|Ahead of print publication
A comparative evaluation of rapid card and widal slide agglutination tests for rapid diagnosis of typhoid fever
Noor Jahan1, Razia Khatoon2, Priyanka Mishra3, Sudhir Mehrotra4, Siraj Ahmad5
1 Department of Microbiology, Integral Institute of Medical Sciences and Research, Lucknow, Uttar Pradesh, India
2 Department of Department of Microbiology, Hind Institute of Medical Sciences, Sitapur, Uttar Pradesh, India
3 Department of Microbiology, Integral Institute of Medical Sciences and Research, Integral University, Lucknow, Uttar Pradesh, India
4 Department of Medicine, Integral Institute of Medical Sciences and Research, Integral University, Lucknow, Uttar Pradesh, India
5 Department of Community Medicine, Integral Institute of Medical Sciences and Research, Integral University, Lucknow, Uttar Pradesh, India
|Date of Submission||10-Jun-2020|
|Date of Decision||10-Jun-2020|
|Date of Acceptance||07-Jul-2020|
Department of Microbiology, Integral Institute of Medical Sciences and Research, Integral University, Lucknow - 226 026, Uttar Pradesh
Source of Support: None, Conflict of Interest: None
Background: Typhoid fever is a serious public health problem. It causes severe systemic infection in lesser developed areas of the world. Although blood culture is regarded as the gold standard for diagnosis, it is a time taking procedure. An early and accurate diagnosis is necessary for an effective treatment. Aims: The present study was done to comparatively evaluate rapid card and Widal slide agglutination tests for rapid diagnosis of typhoid cases. Settings and Design: The study design was a cross-sectional descriptive study done over a period of 6 months from January to June 2018. Materials and Methods: A total of 265 patients suspected of typhoid fever who gave their consent were included in the study whose blood samples were tested by both rapid card and Widal slide agglutination tests. Statistical Analysis Used: The collected data were analyzed using SPSS Data Editor Software version 20. Percentage of variables was calculated. Results: Of 265 patients, 97 patients were positive by the Widal slide test, whereas 113 patients were positive by the rapid card test, with 96.9% sensitivity and 88.7% specificity. Of 113 positives, 83 cases were positive for immunoglobulin M (IgM) only, whereas 30 cases were positive for both IgM and IgG. Conclusion: Rapid card test is a simple and easy to perform the diagnostic test for rapid detection of typhoid cases with an additional advantage of separate determination of IgM and IgG antibody, thereby aiding in identification of current infection and previous exposure so that appropriate and timely treatment could be given to the patients.
Keywords: Rapid card test, rapid diagnosis, typhoid fever, Widal slide agglutination test
|How to cite this URL:|
Jahan N, Khatoon R, Mishra P, Mehrotra S, Ahmad S. A comparative evaluation of rapid card and widal slide agglutination tests for rapid diagnosis of typhoid fever. Med J DY Patil Vidyapeeth [Epub ahead of print] [cited 2021 May 10]. Available from: https://www.mjdrdypv.org/preprintarticle.asp?id=315471
| Introduction|| |
Salmonella More Details enterica subspecies enterica serotype Typhi, a Gram-negative bacillus, is causative agent of typhoid fever. It is a major cause of morbidity and mortality worldwide with an estimated 16.6 million new infections annually. Typhoid fever shows malignant effects if untreated. The disease is unique to humans and characterized by malaise, fever, anorexia, nausea, vague abdominal discomfort, transient rash, dry cough, and myalgia. These are followed by the coated tongue, tender abdomen, splenomegaly, hepatomegaly, bradycardia, and leukopenia. The most prominent complications are intestinal hemorrhage and perforation. Infection occurs in all age groups with higher incidence and variable clinical presentation in children. Both relapse and re-infection are common in typhoid and occur in <10% of cases. Re-infection can only be distinguished from relapse by molecular typing.
The variability is shown in incubation period of these Gram-negative bacteria from 7 to 14 days depending on the infective dose.
India is the second most populous country of the world with little access to modern diagnostic tools, with majority inhabiting the rural areas. Blood culture and Widal test are the routine laboratory tests commonly employed in all clinical laboratories. In India, Widal test has been used for the diagnosis of typhoid fever as it is a serological test with moderate specificity and sensitivity. However, a fast, reliable, and easy to perform serodiagnostic test with a higher sensitivity and specificity than a Widal test is required for rapid diagnosis and management of typhoid cases.
