It is essential in the appropriate husbandry and management of laboratory animals to improve the welfare of the animal and the reliability of the experimental data and to protect the health and safely of those participating in husbandry and management. In this interest it is necessary to equip and maintain surroundings for each species in accordance with the specific needs of the species and to always strive for compassionate management and husbandry. After all, all who carry out experiments with animals have an obligation and responsibility to cooperate in appropriate management and husbandry. Here, the Management Committee of the Research Center for Animal Life Science at Shiga University of Medical Science has determined the Guidelines for the Husbandry and Management of Laboratory Animals along the following lines.
1. Husbandry matters
Cage and other husbandry equipment
The cage and other husbandry equipment are the most important factors in the physical and social environment of the laboratory animal. When designing and using husbandry equipment the following points must be followed in order to give the animal peace of mind, to satisfy the requirements of the research, and to preserve uniformity in experimentation.
･ The construction must allow the animal to move freely, assume appropriate posture, and relax.
･ Be able to maintain agreeable living conditions.
･ Must contain the animal in a way that both prevent escape and is safe.
･ Provide easy access to feed and water.
･ Properly ventilate air.
･ Satisfy the animals biological needs for maintaining body temperature, breeding, defecating and urinating.
･ Provide proper sanitation and dryness.
･ Not cause undue physical restriction.
･ Protect the animal from harmful substances.
Husbandry equipment design that maintains the animals health, will be convenient for the research facility, is durable and will remain useful for a long time, and will prevent as much as possible cross-infection of adjoining areas is wanted. The cage should be easy to provision, have sides that are smooth and impermeable, have few protuberances, corners, or rims where debris and moisture can accumulate and have surfaces that are easy to disinfect. Further more, it should allow easy observation of the interior that does not disturb the repose of the animal inside. Feeders and drinkers must be easily interchanged, washed, and replenished. Cages, exercise runs, pens, etc. must be maintained in a fashion that is does not injure the animal, maximizes agreeability, and allows easy disinfection.
Any interaction of groups or any formation that allows contact between individuals is included in the social environment. The effect of the social environment on confined laboratory animals is very difficult to optimize in comparison to the physical environment because it depends on the breed and the individual animalﾕs history and is not standard. However, from experience and knowledge it is possible to some extent to establish norms.
In the appropriate institutionalization of the social environment it is necessary to decide whether to house the animal singly or in a group based on whether it lives independently or lives in a group in nature. For animals who get along well, group housing is feasible, but population density, dispersal room, familiarity of the animals in the group before housing, age, gender differences, and social ranking must be taken into account. In the organizing canines, primates and other highly social mammals, behavioral and physiological alteration can arise from the integration of the group or introduction of new member, so it is desirable to avoid as much as possible alteration. Especially in the case of long term housing, surroundings appropriate to the species must be carefully considered.
Cage housing has many influences on animals but there is no method for calculating the ideal size of the cage. We must rely on the experience based judgment of the specialist. Even if all factors influencing the animal in the cage could be elucidated and evaluated, it is difficult to establish one ideal, perfect formula. The minimum necessary space for experimental animals is shown in Table. 1. The table shown here can be thought of as being calculated from approved limits by rational hypothesis (Reference Data, NIH Guideline, 1985).
Table 1. Minimum Space Recommendations for Laboratory Animals
|Species||Weight||Type of Housing||Floor Area per individual||Height a|
|Mouse||10 g||cage||38.71 cm2||12.7 cm|
|Rat||100 g||cage||109.68 cm2||17.78 cm|
|Hamster b||60 g||cage||64.52 cm2||15.24 cm|
|Guinea pig b||350 g||cage||387.12 cm2||17.78 cm|
|Rabbit b||60 kg||cage||0.14 m2||35.56 cm|
|Cat||4 kg||cage||0.28 m2||60.96 cm|
|Dog c||15 kg||cage||0.74 m2||81.28 cm|
|group 1||4 kg||cage||0.15 m2||50.80 cm|
|Chickin||0.25 kg||cage||0.023 m2||e|
Note to Table 1.
a. The distance from the floor of the cage to the upper end.
b. Nursing mother requires additional space.
c. Depending on the individual or types stature, some cases require modification (Height that permits the animal to maintain a comfortable posture, and a side of the floor space equal to the length of the animal, nose to tip of tail plus 15.24 cm is necessary).
