Diagnosis Of Leptospirosis Research Articles

Detection of anti-leptospiral antibodies using recombinant ErpY-like lipoprotein based latex agglutination test for serodiagnosis of animal leptospirosis.

Precise and timely diagnosis is essential to prevent severe outcomes of leptospirosis in humans and animals. Existing diagnostic methods face challenges and limitations, underscoring the need for novel, field-applicable screening, and diagnostic tests/assays. This study evaluates the diagnostic utility of a recombinant ErpY-like lipoprotein (rErpY-LIC11966) in a latex agglutination test (LAT) for diagnosis of animal leptospirosis. The ErpY gene sequence from Leptospira interrogans serovar Pomona, excluding the signal peptide, was amplified, cloned into the pETite vector, and expressed in Escherichia coli. The expressed rErpY (∼16kDa) was characterized by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis and Western blot using Leptospira-specific standard sera. To assess the diagnostic potential of rErpY, Ni-NTA affinity-purified protein was used to sensitize latex-coated beads (0.8µm colour beads), which were then employed in the LAT for standardization and optimization with standard positive and negative sera. For evaluation, the rErpY-LAT was tested on serum samples from 177 suspected animal cases and compared to the microscopic agglutination test. It showed a relative diagnostic sensitivity of 90.6%, a specificity of 89.1%, and an overall accuracy of 90%. This study proposes rErpY-LAT as a field testing/screening diagnostic tool for preliminary serodiagnosis of leptospirosis, highlighting the potential of recombinant protein-based assays to address current diagnostic challenges.

The Potential of Eight Plasma Proteins as Biomarkers in Redefining Leptospirosis Diagnosis.

Leptospirosis, a notifiable endemic disease in Malaysia, has higher mortality rates than regional dengue fever. Diverse clinical symptoms and limited diagnostic methods complicate leptospirosis diagnosis. The demand for accurate biomarker-based diagnostics is increasing. This study investigated the plasma proteome of leptospirosis patients with leptospiraemia and seroconversion compared with dengue patients and healthy subjects using isobaric tags for relative and absolute quantitation (iTRAQ)-mass spectrometry (MS). The iTRAQ analysis identified a total of 450 proteins, which were refined to a list of 290 proteins through a series of exclusion criteria. Differential expression in the plasma proteome of leptospirosis patients compared to the control groups identified 11 proteins, which are apolipoprotein A-II (APOA2), C-reactive protein (CRP), fermitin family homolog 3 (FERMT3), leucine-rich alpha-2-glycoprotein 1 (LRG1), lipopolysaccharide-binding protein (LBP), myosin-9 (MYH9), platelet basic protein (PPBP), platelet factor 4 (PF4), profilin-1 (PFN1), serum amyloid A-1 protein (SAA1), and thrombospondin-1 (THBS1). Following a study on a verification cohort, a panel of eight plasma protein biomarkers was identified for potential leptospirosis diagnosis: CRP, LRG1, LBP, MYH9, PPBP, PF4, SAA1, and THBS1. In conclusion, a panel of eight protein biomarkers offers a promising approach for leptospirosis diagnosis, addressing the limitations of the "one disease, one biomarker" concept.

Accuracy of the Dual Path Platform (DPP) rapid test for the diagnosis of leptospirosis: A multi-center study in six Brazilian states

Leptospirosis is a zoonotic disease with significant global impact and a challenging diagnosis. The utilization of adequately validated rapid tests is relevant for the opportune identification of the disease and for reduction in fatality rates. The present study analyzes the accuracy and reliability of the Dual Path Platform (DPP) assay -produced in Brazil by the Oswaldo Cruz Foundation (Fiocruz)- for diagnosing leptospirosis. Firstly, a serological panel was constructed in the Brazilian Reference Laboratory for Leptospirosis using samples routinely handled by reference laboratories of six Brazilian states. It consisted of 150 positive (according to MAT and IgM-ELISA) and 250 negative samples for leptospirosis. Subsequently, the panel samples were distributed to the reference laboratories for the performance of DPP assays in triplicate. Different measures were used in the assessment of diagnostic quality. Predictive values were estimated for different pre-test probability settings. Sensitivities varied between 67.33 % and 74.00 % and specificities between 93.20 % and 98.40 % in the states, and there were adequate agreements between them. Accuracies were lower for the samples of patients with less than 7 days of symptoms. In contexts of prevalence values up to around 25 %, positive and negative predictive values were around 90 %. However, in situations of high pre-test probabilities, NPVs were low. This study improves understanding of the use of DPP in diagnosing leptospirosis, particularly its application in healthcare settings. As long as the time of symptoms onset and clinical and epidemiological contexts are adequately considered for the interpretation of results, DPP is a valid option to be used in the leptospirosis diagnostic routine.

