Seroprevalence of scrub typhus among patients with acute febrile illness: A study from northern districts of West Bengal

ABSTRACT

Background:

Scrub typhus is a re-emerging acute febrile illness with a high case fatality rate. Early clinical features are nonspecific and resemble any acute febrile illness, with or without skin rash creating clinical dilemmas. Untreated or undiagnosed cases can be complicated with myocarditis, pneumonia, acute kidney injury, meningoencephalitis, gastrointestinal bleeding, and multi-organ dysfunction syndrome. Antibody-based serological tests or IgM Enzyme-Linked Immunosorbent Assay against Orientia tsutsugamushi is a standard test for the diagnosis of scrub typhus within the first week of illness.

Aim:

The study aimed to determine the prevalence and epidemiological pattern of scrub typhus in acute febrile illness cases in northern districts of West Bengal, India.

Material and Methods:

Blood samples of patients suffering from acute febrile illness for more than five days duration were collected from different districts of the northern part of West Bengal and IgM capture Enzyme-Linked Immunosorbent Assay was performed at the Microbiology laboratory of North Bengal Medical College over the period, January 2019 to December 2023.

Result and Conclusion:

12.09% of the total sample tested were positive for IgM antibodies (n = 4721). Most cases were between 21 years to 60 years of age with a higher prevalence in males. The number of cases increased during the monsoon and later months. The overall death rate was 7.53%. Early diagnosis is essential for early treatment and improved survival. Surveillance and vector control strategies can effectively control the disease burden.

Introduction

Scrub typhus is a neglected tropical zoonotic disease characterized by an acute febrile illness. It is caused by Orientia tsutsugamushi, and transmitted by the bite of leptotrombidium mites. It is a zoonotic disease that witnesses a case fatality rate of as high as 40% in untreated cases.[1] The pathognomonic feature is a visible necrotic lesion known as eschar at the site of chigger bite (Leptotrombidium spp.), it is absent in 10%–20% of cases. However, its absence does not rule out the infection and the presence or absence of eschar does not alter the severity or the disease outcome.[2,3] This neglected tropical disease is spread throughout the Asia-Pacific belt, some parts of Africa and America, with most cases in Southeast Asia.[4,5,6]

India is an integral part of the Asia-Pacific tsutsugamushi triangle, it was assumed that this disease was only confined to the sub-Himalayan belt in India, but it spread out as an epidemic in Assam and West Bengal during the Second World War. The northern part of India is prone to Scrub typhus, but recent outbreaks have been seen in the sub-Himalayan belt starting from Jammu to Assam, the Northern part of West Bengal and Nagaland.[7,8] Many cases of acute encephalitis syndrome associated with Scrub typhus have been found in West Bengal, Tamil Nadu, Pondicherry, Rajasthan, Uttarakhand, Karnataka, and Kerala.[9,10,11,12,13,14] According to the passive national surveillance system in India, seroprevalence of Scrub typhus infection varies between 9.3% to 27.9% and constitutes about 25.3% of all acute febrile illnesses, with a mortality rate of 30% in untreated cases.[15,16,17]

It is associated with severe complications such as sepsis, acute respiratory distress syndrome (ARDS), hepatitis, myocarditis, pneumonia, acute kidney injury (AKI), meningoencephalitis, and gastrointestinal (GI) bleeding.[1,10] Although the response to drugs such as Doxycycline is excellent, it is often misdiagnosed or underdiagnosed because of mimicry of flu-like symptoms and the absence of eschar. Proper surveillance and research for accurate and early diagnosis and treatment can easily prevent complications and mortality. Serology is the mainstay of diagnosis, of which there are Weil Felix, Enzyme-Linked Immuno-Sorbent Assay (ELISA), and Immuno-Fluorescent Assay (IFA). Among molecular tests, there is a Polymerase Chain Reaction (PCR) from eschar biopsy. IFA is labor-intensive and requires fluorescent microscopy and trained personnel, whereas eschar biopsy is challenging, and Weil-Felix has poor sensitivity.[9,10] Conventional nested or real-time PCR can be used for the diagnosis of scrub typhus. qPCR targeting 16S rRNA gene, 60-kDa HSP GroEL gene, and 40-kDa HtrA OMP gene is faster with higher sensitivity and specificity. PCR can be performed from eschar, whole blood clots or buffy coat. PCR assay remains positive throughout the rickettsemia. Compared with the sensitivity and specificity of IgM ELISA with PCR, ELISA is more cost-effective for program purposes, and qPCR is preferred for the confirmation of diagnosis in reference laboratories and diagnosis in the early stages of infection. Because of the high degree of genetic variations, improving specificity by detecting multiple genes by molecular method is also the need of the hour.[18,19,20] For all these reasons, ELISA being more accurate than IFA, is the most widely accepted and utilized mode of rapid diagnostic test, especially in endemic areas.[21,22,23,24] The Indian Council of Medical Research also recommended IgM Capture ELISA with a cut-off Optical Density (OD) of 0.5 with a sensitivity of 93% and specificity of 91%, as the most sensitive test for the Indian population.[9,25]

