Chapter 2: Manage laboratory animals

Laboratory animal husbandry is a cornerstone of applied biology, underpinning research in fields such as medicine, agriculture, and biotechnology across Kenya. Proper management of laboratory animals ensures not only the reliability of experimental results but also the highest standards of animal welfare, which is increasingly scrutinized by regulatory bodies and the public. Professionals working in research institutions, universities, and pharmaceutical companies must be adept at sourcing, breeding, and caring for laboratory animals according to both scientific and ethical guidelines. This chapter explores the comprehensive practices required to manage laboratory animals effectively, focusing on practical procedures relevant to Kenyan research settings.

2.1 Animal Sourcing, Breeding, and Care

A research colony is only as good as the animals that enter it and the care they receive afterwards. Healthy animals of known origin, correct breeding records, and daily observation make results repeatable and keep the colony free of disease. The work covers where animals come from, how they are handled and sexed, how they breed, how their numbers are controlled, how substances are given to them, which diseases threaten them and which laws apply. Example: a teaching laboratory that buys mice of unknown health status and puts them straight into the breeding room risks losing the whole colony to one infection.

2.1.1 Sourcing and Handling of Animals

Sourcing decides the genetic quality and disease status of the colony, and handling decides how calm the animals are and how reliable the data are. Both are planned before animals arrive.

Sources of laboratory animals

• Accredited breeding facilities: specialised breeders that supply animals of defined strain and health status, with health reports. They are the preferred source for rodents and rabbits used in research.
• Institutional breeding colonies: a research or teaching institution may breed its own animals, which gives control of genetics, age and health status.
• Approved suppliers: a commercial supplier may be used only after its health records and housing conditions have been inspected.
• Field collection: wild animals are trapped for ecological or zoonotic disease studies only under a research permit and an ethics approval. They carry parasites and infections, so they are quarantined and handled under strict biosecurity.
• Farms and breeders of non-standard species: used only when a study needs a species or breed that no laboratory supplier keeps.

Criteria for selecting animal sources

• Health status: animals should come from sources with a health report that shows freedom from the infections that matter for the study.
• Genetic background: a defined strain and a pedigree reduce variation between animals.
• Age, sex and weight: the study protocol sets these and the supplier must be able to meet them.
• Legal and ethical compliance: the supplier must hold the permits required for keeping and moving animals.
• Reliability of supply: regular delivery dates let the technician prepare cages and avoid holding animals longer than planned.

Principles of safe handling

• Calm, confident restraint: animals that are held firmly but gently struggle less, and bites and scratches become rare. Use the method for the species described below.
• Personal protective equipment: gloves, a laboratory coat and, where needed, a mask protect the handler from bites, allergens and zoonotic infections.
• Transport and acclimatisation: animals travel in ventilated, escape-proof containers at a suitable temperature, then rest for several days to adjust to the new room before any experiment starts.
• Quarantine: new arrivals are held in a separate room and observed for disease before joining the colony.
• Low noise and no sudden movement: these reduce stress, and stressed animals give less reliable results.
• Records: each animal's origin, arrival date, identification number and health observations are written in the colony record.

2.1.1.1 Laboratory Rats

Rats are larger and more robust than mice, and they are calm and easy to train, which suits repeated handling and blood sampling. Sourcing is from accredited breeders or from the institution's colony, and rats are supplied by strain and weight. To lift a rat, place one hand over the back and around the chest with the thumb and fingers behind the forelegs, and support the hindquarters with the other hand. Rats that are used to people can be lifted by the base of the tail while the other hand supports the body, but they must never be held by the tail tip. A rat that is startled may bite, so approach slowly and let it smell the hand first.

2.1.1.2 Rabbits

Rabbits are large animals with strong hind legs and a fragile spine. A struggling rabbit can break its own back if it is held badly, so the hindquarters must always be supported. Sourcing is from breeders that supply specific-pathogen-free stock of a stated breed and age, and arrivals are quarantined because rabbits can carry respiratory and intestinal infections. To lift a rabbit, slide one hand under the chest and the other under the hindquarters and hold the body close to the handler. A rabbit must never be lifted by its ears or by the scruff alone. Wrapping the animal in a towel calms it and protects the handler from scratches.

