Leptospiral uveitis- “Transition ‘from epidemic to endemic form” difficulties in laboratory confirmation

Purpose: Leptospirosis is a waterborne zoonotic disease that primarily causes systemic illness, followed by uveitis. After heavy flooding in Madurai district, an epidemic outbreak of systemic and ocular leptospirosis occurred in 1994. Our data shows a transition to endemicity after each epidemic. Aim: The aim of this study is to report the clinical signs, epidemic outbreaks, and persistent endemicity of leptospiral uveitis, as well as the diagnostic dilemmas associated with it. Methods: A retrospective analysis of clinical signs was conducted using medical records of leptospiral uveitis patients over a period of 27 years (1994–2020) in a tertiary care eye hospital. The clinical workup of uveitis included a detailed clinical history, systemic, and ophthalmic examination. Microagglutination tests (MATs) was done at the Centers for Disease Control and Prevention (CDC) in Atlanta and later in our regional laboratory. Serum samples were collected from human systemic leptospirosis cases and a small group of animals in and around Madurai. Results: The first epidemic outbreak resulted in 200 seropositive patients. Subsequent epidemic outbreaks occurred in 1997, 1998, 2001, 2005, and 2012, with Madurai experiencing multiple outbreaks. However, the disease remained endemic, with 25–50 patients being observed per year in between the peaks. Ocular examination revealed acute non?granulomatous uveitis (94.9%), pan uveitis (59.8%), vitreous inflammatory reaction (55.4%), retinal vasculitis (29.5%), disc hyperemia (20.9%), and hypopyon. (16.2%). New serovars emerged every year, resulting in decreased sensitivity of the MAT. Over time, the MAT started to miss diagnoses. Conclusion: The persistent endemicity of leptospiral uveitis emphasizes the need for accessible diagnostic tests. The low performance of the MAT can be attributable to the use of an older panel. The incorporation of new isolates in the MAT by a national laboratory will improve the accuracy of diagnosis

Diagnosis of canine leptospirosis: evaluation of two PCR assays in comparison with the microagglutination test / Diagnóstico de leptospirose canina: avaliação de dois ensaios de PCR em comparação com o teste de microaglutinação

Canine leptospirosis is definitely diagnosed by demonstrating seroconversion in paired serum samples from the acute and convalescent period by the microagglutination test (MAT). However, the application of a polymerase chain reaction (PCR) assay can provide earlier confirmation of suspected cases. The objective of this study was to evaluate two PCR assays used in diagnosis of human leptospirosis (lipL32 real-time PCR and rrs conventional PCR) in cultured microorganisms and experimentally contaminated samples (whole blood, serum, urine), and investigate their applicability in clinical samples from dogs with presumptive diagnosis of leptospirosis by using the MAT as a reference. The analytical sensitivity of the lipL32 real-time PCR was 1 genome equivalent per reaction, whereas that for the rrs conventional PCR was 10 genome equivalents per reaction. Both assays amplified the pathogenic strains but were negative when evaluating the DNA of other microorganisms that may be present in clinical samples. The lipL32 real-time PCR detected 100 bacteria/mL in whole blood samples, 1000 bacteria/mL in serum samples and 10 bacteria/mL in urine samples, whereas the rrs conventional PCR detected 1000 bacteria/mL in whole blood and serum samples and 100 bacteria/mL in urine samples. Seven out of the 51 samples from dogs with presumptive diagnosis of leptospirosis were considered as confirmed cases. ThelipL32 real-time PCR detected positive results in six of the seven confirmed cases, whereas the rrs conventional PCR detected four. The PCR assays evaluated proved to be useful diagnostic tools in the confirmation of canine leptospirosis when used together with the MAT.(AU)

O diagnóstico definitivo da leptospirose canina é geralmente realizado demonstrando a seroconversão em amostras do paciente no período agudo e de convalescença por serologia. No entanto, a aplicação de técnicas de PCR pode contribuir para a confirmação de casos suspeitos num período de tempo mais curto. O objetivo deste estudo foi avaliar dois ensaios de PCR publicados em humanos (PCR-lipL32 em tempo real e PCR-rrs convencional) em culturas puras e em amostras de sangue com anticoagulante, soro e urina experimentalmente contaminados. Posteriormente, investigamos a utilidade de ambos os ensaios de PCR em amostras clínicas de cães com suspeita de leptospirose tomando a técnica de microaglutinação (MAT) como referência. A sensibilidade analítica foi de 1 e 10 genoma equivalente por reação para PCR-lipL32 em tempo real e para PCR-rrs convencional, respectivamente. Ambos os ensaios amplificaram corretamente as 14 estirpes patogênicas, mas foram negativos para avaliar o ADN de outros microrganismos que poderiam estar presentes em amostras clinicas. Em nas amostras experimentalmente contaminadas PCR-LipL32 em tempo real detectou 100 bactérias/mL em sangue total, 1000 bactérias/mL em soro e 10 bactérias/mL em urina. Enquanto o PCR-rrs convencional detectou 1000 bactérias/mL em sangue total e soro e 100 bactérias/mL na urina. Dos 51 cães suspeitos, sete foram considerados casos confirmados pela MAT. O PCR-lipL 32 em tempo real detectou seis dos sete casos confirmados, enquanto o PCR-rrs convencional foi positivo em quatro deles. As técnicas de PCR avaliadas provaram ser uma ferramenta de diagnóstico útil na confirmação de casos clínicos caninos quando utilizados em conjunto com a técnica MAT.(AU)

