RESUMO
Leptospirosis, a neglected zoonotic disease, is caused by pathogenic spirochetes belonging to the genus Leptospira and has one of the highest morbidity and mortality rates worldwide. Vaccination stands out as one of the most effective preventive measures for susceptible populations. Within the outer membrane of Leptospira spp., we find the LIC12287, LIC11711, and LIC13259 lipoproteins. These are of interest due to their surface location and potential immunogenicity. Thorough examination revealed the conservation of these proteins among pathogenic Leptospira spp.; we mapped the distribution of T- and B-cell epitopes along their sequences and assessed the 3D structures of each protein. This information aided in selecting immunodominant regions for the development of a chimeric protein. Through gene synthesis, we successfully constructed a chimeric protein, which was subsequently expressed, purified, and characterized. Hamsters were immunized with the chimeric lipoprotein, formulated with adjuvants aluminum hydroxide, EMULSIGEN®-D, Sigma Adjuvant System®, and Montanide™ ISA206VG. Another group was vaccinated with an inactivated Escherichia coli bacterin expressing the chimeric protein. Following vaccination, hamsters were challenged with a virulent L. interrogans strain. Our evaluation of the humoral immune response revealed the production of IgG antibodies, detectable 28 days after the second dose, in contrast to pre-immune samples and control groups. This demonstrates the potential of the chimeric protein to elicit a robust humoral immune response; however, no protection against challenge was achieved. While this study provides valuable insights into the subject, further research is warranted to identify protective antigens that could be utilized in the development of a leptospirosis vaccine. KEY POINTS: ⢠Several T- and B-cell epitopes were identified in all the three proteins. ⢠Four different adjuvants were used in vaccine formulations. ⢠Immunization stimulated significant levels of IgG2/3 in vaccinated animals.
Assuntos
Anticorpos Antibacterianos , Vacinas Bacterianas , Leptospirose , Lipoproteínas , Animais , Leptospirose/prevenção & controle , Leptospirose/imunologia , Lipoproteínas/imunologia , Lipoproteínas/genética , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/genética , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Cricetinae , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito B/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/genética , Adjuvantes Imunológicos/administração & dosagem , Imunoglobulina G/sangue , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/genética , Leptospira interrogans/imunologia , Leptospira interrogans/genética , Proteínas da Membrana Bacteriana Externa/imunologia , Proteínas da Membrana Bacteriana Externa/genética , Vacinação , Imunidade Humoral , Leptospira/imunologia , Leptospira/genética , Imunogenicidade da VacinaRESUMO
A simple IgG-speciï¬c 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 90â¯% of the animals by IgG-ELISA and 60â¯% by MAT, while after booster, antibodies were detected in 100â¯% and 80â¯% 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.
Assuntos
Testes de Aglutinação , Anticorpos Antibacterianos , Vacinas Bacterianas , Doenças dos Bovinos , Ensaio de Imunoadsorção Enzimática , Imunoglobulina G , Leptospirose , Animais , Leptospirose/veterinária , Leptospirose/diagnóstico , Leptospirose/imunologia , Bovinos , Doenças dos Bovinos/microbiologia , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/sangue , Ensaio de Imunoadsorção Enzimática/veterinária , Anticorpos Antibacterianos/sangue , Vacinas Bacterianas/imunologia , Imunoglobulina G/sangue , Testes de Aglutinação/veterinária , Leptospira interrogans/imunologia , Leptospira/imunologia , Vacinação/veterinária , Testes Sorológicos/veterináriaRESUMO
BACKGROUND: Rodents are recognized as major reservoirs of numerous zoonotic pathogens and are involved in the transmission and maintenance of infectious diseases. Furthermore, despite their importance, diseases transmitted by rodents have been neglected. To date, there have been limited epidemiological studies on rodents, and information regarding their involvement in infectious diseases in the Republic of Korea (ROK) is still scarce. METHODOLOGY/PRINCIPAL FINDINGS: We investigated rodent-borne pathogens using nested PCR/RT-PCR from 156 rodents including 151 Apodemus agrarius and 5 Rattus norvegicus from 27 regions in eight provinces across the ROK between March 2019 and November 2020. Spleen, kidney, and blood samples were used to detect Anaplasma phagocytophilum, Bartonella spp., Borrelia burgdorferi sensu lato group, Coxiella burnetii, Leptospira interrogans, and severe fever with thrombocytopenia syndrome virus (SFTSV). Of the 156 rodents, 73 (46.8%) were infected with Bartonella spp., 25 (16.0%) with C. burnetii, 24 (15.4%) with L. interrogans, 21 (13.5%) with A. phagocytophilum, 9 (5.8%) with SFTSV, and 5 (3.2%) with Borrelia afzelii. Co-infections with two and three pathogens were detected in 33 (21.1%) and 11 rodents (7.1%), respectively. A. phagocytophilum was detected in all regions, showing a widespread occurrence in the ROK. The infection rates of Bartonella spp. were 83.3% for B. grahamii and 16.7% for B. taylorii. CONCLUSIONS/SIGNIFICANCE: To the best of our knowledge, this is the first report of C. burnetii and SFTSV infections in rodents in the ROK. This study also provides the first description of various rodent-borne pathogens through an extensive epidemiological survey in the ROK. These results suggest that rodents harbor various pathogens that pose a potential threat to public health in the ROK. Our findings provide useful information on the occurrence and distribution of zoonotic pathogens disseminated among rodents and emphasize the urgent need for rapid diagnosis, prevention, and control strategies for these zoonotic diseases.