2.1.1.3 Mice

Mice are small, widely used laboratory mammals. They are supplied in many inbred, outbred and genetically modified strains, so the strain name and health report must be checked on arrival. Mice are picked up by the base of the tail, never the tip, and then placed on a rough surface such as the cage lid or the sleeve, from where they can be held in a cupped hand or restrained gently by the loose skin at the neck and shoulders. Handling with a tunnel or cupped hands causes less fear than tail lifting. Males from different litters often fight when housed together, so group composition is planned.

2.1.1.4 Guinea Pig

Guinea pigs are gentle but nervous animals that may freeze or suddenly run when startled. Any group that arrives needs fresh vitamin C in its diet, as explained under Laboratory Animals. Sourcing is from breeders that supply animals of defined strain and health status. To lift a guinea pig, scoop it up with one hand under the chest and place the other hand under the hindquarters, holding it against the body so that it feels secure. Quiet speech and slow movements help, and the animal should never be dropped, as its legs and spine are easily injured.

2.1.1.5 Hamster

Hamsters are small animals, and several species are kept in laboratories. They are often active at night, and one that is disturbed when asleep may bite. Sourcing is from accredited breeders of a stated species and strain. A hamster is lifted by scooping it with cupped hands from the sides and underneath, after it has been woken gently and allowed to sniff the handler. Hamsters should not be grabbed from above. Adult Syrian hamsters are kept in separate cages because they fight, while some dwarf species can be kept in pairs or small groups.

Common handling errors

• Rough handling or holding an animal without enough support, which causes injury, fear and unreliable data.
• Lifting a rodent by the tip of the tail, which can strip the skin.
• Failing to quarantine new arrivals, which can bring disease into the whole colony.
• Handling different animals with the same gloves, which spreads infection between cages.

2.1.2 Sexing

Sexing means finding out whether an animal is male or female. It is done at weaning to set up breeding groups, to separate the sexes and to meet a study's protocol. A mistake at weaning causes unplanned litters.

Why accurate sexing matters

• Experimental design: many studies need animals of one sex, because hormones change responses to drugs and infection.
• Breeding programmes: correct pairing gives planned litters.
• Population control: separating males and females at weaning prevents accidental breeding and overcrowding.
• Welfare: males of some species fight when housed together, so cage groups are made up by sex.
• Records: the sex of each animal is part of its identification record.

Methods of sexing laboratory animals

• Anogenital distance: in mice, rats and hamsters, the distance between the anus and the genital opening is clearly longer in males than in females of the same age. Comparing two littermates side by side makes the difference easier to see.
• Genital shape: in a female, the anus and genital opening are close together, and the genital opening is a slit. In a male, a round penile papilla is visible and the testes may be seen after puberty.
• Nipples: in mice and rats, nipples are visible along the belly of the female but not of the male, particularly after the animals are about two weeks old.
• Gentle pressure around the genital opening: in rabbits and guinea pigs, light pressure on either side of the opening causes the penis to protrude in a male and shows a slit in a female. A female guinea pig's opening forms a Y shape.
• Testes: in adult rabbits, guinea pigs and hamsters, the testes can be seen or felt, and the hamster male's rump looks more rounded.
• Genetic testing: where external signs are unclear, polymerase chain reaction testing for sex-linked genes confirms the sex of genetically modified animals.

Steps for sexing a young rodent

1. Wash and dry hands and put on clean gloves.
2. Lift the animal by the correct method for the species and place it in a clean container with a good light.
3. Hold it so that the underside of the body and the tail base can be seen, and handle it for as short a time as possible.
4. Compare the distance between the anus and the genital opening, and note the shape of the genital opening.
5. Look for nipples and, in older animals, the testes.
6. Check the result against a second animal of the same age if there is any doubt, and enter the sex in the record.
7. Return the animal to its cage at once and change gloves before handling another cage.

2.1.3 Characteristics of Breeding Animals

Breeders are chosen carefully because their health and genes pass to every animal in the colony. Selection is repeated as breeders age.