Seroprevalencia de leptospirosis en pacientes con síndrome febril no palúdico / Seroprevalence of leptospirosis in patients with non-malarial febrile syndrome

Introducción: la leptospirosis es una enfermedad zoonótica transmitida por varias especies de animales domésticos y silvestres que actúan como reservorios del agente causal y que afecta humanos que habitan áreas urbanas y rurales en el mundo. Objetivo: determinar la seroprevalencia de anticuerpos contra Leptospira sp. y los serogrupos dominantes, en pacientes con síndrome febril no palúdico del departamento del Meta, Colombia. Métodos: se realizó un estudio prospectivo de corte trasversal. La población la conformó todo paciente febril que asistiera a un hospital en la ciudad de Villavicencio, entre mayo de 2013 y junio de 2014. Los sueros pareados fueron procesados utilizando la prueba de microaglutinación con los serogrupos: Australis, Ballum, Bataviae, Canicola, Cynopteri, Panama, Pyrogenes, Sejroe, Semaranga. Se practicaron pruebas para diagnóstico de dengue (detección de anticuerpos IgM e IgG por ELISA), rickettsiosis (detección de anticuerpos IgG por inmunofluoresencia indirecta), hantavirosis (detección de anticuerpos IgG por ELISA). Resultados: de los 100 sueros estudiados, 29 resultaron positivos para leptospirosis. Los serogrupos más prevalentes fueron Canicola y Ballum. Conclusiones: los resultados de la prevalencia de leptospirosis muestran que esta enfermedad es subregistrada en la zona, por lo que sería recomendable que estuviera dentro del diagnóstico diferencial de los síndromes febriles(AU)

Introduction: leptospirosis is a zoonotic disease transmitted by several species of wild and domestic animals, which serve as reservoirs of the causative agent. The disease affects humans from urban and rural areas of the world. Objective: determine Leptospira sp. antibody seroprevalence and dominant serogroups in patients with non-malarial febrile syndrome from Meta Department in Colombia. Methods: across-sectional prospective study was conducted. The study population was all the febrile patients attending a hospital in the city of Villavicencio from May 2013 to June 2014. The paired sera were processed using the microagglutination test with the following serogroups: Australis, Ballum, Bataviae, Canicola, Cynopteri, Panama, Pyrogenes, Sejroe and Semaranga. Diagnostic tests were performed for dengue (IgM and IgG antibody detection by ELISA), rickettsiosis (IgG antibody detection by indirect immunofluorescence), and hantaviral disease (IgG antibody detection by ELISA). Results: of the 100 sera studied, 29 were positive for leptospirosis. The most prevalent serogroups were Canicola and Ballum. Conclusions: prevalence results for leptospirosis show that the disease has been under-recorded in the area. It is thus advisable to include it in the differential diagnosis of febrile syndromes(AU)

Cepario autóctono de leptospiras en la prueba de micro - aglutinación / Micro - agglutination test of Autogenous leptospirosis

Introducción: leptospirosis. Zoonosis más frecuente en Cuba, como enfermedad infecto-contagiosa. Objetivos: identificar las cepas de leptospiras aisladas localmente, para ser empleadas en la prueba de microaglutinación. Apreciar su comportamiento serológico frente a las cepas de referencia. Método: estudio experimental, de sueros de pacientes presuntivos de padecer Leptospirosis, con 60 muestras, en el Instituto Carlos J. Finlay, de La Habana. Identificación de 40 sueros reactores a la hemaglutinación pasiva. Identificación de 34 cepas, de un total de 250 aislamientos, a partir de hemocultivos, de pacientes con antisueros policlonales a serogrupos y anticuerpos monoclonales a serovar. Se conformaron cinco cepasrepresentativas de los serovares: Ballum Ballum, Canicola Canicola, Pomona Pomona, Hebdomadis Wolffi e Icterohaemorrhagiae Copenhageni, como las de mayor circulación en la región. Se realizó la prueba de microaglutinación con las cepas aisladas localmente, y las de referencia de forma paralela. Resultados: mayor reactividad con las cepas locales, y conformación de cinco cepas representantes de los serovares: Ballum Ballum, Canicola Canicola, Pomona Pomona, Hebdomadis Wolffi e Icterohaemorrhagiae Copenhageni, como las de mayor circulación en la región. Se encontró una concordancia de 86,36% en la reactividad. Conclusiones: los resultados mostraron mayor reactividad con las cepas locales que con las de referencia, elevada concordancia e incremento de los títulos de anticuerpos de los sueros reactores usando cepas locales, y disminución del promedio de las reacciones cruzadas cuando se utilizaron las cepas locales. Se demostró su utilidad en la evaluación de estudios inmunológicos.

