Accuracy of AccessBio Immunoglobulin M and Total Antibody Rapid Immunochromatographic Assays for the Diagnosis of Acute Scrub Typhus Infection

Using archived samples, we assessed the diagnostic capacity of a rapid immunochromatographic test (ICT) for the detection of Orientia tsutsugamushi IgM and total antibodies to aid with the diagnosis of acute scrub typhus infection in febrile patients in Laos. The sensitivity and the specificity of the ICT for the detection of IgM were 96.8% (121/125 samples; 95% confidence interval [CI], 92.1 to 99.1%) and 93.3% (98/105 samples; 95% CI, 86.7 to 97.3%), respectively. For the detection of total antibodies, the sensitivity was 97.6% (122/125 samples; 95% CI, 93.1 to 99.5%), but the specificity was much lower, at 71.4% (75/105 samples; 95% CI, 61.8 to 79.8%).Scrub typhus, caused by Orientia tsutsugamushi, is an important acute febrile illness in the Asia-Pacific region. As very few health facilities have accessible accurate diagnostic tests, the diagnosis of scrub fever must be based on clinical features. However, this is difficult because the clinical symptoms and signs are similar to those of many other febrile diseases, such as murine typhus, leptospirosis, and dengue virus infection. The diagnosis of scrub typhus infection has relied on the detection of O. tsutsugamushi antibodies during the acute phase of the disease, and the “gold standard” assay is the indirect immunofluorescence antibody assay (IFA) (9). The development of rapid, diagnostic tests by the use of immunochromatographic test (ICT) technologies has provided a mechanism for point-of-care serological testing. The objective of the study described here was to assess the diagnostic capacities of two commercial rapid ICTs for the detection of O. tsutsugamushi IgM and whole antibodies to aid with the diagnosis of acute scrub typhus infection by the use of stored, characterized sera collected from febrile patients in the tropical environment of the Lao People''s Democratic Republic (Laos) and Thailand where scrub typhus is endemic.     

Diagnostic Accuracy of the InBios Scrub Typhus Detect Enzyme-Linked Immunoassay for the Detection of IgM Antibodies in Northern Thailand

In this study, we examined the diagnostic accuracy of the InBios Scrub Typhus Detect IgM enzyme-linked immunosorbent assay (ELISA) and determined the optimal diagnostic optical density (OD) cutoffs for screening and diagnostic applications based on prospectively collected, characterized samples from undifferentiated febrile illness patients in northern Thailand. Direct comparisons with the serological gold standard, indirect immunofluorescence assay (IFA), revealed strong statistical correlation of ELISA OD values and IFA IgM titers. Determination of the optimal ELISA cutoff for seroepidemiology or screening purposes compared to the corresponding IFA reciprocal titer of 400 as previously described for Thailand was 0.60 OD, which corresponded to a sensitivity (Sn) of 84% and a specificity (Sp) of 98%. The diagnostic performance against the improved and more-stringent scrub typhus infection criteria (STIC), correcting for low false-positive IFA titers, resulted in an Sn of 93% and an Sp of 91% at an ELISA cutoff of 0.5 OD. This diagnostic ELISA cutoff corresponds to IFA reciprocal titers of 1,600 to 3,200, which greatly reduces the false-positive rates associated with low-positive IFA titers. These data are in congruence with the recently improved serodiagnostic positivity criteria using the Bayesian latent class modeling approach. In summary, the InBios Scrub Typhus Detect IgM ELISA is affordable and easy-to-use, with adequate diagnostic accuracy for screening and diagnostic purposes, and should be considered an improved alternative to the gold standard IFA for acute diagnosis. For broader application, regional cutoff validation and antigenic composition for consistent diagnostic accuracy should be considered.     

Nested polymerase chain reaction on blood clots for gene encoding 56 kDa antigen and serology for the diagnosis of scrub typhus