Development of a new accurate lateral flow immunoassay for diagnosis of human leptospirosis.

The current diagnostic methods for leptospirosis diagnosis are technically complex and expensive, with limited applicability to specialized laboratories. Furthermore, they lack diagnostic accuracy in the acute stage of the disease, which coincides with a period when antibiotics are highly effective. New simple and accurate tests are mandatory to decentralize and improve diagnosis. Here, we introduced a new lateral flow immunoassay (Lepto-LF) for human leptospirosis. We conducted a double-blinded assay using 104 serum samples from patients with confirmed or discarded diagnosis for leptospirosis. The diagnostic performance of Lepto-LF was estimated across different ranges of days from onset of symptoms (dpo), considering the diagnostic algorithm as reference standard. Additionally, it was compared with the screening methods enzyme-linked immunosorbent assay (IgM-ELISA) and the slide agglutination test using temperature-resistant antigen (SATR). Lepto-LF exhibited perfect diagnostic performance with a Youden´s index J = 1 from 6 dpo in the acute phase. IgM-ELISA gave slightly lower accuracy with J = 0.91 and 95.5% of both sensitivity and specificity; while SATR showed a markedly inferior yield (J = 0.41, sensitivity = 95.5%, specificity = 45.5%). The performances remained consistent in the convalescence phase of the disease (> 10 dpo). Lepto-LF was found to be a reliable test for simple, rapid and early diagnosis of leptospirosis, resulting a promising tool for decentralizing leptospirosis diagnosis and enabling timely treatment of patients. In addition, Lepto-LF may be employed as confirmatory test, especially in remote areas and vulnerable contexts where the standard MAT is not available.

Development and Evaluation of a Noninvasive Microfluidic-Based Paper Analytical Device for Leptospirosis Diagnosis.

Leptospirosis is a re-emerging infectious disease that presents a diagnostic enigma for clinicians with frequent misdiagnosis due to lack of rapid and accurate diagnostic tests, as the current methods are encumbered by inherent limitations. The development of a diagnostic sensor with a sample-in-result-out capability is pivotal for prompt diagnosis. Herein, we developed a microfluidic paper-based analytical device (spin-μPAD) featuring a sample-in-result-out fashion for the detection of Leptospira specific urinary biomarker, sph2 sphingomyelinase, crucial for noninvasive point-of-care testing. Fabrication of paper devices involved precise photolithography techniques, ensuring a high degree of reproducibility and replicability. By optimizing the device's configuration and protein components, a remarkable sensitivity and specificity was achieved for detecting leptospiral sph2 in urine, even at low concentrations down to 1.5 fg/mL, with an assay time of 15 min. Further, the spin-μPAD was validated with 20 clinical samples, suspected of leptospirosis including other febrile illnesses, and compared with gold standard microscopic agglutination test, culture, Lepto IgM ELISA, darkfield microscopy, and Leptocheck WB spot test. In contrast to commercial diagnostic tools, the spin-μPAD was noninvasive, rapid, easy to use, specific, sensitive, and cost-effective. The results highlight the potential of this innovative spin-μPAD for an efficient and dependable approach to noninvasive leptospirosis diagnosis, addressing critical needs in the realms of public health and clinical settings.

