Biotechnology

Animal Cell biotechnology

Biomarkers for animal disease diagnosis

Paper No. : 09 Animal Cell Biotechnology

Module :28 Biomarkers for animal disease diagnosis

Principal Investigator: Dr Vibha Dhawan, Distinguished Fellow and Sr. Director

The Energy and Resouurces Institute (TERI), New Delhi

Paper Coordinator: Dr. Minakshi, Professor & Head, Lala Lajpat Rai University of Veterinary & Animal Sciences, Hisar

Content Writer: Dr. Hari Mohan, Assistant Professor, Maharshi Dayanand University, Rohtak

Paper Reviewer: Dr. Minakshi, Professor & Head, LalaLajpatRai University of Veterinary & Animal Sciences, Hisar

Co-Principal Investigator: Prof S K Jain, Professor, of Medical Biochemistry

JamiaHamdard University, New Delhi

Biotechnology

Animal Cell biotechnology

Biomarkers for animal disease diagnosis

Description of Module

Subject Name Biotechnology

Paper Name Animal Cell Biotechnology

Module Name/Title Biomarkers for animal disease diagnosis

Module Id 28

Pre-requisites Basic knowledge of biomolecules, immunology and molecular biology

Objectives 1. To understand the concept of biomarker

2. To know the application of biomarker in animal disease diagnosis

3. Different types of biomarkers

4. Future prospects of biomarker discovery

Keywords Biomarker, Validation, ELISA, Biosensor, Transcriptomics, Proteomics

Biotechnology

Animal Cell biotechnology

Biomarkers for animal disease diagnosis

Table of Content:

1.) Introduction

2.) Ideal Biomarker characteristics

3.) Applications of Biomarker

4.) Type of Biomarkers in disease diagnosis

4.1.) Imaging based biomarkers

4.2.) Biochemical biomarkers

4.3.) Immunological biomarkers

4.3.1.)Types of Immunoassays based biomarkers

4.4.) PCR based biomarkers

4.5.) Loop mediated isothermal amplification (LAMP)

4.6.) Sensor based biomarkers

4.7.) Array based

5.) Discovery of biomarkers

6.) Role of Bioinformatics in Biomarker Identification

7.) Summary

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Animal Cell biotechnology

Biomarkers for animal disease diagnosis

1. Introduction

Since the era of human civilisation, animals continued to be best source of diet as meat and

milk, wearing items from their skin and wool, medicinal items from their blood etc. and most

importantly companion for recreation. They also contribute significantly to the economy of

farmers and those related with farm business. To achieve these benefit and also for welfare of

animals, the health of animal is of paramount importance. Animals undergo different

disorders in their life cycle which affect their health. Any deviation from the normal

physiological conditions which adversely affect the well being of animals and produce

specific symptoms or malfunctioning of the body's normal homeostatic processes is called

disease. These diseases broadly occur because of following reasons:

a) Because of deficiency of any nutrient or vitamin or mineral called deficiency disease.

b) Any disorder in normal metabolism called metabolic disorders. It may be genetic or

acquired.

c) Due to entry of some toxic material like snake bite, poisoning etc.

d) Due to attack of infectious organisms like bacteria, virus, parasite, fungus.

e) Any abnormal change in anatomy/ structure of organs in animals

Whenever disease attacks animals, it leads to production of physiological signals within the

patient. These physiological changes depend on the stage of the disease. Presence of

etiological agent, physiological changes etc are used as marker for disease diagnosis by the

physician. Many times due to some other interactions like effect of environment, stress,

immune-suppression, faulty managemental practices, secondary invaders etc. present a

complex state, in which it will become difficult to diagnose actual cause (etiology) of the

disease. Further conventional techniques of disease diagnosis are less sensitive, cumbersome

and time consuming. In between the suffering of animal and financial loss to farmer

continues. The condition may become graver if such disease is contagious and spread to other

animals of the farm or near-by location. Therefore, biomarkers which are rapid, sensitive, and

specific will help doctor to diagnose the cause of disease immediately and hence treatment

will be more accurate and faster recovery may be achieved. This will prevent unnecessary

suffering of animal, spread of disease to others and financial loss to farmers.