Purpose: Scrub typhus is a zoonotic illness endemic in the Asia-Pacific region. Early diagnosis and appropriate management contribute significantly to preventing adverse outcomes including mortality. Serology is widely used for diagnosing scrub typhus. Recent reports suggest that polymerase chain reaction (PCR) could be a rapid and reliable alternative. This study assessed the utility of these tests for scrub typhus diagnosis. Materials and Methods: Nested PCR to detect the 56 kDa antigen gene of O. tsutsugamushi was performed on blood clots from 87 individuals with clinically suspected scrub typhus. Weil-Felix test and scrub typhus IgM ELISA were performed on serum samples from the same patients. As a gold standard reference test was not available, latent class analysis (LCA) was used to assess the performance of the three tests. Results: The LCA analysis showed the sensitivity of Weil-Felix test, IgM ELISA and PCR to be 59%, 100% and 58% respectively. The specificity of ELISA was only 73%, whereas those of the Weil-Felix test and PCR were 94% and 100% respectively. Conclusion: Nested PCR using blood clots while specific, lacked sensitivity as compared to IgM ELISA. In resource-poor settings Weil-Felix test still remains valuable despite its moderate sensitivity.     

Assessment of Two Immunoassays for Detection of IgM Antibodies to Scrub Typhus Using a Serum Panel

Laboratory tests are necessary for diagnosis of scrub typhus (ST) especially in the absence of the distinctive eschar. Performance of an ELISA and ICT (immunochromatography) to detect IgM antibodies to scrub typhus was assessed using a panel of 346 sera chosen from healthy individuals, those with scrub typhus and scrub-typhus like illness. A sensitivity of 98.7% for ST IgM ICT and 97.4% for ST IgM ELISA was observed while specificity was 96.3% for ICT and 95.9% for ELISA. As excellent concordance (98.8%) was noted between the two assays, IgM ICT can be used for rapid diagnosis of scrub typhus.     

Evaluation of an Immunoglobulin M Capture Enzyme-Linked Immunosorbent Assay for Diagnosis of Orientia tsutsugamushi Infection

To differentiate scrub typhus from other acute febrile diseases, a rapid and reliable serological diagnosis is important. We developed an immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA) for diagnosis of recent Orientia tsutsugamushi infections in humans. The 56-kDa major outer membrane protein of O. tsutsugamushi is well known as the most immunodominant antigen in scrub typhus. The test is based on the use of the biotinylated recombinant 56-kDa protein of O. tsutsugamushi Boryong, Bor56, which was expressed as a fusion protein with a maltose-binding protein in Escherichia coli. In the test, the serum IgM antibodies were captured by anti-human IgM antibodies coated onto a microtiter plate. The captured IgM antibodies were revealed through sequential addition of biotinylated Bor56 antigen and peroxidase-conjugated streptavidin to the plate. The IgM capture ELISA was compared with the immunofluorescence antibody assay (IFA) by testing 176 serum samples from patients with diagnosed cases of rickettsial disease and patients with other acute febrile diseases. Of the 81 IgG IFA-positive samples, 78 tested positive (sensitivity, 96.3%) and all 31 IgM IFA-positive samples tested positive (sensitivity, 100%) by the IgM capture ELISA. The specificity of the IgM capture ELISA was 99%, and 1 of the 95 IFA-negative samples was positive in the assay. These results strongly suggest that IgM capture ELISA using the recombinant Bor56 antigen is a reliable and detailed method for the detection of early O. tsutsugamushi infection.     

Prospective Evaluation of Commercial Antibody-Based Rapid Tests in Combination with a Loop-Mediated Isothermal Amplification PCR Assay for Detection of Orientia tsutsugamushi during the Acute Phase of Scrub Typhus Infection

Samples from 160 prospectively recruited febrile patients with typhus-like illness in an area of Thailand (Chiang Rai, northern Thailand) where scrub typhus is endemic were used to evaluate the diagnostic capabilities of four rapid immunochromatographic tests (ICTs) for the detection of Orientia tsutsugamushi IgM and total antibodies during acute scrub typhus infection. Of the 160 cases, 54 (34%) had been confirmed to have scrub typhus using the reference scrub typhus infection criteria (STIC), i.e., positive cell culture isolation, an admission IgM antibody titer of ≥1:12,800, a 4-fold rising IgM antibody titer, and/or positivity for ≥2 out of 3 PCR gene targets). The ICTs gave the following sensitivities and specificities: the Panbio IgM ICT, 46% (95% confidence interval [CI], 33 to 60) and 95% (95% CI, 89 to 98), respectively; the Standard Diagnostics IgM ICT, 68% (95% CI, 60 to 75) and 73% (95% CI, 68 to 78), respectively; the AccessBio IgM ICT, 56% (95% CI, 48 to 63) and 90% (95% CI, 87 to 94), respectively; and the AccessBio total antibody ABt ICT, 61% (95% CI, 53 to 68) and 68% (95% CI, 63 to 73), respectively. An isothermal loop amplification (LAMP) PCR assay for scrub typhus demonstrated a sensitivity of 52% (95% CI, 38 to 66) and a specificity of 94% (95% CI, 88 to 98). This study has revealed the diagnostic limitations of antibody-based assays in an acute care setting. However, the combination of ICTs with LAMP usually increased sensitivity with a minimal reduction in specificity. The best combination, the Panbio IgM ICT and LAMP, resulted in a sensitivity of 67% (95% CI, 53 to 79) and a specificity of 91% (95% CI, 83 to 95). The combination of antibody-based assays with DNA- or antigen-based tests shows promise for improved diagnostic sensitivity.     