Acute ST elevation myocardial infarction (STEMI) in a patient with leptospirosis; a therapeutical dilemma – A case report

A 45-year-old previously healthy farmer was seen at the Emergency Department with ischemic type chest pain. He had a history of an acute febrile illness for four days. The ECG showed an acute anterior myocardial infarction. His laboratory investigations revealed thrombocytopenia, a raised serum creatinine level and significantly raised high-sensitive cardiac Troponin I. A provisional diagnosis of acute STEMI with co-existing leptospirosis was made. The patient was managed conservatively with antibiotics, statins, and aspirin. Neither thrombolysis nor primary coronary intervention was attempted due to high risk of bleeding. Diagnosis of leptospirosis was confirmed later by markedly elevated leptospira antibodies. After recovery, he underwent coronary angiogram which revealed a totally occluded left anterior descending artery and is awaiting myocardial viability assessment with dobutamine stress echocardiography before PCI. We report this uncommon presentation of acute myocardial infarction in a patient with leptospirosis, as it causes a considerable therapeutic dilemma especially in a resource-poor setting.

Study on the interference of vaccine antibodies with the serological diagnosis of leptospirosis in cattle

A simple IgG-specific ELISA for Leptospira spp. was compared with the microscopic agglutination test (MAT) to detect IgG antibody responses to a commercial vaccine in cattle. We used an enzyme-linked immunosorbent assay (ELISA) with sonicated Leptospira interrogans serovar copenhageni M 20. After initial vaccination, specific antibodies against Leptospira spp. were detected in 90 % of the animals by IgG-ELISA and 60 % by MAT, while after booster, antibodies were detected in 100 % and 80 % of the animals by IgG-ELISA and MAT, respectively. Both serological MAT and ELISA tests revealed interferences of vaccine antibodies. Disease diagnosis with ELISA and MAT methods should be made two and a half months and four months, respectively, after vaccination to avoid interference of vaccine antibodies. On the other hand, our results suggest that IgG-ELISA may be a useful method to assess the development of IgG antibodies induced by Leptospira vaccine.

Advancing serologic diagnosis: assessing the efficacy of rErpY-like protein in human leptospirosis detection.

Leptospirosis is a globally distributed infectious disease caused by pathogenic spirochetes of the Leptospira genus, often overlooked. It is estimated that the disease affects approximately one million people annually, resulting in more than 58,900 deaths. The gold standard for serodiagnosis of leptospirosis is the Microscopic Agglutination Test (MAT). However, the limitations of this technique necessitate the exploration of alternative diagnostic methods. In this study, we evaluated the ErpY-like recombinant protein (rErpY-like) in the development of a serologic diagnostic assay for human leptospirosis. Eighty-six human sera samples, characterized by MAT, underwent evaluation through indirect IgM-ELISA and IgG-ELISA. The sensitivity and specificity values obtained from IgM-ELISA were 60% and 76%, respectively, while those from IgG-ELISA were 96.4% and 100%, respectively. The use of the rErpY-like protein in both IgM-ELISA and IgG-ELISA proves to be a sensitive and specific method for antibody detection. This could potentially serve as a valuable alternative tool in the diagnosis of human leptospirosis.

Accuracy of rapid lateral flow immunoassays for human leptospirosis diagnosis: A systematic review and meta-analysis.

In the last two decades, several rapid lateral flow immunoassays (LFIs) for the diagnosis of human leptospirosis were developed and commercialized. However, the accuracy and reliability of these LFIs are not well understood. In this study, we aimed to evaluate the accuracy of leptospirosis LFIs as well as the factors affecting the test efficiency using systematic review and meta-analysis. Original articles reporting the accuracy of human leptospirosis LFIs against microagglutination tests (MAT) or immunofluorescent assays (IFA) were searched from PubMed, Embase, and Scopus, and selected as per pre-set inclusion and exclusion criteria. A total of 49 data entries extracted from 24 eligible records published between 2003 and 2023 were included for meta-analysis. A meta-analysis was performed using STATA. The quality of the included studies was assessed according to the revised QUADAS-2. Only nine studies (32.1%) were considered to have a low risk of bias and no concern for applicability. Pooled sensitivity and specificity were calculated to be 68% (95% confidence interval, CI: 57-78) and 93% (95% CI: 90-95), respectively. However, the ranges of sensitivity (3.6 - 100%) and specificity (53.5 - 100%) of individual entries are dramatically broad, possibly due to the heterogeneity found in both study designs and LFIs themselves. Subgroup analysis demonstrated that IgM detection has better sensitivity than detection of IgG alone. Moreover, the test performance seems to be unaffected by samples from different phases of infection. The pooled specificity of LFIs observed is somewhat acceptable, but the pooled sensitivity is low. These results, however, must be interpreted with caution because of substantial heterogeneity. Further evaluations of the LFIs with well-standardized design and reference test will be needed for a greater understanding of the test performance. Additionally, IgM detection type should be employed when leptospirosis LFIs are developed in the future.

