Specific pathogen free and gonobiotic animals RASHTRIYA KRISHI Volume 11 Issue 1 June, 2016 47-50

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Production and management of specific pathogen free and

Specific pathogen free animals are those animals in 2005. People for the ethical treatment of animals
which are free from a group of particular pathogen but (PETA) reported that the National Centre for Laboratory
these animals are not necessarily free from other organism, Animal Sciences (NCLAS) in Hyderabad, supplies
which are not specified, in the conducted experiment. approximately 50,000 animals to laboratories every year.
Gnotobiotic: A word derived from the Greek “gnotos” and Use of specific pathogen free and gnotobiotic
“biota” meaning known flora and fauna. An animal stock animals: These animals are used in certain conditions of
or strain derived by aseptic cesarean section (or sterile body which become severe by secondary complication.
hatching of eggs) which are reared and continuously Ex. Wound. In India, among rodent group of animals e.g.
maintained with germfree techniques. According to mice, rat, G. pig, rabbit, mice are predominantly used in
International Committee on Laboratory Animals (ICLA) most of cases followed by others. SPF animals are very
“Specific Pathogen Free (SPF) animals, which are free much useful in experiments which are carried out for
of specified micro-organism and parasites but not longer period.
necessarily free of on the ones which are not specified.” Mouse: Most frequently used. Pharmacology, genetics
Synonyms of SPF animals are –Disease free animals, of mammals, virology, models of human diseases.
healthy animals, pathogen free animals, clean animals, Rat : Physiology of cognitive processes, behaviour, models
caesarian derived animals. Historically, the concept of of diabetes.
gnotobiotic experimentation is credited to Pasteur’s efforts Rabbit : Serology, insulin quantification, pyrogens
in 1885 Nonhuman primates are important for disease quantification,.
investigation, therapy and vaccination (Schmidt, 1972 and Guinea-pig : In microbiology and serology, physiology of
Dormant et al., 1990). Within the past 20 years there has the auditory system.
been a widespread interest in the artificial rearing of animal Hamster : Genetics.
removed from the dam at or near the end of gestation and Frog : Physiology of blood circulation, electrophysiology.
keep isolated from conventionally reared animals. There Fish : Molluscs, insects.
are two primary reasons for producing animals under such These animals are used in studying of defense
conditions. The 1st of these is to break the cycle of the mechanism of the body. These animals are used in studying
some of the infectious disease organism present the ageing process of individual in animals and human
commonly transmitted from animal to animal. The 2nd is being. In body relationship between different microflora
to provide more uniform experimental animal for many can be study by using the specific pathogen free and
phase of basic and applied research by reducing one of gnotobiotic animals. These animals may be used as the
the variables diseases. It has been estimated that steril organs and tissue for different investigations and
approximately 20 million animals are being used for testing research. Different diet related researches and its reaction
and are killed annually; about 15 million of them are used can be estimated by using these animals.
to test for medication and five million for products. Purchase and techniques for the SPF and gnotobiotic
Different factors may may affects experimental animals animals :
which results in change in experiments (Melby, 1983; Small, Before the purchasing these animals we should get
1983). China has become one of the biggest country using the important information about these animals. We should
lab animals and highest number of lab animals are found select a defined specific pathogen list for the stock.
in China.(e.g., specific pathogen free; genetically modified) Different diagnostic and detection methods should apply
increasingly used in scientific research-16 million a year, during the purchasing. Previous screening and screening
compared to 12 million in the 25 European Union countries test organization should be cleared. By the use of history