Spectrum of Infections in Acute Febrile Illness in Central India

Introduction: Infectious agent when enters in the host results in febrile illness. This may lead to increase in morbidity or even mortality in undiagnosed/untreated cases. There are many aetiological agents which lead to acute febrile illness. Among these aetiological agents, important is bacterial or viral aetiology. Objective: The objective of this study is: (i) To know the aetiological agents responsible for acute undifferentiated febrile illness (AUFI) by serological test or by bacterial culture and (ii) To know the clinical profile of AUFI. Methodology: A total of 270 patients were enroled in the study with a history of AUFI admitted in medicine and paediatric department from January 2015 to November 2016 of tertiary care hospital of central India. Blood sample was collected for blood culture, clot culture and serological tests for immunochromatographic tests (ICTs) and ICT-positive results were confirmed by respective enzyme-linked immunosorbent assay (ELISA). All negative serum samples by immunochromatography were retested for disease-specific ELISA as scrub typhus, dengue and leptospirosis. Results: Out of 270 patients, 127 (47%) were of scrub typhus, 33 (12%) were malaria cases, 47 (17.40%) were dengue, 12 (4%) were enteric fever, 5 (2%) were leptospirosis, undiagnosed were 18 (6.66%) and other infections (viz viral, urinary tract infection, upper and lower respiratory tract infection and acute gastroenteritis) accounts for 28 (10.37%) cases. We have also noticed that there was co-infection of scrub typhus and dengue, leptospirosis and scrub typhus. Conclusion: It is important to know the cause and clinical profile of AUFIs for their proper management also it will help to prevent morbidity and mortality in AUFI cases.     

Evaluation of an immunoglobulin M capture enzyme-linked immunosorbent assay for diagnosis of Orientia tsutsugamushi infection

To differentiate scrub typhus from other acute febrile diseases, a rapid and reliable serological diagnosis is important. We developed an immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA) for diagnosis of recent Orientia tsutsugamushi infections in humans. The 56-kDa major outer membrane protein of O. tsutsugamushi is well known as the most immunodominant antigen in scrub typhus. The test is based on the use of the biotinylated recombinant 56-kDa protein of O. tsutsugamushi Boryong, Bor56, which was expressed as a fusion protein with a maltose-binding protein in Escherichia coli. In the test, the serum IgM antibodies were captured by anti-human IgM antibodies coated onto a microtiter plate. The captured IgM antibodies were revealed through sequential addition of biotinylated Bor56 antigen and peroxidase-conjugated streptavidin to the plate. The IgM capture ELISA was compared with the immunofluorescence antibody assay (IFA) by testing 176 serum samples from patients with diagnosed cases of rickettsial disease and patients with other acute febrile diseases. Of the 81 IgG IFA-positive samples, 78 tested positive (sensitivity, 96.3%) and all 31 IgM IFA-positive samples tested positive (sensitivity, 100%) by the IgM capture ELISA. The specificity of the IgM capture ELISA was 99%, and 1 of the 95 IFA-negative samples was positive in the assay. These results strongly suggest that IgM capture ELISA using the recombinant Bor56 antigen is a reliable and detailed method for the detection of early O. tsutsugamushi infection.     

Sensitive microplate enzyme-linked immunosorbent assay for detection of antibodies against the scrub typhus rickettsia, Rickettsia tsutsugamushi.