Hence, keeping the above facts in mind, the present study was done to compare the efficacy of rapid card test with the Widal slide agglutination test for the rapid diagnosis of typhoid fever among febrile cases attending our tertiary care hospital.
| Materials and Methods|| |
A cross-sectional study was done over a period of 6 months from January to June 2018, among all febrile patients clinically suspected of having typhoid fever and attending outpatient department as well as those admitted in the wards of Integral Institute of Medical Sciences and Research, Lucknow. Patients who were severely ill and suffering from other enteric diseases, those on antibiotics, and who were recently vaccinated and nonconsenting patients were excluded from the study. The present study was approved by the Institutional Ethical Committee letter number IEC/IIMSR/2018/12 conducted on April 04, 2018. A predesigned questionnaire was used to obtain the sociodemographic information of the patients included in the study. A total of 265 patients suspected of typhoid fever who gave their consent were included in the study whose blood samples were received in microbiology laboratory and tested by both rapid card test and Widal slide agglutination tests.
All the 265 blood samples were centrifuged and serum was separated. The sera were subjected to a Widal slide agglutination test using TYDAL Widal antigen set (Tulip Diagnostics Private Limited) according to the manufacturer's instruction [Figure 1]. Briefly, for screening, one drop of positive control was placed onto a reaction circle of the slide. Fifty microliter of physiological saline was placed in the next circle. One drop of the patient's serum was placed upon the required number of reaction circles of the slide. One drop of appropriate TYDAL antigen (Salmonella Typhi O antigen) suspension was added in each circle containing positive control and physiological saline, followed by the addition of one drop of Salmonella Typhi O antigen, Salmonella Typhi H antigen, Salmonella paratyphi AH antigen, and Salmonella paratyphi BH antigen suspension to the reaction circles containing patient's serum. The contents were mixed uniformly over the entire circle using separate mixing sticks. The slide was rocked gently for 1 min and observed for agglutination macroscopically. The sample positive for agglutination indicated the presence of corresponding antibody in the patient's serum and it was further titrated using the semi-quantitative method as per the manufacturer's recommendation. A positive agglutination was observed during the acute and convalescent period of infection. Due to cross-reactivity with other infectious agents, false-positive results may be produced. Both the screening and semi-quantitative tests were performed in the study.
In the present study, a rapid card kit used was OnSite rapid card by CTK Biotech [Figure 2]. The rapid card test utilizes the principle of lateral flow chromatographic immunoassay on a nitrocellulose membrane containing two test lines (G and M lines) and a control line (C line). The immunoglobulin (IgM) antibody, if detected, was suggestive of a current infection, while the IgG antibody indicated previous exposure to infection. The test was performed as per the manufacturer's instruction. Briefly, about 1 drop (30–45 µl) of the patient's serum sample was dispensed into the sample well followed by the addition of 1 drop of sample diluent into the center of the sample well. The result was read at 15 min. Formation of pink to pink-purple color band at the test region, in addition to the presence of C–line, indicated positive IgM or IgG result. The presence of C-line and absence of colored band on test lines indicated a negative result.
The collected data were analyzed using SPSS Data Editor Software Chicago, version 20. Percentage of variables was calculated. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of rapid card test were also calculated.
| Results|| |
A total of 265 febrile patients suspected of having typhoid fever were included in the study. [Table 1] shows that of 265 patients, the maximum number of patients belonged to the age group of 20–29 years (28.7%), followed by 19.4% patients who belonged to 0–9 years, while the least number of patients (1.1%) belonged to the age group of 70–79 years.
|Table 1: Distribution of febrile patients included in the study according to their age (n=265)|
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As shown in [Table 2], majority of patients (53.2%) were female, belonged to rural areas (68.7%), were inpatients/admitted patients (56.6%), and belonged to lower socioeconomic class (49.8%).
|Table 2: Distribution of patients according to their sex, residence, registration status and socioeconomic status (n=265)|
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All the 265 blood samples were tested in the microbiology laboratory by the Widal slide agglutination test, in which 97 (36.6%) samples were found to be Widal positive [Figure 3], while 168 (63.4%) samples were found to be Widal negative. All the samples were negative for Salmonella paratyphi AH and Salmonella paratyphi BH antigen. It was found that of 97 positive samples, majority (19.2%) were positive only for Salmonella Typhi O antigen denoting acute infection among these patients, as shown in [Table 3].
|Table 3: Distribution of results of Widal slide agglutination test according to the Antigen tested (n=265)|
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The 265 samples were also tested by a rapid card test which showed that 113 (42.6%) samples were positive [Figure 4], while 152 (57.4%) samples were negative by rapid card test. Of 113 positives, samples positive only for IgM antibodies were 83 (31.3%), whereas 30 (11.3%) samples were found to be positive for both IgM and IgG antibodies, as depicted in [Table 4].