Characteristics of suitable breeding animals

• Good health: free from the infections and parasites on the facility's health list.
• Proven fertility: animals with a record of successful mating, normal litter size and good rearing of young.
• Appropriate age: young adults at full sexual maturity, neither too young nor too old.
• Sound conformation: normal body structure and no deformities of the teeth, limbs, spine or eyes.
• Calm temperament: animals that are not aggressive to the mate or the young.
• Known pedigree: records of ancestry that avoid mating close relatives, except where an inbred strain is being maintained on purpose.
• No hereditary disorders: animals or families that carry inherited disease are removed from breeding.
• Regular reproductive performance: females with regular cycles, good milk and good maternal care, and males with active mating behaviour.
• Adaptability: animals that settle in laboratory conditions and eat and drink normally.

Reviewing breeders

• Review breeding records regularly and replace breeders whose fertility or health declines.
• Record offspring survival and growth. Poor results in many litters from one pair can indicate a hidden genetic or health problem in the parents.

2.1.4 Oestrous Cycle

The oestrous cycle is the repeating series of reproductive changes in a sexually mature female mammal that ends in a period of sexual receptiveness called oestrus, or heat. Knowing the stage lets the technician place the male with the female at the best time. Cycle length varies by species. Mice and rats cycle about every four to five days, hamsters about every four days and guinea pigs about every 15 to 17 days. Rabbits do not have a regular cycle: they are induced ovulators, which means that ovulation follows mating and they are receptive for most of the time.

Phases of the oestrous cycle

• Pro-oestrus: follicles in the ovary grow and oestrogen rises. The female is approaching receptivity.
• Oestrus: the female accepts the male and ovulation occurs. This is the best time for mating.
• Metoestrus: follows ovulation. The corpus luteum forms and the female no longer accepts the male.
• Dioestrus: the longest phase in rodents, when the reproductive tract rests before the next cycle.
• Anoestrus: a period without cycles, caused by season, young or old age, poor nutrition, stress or illness.

Importance of monitoring the oestrous cycle

• Timed mating: pairing at oestrus raises the proportion of females that become pregnant.
• Experimental timing: studies of reproduction and toxicology need females at a known stage.
• Colony planning: predicting births helps to plan cages, space and feed.
• Health: irregular or absent cycles may signal disease, poor diet or stress.
• Welfare: correct timing avoids repeated failed matings and the stress that goes with them.

Methods of detecting the stage of the cycle

• Vaginal cytology: a small amount of saline is flushed into the vagina and drawn back, and the fluid is examined as a wet or stained smear under the microscope. Pro-oestrus shows mostly nucleated epithelial cells, oestrus shows flat cornified cells without nuclei, metoestrus shows cornified cells and white blood cells, and dioestrus shows mainly white blood cells.
• Behaviour: a female in oestrus may show lordosis, an arched back and raised hindquarters, when touched or approached by a male.
• Appearance of the vulva: in rats and mice, the vulva becomes moist and swollen at pro-oestrus and oestrus and pale and dry at other stages.
• Vaginal plug check: after mating, a plug of coagulated semen may be seen in the vagina of mice, and its presence confirms mating.
• Hormone assays: hormone levels in blood or urine give precise staging in research studies.

2.1.5 Gestation Period

Gestation is the time from fertilisation to birth. It varies by species, and a technician uses it to predict the date of birth, to prepare cages and nesting material and to plan procedures.

Typical gestation periods

Species	Gestation period
Mouse	19 to 21 days
Rat	21 to 23 days
Hamster	15 to 18 days
Rabbit	28 to 32 days
Guinea pig	59 to 72 days

Factors affecting gestation length

• Species and strain: inbred strains differ slightly in length.
• Litter size: large litters are often born slightly earlier than small ones.
• Age and health of the female: disease and old age can cause complications.
• Nutrition and stress: poor feeding and disturbance can interfere with pregnancy and cause abortion.
• Number of previous pregnancies: first pregnancies may differ slightly from later ones.

Importance of knowing gestation periods

• Breeding management: the date of mating gives the expected date of birth.
• Preparation: a clean cage, nesting material and extra feed are set out before the birth.
• Experimental design: timed pregnancies are used in studies of development and toxicology.
• Welfare: pregnant females are kept in quiet rooms with little handling in the last days.
• Records: accurate dates of mating and birth are needed to explain any problems.

Pregnancy care

• Overcrowding pregnant females causes stress, fighting and loss of litters.
• A wrong mating date wastes the preparation and spoils planned experiments, so mating dates are always recorded.