A microtiter enzyme-linked immunosorbent assay (ELISA) has been developed for the titration of antibodies against scrub typhus in human and animal sera. Scrub typhus rickettsiae were grown in monolayers of irradiated mouse LM3 cells and separated from host cell materials by differential centrifugation, filtration through a glass filter (AP-20, Millipore Corp.), and isopycnic banding in Renografin density gradients. The scrub typhus ELISA antigens were obtained from the purified viable rickettsiae by French pressure cell disruption and addition of 0.2% Formalin to the soluble extract. Antisera prepared in rabbits against the prototype Karp, the Kato, and the Gilliam strains of scrub typhus were used to standardize the ELISA and to compare its sensitivity and specificity to that of the indirect fluorescent antibody test (IFA). ELISA titers were measured as the greatest serum dilution showing an optical density 0.25 above controls or by the optical density achieved at a fixed serum dilution. The IFA and ELISA end point titers were quite similar, and all three measures of titer had comparable specificity for the strains of scrub typhus. No cross-reactions between the typhus and scrub typhus wera were observed by ELISA. Both the immunoglobulin M (IgM) and IgG antibody titers of 12 sequential sera from four patients with scrub typhus were obtained by IFA and ELISA. The IFA and ELISA end point titers for IgM and IgG had correlation coefficients of 0.91 and 0.97, respectively, whereas the ELISA optical density values at a serum dilution of 1:100 had slightly lower correlations with IFA titers (0.80 and 0.94). Early rising IgM titers followed by rising IgG titers were demonstrated by ELISA in three patients with primary scrub typhus infections, whereas the IgG response predominated in a patient with a reinfection. It is concluded that the ELISA for scrub typhus is a very satisfactory alternative to the IFA test.     

A rapid, sensitive and reliable diagnostic test for scrub typhus in China

Purpose: To evaluate the performances for detection of IgM and IgG antibodies to Orientia. tsutsugamushi (Ot) using a gold conjugate-based rapid diagnostic test (RDT). Materials and Methods: The RDT employing mixture recombinant 56-kDa proteins of O. tsutsugamushi and the mIFA assay was performed on 33 patients from Fujian and Yunnan province respectively and 94 positive sera (36 from Hainan province and 58 from Jiangsu province) from convalescent stages of the patients with scrub typhus respectively and 82 negative sera from healthy farmers from Anhui province and Beijing City respectively in 2009. A comparison of the RDT and mIFA assay was performed by using the χ² test and the P level of ≤0.05 was considered to be significant. Results: Among these 94 positive sera from convalescent stages of the illness and 82 sera from control farmers, the specificity of RDT was 100% for both IgM and IgG tests. In 33 cases with scrub typhus, 5 cases were positively detected earlier by RDT than by mIFA for the IgM test, and 2 cases were positive for the IgG test. The sensitivities of RDT were 93.9% and 90.9% for IgM and IgG, respectively. Considering IgM and IgG together, the sensitivity was 100%. The geometric mean titre (GMT) of IFA and the RDT assay in diluted sera from confirmed cases were 1:37 versus 1:113 respectively (P<0.001) for IgM test and 1:99 versus 1:279 respectively (P<0.016) for IgG. Conclusions: The RDT was more sensitive than the traditional IFA for the early diagnosis of scrub typhus and was particularly suitable for use in rural areas.     

Evaluation of an in-house PCR for diagnosing scrub typhus along with a preliminary study of genotypic characterization of Orientia tsutshugamushi circulating in Chhattisgarh: A prospective clinico-microbiological study

PurposeScrub typhus, caused by Orientia tsutsugamushi (O. tsutsugamushi) present nonspecific clinical features during manifestation of acute undifferentiated febrile illness (AUFI) to render its early diagnosis difficult. Accordingly, this study was undertaken to assess an in-house groEL PCR versus IgM ELISA for the diagnosis of scrub typhus and to genotypically characterise the randomly selected scrub typhus positive cases.MethodsBlood samples, collected from two hundred twenty one (221) AUFI cases were subjected to groEL PCR and IgM ELISA for diagnosis of scrub typhus. Eleven randomly selected PCR positive cases were processed for DNA sequencing to determine the genetic diversity of O. tsutsugamushi in Chhattisgarh.ResultsScrub typhus prevalence of 35.2% were detected among AUFI cases using both in-house groEL PCR and IgM ELISA. PCR alone showed sensitivity, specificity, positive and negative predictive values of 66.6% (CI: 55.08–76.94), 100% (CI: 90 to 100),100% (CI: 93.15 to 100) and 57.37% (CI: 44.05 to 69.96) while for IgM ELISA, these parameters were 62.8% (CI: 51.13–73.50), 100% (CI: 90 to 100), 100% (CI: 92.75 to 100) and 54.68% (CI: 41.75 to 67.18) respectively. PCR and ELISA could detect scrub typhus in 37.2% and 33.3% cases, when tested alone. groEL PCR detected the O. tsutsugamushi throughout the course of infection. Phylogenetic analysis depicted 5 of 11 positive cases belonged to Kuroki, Japan strain of O. tsutsugamushi, followed by Gilliam and Karp strain in 4 and 2 cases respectively.ConclusionScrub typhus should be considered in differential diagnosis of AUFI. groEL PCR may aid on to IgM ELISA test for optimum laboratory diagnosis of scrub typhus by its implementation especially in seronegative cases. Predominance of Kuroki-like strain followed by Gillian and Karp strains of O. tsutsugamushi in Chhattisgarh confirm variable geographical distribution of O. tsutsugamushi and provide the baseline epidemiological data which will eventually be used to help the researchers for developing better diagnostic tests and vaccine covering the predominant genotypes.     

