2.3 Introduction to Parasitology

Pathogenesis

• The dynamics of any disease process.

• varies according to species and quantity of parasites as well as parasite-host adaptation and host responses.

Traumatic Damage

Examples:

• Small lesions resulting from bite of mosquitoes

• Entry of infective larvae of hookworms into the skin causes physical damage

Lytic Necrosis

disintegration of a cell by disruption of its plasma membrane to create cell injury

Entamoeba Histolytica

• releases enzymes that lyse tissues use for nutritional needs.

• These enzymes enable the parasite to penetrate the tissues of the colon, produce ulcerations in the colon, and extra intestinal viscera.

Stimulation of Host Tissue Reaction

response of the host’s living tissues to the altered conditions stimulated by the parasite Observed in majority of animal parasites

• Cellular Proliferation

• Infiltration Stimulation of red blood cell production

• Systemic Increase

Stimulation of Host Tissue Reaction

• Reaction may be in form of:

Toxic or Allergic Phenomena

• are the manifestations of an altered reaction of the organism

Stimulation of antibody production

Proteins or other metabolites produced by parasites may lead to hypersensitivity or allergic reactions due to:

Vermicularis

where an allergic reaction occurs in the anus as response to the female worm & its eggs leading to its most prominent manifestation of pruritus ani.

• Source of Infection

• Mode of Transmission

• The Infective Stage

• The Pathogenic Stage

• The Diagnostic Stage

General Lifecycle of Parasites

The Infective Stage

morphologic form that infects humans

The Pathogenic Stage

morphologic form that is responsible for the pathology produce leading to clinical manifestations

The Diagnostic Stage

morphologic form that can be detected through laboratory methods

• Protozoa

• Helminths

Classification of Parasites

Protozoa

single-celled

• Amoeba

• Flagellates

• Sporozoa

• Ciliates

Protozoa

Pseudopodia

Amoebae

• move by means of _________

Flagellates

equipped with one or more whip-like flagella that enable them to move

Sporozoa

do not possess any organ for motility

Ciliates

possess rows or patches of cilia that serve as their organs of locomotion

Phylum Sarcomastigophora

Flagellates: Other term

Phylum Apicomplexa

Sporozoa: Other term

Phylum Ciliophora

Ciliates: Other term

Helminths

multicellular metazoa

• Nemathelminthes

• Platyhelminthes

• Trematoda

• Cestoda

Helminths

Nemathelminthes

roundworms

Platyhelminthes

flatworms

Trematoda

flukes

Cestoda

tapeworms

1. Knowledge of proper specimen

2. Timing of specimen collection

3. Proper collection of specimens

4. Proper labelling of the container

Laboratory Diagnosis of Parasitic Infections

• Specimen Collection, Handling, Transport

Mouth

Specimen Collection, Handling, Transport:

• Knowledge of proper specimen

  • Most common entry

Anus

Specimen Collection, Handling, Transport:

• Knowledge of proper specimen

  • Most common portal of exit

Stool

Specimen Collection, Handling, Transport:

• Knowledge of proper specimen

  • Proper specimen to collect

Within 30 mins.

Specimen Collection, Handling, Transport:

• Timing of specimen collection

  • Time frame recommended

Maximum of 24 hours after collection

Specimen Collection, Handling, Transport:

• Timing of specimen collection

  • For formed stools with cyst forms

Preservatives

• formalin

• polyvinyl alcohol

Specimen Collection, Handling, Transport::

• Timing of specimen collection

  • Need to be added if specimen cannot be added right away:

Trophozoite

Specimen Collection, Handling, Transport:

• Timing of specimen collection

  • diagnostic stage for most protozoans that is found in liquid stool.

2-5g

Specimen Collection, Handling, Transport::

• Proper collection of specimens

  • Amount of stool needed

Collection of stool from toilet bowl water since some parasites may be destroyed by water

Specimen Collection, Handling, Transport::

• Proper collection of specimens

  • Avoid

Clean, water-light container that is covered tightly

Specimen Collection, Handling, Transport:

• Proper collection of specimens

  • Where to collect it

Information of the patient including

• history of travel and

• clinical findings

Specimen Collection, Handling, Transport:

• Proper labelling of the container

  • Includes

Ziplock bag for transport to laboratory

Specimen Collection, Handling, Transport:

• Proper labelling of the container

  • Must be placed at

Universal precautions (ex. Wearing gloves)

Specimen Collection, Handling, Transport:

• Proper labelling of the container

  • Follow

1. Direct wet preparation of direct wet mount

2. Concentration Methods

3. Permanent Stains

After Submitting the Specimen

• Microscopic Examination

  • Divided into three groups and uses microscope with an ocular micrometer

• Sedimentation

• Zinc Sulfate Flotation

Concentration Methods: Two Types of Techniques

Sedimentation: Principle

This is based on specific gravity – parasites are heavier than the solution used and thus settle in the sediment of the tube while the fecal debris which are lighter will rise to the upper layers of the test tube.