Biomarkers are methods or tools working as indicators of biological processes and

pathological states or causative agent that can be used objectively to measure and evaluate

normal or abnormal biological processes, presence of causative agent for assessing the risk or

presence of a disease. These tools or methods are specific for a particular disease like

detecting level of glucose will indicate about diabetics in animals.US FDA (2001) defines a

biomarker as a characteristic i.e. objectively measured & evaluated as an indicator of normal

biologic, pathogenic or pharmacologic responses to therapeutic intervention. WHO (2010)

defines Biomarkers as almost any measurement reflecting an interaction between a biological

system and a potential hazard, which may be chemical, physical, or biological; The measured

response may be functional and physiological, biochemical at the cellular level or a molecular

interaction.

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Biomarkers for animal disease diagnosis

2. Ideal Biomarker characteristics

1. It should be obtained from readily accessible body fluids like blood, plasma, urine,

sweat, saliva or hair and faeces.

2. Signal generated from biomarker must be accurate, sensitive and specific for

concerned disease.

3. It should not alter with other disorders, in different environment and other normal

conditions like age, sex, breed, strain etc.

4. Biomarker data/ signal generated must be reliably and reproducible when repeated.

5. It should not change much in wide type of population.

6. Technique or tool must be time effective and can easily be operated.

7. Biomarker results should be easy to interpret.

8. It must be cost-effective.

Once a potential biomarker has been found, it’s validated for its efficiency before it is

commercialised for mass uses. For biomarker validation following points are considered:

1. It should give reproducible result in population across geography within breed or

species concerned.

2. It should give same result in all sex and ages.

3. It is also validated that for which kind of specimen and sample it is useful like serum,

tissue, urine etc.

4. It is also ascertained that what conditions are required for sample collection like

temperature, pH etc.

5. Validation is also done at different physiological conditions like lactation, pregnancy

and hormonal status of body like different phase of oestrus cycle etc.

6. Sensitivity and specificity of test is also validated.

7. Similarly temperature and duration of storage of biomarker is also determined.

8. Effect of animal handling, managemental style, diurnal variation, diet etc is also been

studied to validate the biomarker.

3. Application:

Application of Biomarkers in Disease Diagnosis: Biomarkers are used for various purposes

like drug development, diagnosis of environment contaminates, safety limit, food

contaminates detection, disease diagnosis etc.

1. It helps to identify the disease, for example higher glucose level on consistent basis

indicates diabetics.

2. It helps to identify causative agent of disease like bacteria, virus etc.

3. It helps to identify toxicant or poison taken by animals

4. It helps to identify level of recovery after treatment

5. It helps to predict outcome of disease.

6. It helps to quantify the disease condition

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Biomarkers for animal disease diagnosis

4. Type of Biomarkers in disease diagnosis:

1. Imaging biomarkers

2. Biochemical biomarkers

3. Immunological biomarkers

4. PCR based biomarkers

5. Sensor based biomarkers

6. Array based

Figure 1: Different types of biomarkers based on the principle of working

4.1. Imaging based biomarkers: These are the biomarkers detectable as image. They detect

or diagnose diseases based on change in images of particular organ or tissue in disease

condition with respect to normal condition. Here either different instruments like X-ray, CT,

Electroencephalography, Magnetoencephalography, and MRI or probes link to a targeting

moiety are used to get the image of organ, tissue or cell. The oldest imaging technique

was X-ray based technique applied by Wilhelm Röntgen in 1895. In animal system X-Ray

based technique is generally used to study disease related with bone deformities like fracture,

while Electroencephalography (EEG) is used for study of brain related disease like trauma of

brain. Magnetic resonance imaging (MRI) is used to diagnose disease or problem related with

tumors, bleeding, injury, blood vessel diseases in body. Similarly ultrasound is generally used

to diagnoses diseases related with reproductive organ like anestrum and development related

disorder in foetus like detection of placental abnormalities, large offspring syndrome, still

birth etc. CT scan uses magnetic field; pulses of radio wave energy is used in MRI scan;

ultrasonic waves are used in Ultrasound imaging machine and electric impulse is recorded in

EEG to make pictures of organs and structures inside the body.