The temperature and humidity of tropical regions are appropriate for chigger activity and pathogens transmission in the mites. Certain types of occupational and environmental factors are directly related to the endemicity of the disease in certain regions, such as exposure to rodents, bushy areas, woods, farmers, agricultural workers, distribution and density of population, and socioeconomic factors.[1,23] The majority of cases are related to outdoor activity, working on a farm and forest and rural people are more affected.[1,14,23] Studies from India also showed a post-monsoon rise in cases with the peak from July to November and a sudden decline in cases post-December.[14,21,23]

Disease outbreaks were observed in West Bengal in the 1960s and after a long time gap, a scrub typhus outbreak occurred in 2005 in Darjeeling and Kurseong which was intensively researched.[26] Though it is widespread in India and the Northern districts of West Bengal, specific prevalence data are not available in recent years. The present study aimed to detect the prevalence and epidemiological pattern of Scrub typhus in the northern districts of West Bengal from January 2019 to December 2023.

Material and Methods

Study setting

The study was conducted in the Department of Microbiology, North Bengal Medical College, Darjeeling, West Bengal, India. The Tertiary Care Hospital caters to all districts of North Bengal and the adjacent parts of Bihar, Assam, and Sikkim state. This region is a humid part of the Himalayas with forest and the highest rainfall of about 300 mm with an annual temperature range of 10 degrees Celsius. The areas include tea gardens, farms, rain forests, hills, and sub-Himalayan planes covered with grassland and forest that act as environmental niches for the mites.[1,14] There are regular reports of cases of scrub typhus by the Integrated Disease Surveillance Programme under the West Bengal State Surveillance Unit from this part of the country.

Study technique

It is a retrospective cross-sectional study conducted in the period January 2019 to December 2023.

Sample size calculation

Calculation done on the standard formula for a cross-sectional study: n = [Z2 α/2 P (1 − P)]/e2, where n = the sample size needed, Z = the value from the standard normal distribution corresponding to the desired confidence level (Z = 1.96 for 95% CI), P = the expected true proportion, and e = the desired precision. Under the assumption of P = 0.28 [18,19,20] and e = 0.04, a sample size of 485 was calculated.[27]

Inclusion criteria

1. Fever for more than 5 days

2. Admitted with Acute Encephalitis Syndrome

3. Consent for inclusion is available.

Exclusion criteria

1. Patients with other established causes of pyrexia or acute encephalitis syndrome and not related to infectious etiology, such as immunosuppressed patients, hematological or autoimmune disease, malignancy, etc.

2. Incomplete case records

3. Patients’ refusal to consent to the study.

Sample processing

A detailed history was taken and noted in case sheets. Blood was collected aseptically in a clot vial and serum separation was done by centrifugation (3000 rpm for 8 min). Samples were transported to the Microbiology laboratory immediately, maintaining a cold chain with the Case request forms containing all epidemiological and clinical details. A sample was preserved at −4°C if not processed on the same day and at −20°C if not processed immediately. A written informed consent was obtained for inclusion in the study and preservation of the sample for future research. Scrub typhus-specific IgM antibody ELISA test was performed using a Scrub typhus IgM Microlisa kit (Standardized kit). A cut-off OD >0.5 was considered positive for Scrub typhus according to the recommendations for cut-off titer in the kit literature. After the test serum was preserved at −70°C for future requirements. The study was approved by the Institutional Ethics Committee.