Comparative Accuracy of the InBios Scrub Typhus Detect IgM Rapid Test for the Detection of IgM Antibodies by Using Conventional Serology

This study investigated the comparative accuracy of a recombinant 56-kDa type-specific antigen-based rapid diagnostic test (RDT) for scrub typhus for the detection of IgM antibodies by using conventional serology in well-characterized serum samples from undifferentiated febrile illness patients. The RDT showed high specificity and promising comparative accuracy, with 82% sensitivity and 98% specificity for samples defined positive at an IgM indirect immunofluorescence assay positivity cutoff titer of ≥1:1,600 versus 92% and 95% at ≥1:6,400, respectively.     

Evaluation of nested PCR and loop mediated isothermal amplification assay (LAMP) targeting 47 ​kDa gene of Orientia tsutsugamushi for diagnosis of scrub typhus

PurposeDiagnostic testing, in particular early detection, is critical for scrub typhus, as most infected individuals have nonspecific symptoms that are easily confused with dengue and malaria. PCR and LAMP offer an alternative DNA amplification method for detection of Orientia tsutsugamushi. Detection of Orientia tsutsugamushi DNA by targeting the 47-kDa gene using nested PCR and LAMP for diagnosis of scrub typhus.MethodsA cross-sectional study in a tertiary care hospital in central India. The present study was done on a total of 274 patients with fever of five days or more and negative for other causes of fever viz. malaria, dengue and enteric fever. From each patient 5 ​ml of blood samples was collected in EDTA vial for molecular tests (PCR and LAMP) and in plain vial for serological tests (IgM IFA).The data was entered in Excel sheet and 2 ​× ​2 tables were created to find sensitivity, specificity, positive and negative likelihood ratios, disease prevalence, positive and negative predictive values and accuracy.ResultsPCR showed a sensitivity of 29.73% while the sensitivity of LAMP was 16.22%. The specificity of nested PCR and LAMP was very high, 99.58% and 99.16% respectively. The diagnostic accuracy of nested PCR (90.15%) was found to be marginally better than LAMP (87.96%).ConclusionsFor the treatment of scrub typhus, a gene-based diagnostic test would enable earlier and more accurate detection of the causative agents of the disease than serology in admission samples of patients with acute febrile illness in endemic areas.     

Scrub typhus and spotted fever among hospitalised children in South India: Clinical profile and serological epidemiology

Background: Rickettsial infections are re-emerging. In India, they are now being reported from several areas where they were previously unknown. Objectives: The objective of this study was to describe the epidemiology, clinical profile and outcome of serologically-confirmed scrub typhus and spotted fever among children in a tertiary care hospital in Bengaluru. Materials and Methods: Hospitalised children aged <18 years, with clinical features suggestive of rickettsial disease admitted between January 2010 and October 2012 were included prospectively. Diagnosis was based on scrub typhus and spotted fever-specific IgM and IgG by enzyme-linked immunosorbent assay (ELISA). Results: Of 103 children with clinical features suggestive of rickettsial illness, ELISA test confirmed 53 cases for scrub typhus, 23 cases for spotted fever group and 14 with mixed infection. The average age was 7.3 (±3.9) years and 44 (71.0%) children were male. Majority of cases were from Karnataka (50%), Andhra Pradesh (32.3%) and Tamil Nadu (17.7%). Common clinical features included fever (100%, average duration 11 days), nausea and vomiting (44%), rash (36%); eschar was rare. Compared to the ELISA test, Weil-Felix test (OX-K titre of 1:80) had a sensitivity and specificity of 88.7% and 43.9%, respectively. Treatment with chloramphenicol or doxycycline was given to the majority of the children. Complications included meningoencephalitis (28%), shock (10%), retinal vasculitis (10%) and purpura fulminans (7%). Conclusions: These findings suggest that the burden of rickettsial infection among children in India is high, with a substantially high complication rate. Rickettsial-specific ELISA tests can help in early diagnosis and early institution of appropriate treatment that may prevent life-threatening complications.     

