Animal husbandry practices

Different Methods of Animal Identification Techniques

September 25, 2023

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Different Methods of Animal Identification Techniques

Scientific investigations, preclinical research, and pharmacological studies use a number of laboratory animals as subjects. Therefore, proper animal identification becomes a necessity. Animal identification techniques not only help keep a record of the animals but also aids communication between the healthcare professionals and researchers. Furthermore, the identification systems help experimenters comply with regulations of animal care institutions and research protocols, as animal safety and comfort are prime considerations in research involving laboratory animals.

An identification system should minimally include the researcher’s name, animal’s strain or breed, animal’s age, and source. Basic information like the above-mentioned is usually maintained on the cage cards. However, cage cards are useful when group housing is done. Whereas, when individual identification is required, ear tags, ear notch or tattoos are preferred techniques. The choice of identification method depends upon the housing type, and research protocol used. The identification method should ideally be inexpensive, permanent, and comfortable for animals and without any adverse effects. Following are the identification procedures used for laboratory animals specifically rodents.

Temporary methods

There are some simple temporary methods to mark the animals like tail marking with pen, clipping hair, and fur dying. Pen marks last for 1-2 days while hair clipping can identify the subjects up to 14 days. The temporary nature of these methods outweighs their benefits like easy application, stress-free, and low-tech.

Permanent Methods

Ear Notch/Ear Punch

Ear notching or ear punching is one of the simplest animal identification techniques used for rodents. This technique is inexpensive and effortless that does not require any specific training or equipment. The only requirement of this technique is an ear puncher/notcher. There are two types of punchers available: thumb punch and scissor punch. The thumb punch is preferred in mice while the scissor punch can be used for both mice and rats.

The procedure of ear punching for animal identification is discussed as follows:

The animal is first restrained either manually or with the help of a restrainer. In rats and mice, ear pinna can be punched after two weeks of age. The site of punching is preselected and marked. The ear puncher is placed on the preselected site and quickly engaged so that a definite and firm cut can be obtained. Mostly ear punching codes range from 1-9, however, if 10-99 are to be numbered, then ‘tens’ digits are marked on the left ear while ‘ones’ digits are marked on the right ear. After safe punching, the animal is returned to its cage.

Advantages and Limitations

Ear punching is inexpensive yet simple.
It does not demand specialized training or equipment.
It does not require anesthesia before the procedure.
Ear punching can provide DNA for genotyping by polymerase chain reaction (PCR).
The main disadvantage of the technique is that numbers fade after several weeks because of healing.
Fighting may give similar notches that make individual variation illegible.

Ear Tags

Ear tagging is also one of the uncomplicated and inexpensive animal identification techniques. This technique is permanent unless the tag is removed or fallen off. Labels are placed vertically at the base of the ear beyond the cartilage and in distal one-third of the pinna. Numbering on cards can range from hundreds to thousands depending upon the research or laboratory requirements.

The protocol followed for ear tagging is:

The animal is restrained manually.
The metal tag is grasped between the thumb and forefinger.
A coupling device is positioned near the base of the caudal one half of the ear.

Note: Tag should not impinge the ear canal.

Firmly press the thumb and forefinger together to couple the tag.
After tagging, return the animal to its cage safely.

Advantages and Limitations

Ear tagging is an inexpensive and straightforward procedure for animal identification.
Local anesthesia is required to ensure the proper placement of tags.
Infection at insertion sites may lessen the popularity of ear tagging among researchers.
Tags may dislodge or get entangled in the cages or wire-bar lids.
Improper tag placement can fold the pinna which may lead to inflammation and discomfort for the animal.
The metallic tags may become heavy for smaller rodents and cause stress.

Tattooing

Tattooing is a permanent technique used for animal identification. Equipment for tattoos consists of a pair of micro-tattooing forceps that can hold a hypodermic syringe and needle. It can be done on body parts. However, tail and ventral paw pads are the most appropriate sites for tattooing as they ensure proper vision.

The procedure of tattooing is discussed below:

Micro-tattoo forceps are prepared with tattooing paste according to the instructions. The animal is restrained manually to avoid shaking, and then animals are anesthetized with Ketamine. With a 27-gauge hypodermic needle attached to a 1-ml tuberculin syringe, tattoo ink is injected on the pre-selected site, and the animal is marked. The animal is safely returned to its cage.