Probe link imaging techniques is used for molecular imagingfor in vivo visualization,

characterization and measurement of biological processes at the molecular and cellular levels.

Here signal generating probe is attached to a targeting moiety that target to bio-marker. Here

molecular probes (like florescent material, radioactive material etc.) are used which generate

signal when attached to the target moiety. This type of biomarker is generally used for

tumour detection where tumour cell express certain marker on their cell surface against which

these probes are used. So, in general for action of molecular probes the marker must be

present on cell surface in high number and interaction between marker and targeting moiety

must be strong and specific.

4.2. Biochemical biomarkers: In any disease, there is always deviation from normal

physiology of organism. Whenever this deviation produces significantly decrease or increase

in concentration of biomolecules or body cell produces new biochemical, this biochemical

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Biomarkers for animal disease diagnosis

product can act as a potential biomarker for such disease. Generally such changes occurs in a

number of diseases but sometime such change occur in specific kind of disease and such

biochemical change act a biomarker for such disease. This biomolecule may be metabolic

product or some enzyme can be expressed in some cell or body fluid and are quantified or

detected by some kind of chemical reaction. This quantification is generally done by

colorimetric reaction by using spectrophotometer or biochemical analyser or by ELISA plate

reader. Till now, this diagnostic marker is most commonly used method to identify or

confirm the disease in animal system. There are various metabolites or enzymes are reported

till now which indicates about a particular disease or organ disfuntion like, Cathepsin D is

associated with progression of rheumatic arthritis, high glucose in blood indicative of

diabetes, amylase and lipase enzyme level in pancreaititis, Cystatin C for acute phase protein

(C-reactive protein(СRP) and amyloid protein A) for inflammation response, GDH level

increases in hepatocellular damage, CKincreases markedly in rhabdomyolysis and aortic

thromboembolism, prothrombin for liver synthesis test, blood urea nitrogen (BUN) and

creatinine for renal function. These biochemical tests are generally used in combination as

very few of them have diagnostic values independently.

4.3. Immunoassays based biomarkers: Immunoassay use antigen-antibody reaction to

detect either antigen or antibody in question. This assay use either radioactive compound or

colour forming reagent or fluorescence or chemiluminescence reagent attached to antibody or

antigen against the targeted molecule (antigen like hormones or antibody formed against

infection). These chemicals give visual/ electric/ radioactive signal which is either visualised

or recorded to qualitative or quantitative determination of molecule in question. This assay is

used to quantify the hormone or other chemicals against which antibody can be raised in

different animal and can also be used to confirm the causative agent like bacteria, virus, and

parasite. Generally immunoassay based kit is made on principle of radioimmunoassay (RIA)

or Enzyme Linked ImmunoSorbant Assay (ELISA) or its modification. The radio-

immunoassays technique was first performed by Berson and Yalow (1959) for estimation of

insulin. Whereas, ELISA was first introduced by Engvall and Perlmann, (1971) and is

presently most widely used tool to measure a wide range of soluble molecules including

infectious diseases (viral, bacterial, parasitic, fungal), hormones, fetal proteins, toxins, serum

proteins, drugs and snake venom, etc.

All these techniques are used either to detect level of hormone to diagnose hormonal disorder

like anestrum, repeat breeder, hypo-thyroids, grave disease etc. in animals or to detect the

causative agent such as bacterial disease (Brucella, Chlamydia Corynebacterium, E. coli and

its different strain), Viral diseases (Swine fever, Aleutian disease, Avian adenovirus, Avian

encephalomyelitis, Avian infectious bronchitis, Avian leukosis, Aujeszky's disease, Blue

tongue, Bovine leukosis, Bovine parvovirus, Canine adenovirus, Canine distemper,

Coronavirus, Equine infectious anaemia, Equine infectious peritonitis, Feline leukemia, Foot

and Mouth Disease) and so on. The drawback of these assays is that sometime it give false

positive result more-importantly when animal had undergone vaccination against same

diseases. Similarly to diagnose we need a minimum quantity of protein (titer) in biological

fluids which if absent will give false negative result. This generally happen in sub-clinical

cases.