Seroprevalence of Scrub typhus at a tertiary care hospital in Andhra Pradesh

Introduction: Scrub typhus is a rickettsial infection which is caused by Orientia tsutsugamushi and transmitted by the bite of the chigger of a mite. Delay in diagnosis can be fatal otherwise the treatment is simple, doxycycline being the drug of choice. Indirect immunoflurescence is considered gold standard but it is not used in India as it is costly and also not available. There is need for rapid, economic and simple test for the diagnosis of scrub typhus. This study was taken up to study the seroprevalence of scrub typhus in Andhra Pradesh and to compare two commonly used serological methods; rapid test and IgM ELISA. Materials and methods: This is a prospective study in which 100 serum samples from clinically suspected cases collected over a period of 3 months were processed for the detection of IgM antibodies for scrub typhus by ELISA and Rapid test. Samples were also tested for leptospirosis and dengue fever which the other common causes of fever prevalent in this region. Results: Total number of samples processed was 100 of which 52 were males and 48 females. Among the hundred samples 39 were seropositive. Positivity was higher in the age group of patients between 16 and 30 yrs of age. There was 97% correlation between ELISA and rapid method. Of the 100 samples only three samples positive by ELISA were negative by rapid method. Fever was the most common manifestation and there was no eschar and no mortality reported. Conclusion: Scrub typhus should be included in the differential diagnosis of fever of unknown origin along with dengue, malaria and leptospirosis which are the other common endemic infections in this part of the country.     

Possibility of scrub typhus in fever of unknown origin (FUO) cases: An experience from Rajasthan

Purpose: Fever of unknown origin (FUO) has multiple causes. Scrub typhus is less known cause of FUO in India. The present study reports a recent epidemic of scrub typhus amongst cases of FUO from different areas of Rajasthan, India. There was high mortality in undiagnosed cases of FUO which lead to the diagnosis of scrub typhus. Objective: To study the possibility of scrub typhus as a causative factor in FUO cases by qualitative detection of IgM antibodies with ELISA. Materials and Methods: From September 2012 to December 2012, 271 serum samples of FUO cases were analysed for IgM antibodies to Orientia tsutsugamushi along with dengue, malaria, typhoid, tuberculosis and brucellosis. Results: Scrub typhus IgM antibodies by ELISA were detected in 133 (49.1%) patients. Scrub typhus positivity was significantly higher among female in comparison to males (P < 0.05). Maximum positivity of scrub typhus was found in females of 46-60 years age group. The laboratory parameters were abnormal in most of the patients as evident by thrombocytopenia (63%), deranged liver functions (56%) and renal functions (25%). Conclusion: The present study emphasises the importance of scrub typhus among cases of FUO especially after rainy season and during early cooler months. The study also highlights the significance of ELISA method for rapid and early reporting and ruling out scrub typhus in FUO cases.     

Co-infection of scrub typhus and leptospirosis in patients with pyrexia of unknown origin in Longding district of Arunachal Pradesh in 2013

Background: Scrub typhus and leptospirosis are bacterial zoonotic disease causing high morbidity and mortality. The seasonal outbreak of pyrexia is common in Arunachal Pradesh (AP); many times the disease remains undiagnosed. Objective: An outbreak of pyrexia of unknown origin (PUO) occurred in Longding district of Arunachal Pradesh in 2013, with 108 deaths, which was investigated to elucidate the cause of illness. Methodology: Blood samples from the affected region with acute pyrexia were collected, and screened for the malaria parasite, scrub typhus IgM and leptospira IgM. Results: Scrub typhus IgM was reactive in 97% (30/31), and 25% (8/31) cases were co-infected with leptospira. Incidentally, scrub typhus reactive (67%) and leptospira co-infection (62.7%) were higher in females. Record of previous 3 years (2011–2013) from Longding, Community Health Centre showed an increase in indoor pyrexia cases by 2-fold or more during October and November. Conclusion: The present study is the first report of co-infection of scrub typhus with leptospirosis from Northeast India. Medical officers in this region should take scrub typhus and leptospirosis in their differential diagnosis of patients with PUO for early diagnosis and effective treatment.     