HIND AGRICULTURAL RESEARCH AND TRAINING INSTITUTE

the Surveillance programme and disease history should Staff : A person should be in good health, active, intelligent
be carefully monitored. and well trained. Smoking eating drinking and eating should
The principle is that obtain animals from a stage in be prohibited in all working area. Person working with
their life cycle when they are either a minimum number this should be well aware and well understood of the
of contamination or not at all. operations aspects of the unit. Personnel are required to
Caesarian technique : The placenta acts as a very take a shower while entering and then don sterilized
efficient filter and prevents the fetus from becoming garments including hair nets, face masks, and jump suits.
infected with most bacteria, virus and parasites. The object Personnel facilities include staff and record room, sufficient
is to remove and free from the conventional pathogens. changing room, decontamination area and first aid.
– Normal parturition is delayed by giving daily injection Boot cleaning and disinfection : Visible organic material
of progesterone to the mother during the last three days may be removed from boots. Boots may be disinfected
of pregnancy. by soaking in a clean bath of an appropriate disinfectant .
– In case of bird fertilized eggs can be passed directly It is important to frequently empty, clean, and refill the
from the outside via the germicidal trap, to the interior boot bath to prevent it from being contaminated with
where hatch normally. organic matter. Disposable boots may be used.
– The pregnant dam is prepared for surgery by Training for SPF animal care and use : All SPF animal
removing the hair from abdomen by shaving. care and use personnel must be trained on SPF protocols.
– In case of small animals like rat and mice cervical All SPF trained personnel must follow the SPF care and
dislocation is followed which is quick and humane. For use guidelines at all times.
large spp. Halothane/oxygen mixture can be used for Veterinary rounds : The veterinary staff makes regular
anaesthesia. Hysterectomy used for obtaining young from rounds through the facility to observe the animals, their
the dam. housing conditions and husbandry procedures. All animals
– Gravid uterus is put in a sterile plastic box which in SPF care are observed daily by an animal care staff.
containing some sterilizing will kill microorganism. The Each area of the facility is also assigned a veterinary
uterus is opened dries the fetuses. The fetal membranes technician and an area veterinarian.
removed, leave the placenta in contact at the umbilicus Decontamination of the room : The room should be
for a short time while respiration has been initiated. properly sealed and left over night after fumigation and
– Young one usually requires stimulation by gentle thereafter should be properly ventilated The different
with sterile gauze and drying of the nostril. methods adopted by animal houses depend upon the
– Food is given by hand day and night until they are facilities available, the cost, the simplicity and efficiency
already inside the barrier. of the procedure. When liquid formalin is used 1 ml of 10
Hand rearing : The animals maintained at a temperature per cent solution for every cubic foot of the room space is
33-350C and the humidity kept at 50 per cent or higher required. The room temperature should be at least 180C
The hand rearing of new young like mice is difficult. So with relative humidity about 80 per cent. The sterilization
we can fed the new born mice with the help of rubber of all consumable is very essential Formaldehyde gas may
nipple or stomach tube which stimulated to pass urine and be produced by exothermic reaction. Usually two parts
faeces. of formalin are added to one part of crystals of potassium
Foster nursing : The foster nursing of surgically derived permanganate. When liquid formalin are added to one part
pups is possible in a barrier room. We should introduce a of crystals of crystals of potassium permanganate.
number of good foster mother, who should be mated so Animal room : Animal room should be protected against
as to deliver 1-3 day prior to the date of surgery. The new ingress by pets such as wild rodents and insects. Holes
born young should put into warmed receptacle containing created should be sealed. Adequate arrangement should
fostering mother. After half an hour foster mother is be provided to the receipt or incoming animals. Baker
removed from the litter and put with the new young ones. (1979) recommended a noise intensity of 85 db.Animal
Care and management brought into an animal house should not put at risk animals
– Discipline of the whole operation of SPF unit is that which are already there. In cage rearing system housing
of preventing reinfection and invasion of the clean area temperature may affect ed by the nesting materials and
by pathogen. A persons working inside should be clean type of choosen animals (Woods, 1980 and Corning, 1992).
because they act as carrier for many diseases. Where surgery is to be performed suitable operation
Rashtriya Krishi | Vol. 11 (1)| June, 2016 48 HIND AGRICULTURAL RESEARCH AND TRAINING INSTITUTE
 PRODUCTION & MANAGEMENT OF SPECIFIC PATHOGEN FREE & GNOBIOTIC ANIMALS