Leptospirosis in a patient with cardiac manifestation: A case report study and literature review

Key Clinical MessageCommon cardiac arrhythmias seen in patients with leptospirosis are usually atrial fibrillation or first‐degree atrioventricular block, with bradyarrhythmia being rare in this group. It is essential to prioritize the examination of the patient's medical background, clinical symptoms, and comprehensive physical evaluation in order to promptly identify and address the patient's condition.AbstractLeptospirosis, a zoonotic disease that is widespread worldwide, has a significant impact on tropical areas and can affect various organs throughout the infection. During the initial stage, symptoms are typically non‐specific. Although cases of all three cardiac layers being affected have been reported, issues with the conduction system are especially significant in the early phase of the disease. The most frequent discoveries in these patients are atrial fibrillation or first‐degree atrioventricular block, with bradyarrhythmia being rare. We describe a 37‐year‐old male farmer who initially sought medical attention for general symptoms that had been deteriorating despite receiving outpatient treatment for 3 days for a presumed diagnosis of influenza. During his initial assessment, he exhibited sinus bradycardia, anemia, leukocytosis, elevated levels of direct and total bilirubin, and abnormal liver function test results. Through thorough history‐taking, physical examination, and laboratory analyses, a diagnosis of leptospirosis was conclusively established for him. Focusing on the patient's medical history, clinical manifestations, and thorough physical assessment is crucial for promptly diagnosing and treating patients. This becomes particularly significant for individuals who exhibit atypical symptoms, exemplified by our patient presenting with nonspecific indications and cardiac issues manifested as bradycardia.

Severe leptospirosis in tropical and non-tropical areas: A comparison of two french, multicentre, retrospective cohorts.

Leptospirosis is an anthropozoonosis that occurs worldwide but is more common in tropical regions. Severe forms may require intensive care unit (ICU) admission. Whether the clinical patterns and outcomes differ between tropical and non-tropical regions with similar healthcare systems is unclear. Our objective here was to address this issue by comparing two cohorts of ICU patients with leptospirosis managed in mainland France and in the overseas French department of Réunion, respectively. We compared two retrospective cohorts of patients admitted to intensive care for severe leptospirosis, one from Reunion Island in the Indian Ocean (tropical climate) and the other from metropolitan France (temperate climate). Chi-square and Student's t tests were used for comparisons. After grouping the two cohorts, we also performed multiple correspondence analysis and hierarchical clustering to search for distinct clinical phenotypes. The Réunion and Metropolitan France cohorts comprised 128 and 160 patients respectively. Compared with the Réunion cohort, the metropolitan cohort had a higher mean age (42.5±14.1 vs. 51.4±16.5 years, p<0.001). Severity scores, length of stay and mortality did not differ between the two cohorts. Three phenotypes were identified: hepato-renal leptospirosis (54.5%) characterized by significant hepatic, renal and coagulation failure, with a mortality of 8.3%; moderately severe leptospirosis (38.5%) with less severe organ failure and the lowest mortality rate (1.8%); and very severe leptospirosis (7%) manifested by neurological, respiratory and cardiovascular failure, with a mortality of 30%. The outcomes of severe leptospirosis requiring ICU admission did not differ between tropical and temperate regions with similar healthcare access, practices, and resources, despite some differences in patient characteristics. The identification of three different clinical phenotypes may assist in the early diagnosis and management of severe leptospirosis.