2.1.6 Litter Size

Litter size is the number of young born in one pregnancy. It depends on species, strain and the health of the mother, and it helps the technician to plan how many cages and how much feed are needed.

Typical litter sizes

Species	Typical litter size
Mouse	about 5 to 12
Rat	about 6 to 14
Hamster	about 5 to 12
Rabbit	about 4 to 12
Guinea pig	1 to 6, usually 2 to 4

Factors influencing litter size

• Genetic background: some strains are bred for large litters, and others naturally produce few.
• Age of the female: very young and old females tend to have smaller litters.
• Nutrition: a well fed, healthy female rears larger and heavier litters.
• Environment: stress, overcrowding and extreme temperatures reduce litter size and survival.
• Health: infections and parasites cause smaller or weaker litters.

Importance of monitoring litter size

• Colony planning: it predicts how many animals will be available.
• Genetic monitoring: sudden falls in litter size across the colony may point to inbreeding or disease.
• Welfare: very large litters may need to be reduced if the mother cannot rear them, and very small litters may point to a problem.
• Cost: unused surplus animals cost feed, space and labour.
• Valid results: enough animals of the same age and sex must be available for the study.

2.1.7 Population Control Methods

Laboratory animals breed quickly, and a small mistake can lead to overcrowded cages, fighting, disease and wasted animals. Population control keeps the colony at the size that studies need and the facility can house.

Methods of population control

• Separation of the sexes: males and females are housed apart from weaning, except in planned breeding pairs.
• Timed mating: pairs are placed together for a set period and then separated, and the female gives birth alone.
• Breeding records and schedules: each breeding cage has a card that shows dates of mating, birth and weaning, and cages are reviewed every week.
• Limiting the number of breeders: only the number of breeding animals needed to supply studies is kept.
• Early weaning and sexing: young are weaned at the correct age, about 21 days for mice and rats, and sexed at once so that the sexes can be separated before puberty.
• Sterilisation: vasectomy of males or removal of the ovaries and uterus of females is used for animals that must live with others but must not breed.
• Use of surplus animals in other approved studies: where possible, extra animals are offered to another approved study before any other decision is made.
• Humane euthanasia of surplus animals: where no other use exists, surplus animals are killed by an approved humane method. The methods are covered under Euthanasia/Humane Killing.

Practice in the animal house

• Failure to separate the sexes at weaning is the usual cause of unplanned litters and a sudden crowded colony.
• Clear labelling of every cage and complete records stop mistakes in pairing.

2.1.8 Animal Inoculation

Inoculation is the deliberate introduction of a substance into an animal. The substance may be a vaccine, an antigen to produce antiserum, a pathogen in an infection study, or a test compound. Each route gives a different speed of absorption, and the choice depends on the substance, the species and the study.

Routes used in inoculation

The substance, the species and the aim of the study decide the route. The routes used for inoculation (subcutaneous, intradermal, intramuscular, intraperitoneal, intravenous, oral gavage and inhalation) are described in full under Routes of Drug Administration, and the route and volume are always taken from the approved protocol.

Equipment for inoculation

Name	Specification	Use
Syringe	1 mL to 10 mL, graduated, sterile, single use	Drawing up and giving the dose
Needle	Fine gauge, such as 25 G to 27 G for rodents, sterile, single use	Piercing tissue for injection
Gavage tube	Stainless steel or flexible, with a ball tip, sized to the species	Giving doses by mouth without injuring the throat
Restraint device	Cloth wrap, tube or holder, sized to the species	Holding the animal still
Skin disinfectant	70% ethanol	Cleaning the injection site
Sharps container	Rigid, puncture-proof, with a lid	Disposing of used needles without recapping

Steps for safe animal inoculation

1. Check the approved protocol for the substance, dose, volume, route and the animals to be given it, and put on gloves, a laboratory coat and eye protection.
2. Calculate the dose from the animal's weight and check the label, concentration and expiry date of the substance.
3. Draw up the dose using aseptic technique, with a new sterile syringe and needle for each animal, and remove air bubbles.
4. Restrain the animal gently with the method suited to the species and the route.
5. Clean the site with ethanol and allow it to dry.
6. Insert the needle at the correct site and angle for the route and give the dose slowly.
7. Withdraw the needle and place it directly into the sharps container without recapping it.
8. Return the animal to its cage and observe it for pain, swelling, bleeding or other reactions for the time the protocol states.
9. Record the animal number, substance, dose, route, time, site and observations.

