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 Table of Contents  
ORIGINAL ARTICLE
Year : 2021  |  Volume : 8  |  Issue : 3  |  Page : 222-226

A comparative evaluation of Typhidot and Widal tests for the detection of agglutinins against typhoid and paratyphoid bacilli in patients’ sera


Department of Microbiology, N.C. Medical College and Hospital, Israna, Panipat 132107, Haryana, India

Date of Submission30-Jul-2021
Date of Acceptance12-Aug-2021
Date of Web Publication03-Sep-2021

Correspondence Address:
Dr. Gurjeet Singh
Department of Microbiology, N.C. Medical College and Hospital, Israna, Panipat 132107, Haryana.
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/mgmj.mgmj_57_21

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  Abstract 

Background: The term enteric fever includes typhoid fever caused by Salmonella typhi and paratyphoid fever caused by Salmonella paratyphi A, B, and C. Typhoid fever has been virtually eliminated in the developed countries during the past several decades, mainly as a result of improvement in water supply and sanitation, but it continues to be endemic in the resource-limited nations of the world. The control of paratyphoid fever has not been so successful. Bacteriological diagnosis of enteric fever consists of isolation of the bacilli from the patients and the demonstration of antibodies in their sera. Widal testing is cost-effective and gives reports quickly. The Typhidot test is based on the method of immunochromatography test. It is easy to perform, and the results can be preserved for a long time. This study aims to compare the sensitivity and specificity of the Widal test and Typhidot test for the diagnosis of enteric fever. Materials and Methods: This prospective study was carried out at the Department of Microbiology, N.C. Medical College and Hospital, Panipat, Haryana, India, over 6 months from September 2020 to February 2021. In our study, a total of 150 clinically suspected patients were included. First of all, the area for blood collection was cleaned with spirit-soaked cotton and 10 mL of blood was collected with a sterile disposable needle and syringe from a median cubital vein (antecubital vein) of each patient after obtaining informed consent from the patient (from parents in case of age below 18 years). Seven milliliters of blood was immediately transferred to a blood culture bottle by cleaning the cap of the bottle for isolation and identification of causative organisms and 3 mL was transferred to a sterile plain vial for serum separation. Then the qualitative Widal slide agglutination test and Typhidot test were performed on each sample. Results: In the present study, out of the 150 clinically suspected enteric fever cases, maximum positivity 53 (35.33%) was given by the Typhidot test, followed by 36 (24%) given by the qualitative Widal slide agglutination test and 44 (29.33%) given by blood culture for S. typhi and S. paratyphi. Conclusion: Our study highlights the use of Typhidot for early diagnosis of enteric fever and also cross-checks with Widal test and blood culture for the diagnosis of enteric fever. Typhidot tests are easy to perform and less-trained personnel can also perform the test; they have been designed to detect IgM and IgG antibodies to lipopolysaccharide or outer membrane protein antigens of S. typhi and S. paratyphi. It can be useful in the early diagnosis of infection in the initial weeks of primary infection when a single serum sample is available, with a sensitivity and specificity of 90% and 80%, respectively, considering blood culture as a gold standard.

Keywords: Blood culture, sensitivity, specificity, Typhidot test, typhoid fever, Widal test


How to cite this article:
Singh R, Singh G. A comparative evaluation of Typhidot and Widal tests for the detection of agglutinins against typhoid and paratyphoid bacilli in patients’ sera. MGM J Med Sci 2021;8:222-6

How to cite this URL:
Singh R, Singh G. A comparative evaluation of Typhidot and Widal tests for the detection of agglutinins against typhoid and paratyphoid bacilli in patients’ sera. MGM J Med Sci [serial online] 2021 [cited 2021 Dec 2];8:222-6. Available from: http://www.mgmjms.com/text.asp?2021/8/3/222/325548




  Introduction Top


Enteric fever is a potentially fatal multisystem illness, and it comprised typhoid fever caused by Salmonella typhi and paratyphoid fever caused by Salmonella paratyphi A, B, and C. Typhoid fever has been virtually eliminated in the developed countries during the past several decades, mainly as a result of improvements of water supply and sanitation, but it continues to be endemic in the resource-limited nations of the world. The control of paratyphoid fever has not been so successful. The distribution of paratyphoid bacilli shows marked geographical differences. S. paratyphi A is prevalent in India and other Asian countries, Eastern Europe, and South America, S. paratyphi B in Western Europe, Britain, and North America, and S. paratyphi C in Eastern Europe and Guyana.[1]

Enteric fever is endemic in all parts of India. An incidence of 500–980 per 100,000 population has been reported in different studies varying with age and geographical area. Worldwide, 22 million cases are estimated to occur annually, with 600,000 deaths (the highest concentration in Asia). The proportion of typhoid to paratyphoid A is about 10:1. The disease occurs at all ages but it is probably most common in the age group of 5–20 years. The age incidence is related to the endemicity of the disease and the level of sanitation.[1]