Data collection and analysis

Data were compiled and analysed using Microsoft Excel 2021.

Results

A total of 4721 serum samples were collected from the patients presenting with acute febrile illness from both outdoor and indoor admissions from Hospitals in the northern district of West Bengal.

Of these, 571 (12.09%) were positive for Scrub typhus in the study period. Year-wise distribution of positive cases is shown in Table 1. Males were found to be more infected (57.27%) than females (42.73%). [Table 2] Most cases were between 21 to 60 years of age. Age-wise distribution of suspected cases and positive cases are shown in Table 3. It was found that prolonged fever with a mean duration of 10 days was the predominant symptom. The characteristic eschar was present only in 16.7% of cases. 69.5% of cases presented with headache, loss of consciousness, altered sensorium, seizure with meningeal signs, or other features of acute meningitis or meningoencephalitis. Patients may also have lymphadenopathy or hepatomegaly. 8.7% of cases had complications such as hypotension, acute respiratory distress syndrome, acute kidney injury, or multiorgan dysfunction syndrome.

Table 1.

Year-wise positivity and percentage of positive cases during the study period (2019–2023) (n=4721)

Year	Test	Positive	Percentage
2019	546	72	13.19
2020	493	69	14
2021	786	109	13.87
2022	1390	170	12.23
2023	1506	151	10.03
Total	4721	571	12.09

Table 2.

Year-wise Scrub typhus IgM antibody positivity among male and female

Year	Positive	Male	Percentage	Female	Percentage
2019	72	58	80.56	14	19.44
2020	69	39	56.52	30	43.48
2021	109	68	62.39	41	37.61
2022	170	87	51.18	83	48.82
2023	151	75	49.67	76	50.33
Total	571	327	57.27	244	42.73

Table 3.

Age-wise distribution of study population and scrub typhus IgM-positive cases (n=4721)

Age	2019		2020		2021		2022		2023		Total case
	Test	Positive	Test	Positive	Test	Positive	Test	Positive	Test	Positive	Total test	Total positive	Percentage
<10 Yrs	85	9	80	10	172	24	412	39	272	15	1021	97	9.5
11–20 Yrs	115	10	102	11	127	17	182	17	239	19	765	74	9.67
21–30 Yrs	116	19	100	14	133	20	206	27	305	29	860	109	12.67
31–40 Yrs	80	10	66	12	113	14	192	30	213	32	664	98	14.76
41–50 Yrs	65	11	63	8	78	14	142	22	182	21	530	76	14.34
51–60 Yrs	46	7	50	11	72	11	152	24	156	23	476	76	15.97
>60 Yrs	39	6	32	3	73	9	104	11	139	12	387	41	10.59
Total	546	72	493	69	768	109	1390	170	1506	151	4721	517	12.09

The overall death rate is 7.53%. Other positive patients were treated with doxycycline and were discharged from the hospital in stable condition. The year-wise outcome of Scrub typhus IgM-positive cases is shown in Table 4. The number of positive cases increased from July to October and persisted till December, after that both the number of samples and positive cases gradually decreased. The seasonal trends of the disease are shown in Figure 1, which shows a seasonal trend of the disease in the monsoon and late monsoon.

Table 4.

Outcome of positive cases Year-wise

Year	Total positive case	Discharge	Percentage	Death	Percentage
2019	72	69	95.8	3	4.17
2020	69	58	84.1	11	15.94
2021	109	102	93.6	7	6.42
2022	170	158	92.9	12	7.06
2023	151	141	93.38	10	6.62
Total	571	528	92.47	43	7.53

Figure 1.

