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Laboratory Animal Care andUse Program and FacilitiesAssistant Laboratory AnimalTechnician (ALAT) CertificationClass: Birds, Amphibians, FishMatthew K. Hogan BVetMed, MRCVSULAR Resident Veterinarian

Agenda ALAT Training Manual– Chapter 29-Birds– Chapter 30-Amphibians– Chapter 31-FishLaboratory Animal Care and Use Program and Facilities

BirdsLaboratory Animal Care and Use Program and Facilities

Uses in Research logyEmbryologyLaboratory Animal Care and Use Program and Facilities NOT covered under AWA ifbred for research COVERED under PHSpolicy GUIDELINES for care in theGuide and Ag Guide

Taxonomy: Aves Order: Galliformes– Poultry Turkey Chicken (Gallusgallus domesticus)– Robust immunity– Eggs used in Abproduction fluvaccines QuailLaboratory Animal Care and Use Program and Facilities

Taxonomy: Aves Order: Columbiformes– Pigeon White Carneau(Columba livia)– Atherosclerosis– Learning & memory– DoveLaboratory Animal Care and Use Program and Facilities

Taxonomy: Aves Order: Passeriformes– Common name: passerines SparrowCrowCanaryFinch– Zebrafinches (Taeniopygia guttata)» Learning, memory, language developmentmodels (songs)Laboratory Animal Care and Use Program and Facilities

Taxonomy: Aves Order: Psittaciformes– Common name: psittacines Parrots Parakeets/budgies CockatoosLaboratory Animal Care and Use Program and Facilities

Physical Characteristics Adapted for flight– Forelimbs wings– Fused & hollow bones strength, lightness Lightness fragility injury during handlingLaboratory Animal Care and Use Program and Facilities

Unique Biological & PhysicalFeatures Spur– Male galliformes nail notassociated with a toe– On lower limbs, points towardsthe tail– Fighting, defense, aggression Can trim, remove High body temperature (e.g.chicken 107oF) No sweat glands thermoregulation via pantingLaboratory Animal Care and Use Program and Facilities

Feathers Feature not shared with anyother vertebrate Functions:– Flight– Insulation– Water-proofing preen/uropygial gland Molting 1/year– Can occur due to stress,improper lighting, poor nutritionLaboratory Animal Care and Use Program and Facilities

Digestive Anatomy Crop– Expansible esophageal outpouching– Stores food– Crop milk protein, fat, Ab rich material for hatchlings Gizzard– Grinds up food (seeds) Cloaca– One chamber for intestinal, urinary and reproductivecontents/material to empty– Female lay eggs; male evaginates to pass sperm Waste– Thick-white pasty urine– Green-brown fecesLaboratory Animal Care and Use Program and Facilities

Laboratory Animal Care and Use Program and Facilities

Sexing Sexual dimorphism– Size– Feather conformation– Color pattern Other indicators– Color change (male)– Cloacal change– Featherless breast (female)Laboratory Animal Care and Use Program and Facilities

Sexing Surgical inspection/PME– Internal gonads Female– Left side only functional Male– No accessory sex organs or true penisLaboratory Animal Care and Use Program and Facilities

SexingFemaleLaboratory Animal Care and Use Program and FacilitiesMale

Handling Birds are easily alarmed or frightened injury (fly into wall, trampling), overheating Calm, cool, correct handling imperative– Move slowly and deliberatly– Handle, change cages, manipulate in dimlighting calming state– Attempt capture once, if fail, give extendedbreak before attempting againLaboratory Animal Care and Use Program and Facilities

Handling Galliformes Chickens– Can carry upside down by legs– Restrain wings & legs with hands, tuckunder arm for support (pic)– Add’n aids: tape legs together, drapecloth over head/eyes calm Turkeys– DO NOT carry by legs– Stronger, more aggressive thanchickens– One hand encircles humerus of bothwings over back, restrain head withfingers around neck, hold legs withother handLaboratory Animal Care and Use Program and Facilities