DEVELOPMENT AND VALIDATION OF LOOP-MEDIATED ISOTHERMAL AMPLIFICATION (LAMP) FOR DETECTION OF LEPTOSPIR GENETIC MATERIAL

Primer systems were designed and tested and the method of loop-mediated isothermal amplification was validated for rapid diagnosis of leptospirosis. In general, five sets of primers were calculated and analyzed in situ and in vitro, which were ordered for synthesis and chosen for the future as the most promising. Indicators of analytical sensitivity and detection limits of the method were established, the specificity of the method in its tests with homologous and close to taxonomic characteristics heterologous samples of genetic material was proved and completed reproducibility of the method was established under the conditions of its staging on various laboratory equipment and with reagents from different manufacturers. The developed technique of loop-mediated amplification for the diagnosis of leptospirosis was tested on field samples and its diagnostic effectiveness was established.

Production and characterization of immunoglobulin G anti-rLipL32 antibody as a biomarker for the diagnosis of leptospirosis.

Microscopic agglutination test (MAT) for the diagnosis of leptospirosis requires live cultures and is serovar-specific, while polymerase chain reaction (PCR) requires expensive equipment and sample preparation. The rLipL32 protein is conserved and can be used for the production of immunoglobulin G (IgG) anti-rLipL32 antibody, which can be used as a biomarker for leptospirosis diagnosis. This study aimed to produce and characterize an IgG anti-rLipL32 antibody as a biomarker for leptospirosis diagnosis. Escherichia coli rLipL32 was cultured and analyzed by PCR and sequencing. Cultures were used for rLipL32 protein expression and purification and the rLipL32 protein was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The rLipL32 protein was used to produce anti-rLipL32 serum and was analyzed by enzyme-linked immunosorbent assay (ELISA). Serum was purified to obtain IgG anti-rLipL32 antibody and characterized by SDS-PAGE and western blotting. PCR was able to amplify the LipL32 gene from E. coli rLipL32, and sequencing analysis showed 99.19% similarity with pathogenic Leptospira. SDS-PAGE analysis showed a 32-kDa band. ELISA results showed an increase in OD in anti-rLipL32 serum compared to preimmune serum. Western blotting results showed that the IgG anti-rLipL32 antibody was able to bind and cross-reacts with pathogenic Leptospira serovar but not with E. coli or Staphylococcus aureus. IgG anti-rLipL32 antibody has high specificity and sensitivity against Leptospira pathogens. These findings suggest that IgG anti-rLipL32 antibody is a promising biomarker for the diagnosis of leptospirosis.

Seroprevalence and Risk Factors Associated with Leptospirosis in High-Risk Occupational Groups in the State of Gujarat as Determined by IgM ELISA and MAT Test: A Cross-Sectional Study.

Leptospirosis is a reemerging zoonosis affecting humans and animals worldwide in tropical and subtropical countries. The study was conducted to estimate the seroprevalence of leptospirosis in high-risk occupational groups (cattle farm workers, poultry farm workers, and slaughterhouse workers) in the state of Gujarat and to determine related risk factors for leptospirosis. Using an interview-guided questionnaire, a cross-sectional study was conducted involving 123 people in high-risk occupational groups (namely, cattle farm workers, poultry farm workers, and slaughterhouse workers) from five districts of the state of Gujarat, India. The participants' serum samples were screened for antileptospiral antibodies by using Immunoglobulin G (IgG) and Immunoglobulin M (IgM) enzyme-linked immunosorbent assays (ELISAs) followed by a microscopic agglutination test (MAT). The Chi-square (χ2) test and odds ratio were calculated using IBM SPSS Statistics for Windows, version 22 (IBM Corp., Armonk, N.Y., USA) to ascertain the associated risk factors. Regarding demographic information, this study included 104 men and 19 women in high-risk occupational groups. The overall seroprevalence of antileptospiral antibodies in these high-risk workers was 46.3%. The seroprevalence was the highest among poultry farm workers (56.6%), followed by cattle farm workers (54.5%) and slaughterhouse workers (25.6%). The MAT in combination with IgM is adequate for the serological diagnosis of leptospirosis, but we have also employed IgG ELISA to understand the chronic infection in these high-risk occupational groups as exposure was both past and present. Tests were conducted for antibodies against various Leptospira serovars in the major serogroups; frequent serovars included Hurstbridge, Panama, Javanica, Tarassovi, Copenhageni, Pomona, and Weaveri, among others. The significant (P < 0.05) risk factors associated with leptospirosis in these high-risk occupations reported to be living in field areas, the presence of livestock in the neighborhood, working in slaughterhouses, drinking water from natural water resources, contact with animals such as dogs or livestock, and wounds during occupational practice. Considering the high seroprevalence of leptospirosis, high-risk occupational groups should be regularly screened for potential development of the fatal disease. Hurstbridge was found to be the most prevalent serovar in the studied population, followed by Panama, Javanica, and Tarassovi.