Studies that use live pathogens need an approved biosafety level, a trained team and designated cages, and the animals are handled as infectious until the study ends. Unexpected reactions are reported to the supervisor and the veterinarian immediately.

2.1.9 Animal Diseases

Diseases cause suffering, upset experiments and may pass to humans. Laboratory animal diseases are grouped by the type of causal agent. Many infections in healthy colonies are subclinical, which means that animals show no signs, so routine health monitoring is essential.

Signs of disease in laboratory animals

• Changes in body weight and condition: weight loss, a rough coat or a hunched posture.
• Changes in behaviour: lethargy, hiding, aggression or isolation from cage mates.
• Changes in feeding and drinking: loss of appetite or unusual thirst.
• Discharges and swellings: discharge from the eyes or nose, soiled hindquarters, wounds or lumps.
• Breathing signs: sneezing, noisy or laboured breathing.
• Skin signs: scratching, hair loss, scabs or flaky skin.

2.1.9.1 Fungal Diseases

Fungal diseases in laboratory animals are usually skin infections. Ringworm, caused by dermatophytes such as Trichophyton mentagrophytes, is common in guinea pigs and rabbits and causes round patches of hair loss with scaly skin. It spreads by contact and by contaminated bedding, and it is zoonotic, so handlers must wear gloves and wash their hands. In rats and mice with weak immune systems, the fungus Pneumocystis can cause pneumonia, but it is not passed to humans. Control is by quarantine, clean housing, dry bedding and treatment on veterinary advice.

2.1.9.2 Viral Diseases

Viral diseases are often hidden in a colony and spread quickly through air, contact and contaminated equipment.

• Mouse hepatitis virus: a coronavirus of mice that spreads easily and damages the liver and intestines, and it also alters the immune responses of apparently healthy mice.
• Sendai virus: a respiratory virus of mice and rats that causes breathing problems and poor breeding.
• Sialodacryoadenitis virus: a coronavirus of rats that causes swelling of the neck glands and eye problems.
• Rabbit haemorrhagic disease virus: a highly fatal disease of rabbits that causes sudden deaths and bleeding.
• Lymphocytic choriomeningitis virus: carried by mice and hamsters, usually without signs, and it can infect people, so it is a biosafety concern.

There is no cure for most of these infections. Control rests on buying animals from virus-free sources, quarantine, filtered cages, regular testing of sentinel animals and a policy of removing infected animals as advised by the veterinarian.

2.1.9.3 Bacterial Diseases

• Pasteurellosis: caused by Pasteurella multocida in rabbits. It produces nasal discharge, sneezing and abscesses, a condition sometimes called snuffles.
• Murine respiratory mycoplasmosis: caused by Mycoplasma pulmonis in rats and mice, with noisy breathing and head tilting.
• Salmonellosis: caused by Salmonella species, which cause diarrhoea and weight loss and can pass to people.
• Tyzzer disease: caused by Clostridium piliforme, which damages the liver and intestines of rodents and rabbits.
• Bordetella and Streptococcus infections: cause respiratory disease in guinea pigs and rabbits.
• Rat-bite fever: caused by Streptobacillus moniliformis in rats, which can pass to people through bites and scratches.
• Leptospirosis: carried by wild rodents and transmitted through urine, so wild pests must be excluded from the animal house.

Prevention uses quarantine, good hygiene, clean feed and water, control of wild pests and prompt isolation of sick animals. Antibiotic treatment is given only on veterinary advice and with the approval of the study protocol, because it may alter experimental results.

2.1.9.4 Parasites

• Mites: fur mites such as Myobia musculi and Myocoptes musculinus live on mice and cause scratching and hair loss. Ear mites (Psoroptes cuniculi) cause crusts in the ears of rabbits, and Trixacarus caviae causes mange in guinea pigs.
• Lice: Polyplax species live on mice and rats, and other lice live on guinea pigs. They cause itching and blood loss.
• Pinworms: Syphacia and Aspiculuris species are small roundworms in the large intestine of mice and rats. They cause few signs but are common.
• Tapeworms: the dwarf tapeworm Hymenolepis nana infects rats, mice and hamsters and can infect people.
• Protozoa: Eimeria stiedae causes hepatic coccidiosis in rabbits, and Giardia and Spironucleus infect the intestines of rodents.