Assuntos
Anaplasma phagocytophilum , Bartonella , Coxiella burnetii , Zoonoses , Animais , República da Coreia/epidemiologia , Zoonoses/epidemiologia , Zoonoses/microbiologia , Ratos , Coxiella burnetii/isolamento & purificação , Coxiella burnetii/genética , Bartonella/isolamento & purificação , Bartonella/genética , Anaplasma phagocytophilum/isolamento & purificação , Anaplasma phagocytophilum/genética , Roedores/microbiologia , Murinae/microbiologia , Animais Selvagens/microbiologia , Animais Selvagens/virologia , Doenças dos Roedores/epidemiologia , Doenças dos Roedores/microbiologia , Doenças dos Roedores/virologia , Phlebovirus/genética , Phlebovirus/isolamento & purificação , Reservatórios de Doenças/microbiologia , Leptospira interrogans/isolamento & purificação , Leptospira interrogans/genéticaRESUMO
Leptospirosis is a common but underdiagnosed zoonosis. We conducted a 1-year prospective study in La Guaira State, Venezuela, analyzing 71 hospitalized patients who had possible leptospirosis and sampling local rodents and dairy cows. Leptospira rrs gene PCR test results were positive in blood or urine samples from 37/71 patients. Leptospira spp. were isolated from cultured blood or urine samples of 36/71 patients; 29 had L. interrogans, 3 L. noguchii, and 4 L. venezuelensis. Conjunctival suffusion was the most distinguishing clinical sign, many patients had liver involvement, and 8/30 patients with L. interrogans infections died. The Leptospira spp. found in humans were also isolated from local rodents; L. interrogans and L. venezuelensis were isolated from cows on a nearby, rodent-infested farm. Phylogenetic clustering of L. venezuelensis isolates suggested a recently expanded outbreak strain spread by rodents. Increased awareness of leptospirosis prevalence and rapid diagnostic tests are needed to improve patient outcomes.
Assuntos
Surtos de Doenças , Leptospira , Leptospirose , Filogenia , Roedores , Animais , Leptospirose/epidemiologia , Leptospirose/veterinária , Leptospirose/microbiologia , Leptospirose/diagnóstico , Humanos , Venezuela/epidemiologia , Bovinos , Leptospira/genética , Leptospira/isolamento & purificação , Leptospira/classificação , Feminino , Roedores/microbiologia , Adulto , Masculino , Pessoa de Meia-Idade , Doenças dos Bovinos/microbiologia , Doenças dos Bovinos/epidemiologia , Adolescente , Leptospira interrogans/genética , Leptospira interrogans/isolamento & purificação , Leptospira interrogans/classificação , Adulto Jovem , Estudos Prospectivos , Criança , Idoso , Doenças Endêmicas , Zoonoses/epidemiologia , Zoonoses/microbiologia , Pré-EscolarRESUMO
Leptospirosis is a neglected zoonosis for which investigations assessing host-pathogen interaction are scarce. The aim of this study was to compare the severity and bacterial species involved in human cases of leptospirosis on Reunion and Mayotte islands, territories located in the southwest Indian Ocean that have recorded high human leptospirosis incidence but display fairly distinct epidemiological situations. A retrospective multicentric study including all patients over 18 years of age from Mayotte or Reunion with proven leptospirosis was conducted from January 2018 to April 2020. This study collected demographic, geographical, clinical, and biological data. Overall, 490 patients were included, 222 on Mayotte and 268 on Reunion. More patients were hospitalized on Reunion (n = 215, 80%) compared with Mayotte (n = 102, 46%). Severe disease was more common on Reunion (n = 75, 28%) than on Mayotte (n = 22, 10%). The dominant Leptospira species on Reunion was Leptospira interrogans (79%) followed by Leptospira borgpetersenii (21%), contrasting with the epidemiological situation on Mayotte where L. interrogans was found in only a minority of patients (10%). The high frequency of severe cases on Reunion could be explained not only by higher comorbidities but also by the higher occurrence of L. interrogans infections compared with Mayotte. Finally, the distribution of cases linked to L. borgpetersenii was found almost exclusively on the west coast of Reunion, raising the potential role of a ruminant reservoir.