4.3.1.Types of Immunoassays based biomarkers:

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Biomarkers for animal disease diagnosis

a) RIA: it is oldest technique among all immunoassay techniques. It is like competitive

ELISA, where a known quantity of antigen is radiolabeled with radioactive material

like I-125 and compete with antigen of biological fluid for limited quantity of

antibody. There is competition between both kind of antigen for antigen-antibody

reaction. The binding percentage of radioactive antigen will be inversely proportional

to amount of antigen present in biological fluid. The radioactivity is measured by

either gamma counter or beta counter depending upon radioactive source and thus

with help of standard curve relative quantity of antigen in serum is known. This is

generally used for assay of hormones like T3, T4, progesterone etc for diagnosing

diseases related with hormonal disorder.

b) ELISA: ELISA is based on antigen-antibody reaction on a solid matrix where a

secondary antibody tagged with an enzyme will bound with complex. This enzyme in

presence of substrate will give colour whose intensity is directly proportional to

enzyme present in complex. The enzyme presence in complex is itself directly

proportional to antigen-antibody complex amount. The colour development is stopped

by acid and optical density is measured. Then with help of standard curve relative

quantity of antigen/ antibody in serum is measured. It is more advantageous than RIA

as it does not have any potential health hazard or does not require very special set up

for disposal of waste and running of test.

It has various type like direct ELISA, sandwich ELISA, indirect ELISA and various

modification like spot ELISA, Ellispot, lateral flow etc. Lateral flow assay is also

known as “hand-held” assays (HHA), as they are very simple to use and require

minimal training, can be performed by the help of simple instructions that include

pictures of positive and negative results. This can be performed even in field

conditions as they are very small and are typically designed on nitrocellulose or nylon

membranes contained within a plastic or cardboard housing. These assay are generally

done for confirmation of disease only, quantification is not possible here. Ellispot

assay is a method of visualising and counting cell types usually from blood samples

that have been labelled with one or more specific markers.

c) Electrochemiluminescence (ECL): In Electrochemiluminescence (ECL), detector

antibody is directly labelled with a chemiluminescent label. It is also like ELISA with

a difference that magnetic beads are coated with capture antibody (ECL) while in

ELISA antibody is coated over the solid phase. Here a sample containing antigen like

serum and other biological fluids are added to a mixture of capture anti body-coated

paramagnetic beads and a Ru-conjugated (ruthenium) detector antibody. After a short

incubation period, the analyzer draws the sample into the flow cell, where it captures

and washes the magnetic beads, and measures the electrochemiluminescence for

qualitative and quantitative detection of antigen.

d) Fluorescent Dyes based immunebioassays: These assays are sandwich-type assays

similar to sandwich ELISA except that that the detector antibodies are directly labeled

with lanthanide chelates such as europium, samarium, terbium, and dysprosium.

.4.4. PCR based biomarkers:

Polymerase chain reaction (PCR) based method and its modifications are generally used to

diagnose diseases caused by biotic agents like virus, bacteria, fungi and parasites. It not only

confirms the causative agents but can also confirm its strain/genotype. PCR based assay is

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Animal Cell biotechnology

Biomarkers for animal disease diagnosis

also done for diagnosis of genetic diseases. This technique is very sensitive, rapid,

inexpensive and can detect very small amounts of genetic material like RNA or DNA from a

variety of samples. Therefore, etiological agents can be detected in very less titre also. This

technique can also be used for uncultivable microbes or VBNC (viable but not culturable).

