TRYPANOSOMES

PHYLUM-SARCOMASTIGOPHORA  
ORDER-KINETOPLASTIDA

FAMILY-TRYPANOSOMATIDAE
CLASS-ZOOMASTIGOPHORA

• Members are all parasitic. They are found in blood, plasma, lymph and tissue fluids of mammals and birds, hence called as haemoflagellates.and may also have Pseudopodia.
• They have a characteristic leaf- like body with a single thread like flagellum pass along the body and remain attached to the body by the undulating membrane. Disease caused  is known as trypanosomosis.
• The nucleus is usually vascular and reproduction is generally by longitudinal BINARY FISSION.The nutrition is HOLOZOIC.
• In man, trypanosomes cause African sleeping sickness and Chagas' disease. In domestic animals, this disease is known as surra which means rotten. They are transmitted cyclically and mechanically by hematophagous flies.
• It affects a wide range of hosts and commonly seen in tropical countries like India, South Africa, and the UAE.
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  Trypanosoma

  
  

  

  

  Trypanosoma in cattle

  
  

  

  Fig.Trypanosoma evensi in blood smear of cattle

  
  

  

  
  

  

  
  

  

  
  

  
  

  

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MORPHOLOGY

• Body is usually elongated flattened leaf like in shape somewhat rounded with a vesicular nucleus(having one nucleus) and  kinetoplast located posterior to blepheroplast.kinetoplast contain DNA.
• they have single flagellum attached to the body by undulating membrane.the flagellum is arise from the BLEPHEROPLAST and passes anteriorly.
• Axoneme arises from kinetosome of basal granule attached to the body by undulating membrane and is continuous as free flagellum.
• Movement may be active or sluggish.
• Trypanosomes vary in shape.eg round,elongated leaf like.

• short and  stout stumpy forms
• long and slender forms ,and
• intermediary forms. Such trypanosomes with varying shape are called polymorphic trypanosomes are polymorphic trypanosomes.
• Some may be uniform in size – monomorphic tryps.

TRANSMISSION OF TRYPANOSOMA

• Multiplication is by longitudinal binary fission. Division starts form kinetoplast followed by nucleus and cytoplasm.
• With a single exception of T.equiperdum of equines,transmission from one vertebrates to another is carried out by blood sucking flies(TABANUS) in which development stages occur (cyclical transmission).
• Some of the trypanosomes are transmitted mechanically in which the infective stages are alive for a few minutes and they have to be transferrec to a new host for successful transmission.e.g. Trypanosoma evansi by Tabanus sp.
• Transmission is either cyclical or mechanical.
• CYCLIC TRANSMISSION
• Cyclical development occurs in the vector and may be in anterior station (salivaria) or in posterior station (stercoraria).
• In the anterior station development, infection is transmitted by inoculation by arthropod vectors when they suck blood from the host. eg. Trypanosoma vivax,T.congolense.T.brucei are transmitted by tse tse flies(GLOSSINA spp.)
• In the posterior station develpoment ,the metacyclic trypomastigotes accumulated in the hind gut are passed in the faeces of the arthropods. Infection of vertebrate host occur by contamination of skin or skin wound. e.g. Trypanosoma cruzi transmitted by REDUVIID BUGS.A relative non pathogenic trypanosomes like T.theileri and T.melophagium are transmitted by TABANUS FLIES.
• Any trypanosomes can be transmitted mechanically without cyclical changes experimentally this can be done by syringe passage.
• In nature this is accomplished by blood sucking insects like Tabanus, Stomoxys, Hippobosca sp., etc. which feed several times on different animals before repletion

DEVELOPMENTAL STAGE

usually 4 types of developmental stages...

• Amastigote (Leishmania like bodies, without flagellum)
  1. The body is spherical.
  2. Flagellum is absent or represented by short fibrils or degenerated into a tiny fibril inside the body.
  3. Kinetoplast is present.
  4. This stage is found in vertebrates and arthropods.
• Promastigote (Leptomonad,with a short free flagellum)
  1. Body is elongated.
  2. Kinetoplast and axoneme are towards the anterior tip of the body with no undulating membrane.
  3. It is mostly seen in invertebrates or cultures.
• Epimastigote (Crithidial,small free flagellum and short undulating membrane)
  1. Kinetoplast with axoneme is anterior to nucleus.
  2. Undulating membrane is short.
  3. This stage is seen in vertebrates but principally a stage in arthropod.
• Trypomastigote (Trypanosomes,complete undulating membrane and free flagellum)
  1. The body is leaf like or blade like.
  2. This is the form seen normally in blood films of infected animals.
  3. It is blade like form, kinetoplast posterior to nucleus and near to posterior extremity.
  4. Undulating membrane is well developed.
  5. Free flagellum is often present.
  6. Found in vertebrate host and also in arthropods.
  7. It is the infective stage for the invertebrate host.