Assuntos
Leptospira , Leptospirose , Leptospirose/epidemiologia , Leptospirose/microbiologia , Humanos , Reunião/epidemiologia , Leptospira/isolamento & purificação , Feminino , Adulto , Masculino , Estudos Retrospectivos , Pessoa de Meia-Idade , Comores/epidemiologia , Leptospira interrogans/isolamento & purificação , Adulto Jovem , Adolescente , Idoso , AnimaisRESUMO
The World Health Organization classifies leptospirosis as a significant public health concern, predominantly affecting impoverished and unsanitary regions. By using the Pensacola Bay System as a case study, this study examines the underappreciated susceptibility of developed subtropical coastal ecosystems such as the Pensacola Bay System to neglected zoonotic pathogens such as Leptospira. We analyzed 132 water samples collected over 12 months from 44 distinct locations with high levels of Escherichia coli (>410 most probable number/100 mL). Fecal indicator bacteria (FIB) concentrations were assessed using IDEXX Colilert-18 and Enterolert-18, and an analysis of water physiochemical characteristics and rainfall intensity was conducted. The LipL32 gene was used as a quantitative polymerase chain reaction (qPCR) indicator to identify the distribution of Leptospira interrogans. The results revealed 12 instances of the presence of L. interrogans at sites with high FIB over various land cover and aquatic ecosystem types. Independent of specific rainfall events, a seasonal relationship between precipitation and elevated rates of fecal bacteria and leptospirosis was found. These findings highlight qPCR's utility in identifying pathogens in aquatic environments and the widespread conditions where it can be found in natural and developed areas.
Assuntos
Microbiologia da Água , Leptospirose/microbiologia , Leptospirose/epidemiologia , Leptospira/isolamento & purificação , Leptospira/genética , Fezes/microbiologia , Leptospira interrogans/isolamento & purificação , Leptospira interrogans/genética , Monitoramento Ambiental/métodos , Chuva , Estações do Ano , Baías/microbiologia , Análise Espaço-TemporalRESUMO
Leptospirosis is a zoonotic disease caused by the pathogenic spirochaetes of the genus Leptospira. It is a public health concern in the Pacific Islands and is considered endemic in Palau. However, information on the genotypes and serotypes of causative Leptospira spp. in the country is limited. In this study, we isolated leptospires and detected antileptospiral antibodies in dogs and pigs. The isolates were characterized using a serological method and whole-genome sequencing. Leptospira interrogans was isolated from five of the 20 symptomatic dogs and one of the 58 healthy pigs. Their serogroups were identified as Icterohaemorrhagiae and Pyrogenes; however, the serogroup of one isolate could not be determined. Anti-Leptospira antibodies were detected in 14.4% (26/181) of the dogs and 20% (10/50) of the pigs. The reactive serogroups in dogs and pigs were almost identical, except for the Panama serogroup. Core genome multilocus sequence typing revealed that five of the six core genome sequence types (cgSTs) were newly identified in this study. The cgSTs from the serogroup Icterohaemorrhagiae isolates belonged to the same group as the Copenhageni and Icterohaemorrhagiae serovars isolated in other countries, whereas no similar cgSTs were identified in the Pyrogenes or unidentified serogroup strains. We demonstrated a high incidence of canine and porcine leptospirosis and identified new L. interrogans genotypes (cgSTs) circulating in Palau. Further investigations are needed to determine whether dogs and pigs serve as maintenance hosts for newly identified L. interrogans genotypes and whether they pose a risk of leptospirosis transmission to humans.