Moreover, in a mixed population, it can confirm the presence or absence of targeted

microorganism. Only precaution needed in this method is to design the primer which is

specific for that microorganism parasite against which we are targeting the PCR. Also along

with isolated genetic material we have to run on negative and one positive control for proper

diagnosis.

For microorganism with DNA as genetic material, we can run conventional PCR while

microorganism with RNA as genetic material, we have to first run reverse transcription

reaction. We can also diagnose these disease causing organisms and their relative

concentration by running Real-time polymerase chain reaction (PCR) which need probes and

are generally more sensitive than PCR. Human Immunodeficiency Virus (HIV), M. bovis,

Brucella abortus, Blue Tongue virus, Salmonella, Legionella, Listeria, verotoxin-producing

Escherichia coli, Giardia and Campylobacter etc. can be detected in animal sera or other

biological fluid or tissue by this technique. Similarly PCR along with sequencing or with

restriction enzyme digestion can also be used to identify diseases caused by mutation. RAPD,

RFLP and DNA fingerprinting techniques are modifications of this technique and widely

used in genetic disease identification. These genetic diseases if detected in early life either

can be treated or animal can be culled so that financial losses to the farmer can be minimized.

Example of genetic disease are genetically susceptible animals for anestrum, post-partum

anestrum, dwarfism, Progressive myelopathy, DAG sperm in cattle, Hip Dysplasia,

degenerative myelopathy etc.

4.5. Loop mediated isothermal amplification (LAMP): It is also a technique similar to

PCR technique and is simple, rapid, highly specific, cost-effective and can be performed to

confirm the causative agent at field level itself where outbreak of disease had taken place. It

is a single tube technique for the amplification of DNA. This methodology can be performed

within 15-60 minutes, by incubating the mixture of samples, primers, DNA polymerase with

strand displacement activity and substrates at a constant temperature (about 60-65°C). To

detect the presence of causative agent no need to run electrophoretic gel but it can be

visualised by naked eye, either change in turbidity, or by photometry or fluorimetry. A

number of kits in humans and animals had been developed for diagnosis of disease caused by

pathogens like Salmonella, Legionella, Listeria, verotoxin-producing Escherichia coli,

Giardia and Campylobacter. Main advantages of LAMP are as follow:

a) The amount of DNA produced in LAMP amplification process (400-800 µg/ml) is

considerably higher than PCR (4-40 µg/ml)

b) We can run 2-3 set of primers at a time so it become more specific and less chance

of false result due to mutation.

c) Even RNA based organism can also be detected. For this reverse transcriptase

enzyme is added instead of DNA polymerase.

4.6. Sensor based biomarkers: Leland C. Clark first demonstrated this technique in 1957 by

developing the first glucose enzyme electrode. Biomarker based on sensor generally exploit

different properties of biological sample which changes during course of disease and can

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Biomarkers for animal disease diagnosis

generate certain signals which can be assayed. These signal can be generated because of

change of pH as in acidosis and alkalosis condition, or because of absorbance change of

certain colour light as in pulse oxymeter sense absorbance of infra-red light or when

secondary antibody is attached with some signal generating agent in immune-assay as

discussed above. As per The International Union of Pure and Applied Chemistry (IUPAC)

definition biosensor is as “a device that uses biochemical reactions mediated by isolated

enzymes, organelles or whole cells to detect the effects of chemical compounds by electrical,

thermal or optical signals”.

So biosensor on basis of principle can be further divided as Optical sensor, Electrochemical

sensors, immunosensor. Imunnosensor are Electrochemiluminescence (ECL) and Fluorescent

dyes based sensor which are already discussed above. Electrochemical sensors including

Potentiometric immunosensor, Amperometric immunosensors and Conductometric

immunosensors. And Optical sensor examples are IR sensor, chemiluminiscence, Total

internal reflection spectroscopy (TIRS), Ellipsometry, Optical dielectric waveguides etc.