  PATHOGENESIS

• Mainly anemia (Hemolytic Anemia).
  1. This may be due to immunological mechanism resulting in haemolysis and erythrophagocytosis.
  2. Trypanosome antigen attaches to RBCs and increases erythrophagocytosis.
  3. Anaemia may result due to reduction in half - life of circulating cells.
  4. Due to immune mechanism - enhanced haemolysis occurs; a hemolytic factor produced resulting in direct hemolysis of RBC.
  5. Anemia is also associated with disorders of clotting like thrombocytopenia and disseminated intravascular clotting ( DIC).
  6.  Damages to blood forming organs by trypanosomes.
• Hypoglycemia
•  Trypanosome absorbs glucose in blood leading to increased production of lactic acid. This leads to less intake of O2 by RBCs resulting in asphyxia. and acidosis.
• Serum potassium level is increased due to destruction of blood cells. Calcium and phosphorous ratio is disturbed.
• The lysed  trypanosomes release endotoxin resulting in toxemia and death.
• Due to the anaemia, mucous membrane is pale.
• Some of the clinical signs are
  1. Petechial hemorrhages and emaciation.
  2. Enlargement of spleen, lymph gland and liver.
  3. Congestion of mucosa of intestine, stomach, kidney and  bone marrow
  4. Oedema of dependant parts is common. Animal will be unable to get up.
  5. Keratitis  and conjunctivitis.
  6. Nervous symptoms.
  7. CSF section of the brain will show perivascular cuffing,  infiltration meningitis and  encephalitis.

TRYPANOSOMA EVANSI

      It is the first trypanosome shown to be pathogenic to mammals identified by Griffith Evans a British Vet in India.



HOST

• Natural hosts-Camel, horse, donkey, mule, ox, goat, pig, dog, water buffalo, elephant , mongoose , deer and other wild animals like fox, hyena and tiger.
• Experimental hosts- Many laboratory animals including mouse , rat, rabbit, guinea pig and chicken.

LOCATION

• Blood and lymph. The disease is called trypanosomosis.
• Name of the disease is different in different countries- most widely used is surra.
• Classical disease entity in Indian sub-continent  and occurs in horses and is known as surra.
• It is a Hindi word meaning rotten (or) putrified.
• Disease in camel is called Gufar; murrina – in panama; dorrengadera in Veninzula.
• T.equinum now regarded as dyskinetoplastic strain of T.evensi causes mal de caderes in horses in South America.

 PREVALENCE -Common in Northern Africa, Asia, Northern and South America

  STRUCTURE 

• Mean length varies considerably in different hosts and geographic strains.
• Typically they are 15-34 µm long with a mean of 24 µm.
• Most ones are slender (or) intermediate in shape. But stumpy forms also occur.
• Sporadic strains without a kinetoplast( dyskinetoplastic)  and visible with light microscope may arise  occasionally and  spontaneously or post-treatment with certain dyes (e.g. Acriflavin) and  drugs such diminazene aceturate.

INFECTION IN CATTLE OR BUFFALOES

• These two animals are  the main reservoirs of infection to equines. Infection is sub- clinical, mild and inapparent.
• Occasional outbreaks of acute disease are reported from many places. This may be due to lowered resistance in carrier animals following debilitating intercurrent diseases like rinderpest (or) FMD ,etc., stress after FMD vaccination and over work in draught animals.
• Introduction of new strain of parasite into newer areas  may result in acute form of the disease.  It appears as epizootic of variable severity. This can be highly fatal.