Assuntos
Doenças do Cão , Leptospirose , Doenças dos Suínos , Animais , Leptospirose/veterinária , Leptospirose/epidemiologia , Leptospirose/microbiologia , Cães , Suínos , Doenças do Cão/microbiologia , Doenças do Cão/epidemiologia , Doenças dos Suínos/microbiologia , Doenças dos Suínos/epidemiologia , Leptospira/genética , Leptospira/imunologia , Leptospira/classificação , Leptospira/isolamento & purificação , Leptospira interrogans/genética , Leptospira interrogans/imunologia , Leptospira interrogans/isolamento & purificação , Leptospira interrogans/classificação , Sorogrupo , Sequenciamento Completo do Genoma , Anticorpos Antibacterianos/sangue , Genótipo , Tipagem de Sequências MultilocusRESUMO
Leptospirosis is an infectious disease with a worldwide distribution, which represents a major challenge in animal production across developing countries, mainly in tropical areas. Horses are particularly susceptible to the disease, presenting manifestations ranging from subclinical to the development of uveitis that compromises the visual health of the animals. In recent years, serological studies have been carried out in equid populations from America, demonstrating high exposure. For this reason, the aim of this study was to demonstrate microbiologically and molecularly the presence of the members of the genus Leptospira in urine samples from equids in an endemic state of leptospirosis in Mexico, and to detect the serological presence of anti-Leptospira antibodies in the sampled animals. For this reason, blood and urine samples were collected from 28 horses and one mule from three localities in the state of Veracruz, Mexico. Urine samples were inoculated in Ellinghausen-McCullough-Johnson-Harris (EMJH) medium, and the recovered isolates were typed using a short Multi Locus Sequence Typing scheme. Amplifications of the expected size were subjected to sequencing, and the recovered sequences were compared with those of reference deposited in GenBank using the BLAST tool. To identify their phylogenetic position, we performed a phylogenetic reconstruction using the maximum likelihood method. Additionally, Microscopic Agglutination test was performed on the serum samples to identify anti-Leptospira antibodies. We recovered 16 urine isolates which tested positive for the presence of Leptospira DNA. The phylogenetic reconstruction and the MLST analysis confirmed the presence of several genotypes of Leptospira interrogans and Leptospira santarosai. An overall serological frequency of 97.1 % was detected. Our results represent the first record of the presence of Leptospira through bacteriological isolates in equids from Mexico.
Assuntos
Anticorpos Antibacterianos , Doenças dos Cavalos , Leptospira , Leptospirose , Filogenia , Animais , Leptospirose/veterinária , Leptospirose/microbiologia , Leptospirose/epidemiologia , México/epidemiologia , Cavalos/microbiologia , Doenças dos Cavalos/microbiologia , Leptospira/isolamento & purificação , Leptospira/genética , Leptospira/classificação , Anticorpos Antibacterianos/sangue , Leptospira interrogans/isolamento & purificação , Leptospira interrogans/genética , Leptospira interrogans/classificação , Tipagem de Sequências Multilocus , Análise de Sequência de DNA , DNA Bacteriano/genéticaRESUMO
The reproductive features of equine leptospirosis are often neglected. Equine genital leptospirosis is characterized as a silent chronic syndrome, and besides abortions, leads to placental abnormalities, stillbirths, and birth of weak foals. This study aimed to study the occurrence of placental abnormalities associated with Leptospira interrogans infection in naturally infected mares under field conditions. The studied herd had a high occurrence of placentitis and abortions. Ten pregnant mares, eight with placental abnormalities on ultrasonography and were selected. Serum and cervicovaginal mucus (CVM) samples were collected for serology and PCR, respectively. Positive samples in lipL32-PCR were submitted to the sequencing of the secY gene. In lipL32-PCR of CVM, five out of 10 (50%) mares were positive and all were characterized as Leptospira interrogans. Our results highlight the presence of placental abnormalities in the reproductive subclinical leptospirosis syndrome. We encourage field veterinarians to include leptospirosis testing in their reproductive management.