Biosensors instrument are made of three basic unit, biological recognition element, signal

transduction unit and signal processing unit. When samples are injected by special opening

and biological assay material in sample attached on a solid-state surface, enabling a reversible

biospecific interaction with the analyte (Biological recognition). Antibodies, receptor

proteins, nucleic acids, enzymes, cells and organelles can act as biological recognition

element for progress of interaction. This reaction will generate signal in form of electric

impulse, pH change, heat change, light change by other chemical and physical methods

which is captured and analysed by a signal transducer. This will give result through

transmitter on display.

Glucose meter is one example of electrochemical sensors which is used to monitor diabetic

condition in dogs and other animals. Here blood taken by instrument through capillary action

in known quantity react with enzyme electrode containing glucose oxidase or glucose

dehydrogenase enzyme attached on media. The enzyme is reoxidized with an excess of a

mediator reagent, like ferricyanide ion, at electrode to generate the electric current. The

current is measured and when analysed to give output in term of concentration. Other

example of such assay method is detection of diabetic ketoacidosis (DKA), and alcohol

amount in blood. Further various kind of biosensor has been made to detect different type of

cancer. Similarly, Thrombin (a coagulation protease generated at sites of vascular injury) can

be assayed by aptamer-based sensor.

4.7. Array based

All diseases cannot be diagnosed on the basis of single test as some may require a battery of

tests for correct diagnosis. In such cases of disease diagnosis, we have to go for number of

individual tests, which is a time consuming process. In such cases, array based diagnostic

tools are highly significant.

An array is an orderly arrangement of samples where matching of known and unknown

DNA/RNA/ Proteins samples is done based on base pairing rules. In microarray, we use solid

support (a microscope glass slides or silicon chips or nylon membrane), on which the spotted

samples (DNA, cDNA, or oligonucleotides, antibody or antigen) known as probes (with

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Biomarkers for animal disease diagnosis

known identity) are immobilized. Hundreds or thousands of such spots are fixed where spot

sizes are typically less than 200 microns in diameter, and are arranged in known manner.

Then after sample is incubated after processing in such a way that complementary binding of

the unknown sequences/ of protein will take place between fixed spot and sample contents,

thus allowing parallel analysis for thousands of gene or protein at a time for gene expression,

gene discovery, protein expression and protein discovery. Depending upon the kind of

immobilized sample used, microarrays are of different kind:

a) Microarray Expression Analysis: In this type of array the cDNA derived from the

mRNA of known genes is immobilized. The mRNA from sample from both the

normal as well as the diseased animal is kept to study relative expression analysis.

Spots with more intensity are over expressed and those which are less intensity are

less expressed. This expression pattern can be studied for analysis of disease. Same

kind of array is also developed for identification of RNA genome pathogens.

b) Microarray for identification of DNA based microorganism: Same approach

above is used where spot is made from one strand of various specific DNA of

microorganisms.

c) Microarray for Mutation Analysis: For this analysis, gDNA is used which may

differ with single nucleotide bases. This used to study Single Nucleotide

Polymorphism (SNP) and detecting them is known as SNP detection and this way

genetic diseases and its mutation are identified.

d) Microarray for protein: This is useful in examining relative abundance of large

number of proteins from two different sources/conditions. Here, antibody is fixed as

spot when antigen have to identified or antigen is fixed as spot when antibody have to

be identified for diagnosis of disease by studying over or under expression of proteins.

All these microarray based disease diagnostic techniques are mostly used for research

purpose only till now. But in future when proper standardization will be performed it

can be used commercially on economic way.

5. Discovery of biomarkers: With advancement of omics techniques, potential biomarker

candidate for diseases diagnosis has become quite promising. Now a day two group of

population one which contains healthy individual (control) group and other population with

suffering group is taken. Then samples are collected from both groups. Sampling choice

depend upon disease concerned for which diagnostic tool has to be developed. Then after

depending upon target molecule under study, appropriate omic tool is selected. If it is

expected to be genetic disease because of gene sequence change, then genomic approach is

selected. Here whole gene of animals under both groups is studied to find out any change in

the gene responsible for disease. Similarly when it is expected that it is because of change in

expression pattern of gene, then expression profile of genes are studied with the help of

transcriptomics pattern and then after un-transcribed reason of gene is studied to find out

which gene is over or under expressing under give disease conditions.