Handling Columbiformes &Passerines Small, swift difficulty insafe capturing Nets, small cage hands Can restrain largepigeon/dove like chicken(pic) Parrots/parakeets:– Restrain head to avoid bites( /- gloves); cover head– Stockinette: wings restrained,head & tail exposedLaboratory Animal Care and Use Program and Facilities

Identification Leg bands– Color-coated and/oralphanumeric– Radiofrequency ID chip(larger birds)– Size & place consideredalong with growth of bird– Checked as part of bird healthexam Wing tags DyesLaboratory Animal Care and Use Program and Facilities

Behavior Easily excited– Respond to sudden noise & movements withvocalizations calm, quiet handling Groups can have strict hierarchy fighting, food possession– Nail trims– Restrict flight Pinioning? Surgical removal of distal tip of wing Wing clip 1st 10 primary feathers; one wing!– Beak mgmt cuttle boneLaboratory Animal Care and Use Program and Facilities

Housing AWA does not address birds Physical Environment– Prevent exposure to wild birds/rodents– Perches – variable size– Adequate space for freely flapping wings Psychological well being– Solid panels between cages if conflicts Environmental– Humidity 45 – 70%– Lighting Wide spectrum or incandescent should be used– Temperature Ventilation important, susceptible to heat stressLaboratory Animal Care and Use Program and Facilities

Housing Guide (poultry):– Air temperature: 16-27oC (61-81oF)– Relative humidity: 30-70% Cage litter:– Species dependent– Cover floor with sand, gravel or shavings– Place low plexiglass frame around perimeter to containlitter Design– Flexibility accommodate other species, group sizes– Avoid mixing species, variable age groups– Away from swine/dogs (noise) & shield from visualdisturbances– Have external heat source/brooder for very young birdsLaboratory Animal Care and Use Program and Facilities

Diet Water– Suspended automatic watering system Feed is species dependent– Commercial formulated pellets– Passeriformes: seeds (canary grass) vegetables,fruits, cooked egg yolks– Psittacines: seeds (sunflower, canary grass) peanuts,monkey biscuits Grit– Necessary, remains in gizzard aids in grinding ofseeds during digestion– Includes oyster shells and granite (autoclave/sterilize) Cuttlebone (passerines & psittacines) beakmaintenance CalciumLaboratory Animal Care and Use Program and Facilities

Husbandry Feed & water containers– Several locations at varying heights reduced aggression– Scatter feed on floor dish to adapt birdto bowl feeding Environmental enrichment– Music– Colored objects/toys– Nesting materialLaboratory Animal Care and Use Program and Facilities

EuthanasiaMethodAcceptableAcceptable w/conditionsAvianIV barbiturates Inhaled anesthetics, CO2,CO, N2, Ar, cervicaldislocation (small birds andpoultry), decapitation(small birds)PoultryInjectedbarbituratesand anestheticoverdoseLaboratory Animal Care and Use Program and FacilitiesCO2, CO, N2, Ar, cervicaldislocation (asanatomically appropriate),decapitation, manual bluntforce trauma, electrocution,gunshot, captive bolt

AmphibiansLaboratory Animal Care and Use Program and Facilities

Amphibians Vertebrates– Scaleless, smooth-skinned, ectothermic– Need aquatic or very moistenvironments– Most begin life as gill-breathing larvae(tadpole) metamorphosisLaboratory Animal Care and Use Program and Facilities

Common Species inResearch Frogs– Bullfrog (Rana catesbeiana) Physiology and acoustics Motion sickness in spacetravel– They can vomit!!!– Leopard frog(Rana pipiens) Hormone studiesLaboratory Animal Care and Use Program and Facilities

Common Species inResearch Frogs– African-clawed frog(Xenopus) Entirely Aquatic Originally used inpregnancy assays Developmental studies year roundreproduction withhormone therapy Genetics & molecularbiology oocytesLaboratory Animal Care and Use Program and FacilitiesX. laevisX. tropicalis

