Saturday, May 18, 2019
Diagnosing Ebola Virus Disease (EVD) shortly after infection can be difficult. Early symptoms of EVD such as fever, headache, and weakness are not specific to Ebola virus infection and often are seen in patients with other more common diseases, like malaria and typhoid fever.
To determine whether Ebola virus infection is a possible diagnosis, there must be a combination of symptoms suggestive of EVD AND a possible exposure to EVD within 21 days before the onset of symptoms. An exposure may include contact with:
- blood or body fluids from a person sick with or who died from EVD
- objects contaminated with blood or body fluids of a person sick with or who died from EVD
- infected fruit bats and primates (apes or monkeys)
- semen from a man who has recovered from EVD
Microscopical examination of faecal specimens
Examine immediately those specimens containing blood and mucus and those that are unformed because these may contain motile trophozoites of E. histolytica or G. lamblia
Examination of dysenteric and unformed specimens
1 Using a wire loop or piece of stick, place a small amount of specimen, to include blood and mucus on one end of a slide. Without adding saline, cover with a cover glass and using a tissue, press gently on the cover glass to make a thin preparation.
2 Place a drop of eosin reagent on the other end of the slide. Mix a small amount of the specimen with the eosin and cover with a cover glass.
Value of eosin: Eosin does not stain living trophozoites but provides a pink background which can make them easier to see.
3 Examine immediately the preparations microscopically, first using the 10 objective with the condenser iris closed sufficiently to give good contrast. Use the 40 objective to identify motile trophozoites, e.g. E. histolytica amoebae or G. lamblia flagellates
Note: The eggs of Schistosoma species and T. trichiura, and the trophozoites of B. coli can also result in specimens containing blood and mucus.
Examination of semi-formed and formed faeces
1 Place a drop of fresh physiological saline on one end of a slide and a drop of iodine on the other end. To avoid contaminating the fingers and stage of the microscope, do not use too large a drop of saline or iodine.
2 Using a wire loop or piece of stick, mix a small amount of specimen, about 2 mg, (matchstick head amount) with the saline and a similar amount with the iodine. Make smooth thin preparations. Cover each preparation with a cover glass.
Important: Sample from different areas in and on the specimen or preferably mix the faeces before sampling to distribute evenly any parasites in the specimen. Do not use too much specimen otherwise the preparations will be too thick, making it difficult to detect and identify parasites.
3 Examine systematically the entire saline preparation for larvae, ciliates, helminth eggs, cysts, and oocysts. Use the 10 objective with the condenser iris closed sufficiently to give good contrast. Use the 40 objective to assist in the detection and identification of eggs, cysts, and oocysts.Always examine several microscope fields with this objective before reporting ‘No parasites found’.
4 Use the iodine preparation to assist in the identification of cysts.
5 Report the number of larvae and each species of egg found in the entire saline preparation as follows:
Scanty . . . . . . . . . . . . . . . . 1–3 per preparation
Few . . . . . . . . . . . . . . . . . 4–10 per preparation
Moderate number . . . . 11–20 per preparation
Many . . . . . . . . . . . . . . . 21–40 per preparation
Very many . . . . . . . . . over 40 per preparation
Identification of larvae: In a fresh faecal specimen, S. stercoralis is the only larva that will be found. It can be easily detected in a saline preparation by its motility and large size. If the specimen is not fresh, S. stercoralis will require
differentiation from hookworm larvae.
Identification of helminth eggs: Eggs are recognized by their:
– colour (colourless, pale yellow, brown),
– morphological features.
Note: Some helminths also have a limited geographical distribution.
Identification of intestinal flagellates: The trophozoite of G. lamblia and its differentiation from non-pathogenic flagellates that can be found in faeces.
Identification of ciliates: B. coli is the only ciliate found in human faeces (rare infection).
Identification of cysts and oocysts: In saline and eosin preparations, protozoan cysts and oocysts can be recognized as refractile bodies (shine brightly when focused). Cysts can be identified by their shape, size, nuclei, and inclusions as seen in an iodine preparation Iodine stains nuclei and glycogen but not chromatoid bodies. Burrows stain or Sargeaunt’s staincan be used to stain chromatoid bodies. The cysts of E. histolytica/E. dispar and those of G. lamblia and their differentiation from non-pathogenic or other species.
Faecal leukocytes: The presence of polymorphonuclear neutrophils (pus cells) in faeces is mainly associated with inflammatory diarrhoea caused by bacteria.
Non-parasitic structures found in faeces: Care must be taken not to report as parasites those structures that can be normally found in faeces such as muscle fibres, vegetable fibres, starch cells (stain blue-black with iodine), pollen grains, fatty acid crystals, soaps, spores, yeasts, and hairs. Large numbers of fat globules may be seen in faeces when there is
malabsorption. Charcot Leyden crystals (breakdown products of eosinophils) can sometimes be seen in faeces (also in sputum) in parasitic infections. They appear as slender crystals with pointed ends, about 30–40 m in length.
Wednesday, May 15, 2019
continuation of part two
read tissue processing part one here
Properties of embedding media
Paraffin wax additives
MODIFIED PARAFFIN WAXES
Alternative embedding media
Embedding tissues in paraffin wax
|10% Formalin||1 h||38°C|
|10% Formalin||1 h||38°C|
|50% Alcohol/formalin||1 h||38°C|
|70% Alcohol||1 h||38°C|
|95% Alcohol||1 h||38°C|
|95% Alcohol||40 min||38°C|
|100% Alcohol||1 h||38°C|
|100% Alcohol||40 min||38°C|
Equipment required for Paraffin section cutting
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