Sedimentation: Procedure

Ethyl acetate is added to a saline- washed formalin-fixed sample in a test tube and then centrifuged

Sedimentation: Advantage

It provides good recovery of most parasites and it is relatively easy to perform.

Sedimentation: Disadvantage

The preparation contains more fecal debris than a flotation technique.

Zinc Sulfate Flotation: Principle

This is based on the differences in specific gravity and the sample debris (in this case heavier thus sinks to the bottom while the parasite is lighter and thus floats upward the top of the tube).

Zinc Sulfate Flotation: Procedure

The zinc sulfate used has a specific gravity of 1.18 1.20 and is used as the concentrating solution.

Zinc Sulfate Flotation: Advantage

It is able to remove more fecal debris, hence will yield a cleaner preparation.

Zinc Sulfate Flotation: Disadvantage

Some helminth eggs are denser and may not float to the upper layer of the test tube

Direct wet preparation of direct wet mount

Purpose:

• To detect the presence of motile protozoan trophozoites; other stages detected include cysts, oocysts, ova, and larvae of worms.

Principle:

• A small portion of unfixed stool is mixed with saline or iodine then studied under the microscope.

Concentration Methods

• To aggregate parasites present into a small volume of the sample that enables the detection of small numbers of parasites that might not be detected in direct wet preparations.

• To remove debris and other contaminants that might interfere with the microscopic examination.

• Concentration techniques can be used on both fresh and preserved specimens. It can be used to detect cysts, oocysts, ova, and larvae of nematodes.

Permanent Stains

Purposes

• This serves as the final step in the microscopic examination for the detection of parasites. It is designed to confirm the presence of cysts and/or trophozoites of protozoans.

Procedure:

• A small amount of the fixed sample is placed on a slide glass and allowed to dry after which it is stained. A cover slip is then placed after which a sealant is applied, thus allowing the sample to remain intact for a longer period.

• Duodenal Material

• Sigmoidoscopy Material

• Cellophane Tape or Scotch Tape Preparation

• Blood

• Cerebrospinal Fluid

• Tissue and Biopsy Specimen

• Genitourinary Secretions

• Sputum

• Eye Specimens

• Skin Nips

• Nasal Discharge or Mouth Scrapings

• Xenodiagnosis

Other Specimens and Laboratory Procedures

Duodenal Material

• Collected through: Nasogastric Tube (NGT) or Enteric Capsule Test

• Purpose Prevent: rapid deterioration of trophozoites (if present)

• Volume: Needed _ > 2mL

Sigmoidoscopy Material

• Collected through: Collect and examine material from the colon

• Purpose: Diagnosis of infection with Entamoeba histolytic

• Procedure: Biopsy of colon material

Cellophane Tape or Scotch Tape Preparation

Purpose: Detect eggs of the pinworm Enterobius vermicularis or tapeworm Taenia app.

Procedure: A length of clear cellophane tape is wrapped around 3 or 4 fingers with the sticky side out, and the tape is pressed against the perineum, then placed onto a glass slide.

Blood

• Collected through: Fingertip of earlobe Purpose Detect presence of blood-borne parasites such as Leishmania

• Procedure: Blood is smeared then strained using Wright’s stain or Giemsa stain

Cerebrospinal Fluid

• Purpose: Diagnose certain amebic infections or for African sleeping sickness

• Procedure: CSF examined to detect parasite motility then Wet preparations are performed

Tissue and Biopsy Specimens

• Collected through: Abscess material taken from the liver

• Purpose: Detect presence of Leishmania, Toxoplasma gondii, Trypanosoma, etc. in tissues

Genitourinary Secretions

• Collected through: Detect the blood fluke Schistosoma haematobium in urine. Also, Trichomonas vaginalis that are isolated from genital secretions.

• Purpose: Urine samples are centrifuged and Genital secretions are collected using a sterile cotton swab

• Procedure: Saline wet preparation is performed to demonstrate trophozoite of parasites

Sputum

Paragonimus westermani, larvae of hookworms, etc

Eye Specimens

Acanthamoeba keratitis, Toxoplasma gondii, and Loa loa

Skin Snips

Skin fluid without bleeding obtained by making a small cut into skin

Nasal Discharge or Mouth Scrapings

E. gigivalis, Trichomonas tenax, Naegleria fowleri

Xenodiagnosis

Special method for diagnosis of Chaga’s disease