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Macmaster counting Technique ppt

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					McMaster egg counting technique:
Dr.Kedar Karki

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Purpose
• The McMaster technique is used for demonstrating and counting helminth eggs in faecal samples. • It is the most widely employed method for this purpose.

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Principle
• The McMaster technique uses a counting chamber which enables a known volume of faecal suspension (2 x 0.15 ml) to be examined microscopically.

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Principle
• Thus, if a known weight of faeces and a known volume of flotation fluid are used to prepare the suspension, then the number of eggs per gram of faeces (e.p.g.) can be calculated.

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Principle
• The quantities are chosen so that the faecal egg-count can be easily derived by multiplying the number of eggs under the marked areas by a simple conversion factor.

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Principle
• The McMaster chamber has two compartments, each with a grid etched onto the upper surface.

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Principle
• When filled with a suspension of faeces in flotation fluid, much of the debris will sink while eggs float to the surface, where they can easily be seen and those under the grid counted.

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McMaster chamber

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McMaster egg counting technique: Equipment
• • • • • Two beakers or plastic containers Balance Tea strainer, cheesecloth or dental napkin Measuring cylinder Stirring device (fork, spatula, tongue depressor)

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Equipment List
• Pasteur pipettes and rubber teats • Flotation fluid (choice of solution dependant on species expected to be present and availability of reagents) • McMaster counting chamber • Compound microscope

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Equipment List

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Procedure step 1
• Weigh 4 grams of faeces and place into a container. Warning!!!Faeces must be fresh.

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Procedure step 2
• Add 56 ml of your chosen flotation fluid.

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Procedure step 3
• Stir the contents of the beaker thoroughly with a fork, tongue depressor or spatula.

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Procedure step 4
• Filter the faecal suspension through a tea strainer or double layer of cheesecloth or dental napkin into the second container.

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Procedure step 5
• Stir the filtrate in container two with a Pasteur pipette. • Using the pipette withdraw a sub-sample as the filtrate is being stirred.

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Stir fluid and fill first compartment of the McMaster counting chamber with the sub sample.

Procedure step 6

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Procedure step 6
• Stir fluid again and fill second chamber with another sub sample.

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Procedure step 7
• It is important to leave the chamber to stand to allow the eggs to float to the surface and the debris to go to the bottom of the chamber.

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Allow the counting chamber to stand for 5 minutes.

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Examination step 1
• Examine the subsample of the filtrate under the compound microscope at 10 x 10 magnification.

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Examination step 2
• Identify and count all eggs within the engraved area of both chambers.

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McMaster egg counting technique: Calculation of results
• The number of eggs per gram can be calculated as follows: • Count the number of eggs within the grid of each chamber, ignoring those outside the squares • Multiply the total by 50 – this gives the eggs per gram of faeces (e.p.g.)

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For example:
• 12 eggs seen in chamber 1 and 15 eggs seen in chamber 2= (12 +15) x 50 = 1350 e.p.g.

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McMaster egg counting technique: Interpretation of results
• When interpreting McMaster results, it must be remembered that a number of factors can influence the occurrence, recognition or numbers of helminth eggs found in a faecal sample. In particular, the number of eggs is not necessarily indicative of the number of worms present. Reasons for this include:
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McMaster egg counting technique: Interpretation of results
• Eggs are produced only by fertile adult female (or hermaphrodite) worms and will, therefore, be absent in immature or single sex infections • The daily output of eggs by fertile females is influenced by host-physiological factors such as stress or lactation ( increased ) or immunity ( decreased ) • Chemotherapy can also affect egg-production e.g. corticosteroids ( increased ) or sub-lethal anthelmintic doses ( decreased )
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McMaster egg counting technique: Interpretation of results
• The concentration of eggs (per gram of faeces) is influenced by the daily volume of faeces being produced by the host, the rate of passage by the ingesta through the intestine, and the distribution of eggs throughout the faecal mass

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McMaster egg counting technique: Interpretation of results
• Some types of eggs are heavier than others and may not float well in solutions of lower specific gravity (e.g. Fasciola) • Some eggs from different species are indistinguishable (particularly trichostrongylids and strongylids). This complicates clinical interpretation as some species (e.g. Haemonchus) produce many more eggs per day than others (e.g. Ostertagia).
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Egg identification (common species)
• • • • • Trichuris ovis Nematodirus battus Nematodirus filicollis Strongyloides papillosus Toxocara vitulorum •

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Ruminant eggs: Trichuris

Trichuris ovis Length 70-80 µm Width 30-42 µm Thick-walled Lemon-shaped with polar plugs Granular contents, no blastomeres

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Ruminant eggs: Nematodirus
Ellipse Poles sharply curved 2-8 blastomeres surrounded by wide fluid-filled cavity Nematodirus battus Length 164 µm Width 72 µm Shell thin, brown, parallel sides

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Nematodirus filicollis
Length 150 µm Width 75 µm Shell thin, colourless

Nematodirus helvetianus Length 212 µm Width 97 µm Shell thin, colourless

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Ruminant eggs: Strongyloides
Strongyloides papillosus Length 47-65 µm Width 25-26 µm Broad ellipse, slightly flattened poles Shell thin, colourless Embryonated, L1 larva present

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Ruminant eggs: Toxocara
Toxocara vitulorum Length 69-95 µm Width 60-77 µm Subglobular Thick albuminous shell Granular contents Pitted surface to shell

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Ruminant eggs: Moniezia
Thick shell Irregular shape, tri- or quadrangular Embryonated with pyriform apparatus

M. expansa [mainly sheep]: Size 50-60 µm Triangular to pyramidal
M. benedeni [mainly cattle]: Size 80-90 µm quadrangular 35

Ruminant eggs: Fasciola
Fasciola hepatica Length 130-145 µm Width 70-90 µm Regular ellipse Thin shell Operculum at one pole Granular yellowish-brown contents filling whole egg

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Ruminant eggs: Paramphistomum
Paramphistomum Length 160 µm Width 90 µm Operculum on one pole Pale grey to greenish colour

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Ruminant eggs: Strongyles

Strongyle Approximately 80 µm long Thin-shelled Broad ellipse Barrel-shaped side walls Blastomeres present, number vary

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