Introduction: leptospirosis, a contagious infectious disease considered the most common zoonosis in Cuba. Objectives: to identify locally isolated leptospirosis strains for micro- agglutination test. To evaluate their serological behavior against the reference strains. Method: an experimental study of 60 presumptive Leptospirosis patients' serum samples. Forty sera reactive to passive hemagglutination. To isolate 250 blood cultures of patients. To show 34 strains identified at the Carlos J. Finlay Institute, with polyclonal antisera to serogroups and monoclonal antibodies to serovar. Five strains represented by the serovars: Ballum Ballum, Canicola Canicola, Pomona Pomona, Hebdomadis Wolffi and Icterohaemorrhagiae Copenhageni, were the most circulating strains in the region. The microagglutination test was performed in parallel form to locally isolated strains and reference strains Results: greater reactivity with the local strains. Five strains representing the serovars: Ballum Ballum, Canicola Canicola, Pomona Pomona, Hebdomadis Wolffi and Icterohaemorrhagiae copenhageni as the most circulating in the region. The reactivity showed 86.36% concordance. Conclusions: there was greater reactivity with the local strains than the reference strains. A higher concordance increased the titers of antibodies of the reactive sera, by the use of local strains. The average of cross reactions decreased when the local strains were used. The evaluation of immunological studies was demonstrated.

Seroprevalencia de leptospirosis humana en un asentamiento del área urbana de la ciudad de Guatemala / Seroprevalence of human leptospirosis in a settlement of the urban area in Guatemala city

Introducción: aunque la leptospirosis es considerada una enfermedad de ambientes rurales, la reciente aparición de epidemias urbanas la hace emerger como un problema en salud pública. En Guatemala (2008), se demostró una seroprevalencia de 51,8 % en áreas rurales, por lo que es importante llevar a cabo estudios en áreas urbanas que permitan establecer el impacto que pudiera tener en la población guatemalteca. Objetivos: determinar la seroprevalencia de leptospirosis humana en un asentamiento ubicado en la ciudad de Guatemala, así como los serovares de Leptospira interrogans circulantes y los factores de riesgo asociados a la exposición con esta bacteria. Métodos: participaron 119 habitantes con 6 años y más de los 2 sexos, que aceptaron, previo consentimiento informado. Con una entrevista estructurada se recolectaron los datos sociodemográficos y las muestras de sangre venosa. La técnica de microaglutinación y ELISA IgG se utilizaron para la detección de anticuerpos. Los sueros se enfrentaron a 20 serovariedades de Leptospira interrogans sensu lato. La prevalencia se determinó con un IC95% y las variables sociodemográficas con chi cuadrado, la razón de prevalencia con Epi Info 3.5.1. Resultados: la seroprevalencia de leptospirosis en la población estudiada resultó de 30,3 %, (IC95%). Los serovares más frecuentes fueron Australis y Lanka (11,1 % ambos). El título más frecuente fue de 1:80 por microaglutinación. En la población se encontraron distintos factores de riesgo, pero ninguno mostró una asociación significativa con la presencia de anticuerpos anti-Leptospira (p> 0,05). Conclusiones: la seroprevalencia de leptospirosis detectada fue de 30,3 % en los habitantes del asentamiento ubicado en la ciudad de Guatemala, la cual es comparable a las áreas urbanas de países en donde esta enfermedad es hiperendémica, por lo que es importante implementar medidas de prevención y de control de la enfermedad en la comunidad, en forma conjunta con la municipalidad del distrito.

Introduction: although leptospirosis is considered a rural environment disease, recent urban epidemics make it emerge as a public health problem. A seroprevalence of 51.8 % has been found in rural areas of Guatemala; therefore it is important to establish the impact that this disease may have on the Guatemalan urban population. Objectives: to determine the seroprevalence of human leptospirosis in a settlement of Guatemala city and also to identify urban Leptospira interrogans serovars and risk factors associated with exposure to the bacteria. Methods: there were selected 119 people aged 6 years old and over from both sexes, who agreed to participate after giving their informed consent. Sociodemographic data were collected and venous blood samples were taken. The microagglutination test (MAT) and ELISA IgG were used to detect antibodies. Sera were tested against 20 serovars of Leptospira interrogans sensu lato. The seroprevalence was determined with a 95% confidence interval, and sociodemographic variables were evaluated with Chi square and Odds Ratio using Epi Info 3.5.1. Results: a 30.3 % seroprevalence of leptospirosis was found in the study population (CI, 95%). The more frequently serovars founded were Australis and Lanka (11.1 % each). The most common titer was 1:80 (MAT). Different risk factors were found, but none showed a significant association with the presence of Leptospira IgG antibodies (p> 0.05). Conclusion: the prevalence of IgG anti-Leptospira antibodies detected in the residents of the settlement is comparable to that of the urban areas of other countries where leptospirosis is hyperendemic. For avoiding outbreaks in these areas, it is important to prevent and control the infection in the community.