Assuntos
Doenças dos Cavalos , Leptospira interrogans , Leptospirose , Doenças Placentárias , Placenta , Complicações Infecciosas na Gravidez , Animais , Cavalos , Leptospirose/veterinária , Leptospirose/microbiologia , Leptospirose/epidemiologia , Leptospirose/complicações , Doenças dos Cavalos/microbiologia , Gravidez , Feminino , Leptospira interrogans/isolamento & purificação , Doenças Placentárias/microbiologia , Doenças Placentárias/veterinária , Doenças Placentárias/patologia , Complicações Infecciosas na Gravidez/veterinária , Complicações Infecciosas na Gravidez/microbiologia , Placenta/microbiologia , Placenta/patologiaRESUMO
Leptospira spp. are bacteria responsible for leptospirosis, a zoonotic disease with considerable impacts on the economy, animal health, and public health. This disease has a global distribution and is particularly prevalent in Brazil. Both rural and urban environments are habitats for Leptospira spp., which are primarily transmitted through contact with the urine of infected animals. Consequently, domestic and wild species can harbor these prokaryotes and serve as infection sources for other hosts. In the context of wild animals, there is a dearth of molecular studies elucidating the roles of various animal and bacterial species in the epidemiology of leptospirosis. Therefore, this study aimed to evaluate the presence of Leptospira spp. DNA in different species of free-living and captive wild animals and to assess the phylogenetic relationships of the identified microorganisms in Rio Grande do Sul, Brazil. The samples were evaluated for the presence of the gene lipL32 by polymerase chain reaction (PCR) and sequencing of the amplified fragment after which phylogenetic analyzes were carried out. DNA from Leptospira spp. was extracted from kidney tissue from wild animals (Mammalia class). Pathogenic Leptospira spp. DNA was detected in 9.6% (11/114) of the samples, originating from nine species of wild animals, including the white-eared opossum (Didelphis albiventris), skunk (Conepatus chinga), geoffroy's cat (Leopardus geoffroyi), margay (Leopardus wiedii), pampas fox (Lycalopex gymnocercus), capybara (Hydrochoerus hydrochaeris), common marmoset (Callithrix jacchus), neotropical river otter (Lontra longicaudis), and european hare (Lepus europaeus). Phylogenetic analysis revealed the presence of Leptospira borgpetersenii and Leptospira interrogans in these animals. This research is the first study contributing to the epidemiology of leptospirosis by identifying L. borgpetersenii and L. interrogans in free-living and captive wild animals in Rio Grande do Sul, Brazil, potentially acting as bacterial reservoirs. Additionally, our findings can inform sanitary measures for controlling and preventing the disease, thereby safeguarding public health.
Assuntos
Animais Selvagens , Leptospira interrogans , Leptospira , Leptospirose , Filogenia , Animais , Brasil/epidemiologia , Leptospirose/microbiologia , Leptospirose/veterinária , Leptospirose/epidemiologia , Animais Selvagens/microbiologia , Leptospira/genética , Leptospira/isolamento & purificação , Leptospira/classificação , Leptospira interrogans/genética , Leptospira interrogans/classificação , Leptospira interrogans/isolamento & purificação , Mamíferos/microbiologia , DNA Bacteriano/genéticaRESUMO
BACKGROUND: Leptospirosis is a zoonosis caused by pathogenic species of bacteria belonging to the genus Leptospira. Most studies infer the epidemiological patterns of a single serogroup or aggregate all serogroups to estimate overall seropositivity, thus not exploring the risks of exposure to distinct serogroups. The present study aims to delineate the demographic, socioeconomic and environmental factors associated with seropositivity of Leptospira serogroup Icterohaemorraghiae and serogroup Cynopteri in an urban high transmission setting for leptospirosis in Brazil. METHODS/PRINCIPAL FINDINGS: We performed a cross-sectional serological study in five informal urban communities in the city of Salvador, Brazil. During the years 2018, 2020 2021, we recruited 2.808 residents and collected blood samples for serological analysis using microagglutination assays. We used a fixed-effect multinomial logistic regression model to identify risk factors associated with seropositivity for each serogroup. Seropositivity to Cynopteri increased with each year of age (OR 1.03; 95% CI 1.01-1.06) and was higher in those living in houses with unplastered walls (exposed brick) (OR 1.68; 95% CI 1.09-2.59) and where cats were present near the household (OR 2.00; 95% CI 1.03-3.88). Seropositivity to Icterohaemorrhagiae also increased with each year of age (OR 1.02; 95% CI 1.01-1.03) and was higher in males (OR 1.51; 95% CI 1.09-2.10), in those with work-related exposures (OR 1.71; 95% CI 1.10-2.66) or who had contact with sewage (OR 1.42; 95% CI 1.00-2.03). Spatial analysis showed differences in distribution of seropositivity to serogroups Icterohaemorrhagiae and Cynopteri within the five districts where study communities were situated. CONCLUSIONS/SIGNIFICANCE: Our data suggest distinct epidemiological patterns associated with the Icterohaemorrhagiae and Cynopteri serogroups in the urban environment at high risk for leptospirosis and with differences in spatial niches. We emphasize the need for studies that accurately identify the different pathogenic serogroups that circulate and infect residents of low-income areas.