Diagnosis uses examination of the coat with a hand lens, tape tests, faecal flotation and microscopy of skin scrapings. Control uses quarantine, treatment of the whole room under veterinary guidance and thorough cleaning of cages, racks and bedding.

Disease prevention and control measures

• Quarantine of new arrivals: held apart until health status is confirmed.
• Health monitoring: sentinel animals are examined and tested at intervals so that infections are found before signs appear.
• Hygiene: regular cleaning and disinfection of cages, racks and rooms.
• Pest control: wild rodents, insects and birds are excluded from the building.
• Handling hygiene: hands are washed and gloves changed between cages.
• Prompt action: sick animals are isolated, and dead animals are sent for post-mortem examination.

Biosafety note

Wear gloves, a laboratory coat, eye protection and a mask when handling sick or dead animals, and wash hands afterwards. Report suspected outbreaks and any bite, scratch or needle injury to the supervisor at once.

2.1.10 Animal Laws

Using animals in research is controlled by laws, institutional rules and ethical standards, so that animals do not suffer without good reason and so that the results can be trusted.

Laws and guidelines relevant to animals in research

• Prevention of Cruelty to Animals Act (Cap. 360): prohibits cruelty and unnecessary suffering to animals and controls experiments on animals through a licence.
• Animal Diseases Act (Cap. 364): controls the movement of animals and the reporting of notifiable diseases.
• Veterinary Surgeons and Veterinary Paraprofessionals Act, 2011: governs the registration and practice of veterinarians and veterinary paraprofessionals, whose advice is sought on animal health in the animal house.
• Wildlife Conservation and Management Act, 2013: requires permits for capturing and keeping wild animals, which applies to field collection for research.
• Science, Technology and Innovation Act, 2013: gives the National Commission for Science, Technology and Innovation (NACOSTI) its role in licensing research and setting research ethics expectations.
• Animal care and use committees: research institutions set up committees, also called animal ethics committees, that review every protocol before animals are used and monitor compliance.
• International standards: institutions follow recognised standards such as the 3Rs, the Guide for the Care and Use of Laboratory Animals and the standards of the World Organisation for Animal Health (WOAH) on the use of animals in research and education. The ARRIVE guidelines set out how animal studies are reported.

Importance of legal compliance

• Animal welfare: rules prevent unnecessary pain and distress.
• Valid science: poorly cared-for animals give unreliable results.
• Accountability: approved protocols and records show who did what and when.
• Funding and publication: funders and journals require ethical approval.
• Public trust: transparent and humane practice protects the reputation of research.

Steps to ensure compliance

1. Learn the laws, the institution's rules and the conditions of each approval.
2. Obtain the research licence, ethics approval and any permit before acquiring animals.
3. Submit a protocol to the animal care and use committee that states the justification, species, numbers, procedures, pain relief and humane endpoints.
4. Carry out only the procedures that the approved protocol lists and only if you are trained for them.
5. Keep records of animal numbers, health, procedures and disposal.
6. Attend training in animal welfare, handling and biosafety, and renew it regularly.
7. Report any breach, unexpected suffering or injury to the committee and the supervisor without delay.

Chapter Summary

This chapter provided a comprehensive overview of managing laboratory animals, beginning with the processes involved in sourcing, handling, and caring for common species such as rats, rabbits, mice, guinea pigs, and hamsters. It explored the importance of accurate sexing and selecting breeding animals with desirable characteristics to ensure healthy laboratory populations. The discussion included detailed explanations of the oestrous cycle, gestation periods, and typical litter sizes for each species, all of which are critical for effective breeding management. Population control methods were outlined to maintain optimal animal numbers and minimize overcrowding. The chapter also addressed procedures for animal inoculation, emphasizing both technique and animal welfare. A thorough review of common animal diseases, including fungal, viral, bacterial, and parasitic infections, highlighted the importance of disease prevention and control in laboratory settings. Finally, the chapter concluded with an overview of relevant animal laws, emphasizing the ethical and legal responsibilities involved in laboratory animal husbandry.