Experimental Infection of Dogs, Cats, Ferrets, and Rodents with Dirofilaria immitis, Brugia malayi, B. pahangi, or Dipetalonema reconditum

Printer Version: Experimental Infection Cats And Dogs

TITLE:  Experimental Infection of Dogs, Cats, Ferrets, and Rodents with Dirofilaria immitis,   Brugia malayi, B. pahangi, or Dipetalonema reconditum

SOP NUMBER:  8.5

REVISION NUMBER:  2

 

1.0       PURPOSE

This standard operating procedure outlines the experimental infection of dogs, cats,  ferrets, and rodents with D. immitis, B. malayi, B. pahangi, or D. reconditum by inoculation of infective third-stage larvae (L3).

 

2.0       PROCEDURES

  • 2.1        Collection of the L3
  • 2.1.1    A stereo dissecting scope and a micropipette (i.e., modified pasteur pipette) are used to remove the infective larvae (L3) from the petri dishes.
  • 2.1.2    The L3 are counted into separate petri dishes for each individual animal.  The number of L3/animal will vary according to species of parasite and animal and the purpose of the study.
  • 2.1.3    After counting is completed, the petri dish containing the L3 is tilted slightly to allow the L3 to settle to the lowest part of the dish.
  • 2.1.4    Excess fluid is removed from the dish using a pasteur pipette, such that less than 1 ml of fluid (containing L3) remains in the dish.
  • 2.1.5    The fluid containing the L3 is pulled up into a 1-cc tuberculin syringe.
  • 2.2        Subcutaneous injection of L3 (≤ 400 L3 / animal)
  • 2.2.1     A 20-gauge, 1.5-inch needle is attached to the syringe and the contents of the syringe are injected subcutaneously in the inguinal area of the animal.  After the initial injection, media is added to the syringe and the solution is injected to ensure that all L3 are removed from the syringe; this step may be repeated several times.  After the last injection, media is added to the syringe and the contents are emptied into a petri dish and examined for remaining L3.  For B. pahangi and B. malayi, the L3 may be injected SC in the dorsum of one or more of the paws of dogs and cats.
  • 2.3        Intraperitoneal injection of L3 (≤ 1,000 L3 / animal)
  • 2.3.1    For infecting rodents intraperitoneally with B. malayi or B. pahangi, inject larvae into the peritoneal cavity as described in 2.2.1.  A 1.0- or 1.5-inch needle may be used.

 

3.0       Inoculum size by animal species, filarial species, and infection route.

  • 3.1        Inoculum size and route of infection may vary by study protocol objective and requirements.
  • 3.2        Guidelines for establishing infections
  • 3.2.1     Dirofilaria immitis
  • 3.2.1.1  Dogs (~35 L3 SC per dog to produce microfilaremic infections; up to 400 L3 SC per dog for donor dogs).
  • 3.2.1.2  Cats (~100 L3 SC per cat to produce transient microfilaremic infections).
  • 3.2.1.3  Ferrets (~15 L3 SC per ferret to produce transient microfilaremic infections; up to 75 L3 SC per ferret for donor ferrets).
  • 3.2.2    Dipetalonema reconditum
  • 3.2.2.1  Dogs (up to 200 L3 SC to produce microfilaremic infections).
  • 3.2.3    Brugia pahangi
  • 3.2.3.1 Dogs and cats (up to 400 L3 SC per animal to produce microfilaremic lymphatic infections).
  • 3.2.3.2 Jirds and other rodents (up to 150 L3 SC to produce microfilaremic lymphatic infections; ~400 L3 SC to produce intraperitoneal infections, particularly in the male jirds).
  • 3.2.3.3 Ferrets (up to 200 L3 SC per ferret to produce transient microfilaremic infections).
  • 3.2.4   B. malayi
  • 3.2.4.1 Cats and dogs (up to 400 L3 SC per animal to produce microfilaremic lymphatic infections).
  • 3.2.4.2 Jirds and other rodents (up to 150 L3 SC to produce microfilaremic lymphatic infections; ~400 L3 SC to produce intraperitoneal infections, particularly in male jirds).
  • 3.2.4.3 Ferrets (up to 200 L3 SC per ferret to produce transient microfilaremic infections).