Microbiology Laboratory Turkey

Mikrobiyoloji Ile Ilgili Tüm Konuların Kısa ve Öz Anlatımları. Microbiology Lab Information.

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MicroLab

endo agar etiketine sahip kayıtlar gösteriliyor. Tüm kayıtları göster
endo agar etiketine sahip kayıtlar gösteriliyor. Tüm kayıtları göster

25 Şubat 2019 Pazartesi

Şubat 25, 2019

Membrane Filter Technique | Membrane Filtration Method (Microbes)

Membrane Filter Technique | Membrane Filtration Method

Microbes are considered as an important microorganisms, its have different types, sizes and uses, Bacteria for example can be found everywhere; it has sizes from 0.2 µm to around 3 µm. Coliform bacteria testing known as the most microbes detected for water, and the presence of Escherichia coli (size: 1 µm) is an indication for the presence of pathogens that leads to water contamination. Membrane Filtration Technique is the popular method for the detection of many microbes, it can detect up to 300 Colony Forming Unit (CFU), and it has the ability to detect different types according to media and sample types.

Membrane Filtration need the following equipment in general for work:
Laminar Flow Hood (Optional).
Vacuum Pump with Rubber Tube.
Suction Flask 1 L.
Nitrocellulose Filter, Pore Size: 0.45 µm.
Microbiology Media.
Sterile Distilled Water.
70% Ethanol.
Flame.
Forceps.
Incubator.
Autoclave.
Tissue Paper.
Sterile cups or Sterile bags.
Suction Funnel.
Sample Collection:
Sample types and collection methods are different, Samples may be water, beverage, swab, sugar, or syrup. In this article an explanation of water sample will done. Water samples are collected from tanks as follow:
Wet piece of tissue paper by 70% ethanol and wipe the valve of the tank.
Open the valve to let some water out to erase any excess of ethanol then close it.
Open the sterile bag or sterile cup in the same time open the valve and take 100 ml then close the valve and the bag / cup.
Transfer the sample to Microbiology lab immediately.
Water Samples can be stored in refrigerator at 4°C to inhibit the growth and the rise in microbial numbers.
Working Procedures:
    Disinfect Laminar flow hood and clean all equipment using 70% ethanol.
    Turn on UV lamp and take time according to laminar hood type.
    Turn on air flow (that will turn off UV lamp), then clean all the tools again by 70% ethanol.
    Enter water sample inside the hood and clean it by 70% ethanol.
    Put the filter on the suction funnel and pour half of the sample then turn on the pump and pour the other half.
    Turn off the pump when all sample content filtered.
    Mount the filter on media plate and incubate it on the proper temperature inside the incubator.




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    13 Kasım 2018 Salı

    Kasım 13, 2018

    ENDO Agar

    ENDO AGAR

    Endo Agar is a selective medium recommended for confirmation of the presumptive test for members of the coliform group from clinical and non-clinical samples.
    There are actually two similar Endo agars, Endo agar and LES Endo agar. They differ mainly in that LES Endo agar has added nutrients that support a wider growth of strains, but otherwise display similar results. They are both Selective & Differential Media.  The selective and differential aspects are due to sodium sulfte, basic fuchsin, and lactose in each medium.  
    They are Selective because they encourage some bacteria to grow while inhibiting others.  Sodium sulfte and basic fuchsin generally inhibits Gram positive bacteria from growing (a few will grow) and generally allowing Gram negative organism to grow (some will not grow).
    • If good growth, generally you have a Gram negative bacteria.  
    • If no growth or if very poor growth, you most likely have a Gram positive.


    Both media are Differential because if lactose is utilized, it will create the intermediate acetaldehyde which reacts with the sodium sulfte and basic fuchsin will color the colonies pink/red from the acids produced during lactose fermentation. These plates differentiate between species capable of utilizing  lactose and those unable to metabolize it.
    • If growth is colorless, beige or not pink, lactose is not utilized and it is probably a Gram negative noncoliform.
    • If good growth that is pink/red, lactose is utilized and the bacteria is probably a Gram negative coliform.
    • If good pink/red growth that has a bright metallic sheen, lactose is highly utilized and the bacteria is probably a Gram negative coliform.
    • (If poor or no growth, recall it is probably a Gram positive organism.)

    Purpose: to select for and isolate Gram negative organisms, to check for the presence of coliforms, and to differentiate among the family of Enterobacteriaceae. Its main use is to detect for fecal contamination especially of water and dairy products. Both the Gram negative selection and the detection of coliforms is imperfect, a small percentage of strains do not act as expected.


    Procedure:  
    • Appropriately label you plates. Mark the dish bottom into thirds and label what species will be in each section.  It is best to label the side of the bottom plate or write on the bottom in tiny letters so that you will be able to observe the growth clearly.
    • Inoculate one third with your unknown streak for isolated colonies if possible.  Isolated colonies work best for this test but it is not essential.
    • Inoculate one third with E.coli, and the other third with Staphylococcus epidermidis.  Generally it is best to keep E.coli well away from the other cultures because it may overgrow them.
    • You will want to compare the growth on these plates to growth inoculated on a general purpose media such as NA. or TSA.
    • Incubate your plates upside down in the incubator at 35-37 C for 1-2 days.
    • Slow growing species may require a day or two of additional growth.

    Results  
    Compare your growth on these plates to growth on a general purpose media. (If no growth on the general purpose media, discard your results and repeat the test.)  Look for the presence or absence of growth and if there is growth if it is reduced from normal.  It there is growth, check to see if the growth is pink or red in color. One predicts that if an organism grows well on Endo Agar, it is most likely Gram negative, otherwise it is most likely Gram positive.  And if the growth is good and pink/red, it is most likely a coliform, otherwise it is likely a noncoliform.  Also look if the colonies have a metallic sheen, if they do the organism highly utilizes lactose.  Incidentally, a major difference between a coliform and a noncoliform in the family Enterobacteriaceae is simply if it can metabolize lactose.


    Notes:
    Should confirm their tentative conclusions from this test by performing a Gram stain and testing for lactose utilization.  Other species of bacteria may be used as positive and negative controls. E.coli will have good pink/red growth with a metallic sheen, Enterbacter aerogenes will have good pink growth, and S.epidermidis will grow poorly if at all.  Instructors may wish to have students test several selective and differential media at the same time and then compare them all to growth on one general purpose media plate to save on the cost. 

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