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Widal test in Typhoid Fever Diagnosis – Procedure And Results

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Why is the Widal test In Typhoid Fever done?

The Widal test is done to screen or diagnose enteric fever (typhoid and paratyphoid fever). The test is widely used in developing countries like India, where the enteric fever is commonly found amongst its population. Maybe you want to know about Widal test procedure and about the role of widal test in Typhoid Fever Diagnosis here. So, Widal Test in Typhoid fever diagnosis has the following role:

Screening: here, the test is done to check the possibility of the infection in a suspected person. Here, only one blood sample is taken for the test, which shows a high level of antibodies (protective proteins) formed against the bacteria.

Diagnosis: when the test is done to confirm the presence of enteric fever. Here, two blood samples are taken from your body 10 to 14 days apart. The first sample is taken during the acute infection, and the 2nd sample is taken during the recovery period. A four-fold increase in the values between the two samples is classically considered diagnostic; however, with some deficiency.

What is the role of the Widal test in typhoid fever diagnosis in the present scenario?

The use of the Widal test for typhoid fever diagnosis  is controversial, with many studies suggesting significant false results.

However, due to some inherent advantages and lack of other quick and reliable tests, it is still in use in many parts of the world. So, the role of the Widal test in typhoid fever diagnosis are different in country and area wise.


Role of the Widal Test in Typhoid fever in developed countries

The widal test in typhoid fever is not used in developed countries like the USA due to the low occurrence of the infection and the availability of more reliable tests such as blood culture.


Roleof the Widal Test in Typhoid fever in endemic areas

• Single blood test: The role of a single blood test in typhoid prone regions, is not considered reliable by many studies due to the following:

o Repeated exposure of a person to a small amount of the salmonella bacteria and its related species.

o Immunization developed from a previous episode of typhoid fever

o False results from other infections such as malaria.

• Two blood tests: The role of two samples for diagnosis is also not used regularly due to inconsistent results and delay in the treatment because of the second blood sample.


Role of Widal test vs. Role of Blood culture

The gold standard test for the diagnosis of typhoid fever is blood culture. The test is

• Costly

• Not available everywhere

• Takes about 3-5 days to confirm the results.

Sometimes with the use of antibiotics, the blood culture results may also get affected.

However, the Widal test in typhoid fever is cheaper, commonly available, easy to perform, and needs minimal equipment. Thus, it is continued to be widely used in the endemic areas for giving a probable diagnosis of typhoid and paratyphoid fever. This can be further supported by a strong clinical suspicion and results of other tests.

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When is Widal Test in Typhoid Fever done?

Widal test in typhoid fever is generally done if you belong to a typhoid fever endemic area (all regions in India) and have a fever of more than 3 days.
This becomes more significant if you have no particular localizing signs and has vague symptoms such as:

Fever- particularly, which increases with passing days in the first week, becoming as high as 104-degree Fahrenheit (step ladder pattern).

• Loss of appetite

• Lack of energy (lassitude)

• General feeling of illness (malaise)

• Headache

• Nausea and vomiting

Abdominal complaints- diarrhea, constipation, pain, and distention.

Enlarged liver and spleen- on examination or ultrasound.

Specific signs- rose spots on skin and relatively lower heart rate. These signs are rare but classic for typhoid fever.

What sample is needed, and how is it collected??

You need to give a blood sample for this test. The sample is collected from the vein of your arm in the following steps:

• The technician or nurse will wrap a band around your arm to minimize the blood flow in the veins causing its engorgement and better visualization.

• He or she will use a little bit of alcohol-soaked in cotton to sterile the targeted area. Following which he will insert a thin needle into the vein to draw the blood. You may feel a little stinging sensation without causing a significant problem.

• The collecting tube or a vial is attached to the other end of the needle where the blood gets collected.

• Following this, the technician will loosen the band around your arm, allowing the blood to flow into the vial.

• After collecting a sufficient sample, he will unwrap the band and press the pricking site with sterile cotton or gauze and apply bandage.

What preparations are required for the test?

You need not to fast or do any special preparation for the test.

What is being tested in the Widal test?

Widal test in Typhoid fever detects and measures the amount of antibodies you develop against the bacteria that causes typhoid fever.