Table 1: List of important biomarkers developed for animal production and disease diagnosis

Sr. No. Disease condition/

application

Biological fluid taken Animal

1 Mastitis Milk, white blood cells Cattle

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Biomarkers for animal disease diagnosis

2 Marek’s disease Blood Chicken

3 Johne’s disease Blood Cow

4 Dilated Cardiomyopathy Cardiac tissue Dog

5 Brain Cancer Neoplastic tissue Dog

6 Ovarian cyst Follicular fluid Cattle

7 Copper toxicity Hepatic tissue Sheep

8. Steroid level Urine Cow

9. Liver cancer Hepatic tissue Fish

10. Kidney diseae Renal tissue Dog

When it is expected to be change in protein concentration or type change then proteomics

approach is used. Metabolomics approach is used when we have to study metabolome profile

changes in tissue or body fluid of both populations to find out metabolite based biomarkers.

In this way, potential candidate biomarkers are selected. These biomarkers are then validated

by different techniques like chromatography and immunoassay based techniques for protein

and metabolites, blotting for protein and gene expression and different types of PCR based

techniques for expression differences or genetic changes. After establishing the specificity of

the candidate biomarker, it become imperative to study its sensitivity and repeatability with

respect to variation in sex, age, breeds, species, condition of disease ( acute, per acute,

clinical, sub-clinical), temperature, pH, biological sample variation and other physiological

and pathological conditions it can be used as biomarker. After that depending upon biomarker

properties the approach for making commercially viable biomarker is selected.

Figure: Strategy for development of biomarker: Biological fluids contain a lot of information about every

type of physiological or pathological condition. It require a rigorous exercise to take that information out in the

form of biomarker and use it in clinical and field settings

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Biomarkers for animal disease diagnosis

6. Role of Bioinformatics in Biomarker Identification

With the rapid advancement in sequencing technologies and decrease in price, there is blast

of information generated in putative biomarker search experiments. This enormous data also

known as big data is very difficult to process manually. For a fruitful outcome from this

complex information, there is need to categorize, define standard for the format, processing

parameter and quality assurance of data sets. Public databases such as GenBank, DDBJ,

EMBL, MSDB have contributed enormously towards the advances made in nucleotide,

transcripts, protein knowledge. Although databases are well developed for DNA, RNA and

proteins but there is requirement of metabolite databases of same standard. Separate

databases for animal disease need to be developed. A number of genome projects on

important animals are either complete or in progression. With the help of bioinformatics tools

and software, it will be more easy and less time consuming to identify putative biomarkers.

7. Summary

Discovery of biomarker has great significance in term of increasing animal production. With

the latest development in Omics techniques, it has become possible to relate the change in

gene expression profile with the phenotypic outcome. These methods may be utilized in

improving the diagnosis and outcome of disease by monitoring the effectiveness of the

treatment strategies. Although these post-genomic era techniques seem very promising in

improving animal health and welfare, but these methods also suffer with technical challenges.

Very few biomarker molecules have made their way from research laboratory to the clinical

or field settings. Apart from the complexity of transcript, metabolite or protein data, issues of

cost per sample, practicality in field condition and acceptability among physician are some

major hurdles. When bringing a potential biomarker from laboratory to field condition, low

specificity and sensitivity may augment the problems. Many researchers do not move to large

scale experiments from their laboratory settings and it is an essential component for proper

validation of biomarker. Therefore, there is an emergent need of refining the laboratory

experiments which are focussed on change in transcripts, proteins or metabolites

concentration in different conditions. It is possible to combine the Omics techniques with the

necessary refinement in the putative biomarker experiment in order to come up with a

feasible and handy biomarker. These strategies hold enormous potential for augmenting

animal welfare, productivity and animal health.