d. Names of primate species in groups arranged by approximate sizes.
group 1: marmosets, tamarins, and infants of various species
group 2: capuchins, squirrel monkeys, and similar species
group 3: macaques and African species
group 4: male macaques and large African species
group 5: baboon and nonbrachiating species of over 15 kg
group 6: great apes and brachiating
Group cages may be substituted for individual cages. For example, infants and juveniles can be housed in group cages. When housing infants and juveniles in group cages, it is necessary to include only animals who can get along well. For animals who have just been socialized for the first time, it is necessary to watch for trauma from fighting. In group cages perches and warm places must be provided. Primate pens and exercise runs must be over 1.8 m in height. For chimpanzee and brachiating primates (orangutan, lemur, spider monkey, woolly monkey, etc.) the ceiling must be high enough that the animal can swing from it without brushing its feet on the ground.
e. There must be enough space to accommodate the birds head when it stands.
For animals brought into a man-made environment, their activity is limited in comparison to that possible in nature. However, even though the form of their activity is changed, the volume of their activity remains unlimited. Finally, the necessity or lack there of to exercise the animal is a case made based on the characteristic of the species, breed, strain, etc., the individual animalﾕs age, history and physical condition, and the nature and the planned length of the study. For example, when keeping brachiating so the animal can brachiate. Or, although dogs are frequently kept in cages, for long studies the animals must be walked in an exercise area outside the cage. Even in short-term studies, it is necessary to walk the animal about once a day.
2. Housing environment
Housing environment conforming to the sundry types of animals and types of activities must be maintained. Rats and mice and other common laboratory animals are highly adaptable and soon adapt to cage housing, but less common animals need special consideration. For example, in some species of voles frequent changing of the cage has been shown to disrupt breeding. Below, areas of concern for common laboratory animals are elaborated.
Micro- and Macro-environment
For the animal, the micro-environment is the temperature, humidity, etc. of the physical environment of the cage or the primary enclosure, whereas the macro-environment is the housing room, or the secondary enclosure. In many cases there are differences between these two environments. Except where the cage has separate air-exchange, temperature, humidity, and concentrations of gases such as carbon dioxide and ammonia concentration, etc. are higher in the cage than in the housing room. Although the difference between the two environments depends on the type of cage or construction of the rack, attention must be paid to the effect of the micro-environment on experimental results.
Temperature and humidity
Since temperature and humidity have influence on the animals metabolism and behavior, they are considered the most important aspects of the physical environment. The range for which the animals respiration rate is lowest, regardless relative change in the temperature of the surroundings, is known as the median temperature region. The optimum dry-bulb hanging thermometer temperature for growth, agreeability, adaptability and reactability is, as recommended from experience, generally lower than the median temperature region. The ideal temperature and humidity range for selected species are shown in Table. 2 below.
Table 2. Recommended dry-bulb temperature and relative humidity for common laboratory animals
|Species||Dry-bulb Temperature（℃）||Relative Humidity（％）|
a: 27-29 is desirable for post operation recovery and nursing cages.
b: For marmosets, tamarins and owl monkeys etc. 60-65% is desirable.
c: This is appropriate for birds older than six weeks. Hatchlings need
a higher temperature.
The purpose of air circulation is to provide appropriate oxygen supply, to disperse the thermal load of lights and machines, gas and particular impurities, and to regulate air exchange. The quality of a micro-environment is evaluated based on the maintenance of primary enclosure temperature and the effectiveness of protection from cross infection by air circulation. The circulation of room air in the animal facility is not to take place without management of toxic gas and particulate content of the air. Further more, it is necessary to consider the regulation of air pressure in the animal and accessory areas. Quarantine, isolation, dirty equipment handling, hazardous materials, and primate housing must all be kept in negative pressure. On the other hand, clean equipment storage, specific pathogen free animal husbandry, etc. must be done in positive-pressure areas.
Together with attentive provision of uniform lighting, it is necessary that the lighting be sufficient for facility management, observation of animals, safe working environment, and the comfort of the animals. Although there is no predetermined strict level of lighting to be used, because extremely high levels affect the retina, a value of 150-300 lx at 40-85 cm from the floor has been determined appropriate from both animal and management standpoints. However, exterior windows, due to humidity, temperature, daylight time, dew formation, etc. are not good for the research facility and man-made light, either white or day-light colored fluorescent, must be used.
