Daños renales en fetos de ratas Wistar infectadas con Leptospira canicola

RESUMEN Introducción: La leptospirosis es una de las zoonosis que en Cuba se presenta de forma endémica, es causante de pérdidas humanas, animales y económicas, por lo que es un problema de salud. Las ratas constituyen un vector transmisor de gran importancia. Trabajos experimentales para estudiar las alteraciones renales fetales en modelo de rata Wistar no se han realizado en Cuba. Objetivo: Identificar las principales alteraciones morfométricas renales en fetos de ratas Wistar infectadas con serovariedad Leptospira canicola, en la Universidad de Ciencias Médicas de Pinar del Rio en el período comprendido entre 2017-2019. Método: Se realizó un estudio experimental en ratas Wistar preñadas: 2 grupos con tres animales cada uno: el grupo experimental con dosis de una concentración bacteriana de 300 millones de leptospiras por mililitros; se inoculó 1 ml a cada una infectándolas con cepas de Leptospira canicola. Se estudiaron los riñones de fetos extraídos por cesárea y se analizaron las variables: longitud renal y peso del riñón. Resultados: Los resultados obtenidos confirmaron los daños renales producidos por la Leptospira canicola tanto en el eje mayor como el menor de ambos riñones, así como variaciones en el peso de las crías infectadas. Conclusiones: La infección por Leptospira canicola en ratas Wistar preñadas provoca alteraciones en los riñones de los fetos lo que causa cambios en la morfometría de los ejes tanto mayor como menor, así como en el peso corporal del feto, lo que conlleva a una disminución del peso renal.

ABSTRACT Introduction: Leptospirosis is one of the endemic zoonoses in Cuba, causing human, animal and economic losses, making it a significant health problem. Rats are a very remarkable vector. Experimental work to study fetal renal alterations in the Wistar rats has not been carried out in Cuba. Objective: To identify the main renal morphometric alterations in fetuses of Wistar rats infected with serovar Leptospira canicola, at the Universidad de Ciencias Médicas de Pinar del Rio in the period between 2017-2019. Method: An experimental study was carried out in pregnant Wistar rats: 2 groups with three animals each: the experimental group with a dose of bacterial concentration of 300 million leptospires per milliliter; 1 ml was inoculated to each one, infecting them with strains of Leptospira canicola. The kidneys of fetuses extracted by cesarean section were studied, and the variables: renal length and kidney weight were analyzed. Results: The results obtained confirmed kidney damage caused by Leptospira canicola both in the major and minor axis of both kidneys, as well as variations in the weight of the infected pups. Conclusions: Leptospiral infection in pregnant Wistar rats causes alterations in the kidneys of the fetuses, bringing changes in the morphometry of both the major and minor axes, as well as in the body weight of the fetus, which leads to a decrease in renal weight.

RESUMO Introdução: A leptospirose é uma das zoonoses endêmicas em Cuba, causando prejuízos humanos, animais e econômicos, tornando-se um problema de saúde. Os ratos são um vetor transmissor muito importante. Trabalho experimental para estudar as alterações renais fetais no modelo de rato Wistar não foi realizado em Cuba. Objetivo: Identificar as principais alterações morfométricas renais em fetos de ratos Wistar infectados com sorovar Leptospira canicola, na Universidad de Ciencias Médicas de Pinar del Rio no período entre 2017-2019. Método: Estudo experimental realizado em ratas Wistar prenhes: 2 grupos com três animais cada: o grupo experimental com dose de concentração bacteriana de 300 milhões de leptospiras por mililitro; 1 ml fora inoculado em cada um, infectando-os com cepas de Leptospira canicola. Foram estudados os rins de fetos extraídos por cesariana e analisadas as variáveis: comprimento renal e peso renal. Resultados: Os resultados obtidos confirmaram o dano renal causado por Leptospira canicola tanto no eixo maior quanto no eixo menor de ambos os rins, bem como variações no peso dos filhotes infectados. Conclusões: A infecção por Leptospira canicola em ratas Wistar prenhes causa alterações nos rins dos fetos, o que provoca alterações na morfometria dos eixos maior e menor, bem como no peso corporal do feto, o que leva a uma diminuição da peso do rim.