Advantages and Limitations

Tattooing is a permanent yet tenable animal identification technique.
It helps the researchers mark the animals even 3 days of age, while ear punching or tagging demand at least 3-weeks.
Anesthesia is necessary.
Tattoos can be illegible on pigmented animals.
Equipment used is expensive and labor-intensive that require proper training.

Electronic Transponders/Microchips

Subcutaneous microchips or transponders are also used for animal identification. Glass-encased passive transponders are usually employed. These microchips are biologically inert and do not need any batteries for charging. Standard subcutaneous injection of approximately 2 mm to 13 mm microchip is quick and easy. A low energy radio signal is generated by the reader that will energize the chip to transmit its code. These codes are used as identification numbers. Time is measured in milliseconds at the receiving end.

The protocol for microchips is as follows:

The animal is restrained manually to avoid shaking and disturbance.
The mid-dorsal region between the shoulder blades is swabbed with 70% ethanol.
A microchip is implanted using a subcutaneous injection technique.
A signal at receiving device is assessed to confirm the transponder’s implantation.
Return the animal to its cage.

Advantages and Limitations

Little chances of error or tampering are expected.
The computer-driven system allows the researcher to scan the individual animals temporally.
The equipment used is quite expensive.
Chips may be too large for smaller rodents and neonates.

Toe Clipping

Toe clipping is the least favored technique for animal identification as it is painful and stressful for the subjects. This method requires the removal of distal phalangeal bone. The procedure followed for toe clipping is discussed as follows:

The animal is restrained with the help of anesthesia or vapor coolant. After restraining, the foot is cleansed with betadine solution. For hemostasis, the experimenter should place a sterile gauze over the site and apply pressure. The researcher should only clip one digit per extremity.

Advantages and Limitations

Toe clipping is a simple yet inexpensive method of animal identification.
Pain and distress outweigh its popularity.
It may lead to an abnormal gait of the animal.

The primary considerations for animal identification systems are animal strains, species, housing method (group or individual), skin pigmentation, duration of the research, and type of study conducted. The identification method should ideally be durable, ethically justifiable, and bio-compatible.

Biometric methods of Animal Identification

The word Biometrics is derived from the Greek bios (life) and metron (measure). Biometrics has long been used to describe the measurement and logging of biological data, such as animal and human populations (tracking of large numbers of similar life forms). Nowadays, particularly after the increase in human terrorism, biometrics is used to describe methods for non-invasive identification of individuals.

Advantages of biometric techniques

Traditional methods for marking animals can potentially affect their behaviour and cause harm, leading to erroneous research results and poor animal welfare (referanse om merking av pingviner).

Any method used to apply a marker to an animal entails some degree of stress related to capture, handling and restraint. In addition, many common marking procedures also involve tissue damage and therefore cause pain, such as branding (heat, cold or chemicals), tattooing, toe clipping, ear notching and tagging. Furthermore, wearing a mark may alter the animal’s appearance, social interaction, other behaviours and ultimately its survival. An ideal method should identify individuals reliably and permanently with no adverse effects on the animals.

Biometric methods have therefore been developed to recognize animals based on physical characteristics or behavioural signs. Some of these methods have been used for some time for reliable identification of humans.

An animal biometric identifier is any measurable, robust and distinctive physical, anatomical or molecular trait that can be used to uniquely identify or verify the claimed identity of an animal (Barron et al., 2009). Therefore a good biometric trait should be easily presented to a sensor and converted into a quantifiable format, should not subjected to changes over time and should differ in the patterns among the general population, the higher the degree of distinctiveness, the more unique is an identifier.

Biometric methods are non-invasive, do not cause pain and do not alter the appearance of the animal. These methods will therefore have no effect on the behaviour and survivability of the animals, except in cases where repeated capture and/or handling is necessary.

Examples of biometric identification methods

Visual patterns

Some species have external characteristics that are easy to recognize and that are unique for each individual animal. These include colour rings on snakes, body markings of zebras, belly patches in geese and eyespots on the wings of butterflies. These patterns can be photographed or filmed and used to recognize individual animals. Problems may occur in the field in different light settings or surroundings, but new techniques including digital photography and videofilimg have reduced these difficulties. Digital images can also be manipulated to make recognition easier. The method is cheap and at its simplest needs no more than paper and pencil. In addition, observations can be made at a distance, reducing the risk of stress and altered behaviour.