Seasonal distribution of positive cases (n = 571)

Discussion

Scrub typhus is a neglected but re-emerging tropical disease. The infection should be suspected in febrile patients with or without features of acute encephalitis. Early diagnosis and treatment with Doxycycline and Azithromycin can prevent complications and improve the outcome. Scrub typhus has been identified as an important cause of acute febrile illness in endemic regions.[1,14] In the Asia Pacific region one million people are affected every year.[5] A previous study by Mallick et al.[14] from this region showed a prevalence of 10.05%. A study by Thakur et al.[23] also showed a similar prevalence of 12%. Studies in the rural areas of Tamil Nadu have shown a seroprevalence of 25%. Particularly in the forest and hilly areas, which is much higher than what was found in the present study.[28] The prevalence of scrub typhus in this study was 12.09% compared to other studies where it varies from 13 to 63%.[29,30,31]

There is a male preponderance in the positive cases, which can be because men are more frequently outside the home, working in fields where they are more prone to the bite of the vector trombiculid mite.[1,23,32] Studies by other authors also showed a trend of increased infections in females, especially in rural areas because of the migration of males into cities for work and the involvement of females in agricultural work and outdoor activities in rural areas.[13,14,21]

The age distribution of diagnosed cases showed the middle-aged group was more commonly infected. 12.67% of cases are within the age group of 21–30 years and the prevalence was gradually increasing with age, i.e. 14.76% and 14.34% in 31–40 years and 41–50 years age groups, respectively, and 15.97% in 51–60 years age group. Other studies from India also showed similar trends, which maybe due to more involvement in outdoor and agricultural activities by adult age groups.[11,13,21,33,34] The age distribution data also varies in different countries. However, 62% of reported cases in Japan are within 51–75 years of age, whereas in China and Thailand, 50% of cases are above 50 years of age.[1,34] However, our previous study showed increased prevalence above 60 years of age which was like the study done by Trowbridge et al.[14,35]

In our study, the positive cases were mainly detected post-monsoon, from July extending up to November and December, which maybe because of an increase in scrub vegetation and growth of vectors in cooler months of the year. According to studies from other parts of the country, in southern regions, the disease is more prevalent in winter whereas it is more prevalent through the monsoon and post-monsoon in northern Himalayan states.[1,11,21,23,33,36,37,38] Our previous study from the same geographical area also showed a similar rise in cases during monsoon and post-monsoon.[14]

Fever is the predominant symptom in the study and only 16.7% of cases were presented with pathognomonic eschar. Other studies also showed acute fever as a common presenting symptom associated with headache, cough, breathlessness, and myalgia.[13,14,23,38] According to previous studies, eschar is rare in Southeast Asian patients, which varies from 10% to 70%. It is also difficult to detect small eschar on dark skin if not examined carefully. It may also be because of variation in infecting serotype or endemic strain circulating in this region.[13,21]

The patients diagnosed and treated early were discharged from the hospital in stable condition. However, the overall mortality in the present study was 7.53%. The deaths maybe because of delayed diagnosis or treatment or maybe drug resistance. It is also associated with the circulatory load of O. tsutsugamushi or the serotype involved.[1,13] The mortality rate due to scrub typhus varies between 6–30%.[23,39] Our previous study in this region showed a mortality rate of 32.76%, which may be because of delayed diagnosis, delayed treatment, or maybe genotypic variation in this geographical location.[14]

Conclusion

As an emerging tropical disease with high mortality due to scrub typhus in this part of the country, it should also be considered one of the important differential diagnoses of acute febrile illness. Detailed history with special emphasis on occupational history and exposure risk and careful examination of all cases of acute febrile illness should be considered as a protocol for early diagnosis of cases, which will eventually lead to prompt antimicrobial therapy since it has shown a very good response. The clinicians practicing in this part of the country should be properly oriented about the diagnosis, treatment, and outcome of the disease and they should be made aware of the prevalence of the disease. The clinician should remain especially cautious during monsoon and post-monsoon when handling any case of acute febrile illness with or without pathognomonic eschar. This would prevent the development of complications and would reduce the overall morbidity and mortality rate. Good surveillance in outbreak areas and early diagnostic strategies can effectively control neglected tropical diseases. Measures to control the vector can also be helpful in the preventive mechanisms.

Ethical policy and Institutional review board statement

Informed consent was obtained from each participant before conducting the study in the prescribed format as approved by the Institutional Ethics Committee.

Institutional Ethics Committee Certificate No: IEC/NBMC/M-04/25/2018 dated 23^rd December, 2018.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.