Detección de Leptospira spp. (Spirochaetales: Leptospiraceae) en muestras ambientales de regiones habitadas por poblaciones vulnerables del norte argentino / Leptospira spp. (Spirochaetales: Leptospiraceae) detection in environmental samples from geographical regions inhabited by vulnerable communities in northern Argentina

Resumen La leptospirosis continúa siendo hoy en día un problema para la salud pública, principalmente en poblaciones de bajos recursos socioeconómicos. En este trabajo se presenta la detección de leptospiras patógenas en muestras ambientales (aguas y barros) provenientes de regiones del norte argentino (provincias de Formosa, Salta, Santiago del Estero, Misiones y Chaco) con variadas características climatológicas habitadas por poblaciones vulnerables. De las 89 muestras analizadas, en el 24,7% fue posible detectar molecularmente la presencia de leptospiras patógenas. La prevalencia por tipo de muestra fue de 27,8% para las aguas y 11,8% para los barros. Todas las localidades muestreadas presentaron al menos una muestra positiva a alguna de las pruebas realizadas, por lo que el presente trabajo refleja la necesidad de profundizar los estudios de la leptospirosis en distintas regiones de la Argentina.

Abstract Leptospirosis remains as a major public health problem nowadays, mainly affecting vulnerable communities with low socioeconomic resources. In this study, the molecular detection of pathogenic leptospires from environmental samples (water and mud) from northern Argentina (Formosa, Salta, Santiago del Estero, Misiones and Chaco provinces) is described. Samples were obtained from regions with varied climatological features, all inhabited by vulnerable communities. From the 89 samples that were analyzed, 24.7% showed molecular evidence of the presence of pathogenic leptospires. Prevalence by sample type was: 27.8% in water samples and 11.8% in mud samples. All the sampled regions showed at least one positive sample. This result highlights the need of further research regarding leptospirosis in different regions of Argentina.

Prevalence and genetic divergence of Leptospira interrogans and Leptospira borgpetersenii in house rats (Rattus rattus) from Peninsular Malaysia

Objective: To evaluate the prevalence and divergence of genetically identified Leptospira spp. in the population of Rattus rattus. Methods: A total of 130 rats were used in this study. The infection within the rats were screened using polymerase chain reaction (PCR)-based diagnosis, with Leptospira genus-specific 16S rRNA primer and pathogenic Leptospira spp. specific LipL32 primer, on both kidney and liver tissues of Rattus rattus to detect the presence of potential Leptospira spp. Results: Out of 130 rats studied, 51 (39.23%) individuals were positive for leptospiral DNA. Basic Local Alignment Search Tool (BLAST) and phylogenetic analysis revealed that both pathogenic Leptospira interrogans and Leptospira borgpetersenii were predominantly identified. Phylogenetically, both genes disclosed similar clustering patterns of tree topologies between the two species. Although both genes were conserved, LipL32 gene portrayed higher nucleotide divergence (5.80%) compared to the 16S rRNA gene (0.60%). Minimum-spanning network displayed several haplotypes that are unique to each species, suggesting a higher degree of subdivision between both species. As for prevalence surveillance, both adult and subadult rats were susceptible to the infection, in which males were the most susceptible. Kidney was notable as the favourable organ for colonisation of leptospires. Rats captured from fresh markets were highly infected with Leptospira spp. (54.28%) compared to those from housing areas (26.47%). Conclusions: Rattus rattus represents an important asymptomatic transmitter of pathogenic leptospires, and hence is of public health concerns.

Prevalence and genetic divergence of Leptospira interrogans and Leptospira borgpetersenii in house rats (Rattus rattus) from Peninsular Malaysia