|Table 4: Distribution of results of Rapid card test according to antibodies. (n=265)|
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The comparative evaluation of rapid card test and Widal slide agglutination test is shown in [Table 5], in which it was found that of 265 patients tested, true positivity by both rapid card and Widal slide agglutination tests was found among 94 (83.2%) patients, whereas 19 (16.8%) patients were detected as positive by rapid card test only. The true negative by both rapid card and Widal slide agglutination tests was found among 149 (98.0%) patients, whereas 3 (2.0%) patients were detected as positive by the Widal slide agglutination test only, which may be due to cross-reactivity. The sensitivity, specificity, PPV, and NPV of rapid card test were found to be quite high when compared to the Widal slide agglutination test, as depicted in [Table 5].
|Table 5: Comparison of efficacy of Rapid card test taking Widal slide agglutination test as standard (n=265)|
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| Discussion|| |
Typhoid fever is caused by Salmonella Typhi. Man is the only known reservoir of infection (cases and carriers). Clinical diagnosis of typhoid fever is difficult, particularly in the early stages. Laboratory facilities are essential to ensure the diagnosis, for the application of appropriate therapy and suitable public health management.
Widal test has been used for over a century in developing countries for diagnosing typhoid fever, but it has a low sensitivity and specificity. The role of Widal is more controversial when other members of Enterobacteriacae show the sharing of O and H antigens and also by other Salmonella serotypes. In the present study, 36.6% (97/265) of the patients were found to be positive by the Widal slide test, this is comparable with another study which shows Widal positivity of 40.67%. In contrast to our study, there were studies showing lower Widal positivity of 14.42% and 22.5%.,. Furthermore, there were studies reporting higher positivity of the Widal test to be 100%, 84.7%, 72.58%, and 74%, as compared to our study.,,,
The rapid card test offers simplicity, speed, economy, and early diagnosis with high sensitivity and specificity. The detection of IgM antibodies suggests acute typhoid in the early phase of infection, while the detection of both IgG and IgM antibodies suggests acute typhoid in the middle phase of infection. Since IgG antibodies can persist for more than 2 years after typhoid infection, the detection of specific IgG antibodies cannot differentiate between acute and convalescent cases.
In the present study, rapid card test showed that 42.6% (113/265) of the patients were positive for typhoid fever. This finding corroborates with other studies which reported positivity by rapid card test to be 51.34%, 48%, and 26%.,, In contrast to our findings, higher positivity of rRapid card test was reported by various other studies as 70%, 74%, and 74.3% positive cases.,,
In the present study, sensitivity and specificity of the rapid card test were reported to be 96.9% and 88.7%. A similar study carried out in India reported rapid card test's sensitivity to be 100% and recommended for its utility in conjunction with Widal test for an early diagnosis of typhoid fever. Another study showed 92.3% sensitivity and 98.8% specificity of the rapid card test. In another study done in a group of typhoid patients in Pakistan, typhoid test (rapid card test) had a comparable sensitivity of 94% and specificity of 77%. Another study showed a rapid card test's sensitivity of 100% and specificity of 76%.
The present study reported that of 265 enrolled patients, the majority were female (141) as compared to male (124). In contrast to our study, another study done in the year 2013 reported that of 200 patients, majority (119) were male as compared to female (81). In a study done in Iraq among 120 patients, majority (77) were male as compared to female (43). In a study done in the year 2016, of 92 patients, majority were male (48) and 44 patients were female. However, another study corroborates our finding as out of 1434 patients included in the study, majority were female (750) as compared to male (684). Another study done in 2015 shows contrasting findings as out of 1371 patients included in the study, majority were male (971) and 400 patients were female. A study done in the year 2013 also reported that of 163 enrolled patients, majority were male (89) and 74 patients were female.
| Conclusion|| |
Even today, the Widal test remains one of the most commonly used tests for diagnosing enteric fever in the developing world. Although slide Widal test being a rapid, convenient and simple test is often performed by most of the laboratories but it has a limitation of having high false positivity. On the other hand, rapid card test is simple, easy to perform, more reliable, and rapid screening test having high sensitivity and specificity as compared to the Widal slide test. Rapid card gives results within 15–20 min with an additional advantage of separate determination of IgM and IgG antibody, thereby aiding in the identification of current infection and previous exposure. This allows physicians to take immediate action and early institution of therapy. It is recommended to be adopted in routine clinical settings for early detection of typhoid fever where limited advance diagnostic facilities are available.
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Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]