Assuntos
Leptospira interrogans , Leptospira , Leptospirose , Sorogrupo , Leptospirose/epidemiologia , Leptospirose/microbiologia , Leptospirose/transmissão , Brasil/epidemiologia , Humanos , Masculino , Adulto , Feminino , Estudos Transversais , Pessoa de Meia-Idade , Leptospira/classificação , Leptospira/imunologia , Leptospira/isolamento & purificação , Adulto Jovem , Adolescente , Leptospira interrogans/imunologia , Leptospira interrogans/classificação , Leptospira interrogans/isolamento & purificação , Fatores de Risco , Estudos Soroepidemiológicos , População Urbana , Anticorpos Antibacterianos/sangue , Animais , Criança , IdosoRESUMO
ECF (extracytoplasmic function) σ factors, members of the σ70-family, are the largest class of alternative σ factors which are stimulated in the presence of specific signals and direct RNA polymerase to transcribe a defined subset of genes. Thanks to them, bacterial pathogens can effectively reprogram their gene expression and, consequently, survive in the host and establish infection in a relatively short time. The number of ECF σ factors encoded within bacterial genomes is different depending on a given species and it reflects the likelihood that these bacteria will encounter harsh environmental conditions. The genome of L. interrogans, a zoonotic pathogen responsible for leptospirosis, is predicted to encode 11 ECF σE-type factors, but none of them have been characterized biochemically to date and their functions are still unknown. Here, we focused on one of the leptospiral ECF σ factors, namely LIC_12757, which was previously found to be up-regulated at elevated temperatures and may be related to the expression of clpB encoding an important L. interrogans virulence factor. We report cloning of the coding sequence of the LIC_12757 gene, its expression with the pET system and biochemical characterization of LIC_12757. By performing EMSA and in vitro transcription assays, we provide strong evidence that LIC_12757 indeed functions as a transcriptional factor that enables RNA polymerase to bind to the specific σE-type promoter and to initiate transcription. Interestingly, we demonstrate that LIC_12757 is autoregulated at the transcriptional level. Our study is a first step towards determining key aspects of LIC_12757 function in pathogenic Leptospira.
Assuntos
Proteínas de Bactérias , Regulação Bacteriana da Expressão Gênica , Leptospira interrogans , Fator sigma , Leptospira interrogans/genética , Fator sigma/genética , Fator sigma/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Leptospirose/microbiologia , Leptospirose/veterinária , Fatores de Virulência/genética , Regiões Promotoras Genéticas , Clonagem MolecularRESUMO
Bacterial caseinolytic protease-chaperone complexes participate in the elimination of misfolded and aggregated protein substrates. The spirochete Leptospira interrogans possess a set of Clp-chaperones (ClpX, ClpA, and ClpC), which may associate functionally with two different isoforms of LinClpP (ClpP1 and ClpP2). The L. interrogans ClpC (LinClpC) belongs to class-I chaperone with two active ATPase domains separated by a middle domain. Using the size exclusion chromatography, ANS dye binding, and dynamic light scattering analysis, the LinClpC is suggested to undergo nucleotide-induced oligomerization. LinClpC associates with either pure LinClpP1 or LinClpP2 isoforms non-preferentially and with equal affinity. Regardless, pure LinClpP isoforms cannot constitute an active protease complex with LinClpC. Interestingly, the heterocomplex LinClpP1P2 in association with LinClpC forms a functional proteolytic machinery and degrade ß-casein or FITC-casein in an energy-independent manner. Adding either ATP or ATPγS further fosters the LinClpCP1P2 complex protease activity by nurturing the functional oligomerization of LinClpC. The antibiotic, acyldepsipeptides (ADEP1) display a higher activatory role on LinClpP1P2 protease activity than LinClpC. Altogether, this work illustrates an in-depth study of hetero-tetradecamer LinClpP1P2 association with its cognate ATPase and unveils a new insight into the structural reorganization of LinClpP1P2 in the presence of chaperone, LinClpC to gain protease activity.