The noise generated by husbandry activity and the animals themselves in the animal facility is difficult to avoid. Cage washing, waste disposal and other such noise-producing activity at least must be taken care of outside of the animal room and, if possible, in an entirely separated area. Also, workers must cooperate in keeping work related noise to an absolute minimum. Rats and mice exposed continuously to levels over 85 decibels suffer hearing loss as well as non-auditory symptoms, such as increases heart beat, respiration, and raised blood pressure. Therefore, primates, canines and other noisy species should be housed separately from rodents and rabbits.
Except where indicated otherwise by the investigation, easily accessible, highly palatable feed meeting the nutritional requirements of the species must be provided daily. Appropriate feeding must not only maintain the health of animals in normal development, but must also meet the special metabolic needs of pregnant and nursing animals. Animal supervisors and animal technicians must pay careful attention to the methods used to preserve the nutritional value of the feed at the manufacture or vendor, and the transportation, storage and handling of the feed in order to avoid spoilage and contamination with chemical pollutants, disease vectors (arthropod, etc.), parasites, and pathogens. Flutes and vegetables for primates rot easily and are sometimes contaminated with biological and chemical agents so special care must be taken. Even if contamination of the food is not great enough to cause symptoms of poisoning, enzyme activity, hormone production and other physiological or biochemical changes in the animal can still be conspicuous. In order to obtain standard data, certified feeds for which the amount of each biological or non-biological foreign particle has been analyzed and measured are necessary. Furthermore, attention must be paid to deterioration of nutrition and change in quality after purchase. Generally speaking, if feed is protected from high temperature, high humidity, and vermin it should be good for six months from the date of manufacture. However, vitamin C used to supplement food for certain species, such as that for guinea pigs and primates, is good for only 3 months from the date of purchase. For species which need vitamin C supplements, vitamin C must be supplied in the feed or given in the form of fruit and vegetables. Although refrigeration can extend the life time of feed, the storage period should be kept as short as possible in every situation.
For feed sterilized by autoclave, the breakdown of nutrients from autoclaving must be considered and feed used as soon as possible. Feed must be kept in a dry, arid, sanitary, vermin-free area protected from contamination by foreign objects. Feed must be stored on top of pallets, shelves, or carts so as there is no direct contact with the floor. Care must be taken to refrigerate fruits, vegetables and other perishable feed. Left over feed must be kept stored in separate vermin-proof containers and not returned to the original container in order to prevent contamination or diffusion of pathogens. Finally, the shipping and storage container should not be taken to the animal area: the container for left over feed may be used to transport feed to the animal area. The temporary container must be routinely disinfected.
Bedding is a necessity in order that pease of mind is given to rats, mice and other small animals. Bedding should be highly absorbent, free of toxic materials harmful to humans and animals, and unpalatable to the animal. Bedding should be sufficient in quantity to keep the animal dry and unsoiled from feces and urine, yet not come in contact with drinkers. Bedding should be stored on pallets, shelving or carts so as not to come in contact with the floor. In housing situations where bedding is not permitted, such as metabolic cages, fiber intake studies, etc. mesh flooring is indicated. A material providing more insulation and comfort than bare metal should be selected for the mesh.
Fresh water free from micro-organisms and chemical contamination, as well as of the proper pH balance and hardness must be provided. For normal husbandry purposes, tap water is acceptable, but depending on the experiment greater purity may be necessary. Moreover, for animals with a defined micro bacteriological status, water percolated through a filter, or chlorine disinfection is necessary. Water bottles and automatic watering systems must be checked daily to ensure proper supply and functioning. Animals must also be taught how to use automatic drinking systems. Replacement of dirty water bottles with freshly cleaned ones is preferable, but in the case dirty water bottles are refilled each one must be returned to the same cage it was taken from.
It is necessary to maintain proper sanitation in any facility where animals are kept. Corridors, animal rooms and storage areas must be cleaned and sterilized at appropriate intervals and debris and hazardous materials removed. The same brooms, mops, buckets and other cleaning equipment must not be used in housing areas for different species.
