Parasitology Lab 1 & 2 QS7 PDF

Introduction to Parasitology, Collection of Stool Sample, Preserve Stool Specimens Lab 1&2 Mazin Mehdi 2024 - 2025 Parasite: the parasite is a living creature (single or multicellular) that depends apart or all its life cycle in symbios...

Introduction to Parasitology, Collection of Stool Sample, Preserve Stool Specimens Lab 1&2 Mazin Mehdi 2024 - 2025 Parasite: the parasite is a living creature (single or multicellular) that depends apart or all its life cycle in symbiosis with another different living creature to get food and in habitation source. Host: the host is the creature that the parasite spends a period or a part of its life inside on the host.  Infection stages of parasites (cause diseases) : The parasite passes with many roles through its life and the infective stage is considered the most important stage especially epidemiology side. This stage is different from parasite to parasite and may be one of the following types 1- Ovum: as in many parasitic worms as in Ascaris worms Schistosoma mansoni and Tape worms. 2- larva: it may be free in living like larva of worms as Hookworms or swim in water as in blood fluke or exist in the blood of the host as Microflora larva. 3- Cyst: as in Fasciola hepatica and some primary parasites like Amoeba dysentery 4- Adult: as in Entamoeba gingivalis and Trichomonas vaginalis. Types of Samples: 1- Blood 2 -Stool and urine 3 -Anal swab, duodenal aspirate, sputum 4 -Biopsy  COLLECTION OF STOOL SAMPLE forms of protozoa and helminthes may be detected from a properly collection and prepared stool specimen. The acceptable amount of stool required for parasite test is 2 to 5 g, often referred to as the size of a walnut.  The conditions of stool sample collection are: 1- Stool specimens should be collected in a clean, watertight container with a tight-fitting lid. 2- Urine should not be allowed to contaminate the stool specimen because it has been known to destroy some parasite. 3- Stool should not be retrieved from toilet bowl water because free-living protozoa and nematodes may be confused with human parasites. In addition, water may destroy select parasites. 4-The specimen container should be labeled with the patient’s name and identification number, the physician’s name, and the date and time of sample collection. 5- Stool samples from patients whose therapy includes barium, bismuth, or mineral oil should be collected prior to therapy or not until 5 to 7 days after the completion of therapy. 6- Collection of specimens from patients who have taken antibiotics or antimalarial medications should be delayed for 2 weeks following therapy. 7-The typical stool collection protocol consist of Three specimens, one specimen collected every other day or total of three collected in 10 days. One exception is in the diagnosis of Amebiasis, in which up to 6 specimens in 14 days is acceptable. 8- It is recommended that liquid specimens be examined within 30 minutes of passage.  Laboratory of medical parasites The accurate laboratory diagnosis for the parasitic diseases is considered on important side to obtain ways to the right treatment and laboratory diagnosis contains the following 1-Direct examination 2- Indirect examination 1- Direct examination Is considered the most important laboratory diagnosis because it is the base of the primary diagnosis for the parasite and watching it under the microscope.this consists the following diagnosis A-Direct stool examination The stool is considered an infection source because it contains on microorganisms that causes diseases so that it Parasites must deal with these samples cautiously and the direct diagnosis for the stool has two ways. Macroscopic Examination Specimens are examined to determine the consistency (hard, formed, loose or watery), color and presence of gross abnormalities such as distinctive odor or the presence of worms, mucus, pus or blood. Microscopic Examination His happens with the using of solution to facilitate detection under microscope. The solution are normal saline and Iodine solution stain that color the parasite nucleus and also using of Hematoxylin stain to clarification of the shape of the parasite specially the primary parasites and diagnosis the stool has two ways. A-Direct wet mount The materials used 1-microscope slide 2-cover slide 3- a stool sample 4-plastic piptte and astick The method of work 1-we take a clean microscope slide 2-we put a drop of liquid stool (when the sample is liquid ). And add the solution like normal saline (in the case that the sample is solid). 