The most obvious biometric marker is the coat pattern of animals which often appears on major body parts as colourations of either fur, feathers, skin or scales. For example, zebras and tigers can be identified from their stripes; cheetahs and African penguins carry unique spot patterns and snakes have coloured rings (Burghardt, 2008).

In a Norwegian study, individuals of the Lesser White-fronted Goose, Anser erythropus, were identified by differences in individual belly patches. The patches were drawn and individuals followed over seven seasons with high accuracy. No individuals were found with similar patches (Øien et al., 1996). Two observers were always present to reduce the risk of mistakes.

Photographic identification has been used since the 1970s to identify aquatic animals such as dolphins and whales (Rugh et al., 1998). Individual bottlenose dolphins can be identified by comparing photographs of their fins, which display curves, notches, nicks and tears. Whales can be distinguished by the callosity patterns on their heads (Wells, 2002).

Nose-prints

This method has been used to identify cattle and was first published by Petersen (1922). The method was developed to avoid the potential for fraud associated with traditional marking methods such as branding, tattooing and ear tags. Both sheep and cattle can be individually identified on the basis of the arrangement and distribution of ridges and valleys on the muzzle (Ebert, 2006).
The method is cheap and simple: ink is applied to the nose and used to make an impression on paper, rather like taking a finger-print from a human. Its accuracy depends, however, on each print being taken in the same way, with the same pressure, ink and type of paper to avoid confusing two animals. They may also be difficult to read due to smearing and they require a trained eye to verify a match. The method is therefore dependent upon the operator’s skills. Nose prints have been shown to be stable over time.

Iris patterns

Iris recognition technology was originally developed for use in humans but has been tested in animals (Musgrave & Cambier, 2002). Iris scanning can be performed rapidly and images can be captured digitally. Its use in animals is limited by the fact that the iris pattern does not stabilise until the animal is several months old and may undergo alteration following injury or infection.

Retinal patterns

The retinal vascular pattern is a unique and distinct biometric trait in animals. It is based upon the branching patterns of the retinal vessels which are present from birth and do not change during the animal’s life. Individual blood vessels in the eye can be detected using a retinal scanner. This pattern can be recorded with a hand-held device about the size of a video camera. Scans from individual animals are registered in a database. Some devices can also measure GPS coordinates. This method can be used when marking cattle and can be compared to nose-prints. The method is also relatively cheap.

Retinal imaging and nose-prints of sheep and cattle were compared by Rusk et al. (1986). Nose-prints are a quicker method than retinal scanning, but retinal scans are easy to analyse for inexperienced operators (Howell et al., 2008). Computer software for the analysis of digital pictures from both retinal scana and nose-prints makes analysis faster, cheaper and more reliable.

Facial recognition

This method has been investigated as an identifier for sheep and was adapted from an independent-components algorithm for human face recognition (Corkery et al., 2007). However, despite the fact that this biometric method has been used by humans for thousands of years, it is difficult to design instruments that can perform facial recognition accurately.

Ear vessel patterns

Inspired by fingerprint identification of humans, the unique blood vessel pattern in the ear of rodents has been studied as a biometric identification method (Cameron et al., 2007). The animal’s ear is photographed from the front while applying backlight to provide a detailed, high-contrast picture of the blood vessels. The branching points of the ear’s blood vessels are automatically detected and compared between two images to identify the individual.

Bite marks

An impression of an animal’s bite marks can be used for identification purposes, in a similar fashion to its use in human forensic medicine. This method is not applicable to all animals and can be difficult to conduct without sedating the individual. For this reason, other methods are preferable.

Saliva sampling

Saliva contains DNA that can be used to recognize individual animals. The method is less invasive than the use of blood sampling to collect DNA.

Movement patterns

It has been suggested that aquatic animals can be identified by analysing their movement patterns using a tri-axial accelerometry device (Shepard et al., 2010). By measuring the movements of animals in three dimensions, their movement patterns can be stored and these can be used to diagnose aberrant behavioural patterns, such as those associated with infections. Accelometery may have the potential to be a powerful tool to produce maps for conservation purposes, where animal movements can be plotted.

Methods of relevance to biometric identification of fish

There are very few published studies where biometric methods have been applied to fish or indeed other aquatic species, for identification purposes. As mentioned above, these have mainly been applied to mammals such as whales and dolphins (Rugh et al., 1998; Wells, 2002). Of the methods described above, external body patterns are likely to be the only biometric methods of any relevance in the foreseeable future: