Widal Test: Definition, Principle, and Procedure

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Widal Test

Now-a days diagnosis has become the basic clinical practice all over the world. As many types of diseases are affecting individuals, and which is becoming new with its origin day by day, diagnosis is very much important for treating the affected individuals.

Widal test is one such method which is used to diagnosis the enteric fever by assuming the symptoms in the individual.

This test is now no longer performed in the any other developed countries like United states, as enteric fever is considered as an endemic and the resources are limited for the rapid use of affordable testing alternatives.

This method is easy to perform, where as the concerns remain about the reliability of the test.

This test is not specific for typhoid fever, it can also be positively used when the person does not have any other infections.

Enteric Fever

Enteric fever is considered as one of the life-threatening illness which is caused by the bacterium, Salmonella enterica serotype Typhi.

It is transmitted my means of contaminated fecal matter. The symptoms associated with this disease includes high fever, fatigue, headache, abdominal pain, constipation, diarrhea, weight loss and sometimes rashes tend to occur, commonly known as rose spots.

Early diagnosis and treatment of this disease is very much essential because this infection leads to serious complications as such severe bleeding in the intestine, perforation in the intestinal tract, etc., which arises within two days of the infection.

Features of Widal Test

In the year 1896, Georges Fernand Widal invented this test as indirect agglutination for enteric fever or undulant fever.

Here the bacteria causing typhoid fever is mixed with the serum which contains specific antibodies are collected from the individual.

In cases of Salmonella infection, the demonstration of this test results in false results. However, test results have to interpreted carefully in order to account for any history of enteric fever, generally people follow typhoid vaccination and general level of antibodies to get rid of this disease.

Typhi dot is another type of test used to diagnosis the typhoid fever, as salmonella typhi gives false results for widal test, other culture techniques like culturing of blood and urine or feces is being allowed.

Basically, widal test is an agglutination test which helps in detecting the presence of serum agglutin’s in the serum of the patients who has typhoid or paratyphoid fever.

When the facilities for culturing is not available widal test greatly helps in diagnosis the typhoid fevers in the endemic regions.

In this test the patient’s serum is tested for O and H antibodies against the antigen suspensions. On testing the serum against the antigen suspensions, salmonella antibodies start arising in the serum at the end of the first week and it rises sharply during the 3rd week of the endemic fever.

Where as in acute typhoid fever, O agglutin’s are detected. After 6 to 8 days of fever, H agglutin’s are detected after 10 to 12 days from the onset of fever.

It is usually preferable that two set of specimens for performing the test at the interval of 7 to 10 days. Salmonella antigens can be performed in slide and through tube techniques.

Principle of Widal Test

Bacterial suspensions which carry the antigen will agglutinate when exposing them to the antibodies of the salmonella species.

Patients suffering from the enteric infection possess antibodies in their serum that reacts and agglutinates in serial doubling dilutes of kills, or colored salmonella antigens in the agglutination test.

The central theme or principle of this widal test is that if homologous antibodies are present in the serum of the patients it will react with the respective antigens that are injected and forms a visible clumping on the test slides or agglutinations in the tubes.

The most commonly used antigens are H and O.

Procedure of Widal Test: Slide Method

– A drop of positive control placed on the slide in the first reaction cycle and further one drop of isotonic saline is pipetted on the next reaction cycle.

– Pipette one drop patient’s serum in the remaining four reaction cycles.

– One drop of widal test antigen “H” is added to the first two reaction circles.

– The antigens “o”, “H”, “AH”, “BH” are added in the respective order to the remaining circles.

– The contents have to be mixed uniformly over the entire reaction circle using separate mixing sticks, then by slitting the slide gently back and forth any agglutination formed is observed visibly.

Interpretation of Widal Test: Slide Method

Agglutination is a positive signal for the enteric fever which indicates the clinically significant levels of the corresponding antibodies in the serum of the patients.

On the other hand, no agglutination is a negative result and it indicates the absence of the clinically significant levels of the corresponding antibodies in the patient’s serum.

Widal Test: Test Tube Method

In widal tests, generally two types of tubes are being used namely Dreyer’s tube and Felicx tube.

Dreyers tube is used for H agglutination and Felix tube is used for O agglutination. But now a days Kahn tubes are used for both the types of agglutinations.

Procedure of Widal Test: Test Tube Method

Let us take four set of Kahn test tubes and label them in the numeric from 1 to 8 for O, H, AH and BH antibody detections.

First, we have to pipette out 1.9ml of isotonic saline solution the first tube. And then 1.0ml of isotonic saline in the remaining tubes.

Then 0.1ml of serum sample to added to the tube 1 and it is mixed thoroughly. Now transfer the diluted serum from tube one and mix it in the tube two using the same technique as of serial dilution and perform the same in the remaining tubes.

At last, add one drop of the respective widal test antigen in all the tubes and mix well and cover the tubes.

Further incubate the covered tubes at 37degree overnight for approximately 18 hours. Dislodge the sedimentation button gently and observe whether the agglutination takes place.

Interpretation of Widal Test: Test Tube Method

The Patients titre using the widal test antigen suspensions in the highest dilution form shows the visible agglutination results.

The sample which shows the titre of 1:100 or more for the O agglutination and 1:200 or more than that for the H agglutination is considered as clinically significant.

Usually, H agglutination is found as more reliable than the O agglutin. Agglutination usually starts appearing in the serum but the end of the first week which rises sharply in the upcoming 2nd and 3rd weeks and after that it declines.

Widal Test Citations


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