Detection of pathogenic Leptospira with rapid extraction followed by recombinase polymerase amplification (RPA) and quantitative polymerase chain reaction (qPCR) assay-A comprehensive study from Sri Lanka.

Leptospirosis is the most widespread zoonosis in the world. The disease is more prevalent in tropical regions where the majority of developing countries are located. Leptospirosis is considered a protean manifestation zoonosis with severity of the disease ranging from a mild febrile illness to a severe and life-threatening illness. Clinical symptoms of leptospirosis overlap with other tropical febrile illnesses. Early, rapid, and definitive diagnosis is important for effective patient management. Since Polymerase Chain Reaction (PCR)-based assays are not readily available in most clinical settings, there is a need for an affordable, simple, and rapid diagnostic test. Quantitative PCR (qPCR) and Recombinase Polymerase Amplification (RPA) were implemented at the Faculty of Medicine, University of Kelaniya, and a prospective study to evaluate RPA for diagnosis of acute phase of leptospirosis was conducted. Results indicate that RPA and qPCR were positive in 81% (98/121) of the total positive and acute clinical samples. Of the 81 positive MAT confirmed patients 60 (74%) and 53 (65%) were positive with qPCR and RPA respectively. Retrospective evaluation revealed a high diagnostic accuracy (sensitivity-70% and specificity-87%) of RPA compared to MAT as the reference gold standard. Results further suggest that there is no significant difference between the two assays, qPCR and RPA-SwiftX (P = 0.40). Laboratory procedures for the extraction and detection by qPCR in the laboratory have been optimized to obtain results within 6 hours. However, the RPA-SwiftX method under field conditions took 35 minutes. The RPA-SwiftX method could replace the qPCR which shows similar sensitivity and specificity. Therefore, RPA established under the current study presents a powerful tool for the early and rapid diagnosis of leptospirosis at point-of-care.

Improved Antibody Detection for Canine Leptospirosis: ELISAs Modified Using Local Leptospiral Serovar Isolates from Asymptomatic Dogs.

Leptospirosis is a zoonotic disease of significant concern for human and animal health, with domestic animals, including dogs, acting as reservoirs for human infection. Serology is widely used for leptospirosis diagnosis, even though the standard microscopic agglutination test (MAT) using a panel of serovars lacks specificity and can lead to detection limitations in certain regions. In this study, we aimed to develop an antibody detection tool for dogs using an indirect enzyme-linked immunosorbent assay (ELISA) with a set of local serovar isolates, including Paidjan, Dadas, and Mini, to enhance the accuracy of leptospirosis surveillance in our region. The specificity and sensitivity of various antigen preparations, namely leptospiral whole-cell protein (WCP), total membrane protein (TMP), and outer membrane protein (OMP), were assessed using sera from infected and non-infected dogs, as well as negative puppy sera. Leptospirosis diagnosis was supported using a genus-specific nested polymerase chain reaction test on all collected sera. Protein preparations were validated using SDS-PAGE and Western blotting analysis. In the results, the standard MAT failed to detect antibodies in any of the dogs confirmed as being infected using PCR and isolation, highlighting its limitations. In contrast, the OMP-based ELISAs using local isolates of Leptospira serovars gave positive results with sera from all infected dogs, and negative results with sera from all dogs from non-endemic areas. IgG titres of infected and unvaccinated dogs from endemically affected areas were significantly higher than those in non-endemic regions. Using the OMP-based IgG/ELISAs with the local serovar Dadas resulted in higher specificity and lower sensitivity than when using the WCP- and TMP-based IgG/ELISAs. Agreement analysis revealed fair and moderate concordance between OMP-based IgG/ELISAs and PCR results, whereas slight and fair agreement was observed between OMP-based ELISAs and the MAT. Overall, the modified OMP-based IgG/ELISAs, utilising relevant local serovar isolates from dogs, demonstrated improved accuracy in detecting leptospirosis in the study area, overcoming the limitations of the MAT. This study highlights the importance of identifying and incorporating these local circulating serovar isolates into serological techniques for leptospirosis diagnosis and surveillance.