Besides frogs! Tiger Salamander (Ambystomatigrinum)– Ophthalmology– Limb regeneration Red Spotted Newt(Notophthalmus viridescens)– Limb and lens regeneration Axolotl (Ambystomamexicanum)– Limb regenerationLaboratory Animal Care and Use Program and Facilities

AnatomyLaboratory Animal Care and Use Program and Facilities

Skin Absorb O2 through moist skin breathing!– Have lungs, no diaphragm Larve breath via gills– Chemicals can permeate skin too Mucous Glands– Slimy protective layer Granular Glands– Head & shoulders– Compounds & chemicals Defense compounds (neurotoxins, hallucinogenics) Magainins (inhibit growth of pathogens) PheromonesLaboratory Animal Care and Use Program and Facilities

Digestion & Excretion Adults:– Carnivores w/short GIT welldeveloped tongue Xenopus no tongue, use front limbs tofeed– Cloaca (like birds!)Laboratory Animal Care and Use Program and Facilities

Sexing Xenopus:– F M (female is pear-shaped)– Male has roughened blacknuptial pads & large thumbs General:– Tympanic membrane 2x larger inmales (bullfrogs)– Species specific matingbehaviorsLaboratory Animal Care and Use Program and Facilities

Tympanic MembraneLaboratory Animal Care and Use Program and Facilities

Mating Induce ovulation in female &stimulate sexual activity in male withhormone injections Egg collection:– Collect before adults eat them– Can collect unfertilized eggs Manual expression SurgeryLaboratory Animal Care and Use Program and Facilities

Development Metamorphosis Neoteny– Retain larval characteristics throughoutlifeLaboratory Animal Care and Use Program and Facilities

Handling & Restraint Only handle when necessary Handle w/moist, powder-free gloves– Nets or jars– Eye protection: cane toads toxic skinsecretions from parotid gland Restraint:– Salamander: grasp head and forelimbsw/one hand, hindlimbs w/other (neverhold by tail!)– Axolotl: never use net!Laboratory Animal Care and Use Program and Facilities

Frog RestraintLaboratory Animal Care and Use Program and Facilities

Identification Cage card may be sufficient Individual ID:–––––Photographs of skin patternsTattoo (fades as skin sheds)Toe-clipping (discouraged; IACUC approval)Freezing (branding-method)PIT passive integrated transponders(microchips)Laboratory Animal Care and Use Program and Facilities

Behavior Cannibalism– Adults may eat eggs, tadpoles, growingadults due to overcrowding, poor diet Sensitivity– Environmental temperature (ectothermic)– Overhandling, too much enrichment– Skin water quality key! Pain, distress, illness– Lethargic, poor appetite– Bacterial infection: ‘red leg’– Parasitic infection: thick, pitted, sloughing ofskinLaboratory Animal Care and Use Program and Facilities

Housing Temperature– Tropical species – 21‐29oC (70‐85oF)– Temperate – 18‐22oC (65‐72oF) High humidity: 80% need moisture Lighting– Full wavelength spectrum Reduce airflow to prevent habitat &animal desiccationLaboratory Animal Care and Use Program and Facilities

Housing: Aquatic SystemsSystemFeaturesStatic No recirculation of tank waterWater change by draining & refillingClean frequently due to waste accumulationReplace old water with same temperature newwaterRecirculating Filters remove waste & debris Less frequent cleaning, but must replace filtersregularlyFlow-through Constants stream of water in and out of tank Fresh water is always available, but must makesure chlorine is reliably removed from thisincoming waterLaboratory Animal Care and Use Program and Facilities

Caging Terrestrial– Moist, misty environment Semi-aquatic– Sloping floor Hiding places– Bark– PVC piping– Pads– Dark-colored cagingLaboratory Animal Care and Use Program and Facilities

Water Quality Monitor & adjust:– Water hardness– Dissolved CO2 level– O2 level Bubbling filtered air through water:– Maintains correct concentration of dissolvedgases Keep light if new hatchlings present No detergents or disinfectants for cleaning– Remnants can be absorbed– Heat sterilization & scrubbing recommendedLaboratory Animal Care and Use Program and Facilities