facilities should be provided including separate preparation the ‘dunk’ tank and there allowed to breed.
area for the animals, equipments and staff. There should Dispatch of animals : Weanling animals, other than
be post operative recovery area. The relative humidity of breeding replacements, are not kept in the rodent breeding
the laboratory animals should around 50 per cent and unit, but are transferred to a stock room in an experimental
ranges between of 40-70 per cent (Clough, 1987). unit. A litter is transferred by placing them in a sterile
Food and bedding stores should be clean dry vermins cardboard box which is then sealed into a polythene bag
and insect free. In addition food stores should be cooled and passed out through the ‘dunk’ tank.
and sunless provided with ventilation. Perishable food The litter then enters the experimental unit through a
should be stored in cool room refrigerators and freezers. further ‘dunk’ tank, where the polythene bag is removed
Floors should be moisture resistant, non-absorbent, and the litter caged. The transport boxes are stored flat
impact resistant, and relatively smooth, although textured and made up as required.
surfaces may be required in some high-moisture areas Biosecurity : Good biosecurity begins with personal
and for some species. cleanliness. Showering or washing facilities and supplies
A vermin free collection area should be provided for should be provided, and personnel should change their
waste prior to its disposal. Special arrangement should be clothing as often as necessary to maintain personal hygiene.
made for handling carcasses and radioactive or other Personnel should not be permitted to eat, drink, apply
hazardous materials. cosmetics, or use tobacco in animal facilities. Visitors should
Each animal room is emptied, cleaned and fumigated be limited as appropriate, and institutions should implement
with formaldehyde and water vapors at least once per appropriate precautions to protect the safety and well-being
year so as to prevent the buildup of bacterial contamination. of the visitors and the animals. It is essential that the
The fumigation is carried out by evaporating a mixture of agricultural animal care staff maintain a high standard of
formaldehyde and water to near dryness by boiling. In rat biosecurity to protect the animals from pathogenic organisms
and mouse house half a liter of formalin (40% that can be transferred by humans. Disposable gear such as
formaldehyde) and one liter of water is allowed for each gloves, masks, coats, coveralls, and shoe covers may be
1000 ft. required under some circumstances. Personnel should not
Inside the SPF unit : leave the work place in protective clothing that has been
Laminar flow hood (LFH) : A unit which provides a worn while working with animals.
sterile work environment by very high efficiency filtration Ventilation :Ventilation, humidity, temperature, lightening
of the air that circulates across the work surface. Room and noise contribute for good science. High level of
air is taken in through the back of the hood and passed Ammonia causes the respiratory problems in the rodents
through the HEPA filter. Sterile air moves across the work (Lindsey et al., 1978). In coming air should be filtered
surface from back to front and is expelled through the from dust particles, when most sources are also removed.
sash opening. Ultraviolet pathway within the dunk tank can be used.
MI cage/unit : A housing unit consisting of a Temperature should be maintained between 10 to 210C.
polycarbonate shoebox-style bottom, stainless steel wire- Number of air changes 5 to 15/ hr. provided adequate
bar lid and a polycarbonate top that holds a permeable ventilation. Heat is removed from the exhaust air by means
filter. Additional items also include bedding, water bottle, of a heat pump unit incorporated in the extract system
food, and cage card holder. and used to heat the water in the breeding unit. Light
Mobile shelf-unit (MSU) : Mobile shelving unit and cover intensity is very important for the laboratory animals
used to temporarily store and transport clean micro isolator because it may influence the aggression and cannibalism
cage units. in the animals (Fall, 1974; Weihe, 1976). Belhorn (1980)
Bath : The polycarbonate cage bottom located in the suggested a light intensity level of 323 lux (30 fc) for
laminar flow hood which contains the disinfectant solution. animals care and management practices.
If the bath becomes cloudy or excessively soiled with feces Materials of biological origin : Materials of biological
or bedding, the solution should be emptied in the sink and origin such as cells, tissues, serum and cultures will be
box rinsed with tap water. damaged or destroyed by autoclaving or gas sterilization
Introduction of animals : Specific-pathogen-free (SPF) techniques. Therefore, they must be tested before they
rats and germ-free mice were purchased; the animals inside are introduced into animals. Please contact Rodent Health
their delivery box passed into the breeding unit through Monitoring.
Rashtriya Krishi | Vol. 11 (1)| June, 2016 49 HIND AGRICULTURAL RESEARCH AND TRAINING INSTITUTE
 KULADIP PRAKASH SHINDE AND SHAILESH KUMAR GUPTA