Relevance of Polymerase Chain Reaction in Early Diagnosis of Leptospirosis.

Leptospirosis is common in India, especially in the southern states. Mortality is high among untreated cases. Diagnosis of leptospirosis remains a challenge in India as polymerase chain reaction (PCR), which is more sensitive than Immunoglobulin M (IgM) is not widely available. This study aimed to find out the difference in diagnostic yield with PCR and IgM in early leptospirosis. This retrospective, single-center study included 67 adults with laboratory-confirmed leptospirosis (IgM, PCR, or both) who presented within 7 days of symptom onset and were admitted to the intensive care unit (ICU). The difference in the diagnostic yield with PCR and IgM ELISA was studied. About 77.6% of the patients tested positive by PCR and 55.2% tested positive by IgM. There was a statistically significant difference in the detection of leptospirosis by PCR and IgM (p-value = 0.036). In the subgroup of patients who presented within 3 days of onset of symptoms, PCR positivity was 90.32% whereas IgM positivity was only 25.8%. Our study showed that the sensitivity of leptospira PCR is significantly higher than IgM in the first week of illness. It also showed that among the subset of patients who died, a majority were detected only by PCR. Since PCR is not widely available, leptospirosis remains underdiagnosed and mortality from the same is underestimated. Polymerase chain reaction, if routinely done along with IgM for all suspected cases of leptospirosis that present within the first week of illness helps in prompt diagnosis and treatment. Sreevalsan TV, Chandra R. Relevance of Polymerase Chain Reaction in Early Diagnosis of Leptospirosis. Indian J Crit Care Med 2024;28(3):290-293.

Fulminant Leptospirosis Presenting with Rapidly Developing Acute Renal Failure and Multiorgan Failure.

Leptospirosis, caused by pathogenic spirochetes of the Leptospira genus, is a common zoonosis in tropical and subtropical regions and can lead to an epidemic following heavy rainfall or flooding. The primary reservoirs of Leptospira include rodents, wild animals, dogs, cats, amphibians, and others, but the brown rat (Rattus norvegicus) remains the main source of human Leptospirosis. Humans are often accidental hosts and they can be infected through cuts, abrasions, mucosa, conjunctiva, or by ingesting contaminated water. The clinical manifestation of leptospirosis can vary from mild, nonspecific symptoms to a fatal outcome involving liver and renal failure, pulmonary hemorrhage, meningitis, and septic shock. The severity of fatal outcomes is likely to be due to virulence factors, host susceptibility, and epidemiological conditions. L. interrogans are associated with high-risk individuals, particularly patients older than 60 years of age in clinical settings. The current case study showed a foreign worker who presented with rapidly deteriorating clinical signs of fever, jaundice, impaired consciousness, and oliguric acute renal failure. Drawing from our experience, it is advisable to consider the possibility of leptospirosis diagnosis in patients who show clinical symptoms such as fever, hepatic failure with jaundice, and acute renal failure. This is particularly important for those individuals with a prior history of pathogen exposure. This case study had a strong suspicion of leptospirosis, which was confirmed by the microscopic agglutination test (MAT) and, later, the patient's recovery following treatment.