Any microbe that enters your body initiates a defense response in your body. This process produces some cells and proteins to kill the microbe. The antibodies are one such proteins that are formed, which circulates in your blood.

The serological tests such as Widal tests detect the presence of these antibodies in your blood to assess your exposure to the infective microbe. Thus, it helps to identify the cause of infection.

The Widal test in typhoid fever detects antibodies against two antigens present on salmonella typhi- the bacteria that causes typhoid fever. The two antigens are “H antigen” present on its hair-like projection and “O antigen” present on its body.


Widal Test Procedure

During the Widal test procedure, a special liquid containing “O” and “H” antigens is mixed with an equal amount of serum (filtered from your blood). If you have antibodies in your blood for the two antigens, then the mixing of the two liquids will form visible clumps. This is called agglutination.

The IgM antibodies against O antigen appears first during the acute typhoid fever while the IgG antibodies against H antigen forms later but stays longer.

There are two types of Widal tests- slide agglutination and tube agglutination tests. The slide test is used for initial screening, and the tube test is used to confirm the results of the slide test.

In both the tests, the liquid agent containing antigens is mixed with equal amounts of serially diluted serum obtained from your blood. The results are scored from 0 to +4 according to the degree of agglutination:

• 0 (no agglutination),

• +1 (25% agglutination)

• +2 (50% agglutination),

• +3 (75% agglutination)

• +4 (100% agglutination).

The least quantity of serum exhibiting a 50% or 2+ agglutination is denoted as the titer (endpoint of serum activity).


The results are stated as:

• No agglutination- in the absence of clumping, or

• Agglutination presents with titers of such as 1:20, 1:40, 1:80 or 1:160, and so on.

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What does the Widal test result mean?

The Widal test in typhoid fever detects the antibodies against “O” and “H” antigen of the salmonella typhi bacteria.

The presence or absence of clumping and the least quantity of serum required to produce it determines the result.


Widal Test
No agglutinationAbsence of antibodies against Salmonella antigens in the blood.
Agglutination presentPresence of antibodies in the blood against salmonella antigens. Titre values of:
1:20, 1:40, 1:80, 1:160, 1:320 with cut off value for:• IgM antibodies: 1:80• IgG antibodies: 1:160


There are several factors that sometimes make the test positive even when you don’t have typhoid fever. This is called a False-positive test, and the causes are as follows:

Other group D salmonella bacteria: Apart from the bacteria (salmonella typhi) that causes typhoid fever, there are many other salmonella bacteria grouped from A to E types. Group D consists of the salmonella typhi with many other bacteria that don’t cause typhoid fever but has the “O” antigen.

Group A and B salmonella bacteria: There are other bacteria from the group A and B that has “O” antigen similar to salmonella typhi.

The Widal test detects and quantifies the antibodies against the O antigen irrespective of type of Salmonella- whether typhoid or non-typhoid Salmonella.

Other microbes: Moreover, there are other microbes such as malaria parasites that may show similar antigen cross reactivity.

Previous infection or immunization: There are also significant chances that a person with a prior history of typhoid infection or repeated trivial exposures may develop antibodies in the blood. This is especially common in endemic areas where people have high chances of a past history of disease.



There are also several reasons when your Widal test comes negative even when you are suffering from typhoid fever. This is called a False-negative test, and the causes are as follows:

Use of antibiotics: if you have already taken antibiotics for your condition prior to getting the Widal test, it may lead to already decreased levels of antibodies in the blood. Thus, leading to reduced titers in the test and false-negative result.

Carrier state: in this state, you may carry salmonella typhi bacteria but may not develop the infection with significant antibodies to be detected in the blood.


Causes of negative Widal testCauses of positive Widal test
• No salmonella typhi infection

• Carrier state

• Prior antibiotic treatment

• Inadequate bacterial antigen in the host to produce antibodies

• Technical errors in the test

• Difference in commercial antigens preparation

• Presence of salmonella typhi infection- typhoid fever

• Prior infection or immunization with Salmonella bacteria antigen.

• Cross-reaction from non-typhoidal Salmonella

• Variability and inconsistently standardized antigen preparation

• Malaria infection

• Other diseases such as dengue

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