Scrub typhus and leptospirosis in rural and urban settings of central India: a preliminary evaluation

Introduction

Acute undifferentiated febrile illnesses (AFIs) are defined as febrile illnesses having a duration of ≤ 2 weeks, rapid in onset, caused by varied pathogens and with no indication of an organ-specific disease.^1–3 They are a cause of considerable morbidity, mortality and economic burden to low-income tropical nations. While dengue, malaria and enteric fever are quite commonly diagnosed, a major subset of AFI that remains generally undiagnosed is attributable to scrub typhus and leptospirosis.

Rickettsial disease is one of the neglected zoonotic diseases of public health importance, ⁴ now recognised as re-emerging. ⁵ Scrub typhus, caused by Orientia tsutsugamushi, is the most common rickettsial infection reported from many parts of Asia and the Pacific islands. ⁶ A prevalence as high as 47%–58% has been reported from south India.^7,8 An important concern is the high case fatality rate, which may be in the range of up to 30%–40% of untreated cases. ⁹ Leptospirosis is a common spirochaetal zoonosis caused by pathogenic strains of Leptospira interrogans and has caused outbreaks in large parts of India. These infections present with non-specific clinical features, indistinguishable on clinical grounds alone, though proper management is characteristically distinct. Therefore, laboratory confirmation of either scrub typhus or leptospirosis is essential.

Data regarding the prevalence of these infections from Chhattisgarh are non-existent. As these infections have been documented in neighbouring states and the environmental factors required for the survival and transmission are clearly present in Chhattisgarh, they should be expected to pose a problem in our state. A pilot study was thus conceived to delineate the prevalence of leptospirosis and scrub typhus in cases of AFI among urban and rural patients seeking medical advice and/or referred to All India Institute of Medical Sciences (AIIMS), Raipur, a tertiary care health institute in central India.

Material and methods

A prospective observational study was conducted between May and December 2018. Consecutive patients aged 1–75 years, with a fever duration of 5–14 days, and with no evident focus of infection following initial clinical evaluation, were recruited after informed consent (or assent, if applicable). Immunocompromised patients and those with malaria, enteric fever, dengue or sepsis were excluded from the study. To ensure a representation from the rural population, study participants were also recruited from Jan Swasthya Sahyog (JSS), a rural healthcare centre in Ganiyari village in adjoining Bilaspur. All patients underwent complete physical examination. Detailed clinical history, risk factors and demographic information were recorded according to a predesigned proforma. Complete blood counts, liver and renal function tests were noted from the patients’ records. Each patient had 5 mL and 3 mL blood samples collected depending on whether they were an adult or a child, respectively. Blood samples from JSS were transported to the Department of Microbiology, AIIMS Raipur in the standard triple-packaging cold transport system, along with duly filled case record forms.

Blood samples were centrifuged at 3000 G for 10 min and serum was aliquoted in two vials and stored at −20℃ for serological and molecular testing (if possible). Widal test for enteric fever and immune-chromatography for dengue and malaria were performed. The samples were subjected to scrub typhus Detect IgM ELISA (InBios International Inc., Seattle, WA, USA) according to the manufacturer’s instructions. This ELISA uses Karp, Kato, Gilliam and TA716 recombinant proteins of the 56-kD outer membrane protein and has a sensitivity of 84% and specificity of 98% for the IgM assay. ¹⁰ Absorbance was read at 450 nm using PR4100 Microplate reader (Bio-Rad Laboratories, Hercules, CA, USA). IgM ELISA was also performed for leptospirosis using Panbio Leptospira IgM ELISA kit (Standard Diagnostics Inc., Republic of Korea) according to the manufacturer’s instructions.

For determination of epidemiological cut-off for both kits, serum samples from 30 healthy age-matched volunteers were collected and tested. The mean +3 standard deviations (SD) was taken as cut-off optical density (OD). The cut-off/span was 0.5 OD for scrub typhus ELISA and 1.7 for leptospirosis ELISA. The remaining serum was stored at −20℃ for further testing.

The study was approved by the Institutional Ethics Committee and Research Cell, AIIMS, Raipur (301/IEC-AIIMSRPR/2017).

Descriptive statistics including frequency, mean and SD were calculated for the demographic data and laboratory parameters. Categorical variables were presented as percentage; continuous variables were presented as mean along with 95% confidence limit. The associations of disease complications and laboratory features with the outcome were analysed by univariate analysis. For all tests, a two-sided P value ≤ 0.05 was considered statistically significant. All statistical analyses were performed using Epi info version 7.2.

In the absence of any prevalence data, the cut-off titre of antibodies to scrub typhus was determined by testing serum samples from healthy volunteers by IgM ELISA (InBios Scrub typhus Detect). The cut-off titre was detected as 0.5 OD which is similar to some other studies.^11,12 As mentioned earlier, in India, IgM ELISA with a cut-off of OD 0.5 has been recommended to be most sensitive test to indicate recent infection with O. tsutsugamushi. ¹⁰

Results

During the study period, 169 patients with AFI were enrolled (80 boys/men, 89 girls/women; mean age = 31 years; age range = 3–83 years). Of the total, 105 (62.15%) samples were from patients of AIIMS, Raipur and 64 (37.87%) were those received from JSS.