Diagnosis of human leptospirosis: systematic review and meta-analysis of the diagnostic accuracy of the Leptospira microscopic agglutination test, PCR targeting Lfb1, and IgM ELISA to Leptospira fainei serovar Hurstbridge

BackgroundLeptospirosis is an underdiagnosed infectious disease with non-specific clinical presentation that requires laboratory confirmation for diagnosis. The serologic reference standard remains the microscopic agglutination test (MAT) on paired serum samples. However, reported estimates of MAT’s sensitivity vary. We evaluated the accuracy of four index tests, MAT on paired samples as well as alternative standards for leptospirosis diagnosis: MAT on single acute-phase samples, polymerase chain reaction (PCR) with the target gene Lfb1, and ELISA IgM with Leptospira fainei serovar Hurstbridge as an antigen.MethodsWe performed a systematic review of studies reporting results of leptospirosis diagnostic tests. We searched eight electronic databases and selected studies that tested human blood samples and compared index tests with blood culture and/or PCR and/or MAT (comparator tests). For MAT selection criteria we defined a threshold for single acute-phase samples according to a national classification of leptospirosis endemicity. We used a Bayesian random-effect meta-analysis to estimate the sensitivity and specificity of MAT in single acute-phase and paired samples separately, and assessed risk of bias using the Quality Assessment of Studies of Diagnostic Accuracy Approach- 2 (QUADAS-2) tool.ResultsFor the MAT accuracy evaluation, 15 studies were included, 11 with single acute-phase serum, and 12 with paired sera. Two included studies used PCR targeting the Lfb1 gene, and one included study used IgM ELISA with Leptospira fainei serovar Hurstbridge as antigen. For MAT in single acute-phase samples, the pooled sensitivity and specificity were 14% (95% credible interval [CrI] 3–38%) and 86% (95% CrI 59–96%), respectively, and the predicted sensitivity and specificity were 14% (95% CrI 0–90%) and 86% (95% CrI 9–100%). Among paired MAT samples, the pooled sensitivity and specificity were 68% (95% CrI 32–92%) and 75% (95% CrI 45–93%) respectively, and the predicted sensitivity and specificity were 69% (95% CrI 2–100%) and 75% (2–100%).ConclusionsBased on our analysis, the accuracy of MAT in paired samples was not high, but it remains the reference standard until a more accurate diagnostic test is developed. Future studies that include larger numbers of participants with paired samples will improve the certainty of accuracy estimates.

Molecular and genetic characterization of LEPTOSPIRA spp. collection strains from the St. Petersburg Pasteur institute based on 16S rRNA gene sequencing data

Leptospirosis is a zoonotic disease found virtually worldwide. Microscopic Agglutination Test with live leptospira (MAT) is the reference method for the serological diagnosis of leptospirosis. MAT is based on assessing serum potential to agglutinate live reference serovar Leptospira maintained at a reference laboratory. At some laboratories having own collections of isolated and reference Leptospira strains applicable for serological diagnosis, those microorganisms are maintained for many years by repeated subculturing, that increases markedly a chance of strain cross-contamination. The lack of adequate quality control for reference strains may affect data of epidemiological studies. Control of Leptospira spp. reference strains purity and stability of their antigenic composition is very important for diagnosis of leptospirosis. The study objective was to compare the 16S rRNA gene nucleotide sequences of some Leptospira strains from the collection of the St. Petersburg Pasteur Institute to with relevant sequences uploaded to GenBank. In this study, 38 Leptospira strains were investigated. Nucleotide sequences of 36 strains were deposited in the international GenBank database, inconsistencies were revealed in two strains. The study found that the control Leptospira strains from the collection of the St. Petersburg Pasteur Institute had minimal dissimilarities from international control strains. The analysis of the resultant 16S rRNA sequences has shown the presence of point mutations, transitions, deletions and insertions, regardless of the strain species. The open leptospira pan-genome demonstrates high genomic variability in species due to the capability of leptospira for lateral gene transfer in order to adapt to changing environmental conditions. The massive acquisition and loss of genes give rise to an increased species diversity. The 16S rRNA gene is suitable for screening diagnostics; however, high level of the fragment similarity and close phylogenetic relationship between different species put bounds to its use in genotyping. The presence of point nucleotide mutations is most likely associated with the evolutionary mechanisms of leptospira, their ability to horizontal gene transfer and crossing-over, including ribosomal genes, but this assumption necessitates additional research. For specimen genotyping it is necessary to select alternative genes with high specificity and sufficient level of nucleotide divergence. The study shows a need for genetic analysis of collection strains in order to control the purity of cultures.