Stem : Most commonly used method of sterilization carried Ed. Harlow, Essex: Longman Scientific and Technical.
by means of double autoclave situated in the barrier with Corning, B.F. and Lipman, N.S (1992). A comparison
one door. of rodent caging systems based on micro-environmental
Sterile diets : Diets are autoclaved and ethylene oxide parameters. Lab. Anim. Sci., 41: 498-503.
fumigation. Water is decontaminated by acidification, hyper
chlorination and/or filter sterilization,. Sometimes nutritive Dormont, D., Livartowski, J., Vogt, G., Chamaret,
value destroyed but sterilization with ionizing radiation and S., Nicol, I., Dwyer, D., Lebon, P., Guetard, D. and
microwave infrared is healthy. Montagnier, L. (1990). Second in vivo passage of HIV-
Screening and control of pathogen : Regular routine 2 rhesus monkeys. In: Animal Models In Aids.
sampling of stock for bacteria and parasites is done. Amsterdam: Elsevier. pp. 63-72.
Serological examination : Each SPF room will have Fall, M.W. (1974). The use of red light for handling wild
serology conducted every 3 months. The serum test will rats. Lab. Anim. Sci., 24: 686-687.
be done annually by a basic panel. The infected animals
will be immediately treated or discarded. Lindsey, J.R., Conner, M.W. and Baker, H.J. (1978).
Routine screening procedures : Swabs from the surface Physical, chemical and microbial factors affecting biologic
of the walls and floors of the breeding unit and samples of response. In: National Research Council/Institute of
the filtered water supply and of the ‘dunk’ tank fluid are Laboratory Animal Resources. Laboratory animal
taken four times at weekly intervals. These are examined housing. National Academy of Sciences, pp. 44-64,
for the number and type of organisms present. Salmonella, Washington, D.C., U.S.A.
Mycobacterium, Bordetella, Pasteurella, Mycoplasma Melby, E.C. JR. and Balk, M.W., (1983).Eds. The
and Corynebacterium. importance of laboratory animal genetics, health, and the
Hazardous wastes : That are toxic, carcinogenic, environment in biomedical research. Ont. Academic Press,
flammable, corrosive, reactive, or otherwise unstable TORONTO.
should be placed in properly labeled containers and
Schmidt, L.H. (1972). Problems and opportunities of
disposed of as recommended by occupational health and
breeding primates. In: Beveridge WIB, Editor. Breeding
safety specialists. In some circumstances, these wastes
Primates. Basel: Karger. p. 1-23.
can be consolidated or blended.
Record keeping : Record keeping is important for- Small, J.D.(1983).Environmental and equipment
– Animal house plans which includes typical floor plans, monitoring. In: Foster, H.L., Small, J.D. and Fox, J.G.,
all fixtures etc. eds. The mouse in biomedical research. III. Normative
– Breeding stock, purchases and sales records. biology, immunology, and husbandry. Ont. Academic Press,
– Minutes of institute animals ethics committee 83-100pp. TORONTO.
meeting. Weihe, W.H. (1976). The effect of light on animals. In:
– Records of sick animals. McSheehy, T.,Ed. Control of the animal house
– Death records. environment.Laboratory animal handbook 7.Buckden,
References : Huntingdon, Laboratory Animals Ltd. pp 63-76pp.,Cambs.,
Belhorn, R.W. (1980). Lighting in the animal U.K.
environment. Lab. Anim. Sci., 30 : 440-450.
Woods, J.E.(1980). The animal enclosure- A micro-
Clough, G. (1987). The animal house: Design, equipment environment. Lab. Anim. Sci., 30: 407-413.
and environmental control. In: Poole, T., Ed. UFAW
(Universities Federation for Animal Welfare) handbook
on the care and management of laboratory animals. 6th Received : 31.03.2016 Revised : 17.04.2016 Accepted : 16.05.2016

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