Bedding must be changed as frequently as is necessary to keep the animal clean and dry. For rats, mice, hamsters and other small rodents, under normal husbandry conditions one to three times a week is sufficient. For larger animals like cats, dogs and monkeys feed bowls must be changed daily. Cages must be hosed down at least once daily, and care must be taken not to get the animal wet while hosing down the cage. Especially for young, care must be taken to move them to a different holding area or not to splash them. In some cases frequent bedding or cage changing can interfere with breeding or the process of the experiment. In this case, the cage may be cleaned as the sanitation of the animal dictates. Cage sterilization must not be carried out in the animal area. Equipment such as the cage, rack,feeder, water bottles, etc. must all be frequently washed and sterilized in order to maintain sanitation and protect the animal from harmful materials. Flat caging for rodents and accessory equipment must be changed as frequently as possible, the racks once a day, and plans should be made to change other animalsﾕ cages at least once every two weeks. Moreover, watering devices such as drinkers enclosed in cages, water bottles, stoppers, etc. must be either steam sterilized or gas sterilized. Since bacteria also breed in automatic watering system, the system must be periodically frushed and cleaned. Corridorrs, animal rooms etc. must be periodical cleaned of garbage and rinsed down, then sterilized with a disinfectant. Suggested concentrations of commonly used disinfectants are shown in Table 3.
Table. 3 Commonly used disinfectants and their characteristics
|Chemicals||Usual concentration||Virus||Spore||Aerobe||Gram positive||Gram negative||Fungi||Characteristics and precautions|
|Ethyl alcohol||70-90%||fair||no||good||good||good||good||Irritant to the skin|
|Formaldehyde fumigation||15-20ml/m3||good||good||good||good||good||good||Irritant, corrosive fumigation|
|Quaternary ammomium salts (Osban, Micro-quat, Tego, etc.)||0.05-0.1%||fair||fair||no||good||fair||good||Irritant, ineffective when used with soap|
|Sodium hydrocholoride (Pyurax, etc.)||100-200-ppm||good||good||good||good||good||good||Effectiveness reduced by organic material|
|Iodine-based (Micro-clean, Isodine, etc.)||50-100ppm||good||good||good||good||good||good||Corrosive, stains, produces poisonous isodine vapor|
|Chlorhexidine (Hibitane, etc.)||0.1-0.5%||fair||fair||fair||good||good||fair||No irritant, low toxicity|
After thoroughly scrubbing the hands in water, they should be disinfected chemically. Where irritability to the skin is a concern, Chlorhexidine, quaternary ammonium salts (detergents), iodine-based disinfectants, and rubbing alcohol are available. Carts, uniforms, smocks, rubber globes, etc. must also be thoroughly rinsed in water then steam sterilized or, when steam sterilization is not possible, they must be soaked or sprayed with quaternary ammonium salt, iodine-based disinfectants, or chlorhexidine. Disposable masks and caps must be discarded immediately after use. Disposable towels used to wipe the hands after disinfection must also be discarded.
Rubbish, excrement, bedding, animal carcasses, and similar soiled materials are environmental pollutants, so sufficient care should be taken to avoid bringing them into contact with humans and they should be discarded as soon as possible. Materials to be discarded should be put in plastic bags or water-proof feed bags, etc. and, in addition to taking care not to spill anything during transportation, should be incinerated as soon as possible. In the case incineration can not be carried out immediately, waste must be temporarily stored. However, the container must be such that noxious odors are not emitted, flies and cockroaches cannot bree and rodents are nor attracted. If vermin are detected, they must be quickly captured and exterminated, their origin determined, and proper measures taken to prevent their return. In the case pesticides are used, care must be taken to eliminate the possibility of side effects to animals being housed and personnel. Used hypodermic syringes, needles, razors, glass, and disposable plastic items should be put in a safe, leakproof, sanitary sharps container for disposal. Items used in contagion research or quarantine where there is a fear of contamination by microorganism should be sterilized and disinfected before disposal.
7. Identification and Record Keeping
The identification of individual animals can be done by room, rack, cage, card, collar, band, hanging tag, attached tag, dye, ear punch, ear clipping, tattoo, etc. The importance of records for individual animals cannot be ignored. The contents of the ID card can range from the sketchy to particular. At the very least the investigatorﾕs name, lab, date the animal was first used and the animalﾕs origin must be included. The need for further individual records is based on the requirements of the investigation. Origin and final disposition of the animal are information of great importance and are indispensable data at times, so they must be carefully recorded.
