is red-orange when the pH is neutral (around pH 7) (Fig.1). If a species ferments mannitol and produces acid, the pH of the medium decreases, and phenol red will become yellow (Fig.1).
- medium remains red-orange after growth: mannitol fermentation negative
- medium is yellow after growth: mannitol fermentation positive
Laboratory Instructions
- Obtain an MSA plate and label it with your name/group name, course section, date, and "MSA".
- Using a sharpie, on the bottom of the Petri plate, divide your plate into three (3) sections. Label the sections accordingly (e.g., Staphylococcus saprophyticus, Staphylococcus epidermidis, and Escherichia coli.
- With a sterile cotton swab, aseptically inoculate each section of the plate with the corresponding bacterial species. Make sure to use a new cotton swab for each species.
- Parafilm your inoculated plate.
- Invert the Petri plate and incubate for 24-48 hours at 37 °C.
- Observe, record, and interpret results.
Methicillin-Resistant Staphylococcus aureus
Variable Staphylococcus species are found on human skin. Staphylococcus epidermidis is part of our normal microbiota and helps prevent colonization by pathogenic species. S. epidermidis is unlikely to causes disease unless the bacteria enters the body through wounds or indwelling devices. However, other species like Staphylococcus aureus are highly contagious and resistant no matter their location on or in the human body. M ethicillin- r esistant S taphylococcus a ureus (MRSA) is a common infection found in healthcare settings that has the potential to cause severe outbreaks if not identified early-on. Untreated MRSA can lead to pneumonia, sepsis, and even death. (CDC, 2024) MRSA is estimated to cause 70,000 infections and 9,000 deaths per year. (CDC, 2024)
One way to distinguish S. epidermidis from S. aureus is by growing specimen cultures on m annitol s alt a gar plates (MSA). Both species can grow on MSA plates as they are halophiles or salt lovers. However, the bacteria will have different effects on the media color, allowing us to differentiate between them.
S. aureus is able to ferment (break down) mannitol present in the MSA plate. This fermentation process produces acid, causing the pH of the agar to lower and turn yellow. S. epidermidis can not break down mannitol and will not change the pH of the media. The media will stay the same original pink/red color. Thus, MSA plates are a quick diagnostic tool to determine if an infection contains S. aureus and could lead to a MRSA outbreak.
Attributions
- "Microbiology Laboratory Manual: Labs, 1.29 Mannitol Salt Agar" by Dr. Rosanna Hartline , West Hills College Lemoore, LibreTexts: Biology is licensed under CC BY-NC 4.0
- Chapter Image: Mannitol salt agar (MSA) with growth of S. aureus, CoNS and no growth of E. coli.jpg by Ajay Kumar Chaurasiya is licensed under CC BY-SA 4.0
- General Microbiology Lab Manual (Pakpour & Horgan) by Nazzy Pakpour & Sharon Horgan is licensed under CC BY-SA 4.0
- Microbiology Labs I by Delmar Larsen is licensed under an undeclared license
- "Microbiology Textbook: Chapter 21, Skin and Eye Infections" by Openstax , Digital ISBN: 13: 978-1-947172-23-4 is licensed under CC BY 4.0
- "Methicillin-resistant Staphylococcus aureus (MRSA): Basics" by Centers for Diseases Control and Prevention , Use of CDC Materials Statement is in the Public Domain
- "Methicillin-resistant Staphylococcus aureus (MRSA)" by Centers for Diseases Control and Prevention , Use of CDC Materials Statement is in the Public Domain