Assuntos
Proteínas de Bactérias , Proteínas de Choque Térmico , Leptospira , Multimerização Proteica , Trifosfato de Adenosina/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Endopeptidase Clp/metabolismo , Endopeptidase Clp/química , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/metabolismo , Leptospira/metabolismo , Leptospira/enzimologia , Leptospira interrogans/enzimologia , Leptospira interrogans/metabolismo , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/química , Ligação Proteica , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/química , ProteóliseRESUMO
Antimicrobial resistance (AMR) is one of the global health challenges of the 21st century. Data regarding AMR mechanisms in Leptospira interrogans, the causative agents of leptospirosis, have been relatively limited. Therefore, our study aimed to identify resistance genes and explore potential resistance mechanisms specific to particular serovars. We conducted a comprehensive analysis of 98 Leptospira strains, representing 10 common serovars, using whole-genome sequencing (WGS) FASTA files. Employing the PATRIC tool from the Bacterial and Viral Bioinformatics Resource Center (BV-BRC), we scrutinized the genomes for AMR genes. Our investigation revealed 32 genes associated with AMR, with 20 key genes consistently prevalent across most strains. Notably, we identified unique efflux pump systems in serovar Pomona, indicating distinctive resistance mechanisms in this serovar. In summary, our findings shed light on the genetic landscape of AMR in Leptospira, uncovering both common and serovar-specific resistance elements. The presence of unique efflux pump systems in serovar Pomona introduces a novel dimension to our understanding of resistance mechanisms. These insights underscore the importance of tailored intervention strategies and collaborative efforts between human and veterinary healthcare professionals, as well as environmental scientists, to address the complex dynamics of leptospirosis and its implications for antibiotic resistance.
Assuntos
Antibacterianos , Farmacorresistência Bacteriana , Genoma Bacteriano , Leptospira interrogans , Sorogrupo , Sequenciamento Completo do Genoma , Leptospira interrogans/genética , Genoma Bacteriano/genética , Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Leptospirose/microbiologia , Humanos , Biologia Computacional/métodos , Testes de Sensibilidade MicrobianaRESUMO
Leptospira interrogans serovar Hardjo is a long slender bacterium of size 0.1-0.3 µm × 5-50 µm. It is one of the major causes of bovine leptospirosis and is of economical importance because of the reproductive failure, still birth, abortion, and reduced productivity in cattle. It is also a zoonotic disease-causing infection in humans characterized by headaches, fever, chills, sweats and myalgia, lethargy, aching joints, pulmonary haemorrhages, and death in severe cases. Control of the disease involves antibiotic therapy, management and vaccination, of which immunization is the cheapest and effective means of disease prevention. The present study was developed to isolate and characterize the outer membrane vesicles of Leptospira interrogans serovar Hardjo and to evaluate their vaccine potential in guinea pig model. The OMVs were isolated from the culture by sonication and ultracentrifugation. In transmission electron microscopy, the isolated OMVs appeared as small spherical structures of 50-200 nm size. In Western blot and indirect ELISA, antibodies specific to OMVs were observed as indicative of a good humoral immune response elicited by L. interrogans serovar Hardjo OMV. The OMV-based Leptospira vaccine was able to prevent kidney lesions and renal colonization compared to the control and bacterin vaccinated group as proven by histopathology and PCR.