8. Emergency, weekend, and Holiday Management
In order to deal with emergencies it is necessary that contact with the facility head, supervisor, and animal technician be possible. The contact persons name and number and an emergency response center number must be prominently displayed at various places in the facility, as well as on every phone. Measures must be taken so that veterinary assistance is available on weekends and holidays as well as regular work days (in addition to the needs of the experiment, it is necessary for the safety and well-being of the animals that all animals be observed daily, to include weekends and holidays).
Appendix 1 Veterinary management
Veterinary management is essential in the planning of animal management, and includes the following articles.
･ Every day all animals must be observed and their health and comfort verified.
･ Disease and outside influences must be prevented, controlled, diagnosed and treated.
･ The appropriate performance of animal handling, restraint, anesthesia, analgesia, and euthanasia must be ensured.
･ Surgical procedure and post -op recovery must be observed.
Veterinary management is the obligation of veterinary personnel with experience in animal science. There is no problem in having non-veterinary personnel carry out these roles, but at appropriate times veterinary personnel must be available to receive reports of any problems and the general condition of the animals health, behavior and comfort. In addition, there are other matters of concern to veterinary management, such as advice in planning animal experiments (suggestions and corrections for the parts of a proposal concerned with veterinary management, animal husbandry, animal welfare, etc.), safety in the work place, storage of hazardous materials, enforcement of safeguards against zoonosis, and ensuring feeding, watering and sanitation procedures are carried out properly.
Disease prevention is an important aspect of veterinary management. Aside from mastering animal handling, there are several points that apply to disease prevention.
All animals must be acquires legally. In evaluating the quality of each suppliers animals, it is necessary to investigate the means of transportation. For each shipment received, all points of the detailed invoice must be verified, various fluid samples taken from the animals, and the animals quarantined and acclimatized to their new environment by means appropriate to the species and housing environment. Since vendor quality control data are used as references in deciding these means, it is absolutely necessary to request a copy of quality control data.
Quarantine and Acclimatization
Quarantine refers to the housing of newly arrived animals separately from resident animals until their health status can be fully evaluated. Through effective quarantine the transmission of pathogens to the resident colony can be prevented.
The vendor quality control situation and animal health prognosis can be considered in setting up quarantine at the University. Even though it is possible to shorten the quarantine period for rodents based on that information, it is still necessary to acclimatize all new animals to the new surroundings before their use in an experiment. Once acclimatized to the new surroundings, the psychological condition and behavior of the animals will stabilize. The acclimatization period for mice, rats, and guinea pigs is about seven> to ten days. For other animals a longer period is believed necesary. For newly arrived animals whose history is unclear on certain points, a quarantine using broader methods taking into account the incubation period of any disease the animal may have must be carried out. The following items are done during the quarantine & acclimatization period;
･ Diagnosis of disease, to include zoonosis, control of the disease, prophylaxis, and treatment.
･ Physiological and nutritional adjustment. ･ When necessary, bathing, medicinal dips, and nail-clipping.
Separation by species, origin, and microbiotic status
The physical separation of species is necessary to prevent the inter-species spread of disease, agitation arising from inter-species conflict, and conduct of the research. Normally, each species is housed in a different room. However, it is possible to house rodents of different species used for similar investigations but similar microbiotic status in the same room if within the room they are separately kept in laminar flow cabinets, clean racks, or filter-toped plastic box cages, etc. On the other hand, even animals of the same species must be housed in separate rooms if they are of different microbiotic status.
Investigation of Disease, Diagnosis, and Treatment
For all experiment animals, disease symptoms, trauma, or unusual behavior should be watched for in daily observations by a staff member familiar with possible problems until the end of the quarantine period. Especial attention should be paid to unexplained deaths and abnormal conditions. As soon as symptoms or trauma are detected the animal must be remanded to veterinary care. Any animals manifesting signs of infection by a contagious disease must be isolated from healthy members of the colony. On the other hand, if it is clear a room, or a group, are all infected, or the possibility they are all infected is high, those animals should be kept where they are while they are diagnosed, treated and a solution is thought out.
Prophylaxis, diagnosis and treatment must use the most advanced, approved method available, and, in verifying diagnoses, in addition to physical observation of symptoms, autopsy, pathological histological investigation, clinical pathological investigation, haematological investigation, microbiological investigation, and clinical medical investigation, etc. must be used.
This guide shall become effective from June 15, 1988.
２. This guide shall become effective from April 1, 2003.