![Page 1: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/1.jpg)
Bacteria and other microbes have particular requirements for growth
When they reside in and on our bodies or in the environment, they harvest their food from us or from the environment
When we grow bacteria in lab, we are essentially creating a captive environment for bacteria – like a bacteria zoo. So we must provide the bacteria we grow in lab with all of the materials that they need to grow
![Page 2: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/2.jpg)
In this lab we learn about different types of media that are used to grow bacteria. Some types of media will grow just about any type of bacteria and the others are more selective and only grow specific types of bacteria
![Page 3: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/3.jpg)
There are generalized media, like (Nutrient agar)that will grow many different types of microbes.
This media is the type most often used to culture bacteria
![Page 4: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/4.jpg)
culture medium that allows the growth types of organisms,
while inhibiting the growth of other organisms
EMB (Eosin Methylene Blue) dyes inhibit Gram (+) bacteria selects for Gram (-) bacteria
Example:
![Page 5: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/5.jpg)
Differentiates between different organisms growing on the same plate Example: Blood agar
culture medium that includes ingredients, such as chemical indicators , that produce observable differences between species of bacteria
![Page 6: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/6.jpg)
This media is differential because:
Certain bacteria produce enzymes (hemolysins…) that act on the red cells to produce either:
Beta hemolysis: Enzymes lyse the blood cells completely, producing a clear area around the colony.
Alpha hemolysis: Incomplete hemolysis produces a greenish discoloration around the colony
Gamma hemolysis: No effect on the red cells.
![Page 7: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/7.jpg)
There are also (selective)
and
(differential)media
![Page 8: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/8.jpg)
MacConkey’s is both a selective & differential media.
1. MacConkey’s is selective media because it inhibits the
growth of some organisms [Gram positive bacteria].
2. MacConkey’s is differential media - “lactose fermenters” bacteria will grow in red colonies while” non-lactose fermenters” will be colorless and clear.
So if there are colonies of bacteria growing on MacConkey’s, it’s understood that they are Gram- If those colonies are colorless, they are not lactose fermenters If the colonies have a pinkish appearance, they are lactose fermenters
![Page 9: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/9.jpg)
![Page 10: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/10.jpg)
left: no lactose fermentation
right: lactose fermentation
MacConkey Agar
![Page 11: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/11.jpg)
FORMS OF CULTURE MEDIA
broth liquid medium
most common growth media for microorganisms are ( nutrient broths ( liquid nutrient medium
![Page 12: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/12.jpg)
Solid media commonly contain 1.5% agar per weight to solidify the liquid media.
After sterilization, the media is poured into sterile Petrie plates.
Agar is liquefies at 100 C and solidifies at 40 C
![Page 13: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/13.jpg)
General Media:
Nutrient Broth and Nutrient Agar
![Page 14: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/14.jpg)
or After autoclaving the media(in tube) for 20 minutes, the tubes are placed in a slanted position to allow the agar to solidify. These tubes are called slants
![Page 15: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/15.jpg)
Slant tubes:
are tubes containing a nutrient medium plus a solidifying agent, (agar-agar. The medium has been allowed to solidify at an angle in order to get a flat inoculating surface
Agar Plate:
are sterile petri plates that are filled with a melted sterile agar medium
Microorganisms grow on the surface of agar plates and slants
![Page 16: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/16.jpg)
140 Micro
Lab 4: Isolation of microbes from
different environments
![Page 17: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/17.jpg)
Isolation of micro organisms from various environments
![Page 18: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/18.jpg)
1. Isolation –separating one species from another.
2. Inoculation – introduction of a sample into a container of media to produce a culture of observable growth.
3. Incubation – conditions that allow growth eg., temperature ,humidity etc..
4. Inspection.
5. Identification.
![Page 19: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/19.jpg)
Environmental Sample Normal Flora Samples
Surface samples are normally taken using sterile swabs
Applying oral sample to surface of agar
![Page 20: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/20.jpg)
![Page 21: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/21.jpg)
Petri plates with media
Soil
Rotten fruit
Yoghurt
Mouth swab etc
Incubator……
![Page 22: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/22.jpg)
Bacteria Fungi
![Page 23: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/23.jpg)
Yoghurt
Mouth
Hand
Isolation of Bacteria
• Put a drop of diluted yoghurt on the solid media plate of bacteria . • Incubate at 37 °C for 1 day.
• Take some saliva with the help of a cotton swab. • Inoculate it on the media and incubate.
•Touch the surface of the solid media plate . • Incubate 37 °C for 1 day. .
![Page 24: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/24.jpg)
Agar plates are stored upside down to prevent condensation.
![Page 25: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/25.jpg)
These plates of bacteria will be incubated at 37° C for 24 hours and then stored at refrigerator until next week when you will observe for results.
![Page 26: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/26.jpg)
Air
Soil
fruit
Isolation of Fungi
• Expose the prepared plate of solid media in air for 5 min • Close the lid and incubate at 28 °C. • After 2-5 days fungus observe for grow
• Sprinkle a pinch of soil on the solid media plate • Close the lid and incubate at 28 °C • After 2-3 days fungus observe for growth.
• Clean the rotten part of fruit with alcohol • Cut a piece and inoculate it on the media plate •Then incubate ,after 2-5 days fungus will grow
![Page 27: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/27.jpg)
These plates of fungus will be incubated straight at 28° C for 24 hours and then stored at refrigerator until next week when you will observe for results.
![Page 28: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/28.jpg)
![Page 29: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/29.jpg)
Bacteria Fungi
![Page 30: Bacteria and other microbes have particular When …fac.ksu.edu.sa/sites/default/files/140_lmml_lrb_1.pdfLab 4: Isolation of microbes from different environments Isolation of micro](https://reader036.vdocuments.us/reader036/viewer/2022071020/5fd4a1566057887b0135cf1e/html5/thumbnails/30.jpg)
الشهرانيشروق .أ