Clinical signs are

• High fever 41⁰C
• Intense excitement alternative to those of severe depression (coma)
• Animals move aimlessly in circles frequently falling down,  show colic, grinding of teeth, eyes staring wide open , breathing hard  and noisy
• Goring against wall apparent blindness stamping of feet following groaning, micturition, profused salivation, twitching of muscles followed by partial loss of senses  and prostration
• Parasitaemia due to factors unknown becomes too high; blood smears are seen with large number of parasites which occlude cerebral capillaries.
• Death may occur in 18 hr to 3 days.
• Sudden death may be mistaken for poisoning or  snake bite or  anthrax.
• In per acute cases death occurs in 2-3 hours. The nervous form of the disease show symptoms as above.
• Sub acute and chronic
  1. The animals look dull, sleepy
  2. lacrimation of eye, progressive emaciation rapid pulse
  3. Intermittent fever, oedema of leg, diarrhoea  and death
  4. Corneal opacity – twitching of muscles below eye
  5. Sub normal temp

INFECTION IN DOGS

• Disease is acute (or) fatal.
• Death in 2-4 weeks.
• Oedema is marked.
• The classical signs are corneal opacity, oedema of larynx resulting in voice changes and similar to that occur in rabies.

CAUSE OF DEATH

To sum - up, death in surra is mainly due to

• High fever, toxaemia
• Anaemia – PCV is less than 25 and 30 and decreased haemoglobin
•  damage of bone narrow  due to trypano toxin
• Increase in erythrocyte sedimentation rate   
• Hypoglycemia –  reduction of blood glucose  level by 30%
•  Exhaustion of glycogen reserves
•  Failure of liver cells  to compensate the loss in glycogen reserve

 DIAGNOSIS - TRYPANOSOMA EVANSI


  

• Clinical diagnosis can be done with history and clinical signs as described.

Laboratory diagnosis

    

• Direct examination or wet film examination
  • It is a quick method of detecting the organism by studying their movement and relative size . Species of trypanosomes involved can be guessed but it is to be confirmed by staining
• Peripheral blood smear examination
  • Thick and thin blood smears at the height of temperature is more desirable.
  • Parasitaemia is common in equines and canines but not readily seen in cattle, buffalo & camel.
  • Smears are stained with any of the GIEMSA OR LEISHMAN stain.
• Lymph node biopsy smears
  • Inject sterile normal saline into lymph node preferably prescapular lymph node with the help of a tuberculin syringe, massage and then aspirate.
  • It is risky to the operator
• Buffy coat smear
  • The suspected blood is spun and the buffy coat is examined for the presence of trypanosomes
• Biological test or animal inoculation test
  • The suspected blood is injected intra peritoneally  into susceptible laboratory animals like white rat, white mice, guinea pigs, rabbits .etc.,
  • Cryptic (or) sub- clinical trypanosomes in blood will multiply in these animals and cause death of these laboratory animals- Mouse: 48-72 hours; guinea pigs- 7 days; rabbits-60 days.
• Culture RPMI-1640,CAM
• Indirect test or Non-specific test to measure alteration of serum proteins
  • Mercuric chloride test
    • Reliable for surra in camels only.
    • This is done by adding one drop of suspected serum into 1: 20,000 solution of mercuric chloride.
    • A white precipitate is formed. In infected camels , this test gives  positive results in  2-3 weeks post infection
  • Stilbamidine test
    • Useful in bovine surra.
    • 0.5 – 2.5 ml of freshly prepared 10 % stilbamidine solution is taken in agglutinating tubes and one drop of inactivated serum is added.
    • In positive cases, coagulation occurs on the surface,begins to settle down in half a minute and dissolves in 5-10 minutes.
  • Formal get test
    • Useful in camel surra.
    • Two to four drops of formalin (40%) is added to 1ml of suspected serum and allowed it to stand .
    • In positive cases, gel formation occurs in 2-3 hrs.
  • Nitric acid test
    • One ml of 1.7 % nitric acid test is added to one drop of suspected serum.
    • In positive cases, white floccules are seen.
• Examination of other fluids
  • Cerebrospinal fluid will be examined in nervous form for the presence of trypanosomes but it is not important in veterinary practice.
  • Aqueous humor will be examined in case of blindness and in corneal opacity.
• Serological test
  • Some of the serological tests standardised are indirect haemagglutination, gel Diffusion, enzyme linked immunosorbant assay, complement fixation test and flourescent antibody test.
• Molecular test polymerase chain reaction (PCR) and PCR- restricted fragment length polymorphism (PCR-RFLP) are available for diagnosis and to know strain variation.