Positive tests for scrub typhus are tabulated (Table 1).

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Table 1.

Prevalence of scrub typhus among urban and rural patients.

Study site	Total cases (n)	Positive cases (n (%))	Boys/men testing positive (n)	Girls/women testing positive (n)
AIIMS (urban)	105	19 (18.1)	3	16
JSS (rural)	64	16 (25)	4	12
Total	169	35 (20.7)	7	28

Girls and women were more commonly affected compared to boys and men (P < 0.001). Seropositivity for scrub typhus was highest in the month of September (18/44 samples tested, 41%) followed by October (10/32 samples tested, 31.2%)

Only one patient tested positive for leptospirosis.

Clinical profile

The mean duration of fever at presentation was 10 days. The most common symptoms were headache (68.57%), extreme weakness (57.14%), myalgia/arthralgia (54.29%), abdominal pain (51.43%), nausea/vomiting (48.57%) and chills (45.71%). The common clinical signs were abdominal tenderness, dehydration, hepatomegaly, pedal oedema and splenomegaly. Rash and lymphadenopathy were present less commonly. Eschar was present in only 5.71%. No co-morbidity was present in significant proportions (Table 2).

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Table 2.

Risk factors, clinical features and laboratory profile of patients with scrub typhus (n = 35).

	Number	%
Risk factors
Pets	9	25.7
Contact with animal carcass, excreta, urine	5	14.3
Exposure to flood	1	2.9
Clinical features
Prodromal symptoms
Headache	24	68.6
Extreme weakness	20	57
Myalgia/Arthralgia	19	54.3
Chills	16	46
Gastrointestinal features
Abdominal pain	18	51.4
Abdominal tenderness	17	48.6
Nausea/vomiting	17	48.6
Hepatomegaly	10	29
Splenomegaly	8	22.9
Loose stool	5	14.3
Icterus/jaundice	4	11.4
Respiratory features
Breathlessness	12	34.3
Cough	7	20.2
Crepitations	2	5.7
Signs of dehydration	11	31.4
Bilateral pitting oedema	8	22.9
Calf muscle tenderness	7	20
Neurological features
Irritability	4	11.4
Neck stiffness	3	8.6
Feeding difficulty	1	2.9
Rash	4	11.4
Decreased urine output	3	8.6
Overt bleeding	2	5.7
Eschar	2	5.7
Lymphadenopathy	2	5.7
Laboratory parameters
Anaemia (Hb <110 g/L)	15	42.9
Thrombocytopenia (platelets <100 × 10/mm3)	13	37.1
SGPT/SGOT (>×2 N)	12	32.3
Hyperbilirubinemia (total bilirubin >22 µmol/L)	3	8.6
Leucocytosis (>11,000/mm3)	3	8.6
Creatinine (>190 µmol/L)	1	2.9

Discussion

Scrub typhus is an emerging public health problem and undoubtedly an important cause of AFIs. These infections may pose a serious public health threat if not diagnosed on time or are misdiagnosed. A low index of suspicion, protean manifestations, and lack of sensitive and specific diagnostic tests contribute to notorious difficulty arriving at a correct diagnosis. Moreover, failure of timely diagnosis and correct treatment leads to significant morbidity and mortality. A simple treatment with doxycycline is effective in treating the patient.

In India, the disease was documented for the first time during World War II among field troops in Assam and West Bengal. Since then, a pan-India presence of scrub typhus has been well documented, particularly from south India and the Himalayan region of northern India.^13–15 Outbreaks have since been reported from many other areas of sub-Himalayan north India and central India.^16,17 Our study reports the prevalence in Chhattisgarh state for the first time.

In our study, 35 (20.7%) patients with AFIs were found positive for scrub typhus, similar to other studies.^10,11,18 The maximum number of positive cases was found in the rainy season.^12,19

Percentages of seropositivity among rural patients tend to be higher than urban patients. ²⁰

Non-specific symptoms, including gastrointestinal, predominate.^12,20–23 Eschar at the site of attachment of the larval mite/chigger is considered pathognomonic of scrub typhus, but occurs in a variable proportion. ²⁴ It is a blackish necrotic lesion resembling a cigarette burn generally found in areas where the skin is thin, moist or wrinkled, and where the clothing is tight such as over the abdomen, groin and legs. ²⁵

It is noteworthy that only one patient out of 169 tested positive for leptospirosis. There are various reports of leptospirosis outbreaks and its prevalence in various parts of India including the Andamans, Tamil Nadu, Karnataka, Maharashtra, Andhra Pradesh and Orissa; a few cases are also reported from northern India.^26–32 With leptospirosis prevalent all over the country, the absence of cases in our study, despite presence of conducible environmental conditions, is noteworthy.

A significant limitation of this study is the relatively small number of cases, which ideally need molecular diagnostic tests for confirmation.

Conclusion

As up to one-quarter of patients with AFIs may be attributable to scrub typhus, serological assays must be included in the diagnostic work-up of patients presenting with AFIs. Tests for leptospirosis are recommended where outbreaks have occurred and where prevalence is known to be significant.