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    NCBA Nosema Check Protocol

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    Christopher Mayack
    1. The document provides instructions for checking bees for Nosema spores using a compound microscope. 2. Bees are collected, frozen to kill them, then their guts are removed and ground in water before being placed on a slide for examination under increasing microscope powers. 3. Under the highest power, Nosema spores should appear round or oval with thick coats, and Nosema ceranae spores are generally oval while Nosema apis are more round.

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    NCBA Nosema Check Protocol

    Uploaded by

    Christopher Mayack
    37 views4 pages
    1. The document provides instructions for checking bees for Nosema spores using a compound microscope. 2. Bees are collected, frozen to kill them, then their guts are removed and ground in water before being placed on a slide for examination under increasing microscope powers. 3. Under the highest power, Nosema spores should appear round or oval with thick coats, and Nosema ceranae spores are generally oval while Nosema apis are more round.

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    1. The document provides instructions for checking bees for Nosema spores using a compound microscope. 2. Bees are collected, frozen to kill them, then their guts are removed and ground in water before being placed on a slide for examination under increasing microscope powers. 3. Under the highest power, Nosema spores should appear round or oval with thick coats, and Nosema ceranae spores are generally oval while Nosema apis are more round.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as DOC, PDF, TXT or read online from Scribd
    Download as doc, pdf, or txt
    37 views4 pages

    NCBA Nosema Check Protocol

    Uploaded by

    Christopher Mayack
    1. The document provides instructions for checking bees for Nosema spores using a compound microscope. 2. Bees are collected, frozen to kill them, then their guts are removed and ground in water before being placed on a slide for examination under increasing microscope powers. 3. Under the highest power, Nosema spores should appear round or oval with thick coats, and Nosema ceranae spores are generally oval while Nosema apis are more round.

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    Attribution Non-Commercial (BY-NC)
    Download as DOC, PDF, TXT or read online from Scribd
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    Checking for Nosema Spores Using a Compound Microscope

    1. First check to see that all of the materials are present and that they were successfully transferred to you (materials identification sheet attached). 2. Collect at least 10 forager bees, place them in a zip lock baggy, and freeze kill (put in freezer for at least 20 minutes) them until you ready for diagnoses. 3. Thaw bees 5 minutes prior to dissection. 4. Take the small scissors from the checked out dissecting kit and decapitate the bee. 5. Then take the fine forceps (tweezers) and pull the stinger and the last abdominal segment out slowly. Typically the entire gut will come out as it is connected to the last abdominal segment. You may have to stick the tweezers inside the abdominal cavity and try to pull the gut out again if the gut breaks at all. 6. If you are having trouble removing the gut from the abdominal cavity (usually the case for old or rotten bees) then just clip off the abdomen where it meets the thorax of the bee and place the whole abdomen in the plastic grinder. 7. Add 2.5 milliliters (about teaspoon) of water per 10 bees to the grinder and grind until the solution looks uniform. If whole abdomen used the exoskeleton will remain intact no matter how much grinding is done, which is fine. 8. Make a wet mount slide by taking one drop of solution and placing it on a glass slide, slowly lower a cover slip over the slide at an angle to prevent air bubble formation. 9. Check to see that the microscope is on the lowest (4x) objective power (see microscope diagram). 10. Take the wet mount slide and place it on the stage of the microscope, in order to do this pull back the stage clip and then slowly release back into position so that the wet mount slide is secured on the stage. Center the slide so that the cover slip is below where the objective lens is. 11. Make sure the microscope is plugged in and switch on the light at the bottom of the microscope. 12. Use the course adjustment knob and the fine focus to put what is in the water into focus. Mature Nosema spores at this magnification will be tiny specs at best.

    13. Then revolve the objective lens mount to switch to the next highest power (10x) and bring the specs that you saw before into focus with the course adjustment and the fine focus. 14. Lastly, revolve the objective lens mount to the next highest power (40x) and then use the fine focus ONLY to bring the spores into focus because using the course adjustment may drive the objective lens into the slide and destroy the lens. 15. When in focus the spores should look uniform, have bluish tinge, and be round oval objects with thick spore coats. Under the highest power (40x) Nosema ceranae spores should look like this if the bee is highly infected

    Nosema apis spores are generally a little bit bigger and they are more roundish than oval. Spores from each species look so similar molecular techniques are needed in the scientific community to confirm the identity of the species. You can clean the slides and cover slips with water and both can be re-used after drying.

    Microscope Diagrams

    Objective lens (10x) Stage clip Objective lens (4x) Stage

    Course adjustment

    Controls to move the stage around

    Fine adjustment

    Control for brightness

    Power Switch

    Materials That Should be Present


    1. Dissecting Kit with probes, forceps, scissors, scalpel, ruler, and scalpel blades 2. Compound Light Microscope 3. Glass Slides 4. Cover Slips 5. Tissue Grinder that includes a plastic tube and a pestle for grinding Scalpel Blades

    Ruler

    Probes

    Forceps

    Scissors

    Scalpel

    Glass Slides

    Cover Slips

    Pestle

    Tissue Grinder

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