Objective: To evaluate the prevalence and divergence of genetically identified Leptospira spp. in the population of Rattus rattus. Methods: A total of 130 rats were used in this study. The infection within the rats were screened using polymerase chain reaction (PCR)-based diagnosis, with Leptospira genus-specific 16S rRNA primer and pathogenic Leptospira spp. specific LipL32 primer, on both kidney and liver tissues of Rattus rattus to detect the presence of potential Leptospira spp. Results: Out of 130 rats studied, 51 (39.23%) individuals were positive for leptospiral DNA. Basic Local Alignment Search Tool (BLAST) and phylogenetic analysis revealed that both pathogenic Leptospira interrogans and Leptospira borgpetersenii were predominantly identified. Phylogenetically, both genes disclosed similar clustering patterns of tree topologies between the two species. Although both genes were conserved, LipL32 gene portrayed higher nucleotide divergence (5.80%) compared to the 16S rRNA gene (0.60%). Minimum-spanning network displayed several haplotypes that are unique to each species, suggesting a higher degree of subdivision between both species. As for prevalence surveillance, both adult and subadult rats were susceptible to the infection, in which males were the most susceptible. Kidney was notable as the favourable organ for colonisation of leptospires. Rats captured from fresh markets were highly infected with Leptospira spp. (54.28%) compared to those from housing areas (26.47%). Conclusions: Rattus rattus represents an important asymptomatic transmitter of pathogenic leptospires, and hence is of public health concerns.

Pesquisa de sorovares de leptospiras em rebanho bovino leiteiro no estado do Piauí, Brasil / Survey of leptospira serovars in a dairy cattle herd in Piauí state, Brazil

Este trabalho teve como objetivo relatar a pesquisa de sorovares de leptospiras em animais de uma fazenda localizada no Município de Buriti dos Lopes, integrante da Bacia Leiteira de Parnaíba, Piauí, Brasil. Foram colhidas amostras de sangue de 18 vacas da raça Girolando que abortaram no período de julho de 2004 a abril de 2005. As amostras foram submetidas ao teste de soroaglutinação microscópica (SAM) utilizando-se antígenos vivos de 23 sorovares de leptospiras patogênicas. 83,33% (15/18) das amostras apresentaram aglutininas anti-leptospira, sendo 53,33% de sorovar Hardjoprajitno (8/15), 6,67% para Hardjobovis (1/15), Wolffi (1/15) e Shermani (1/15) e quatro (4/15) co-agluitinações. A amostra do reprodutor apresentou aglutininas antileptospira para o sorovar Hardjobovis com titulação 1:400 o que sugere que este animal teve participação na disseminação da infecção no rebanho.

This study aimed to report the leptospira serovar varieties in animals from a farm located in the county of Buriti dos Lopes, within the so-called "milk basin" of Parnaíba, state of Piauí, Brazil. Samples of blood were collected from 18 Girolando cows who aborted between July 2004 and April 2005, and a sample was taken from a breeder bull. The samples were subjected to the microscopic agglutination test (SAM), using live antigens of 23 pathogenic leptospira serovars. Serum samples were screened in the dilution of 1:100. Data analysis revealed that 83.33% (15/18) of the samples showed anti-leptospira agglutinins, of which 53.33% were for the Hardjoprajitno serovar (8/15), 6.67% each for Hardjobovis (1/15), Wolffi (1/15) and Shermani (1/15), along with 4 (4/15) coagglutinations. The breeder-bull sample showed anti-leptospira agglutinins for serovar Hardjobovis with titration of 1:400, suggesting that this animal had participated in the spread of the infection in the herd.

EMJH medium with 5-fluorouracil and nalidixic acid associated with serial dilution technique used to recover Leptospira spp from experimentally contaminated bovine semen / Meio de EMJH com 5-fluorouracil e ácido nalidíxico associado a técnica das diluições seriadas usados para recuperar Leptospira spp do sêmen bovino experimentalmente contaminado

Bovine semen experimentally contaminated with Leptospira santarosai serovar Guaricura was submitted tothe modified EMJH medium with 5-fluorouracil (300mg/L) and nalidixic acid (20mg/L), named as ôselectivemediumõ and using the serial dilution technique, in order to evaluate the percentage of recovery of the addedmicroorganism. The selective EMJH medium was found with higher percentage of recovery of leptospirasand minor losses of samples due to contamination with opportunistic microorganisms than the non-selectiveEMJH medium: 151/376 (40.0%) of positive growth; and 38/376 (10.0%) contamination and 58/376 (15%) and129/376 (34.0%), respectively. These results were statistically significant (p<0. 0001; Fisher). Differences werefound when the frequencies of positive leptospires recovery have been compared in the serial dilution technique (10-1 to 10-4) between the selective and non-selective media at different dilution factors. At 1/10th dilution the percentages found were (0%, 0/80) and (38%, 30/80), at 1/100th dilution, (3%, 2/80) and (49%, 39/ 80) and at 1/1,000th dilution, (25%, 20/80) and (50%, 40/80), respectively. The percentage of recovery of leptospires was found to be directly proportional to the dilution used. The methodology of the serial dilution technique (setting at least three dilutions) and the use of selective EMJH medium have been found to be efficient for the isolation of leptospires from the bovine semen samples.