Typhoid fever continues to be a major health problem especially in the Asia Pacific region, the Indian subcontinent, Central Asia, Africa, and South America.[2] Definitive clinical diagnosis of typhoid is unreliable because typhoid fever symptoms mimic other diseases such as fever that are common in this part of the world. Clinical presentations vary tremendously among patients and cover a wide spectrum; hence, there is the need for a good laboratory test.[3],[4],[5],[6],[7],[8],[9],[10]

Laboratory diagnosis of enteric fever includes isolation of causative organisms, which are routinely done on blood culture and feces culture. Bone marrow culture is valuable as it is positive even when blood cultures are negative but the sample collection procedure is painful and required specialized equipment and surgeon and it is time-consuming. Therefore, it can be done only when the other tests are inconclusive. Blood cultures are positive in approximately 90% of cases in the first week of fever, 75% in the second week, and 60% in the third week. Positivity rate declines thereafter and blood culture remains positive in 25% of the cases till the subsidence of pyrexia. The culture of bile is usually positive and may be useful in the detection of chronic carriers. The Widal test and Typhidot test are cost-effective, easy to perform; no specialized equipment is required and less-trained personnel can also perform the tests. In India, Widal tests are carried out at the largest number and are in high demand for the diagnosis of enteric fever.[1],[11],[12]

A key challenge to the effective control of typhoid fever is related to poor diagnosis. Diagnosis of typhoid fever in clinical settings is complicated because of overlapping symptoms with other common infections such as malaria, dengue, and viral enteritis.[13],[14],[15] For proper diagnosis, a test with good diagnostic performance especially in children with febrile diseases[16],[17] is of importance. In addition, the misuse of antibiotics via auto medication makes diagnosis difficult on a clinical basis.[18] The gold standard for diagnosis of enteric fever is blood culture, but this test not only has poor sensitivity in clinical settings but also is time-consuming and expensive for patients and clinics in remote settings in which culture facilities may not be always available and the population is poor.[19] The main diagnostic in such settings is based on the Widal slide agglutination test which is difficult to interpret for several reasons: cross-reactivities, time lag between infection and production of antibodies, and persistence of target antibodies long after treatment with very low correlation with active disease.[20]

The present study was therefore designed to compare the sensitivity and specificity of the Widal test and Typhidot test for the diagnosis of enteric fever accurately when compared with blood culture, which was considered as a gold standard technique.


  Materials and methods Top


This prospective study was conducted in the Department of Microbiology, N.C. Medical College and Hospital, Panipat, Haryana, India, over 6 months from September 2020 to February 2021.

Inclusion criteria

Patients clinically suspected of enteric fever attending tertiary care hospitals were included.

Exclusion criteria

Patients diagnosed with other diseases such as malaria, hepatitis, and dengue were excluded.

Study subject

A total of 150 patients with clinically suspected enteric fever were included in the study.

Ethical clearance

The project was reviewed and approved by the Institutional Ethical Committee of N.C. Medical College and Hospital, Panipat, Haryana, India vide their letter no. NCMCH/EC/2021/002, dated January 5, 2021.

Sample collection

After obtaining informed consent from the patient (from parents in case of age below 18 years), 10 mL of blood is collected by venepuncture under aseptic conditions and 7 mL of blood is transferred into a blood culture bottle containing brain heart infusion broth (BHI) through a hole in a cap by inserting the needle of the syringe rather than opening the bottle; this was done to avoid contamination from the external environment. Blood contains substances that inhibit the growth of the bacilli, and hence it is essential to dilute out these substances by adding 7 mL of blood into 70 mL culture media to make 1:10 dilution of blood and culture media. Blood culture bottles were incubated at 37ºC overnight. Subculture was made on blood agar and MacConkey’s agar plates from that bottle after overnight incubation. The bacterial isolates were confirmed by standard biochemical tests.

The remaining 3 mL of blood is transferred into a well-labeled plain tube and waited for 10 min to allow clotting of the blood. Serum was separated by centrifugation at 3000 RPM for 5 min. Serum was subjected to the qualitative Widal slide agglutination test and Typhidot test.


  Results Top


In the present study, we have included a total of 150 samples as per the inclusion and exclusion criteria, collected blood samples under strict aseptic precaution after obtaining consent from the patients, and performed the blood culture, qualitative Widal slide test, and Typhidot test of all samples. Out of the 150 samples, the male-to-female ratio was 1:2 (55 males and 95 females). Age-wise distribution was the maximum incidence of enteric fever, which was recorded in the age group 41–70 years, followed by 11–30 years and lowest in the age group of 71 and above. The qualitative Widal slide agglutination test showed a positive result in 36 (24%) samples and a negative result in 114 (76%) samples, followed by the Typhidot test in 53 (35.33%) positive and 97 (64.67%) negative samples. Among all samples, 44 (29.33%) were positive by blood culture and 106 (70.67%) were negative for blood culture [Table 1] and [Figure 1].
Table 1: Positivity rate of Widal slide test, Typhidot test, and blood culture

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Figure 1: Percentage of positivity rate of Widal slide test, Typhidot test, and blood culture