3- the cover slid is put calmly to prevent making bubbles inside the sample. B- Construction method It consists of examining by concentration the stool to make the parasite seeing with its stages by the microscope with a huge number in a small area and the concentration way is used when the diagnosis is negative but the symptoms is continuous and this way contains two ways. 1-Flotation The saturated solution with sugar or salt is used in this process. the zinc sulphate solution is considered with intensity of quality 1,18 the perfect liquid to diagnose the cyst and eggs of the worms. *The used materials 1-test tube 2-zinc 3-formalin 4- a sample of stool 5-plastic pipette and stick *Method of work 1-we put 2ml of stool in a clean test tube. 2-we fill the tube with the zinc solution. 3- close the tube firmly and the sample is mixed easily. 4- put the tube for 2min in the centrifuge. 2- sedimentation * The used materials: 1-test tube 2-formaline % 10 3- one of the saturated solutions like NaOH 4- A sample of stool 5-Absorbent , stick , slides and their covers *Method of work: 1-we take a test tube and pnt 2ML of stool by the plastic obsorbent (take into conslidation to mark the sample and give every sample a special number not to mix between the samples ). 2-Complete filling the tube with 7 ML of the formaline solution and add 3ML Ethyl estate. 3- close the tube and shake the sample quietly. 4- open the caver of the tube alittle bit for ventilation and close it and mixing should be in a fast way. 5- we put the tube in the centrifuge for 3 to 4 minutes with speed of 500 cycle per minute 6- after this process it is noticed a mass concentration on the top of the tube it is removed by the stick 7- we throw the solution that exist in the tube with keeping mass concentration in the bottom of the tube Parasite 8- we take a drop of a solution by the absorbent and put it on the slide and add a drop of stool that is con centrated in the bottom of the tube and mix them well after that we put the caver of the slide and examine it under microscope B- Direct blood examination This process is by putting a drop of blood on a clean slide. it is pulls with edge of the slide (spreader) to caver all the slide and we wash the slide by putting methyl alcohol and after that the sample is tight by the pigments by putting 2 or 3 drops of blood in the middle of the spreader as a circle shape after that we let it dry under the room temperature. Stains that are used to firm the blood 1-Leishman stain 2-Giemsa stain 3-field stain  Why we preserve stool specimens? it is best to examine samples as early as possible after collection, but preservation of specimens is necessary when stool specimens delayed in delivery or testing. 1- Preservatives will prevent the deterioration of any parasites that are present. 2- Add one volume of the stool specimen to three volumes of the preservative. 3-The preservatives have different effect on the various stages of the parasites.  Formalin Advantages 1- Suitable for Helminthes Egg , Larvae and protozoan cysts. 2- Used for concentration techniques (sedimentation techniques). 3- Long shelf life. 4- Commercially available. Disadvantages. 1- Permanent stained smears cannot be prepared from formalin preserved fecal specimens. 2- Trophozoites do not preserve well in formalin.  Sodium acetate Acetic acid Formalin (SAF) Advantages 1- Good routine fixative for protozoan cyst and trophozoites, Helminthes Egg, and larvae. 2- SAF is easy to prepare and has a long shelf life (months to years). 3- The SAF fixative contain no mercury compounds. 4- Much less toxic than other preservative. Disadvantages 1- SAF have poor adhesive properties. 2- Permanent stains not as good as with other.  Polyvinyl Alcohole (PVA) Advantages 1- This fixative is recommended for the preservation of the trophozoite and cyst stages of the intestinal protozoa. 2- It is a plastic resin that serves as adhesive for the stool material. 3- Has a long shelf life ( months to years). Disadvantages 1- Different to prepared in the laboratory. 2- Toxic , because by PVA contain mercury compound.  Merthiolate Iodine Formalin (MIF) Advantges 1- Easy to prepare. 2- Combines preservation and staining for most kind and stages found in stool Disadvantges 1- Short shelf life ( due to Iodine). 2- Cannot prepare a permanent stained smear from MIF preserved material. 3- Iodine may cause distortion of protozoa.