Sêmen bovino experimentalmente contaminado com a estirpe Leptospira santarosai Sorovar Guaricura foi utilizado para verificar a porcentagem de recuperação de leptospiras, utilizandoo meio de EMJH com 5-fluorouracil (300 mg/L) e ácido nalidíxico (20 mg/L), denominado ômeio seletivoõ, associado à técnica das diluições seriadas. O meio seletivo EMJH foi encontrado comporcentagem de recuperação mais elevada e com menos perda de amostras devido à contaminação com microorganismos oportunistas quando comparado com o meio EMJH não seletivo, que foram encontrados, respectivamente, com 151/376 (40.0%) de crescimento positivo e 38/376 (10.0%) de contaminação, 58/376 (15%) e 129/376 (34.0%). Estes resultados foram estatíticamente significantes (p<0, 0001; Fisher). Diferenças foram encontradas quando as freqüências de leptospiras recuperadas foram comparadas com a técnica de diluição seriada (10-1 a 10-4) e entre os meios seletivo e não-seletivo, em diferentes níveis de diluição. À diluição de 1/10, as porcentagens encontradas foram (0%, 0/80) e (38%, 30/80); à diluição 1/100, (3%, 2/80) e (49%, 39/80); e à diluição 1/1000, (25%, 20/80) e (50%, 40/80), respectivamente. A porcentagem de recuperação de leptospiras foi diretamente proporcional às diluições empregadas. A metodologia das diluições seriadas (estabelecendo pelo menos três diluições) e o uso de meio EMJH seletivo mostraram ser eficientes no isolamento de leptospiras presentes nas amostras de sêmen bovino.

Clinical trial of leptospires vaccine on its immunogenicity and safety / 예방의학회지

Since the reservior of leptospires organism is consisted of a broad spectrum of animals, the best method of prevention is vaccination. The clinical trial of leptospires vaccine conducted on human volunteer for its immunogenicity and safety. Summarized results are as follows: 1. The Oral temperature among vaccinated group ranged from 36.7 +/- 0.34 degrees C, while in placebo injected group it ranged from 36.4 +/- 0.46 degrees C to 36.7 +/- 0.53 degrees C. There wan no association between vaccination and fever (p<0.05). 2. Mild local reactions revealed in vacciness were swelling (50-75%), Redness(75-90%), and induration(25-40%). Placebo injected group revealed only redness in 12.5% in the 1st injection and 37.5% in the second injection. The duration local reactions on injection site for both vaccinees and placebo groups disappeared within 48 hours. 3. Generalized Symptoms complained by the vaccinees were myalgia (25%), back pain(15%), headache(15%), pruritus(15%), and abdominal pain(10%), whereas placebo group complained of headache(25%), myalgia(12.5%), back pain(12.5%), pain in eyes(12.5%), abdominal pain(12.5%) prutitus(12.5%) and nausea(12.5%). 4. The serological test(MAT) of vaccinees showed geometric mean antibody titer as follows: a. L. icterohemorrhagiae lai, 1 week after 1st vaccination: 22.45, 1 week after 2nd vaccination: 111.23, 3 week after 2nd vaccination: 266.64, b. L. canicola canicola, 1 week after 1st vaccination: 24.62, 1 week after 2nd vaccination: 123.92, 3 week after 2nd vaccination: 276.55, c. L. icterohemorrhagiae copenhageni, 1 week after 1st vaccination: 28.28, 1 week after 2nd vaccination: 128.55, 3 week after 2nd vaccinaton: 247.88, Whereas all of the placebo injected group, showed below 1 : 20 titers. The sero-conversion rate of vaccinees were 100 percent.