Comparison of ELISA and indirect immunofluorescent antibody assay detecting <Emphasis Type="Italic">Coxiella burnetii</Emphasis> IgM phase II for the diagnosis of acute Q fever

A commercially available enzyme-linked immunosorbent assay (ELISA) detecting Coxiella burnetii phase II-specific IgM for the diagnosis of acute Q fever was compared with indirect immunofluorescent antibody assay (IFA). IFA is the current reference method for the detection of antibodies against C. burnetii, but has disadvantages because the judgment of fluorescence is subjective and tiring, and the test is expensive and automation is not possible. To examine whether phase II IgM ELISA could be used as a screening assay for acute Q fever, we compared the sensitivity and specificity of IFA and ELISA. The sensitivity of the IFA and ELISA tests were 100 and 85.7%, respectively, with a specificity of 95.3 and 97.6%, respectively. Because of the high sensitivity and specificity of the ELISA in combination with the practical disadvantages of the IFA, we introduced a new algorithm to screen samples of patients with symptoms of acute Q fever infection.     

A serosurvey of Orientia tsutsugamushi from patients with scrub typhus

Many countries where scrub typhus is endemic use their own cutoff values for antibody titres to differentiate between cured cases and current infections. To establish an antibody titre cutoff value, one needs to investigate the seroprevalence in endemic areas, and the duration of the increase in titre after complete cure. We conducted a follow-up study of anti-Orientia tsutsugamushi antibody titres using indirect immunofluorescence assays (IFA) and passive haemagglutination assays (PHA) in patients with scrub typhus. After the onset of symptoms, IgM antibody titres increased gradually over 2–3 weeks, peaked at about 4 weeks, and started to decrease rapidly between 4 and 5 weeks. At 1-year follow-up, the median IgM value was 1:10. Out of 77 patients who were tested at that time, 36 (47%) had IgM titres ≥1:20, and none had titres exceeding 1:80. Over the first 2 weeks, IgG antibody titres increased sharply, peaked at about 4 weeks and decreased rather gradually thereafter, with a median titre of 1:128 maintained up to the 18th month. At 1-year follow-up, five out of 77 patients (6.5%) had titres ≥1:1,024 and 57% had titres ≥1:128. Based on these results, a cutoff value of ≥1:160 for IgM antibody should differentiate between previous and current infections in endemic areas such as Korea and Japan, where scrub typhus occurs mainly in the autumn.     

Recent outbreak of scrub typhus in North Western part of India

Background: Scrub typhus usually affects previously healthy active persons and if undiagnosed or diagnosed late, may prove to be life-threatening. Diagnosis of scrub typhus should be largely based on a high index of suspicion and careful clinical, laboratory and epidemiological evaluation. Objective: To describe the diverse clinical and laboratory manifestations of scrub typhus diagnosed in Mahatma Gandhi Medical College and Hospital, Jaipur. Materials and Methods: All cases of febrile illness diagnosed as scrub typhus over a period of 3 months were analysed. Diagnosis was based on ELISA test for antibody detection against 56 kDa antigen. Results: Forty-two cases of scrub typhus were seen over a period of 3 months (October, 2012-December, 2012). Common symptoms were high grade fever of 4-30 days duration, cough, haemoptysis and breathlessness. Eschar was not seen even in a single patient. Liver enzymes were elevated in nearly all cases (95.9%). Multiple organ dysfunction syndrome (MODS) was present in 16.66% of our patients (7 out of 42). Hypotension (6 patients, 14.2%), renal impairment (9 out of 15 patients, 60%), acute respiratory distress syndrome (4 patients, 9.52%) and meningitis (4 patients, 9.52%) were some of the important complications. There was a dramatic response to doxycycline in nearly all the patients, but initially when the disease was not diagnosed, seven patients had died. Conclusion: Scrub typhus has emerged as an important cause of febrile illness in Jaipur. Empirical treatment with doxycycline is justified in endemic areas.