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  Discussion Top


The genus Salmonellae is motile Gram-negative of bacilli that infects or colonizes humans, leading to enteric fever, gastroenteritis, septicemia with or without focal suppuration, and the carrier state. The most important member of the genus is S. typhi, the causative agent of typhoid fever. The typhoid bacilli were first observed by Eberth in 1880 in the mesenteric nodes and spleen of fatal cases of typhoid fever and were isolated by Gaffky in 1884.[1],[21] It came to be known as the Eberth–Gaffky bacillus or Eberthellatyphi. Salmon and Smith in 1885 described a bacillus that was believed to cause hog cholera (mistakenly, as it is a viral disease). This bacillus later called S. cholera-suis was the first of a series of similar organisms to be isolated from animal and human beings, the genus Salmonella. It was subsequently realized that the typhoid bacillus also belonged to this group, despite minor biochemical differences, and it was redesignated S. typhi, the genus Eberthella having been abolished.[1],[21],[22]

In the present study out of the 150 samples, the qualitative Widal side agglutination test showed a positive result in 36 (24%) samples and a negative result in 114 (76%) samples. Olsen et al.[23] from Georgia, USA have reported that the sensitivity of the Widal test varies from 73% to 95% and the specificity of the Widal test varies from 73% to 80%. Willke et al.[9] from Turkey reported that using a cutoff of ≥1/200 for the O antigen test performed on acute-phase serum gave a sensitivity of 52% and a specificity of 88% with a positive predictive value (PPV) of 76% and a negative predictive value (NPV) of 71%. He reported that this increased to 90% sensitivity and specificity with a PPV of 88% and an NPV of 93% when the convalescent-phase serum was tested. Keddy et al.[24] from South Africa have reported the poor specificity and poor PPV of the Widal test.

Andualem et al.[25] from Ethiopia studied 270 participants, out of which 186 (68.9%) were females and 84 (31.1%) were males. Seven (2.6%) cases of S. typhi and four (1.5%) cases of S. paratyphi were identified with the total prevalence of typhoid fever of 4.1%. The total number of patients who are indicative of recent infection by either O or H antigen Widal test is 88 (32.6%). They reported that sensitivity, specificity, PPV, and NPV of the Widal test were 71.4%, 68.44%, 5.7%, and 98.9%, respectively.

In our study, out of the 150 samples, the Typhidot test shows positive in 53 (35.33%) and negative in 97 (64.67%) samples. Keddy et al.[24] from South Africa have reported that the Widal test had PPVs of 25.0% and Typhidot IgM of 56.7% and Typhidot IgG of 54.3%. Khoharo[26] from Pakistan reported that the Typhidot test was positive in 74 (97.36%) cases, with a sensitivity, specificity, and PPV of 96%, 89.5%, and 95%, respectively, compared with the Widal test which was positive in 56 (73.68%) cases with a sensitivity, specificity, and PPV of 72%, 87%, and 87%, respectively.

In our study out of the 150 samples, 44 (29.33%) were positive by blood culture and 106 (70.67%) were negative for blood culture. Mukherjee et al.[27] from Odisha reported that the slide test had a sensitivity of 93.33%, specificity of 82.86%, the PPV of 70%, and NPV of 96.67%, and the tube test had a sensitivity of 86.67%, specificity of 98.57%, PPV of 96.30%, and NPV of 94.52%, considering blood culture as the gold standard. Jesudason and Sivakumar[28] from Vellore, India, studied 563 samples and reported that the Typhidot test and blood culture were positive in 36 patients; both the tests were negative for 503 patients. The Typhidot test was positive for 9 patients with S. paratyphi A infection. They also mentioned that the sensitivity and specificity of the test using blood culture as gold standard were 92.3% and 98.8%, respectively, for typhoid fever.


  Conclusion Top


Our study highlights the use of Typhidot for early diagnosis of enteric fever and also cross-checks with the Widal test and blood culture for the diagnosis of enteric fever. The Typhidot test is easy to perform and less-trained personnel can do the test also; it has been designed to detect the IgM and IgG antibodies to lipopolysaccharide or outer membrane protein (OMP) antigen of S. typhi and S. paratyphi. It can be useful in the early diagnosis of infection in the initial weeks of primary infection when a single serum sample is available. The sensitivity and specificity of the Typhidot test were found as 90–92% and 89–90%, respectively. On the contrary, corresponding values for the Widal test were 70–75% and 83–85%, respectively, considering blood culture as a gold standard. Thus, the Typhidot test seems to be a practical alternative to the Widal test for the early diagnosis of enteric fever.


  Recommendation Top


The diagnostic methods for enteric fever include Widal, Typhidot for detection of antibodies, and blood culture for isolation of Salmonella. Widal testing is cost-effective and gives reports quickly. The Typhidot test is based on the method of immunochromatography test. It is easy to perform and the results can be preserved for a long time. Our study recommended the use of Typhidot test for the diagnosis of enteric fever.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

Ethical consideration

The study was undertaken based on samples received in Microbiology Laboratory; therefore, patient consent was not required. Institutional ethical clearance has been obtained vide letter no. NCMCH/EC/ 2021/002, dated January 5, 2021.



 
  References Top

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