Assuntos
Vacinas Bacterianas , Leptospirose , Animais , Cobaias , Leptospirose/prevenção & controle , Leptospirose/imunologia , Leptospirose/microbiologia , Vacinas Bacterianas/imunologia , Modelos Animais de Doenças , Leptospira interrogans/imunologia , Membrana Externa Bacteriana/imunologia , Membrana Externa Bacteriana/metabolismo , Feminino , NanovacinasRESUMO
AIMS: Rat-associated zoonotic pathogen transmission at the human-wildlife interface is a public health concern in urban environments where Norway rats (Rattus norvegicus) thrive on abundant anthropogenic resources and live in close contact with humans and other animal species. To identify potential factors influencing zoonotic pathogen occurrence in rats, we investigated associations between environmental and sociodemographic factors and Leptospira interrogans and Bartonella spp. infections in rats from Windsor, Ontario, Canada, while controlling for the potential confounding effects of animal characteristics (i.e., sexual maturity and body condition). METHODS AND RESULTS: Between November 2018 and June 2021, 252 rats were submitted by collaborating pest control professionals. Kidney and spleen samples were collected for L. interrogans and Bartonella spp. PCR and sequencing, respectively. Of the rats tested by PCR, 12.7% (32/252) were positive for L. interrogans and 16.3% (37/227) were positive for Bartonella species. Associations between infection status and environmental and sociodemographic variables of interest were assessed via mixed multivariable logistic regression models with a random intercept for social group and fixed effects to control for sexual maturity and body condition in each model. The odds of L. interrogans infection were significantly higher in rats from areas with high building density (odds ratio [OR]: 3.76; 95% CI: 1.31-10.79; p = 0.014), high human population density (OR: 3.31; 95% CI: 1.20-9.11; p = 0.021), high proportion of buildings built in 1960 or before (OR: 11.21; 95% CI: 2.06-60.89; p = 0.005), and a moderate number of reports of uncollected garbage compared to a low number of reports (OR: 4.88; 95% CI: 1.01-23.63; p = 0.049). A negative association was observed between median household income and Bartonella spp. infection in rats (OR: 0.26; 95% CI: 0.08-0.89; p = 0.031). CONCLUSIONS: Due to the complexity of the ecology of rat-associated zoonoses, consideration of environmental and sociodemographic factors is of critical importance to better understand the nuances of host-pathogen systems and inform how urban rat surveillance and intervention efforts should be distributed within cities.
Assuntos
Infecções por Bartonella , Bartonella , Doenças dos Roedores , Zoonoses , Animais , Ratos , Ontário/epidemiologia , Infecções por Bartonella/veterinária , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/microbiologia , Bartonella/isolamento & purificação , Bartonella/genética , Doenças dos Roedores/microbiologia , Doenças dos Roedores/epidemiologia , Leptospirose/epidemiologia , Leptospirose/veterinária , Leptospirose/microbiologia , Humanos , Leptospira interrogans/isolamento & purificação , Masculino , Fatores Sociodemográficos , Feminino , Meio AmbienteRESUMO
Leptospira is a genus of bacteria that includes free-living saprophytic species found in water or soil, and pathogenic species, which are the etiologic agents of leptospirosis. Besides all the efforts, there are only a few proteins described as virulence factors in the pathogenic strain L. interrogans. This work aims to perform L. biflexa serovar Patoc1 strain Paris global proteome and to compare with the proteome database of pathogenic L. interrogans serovar Copenhageni strain Fiocruz L1-130. We identified a total of 2327 expressed proteins of L. biflexa by mass spectrometry. Using the Get Homologues software with the global proteome of L. biflexa and L. interrogans, we found orthologous proteins classified into conserved, low conserved, and specific proteins. Comparative bioinformatic analyses were performed to understand the biological functions of the proteins, subcellular localization, the presence of signal peptide, structural domains, and motifs using public softwares. These results lead to the selection of 182 low conserved within the saprophyte, and 176 specific proteins of L. interrogans. It is anticipated that these findings will indicate further studies to uncover virulence factors in the pathogenic strain. This work presents for the first time the global proteome of saprophytic strain L. biflexa serovar Patoc, strain Patoc1. SIGNIFICANCE: The comparative analysis established an array of specific proteins in pathogenic strain that will narrow down the identification of immune protective proteins that will help fight leptospirosis.
Assuntos
Leptospira interrogans , Leptospira , Leptospirose , Humanos , Proteoma/metabolismo , Fatores de Virulência/metabolismoRESUMO
Myalgia is a common symptom of Leptospira infection in humans. Autopsies have reported that muscle tissue shows degeneration and necrosis of the myofibers and infiltration of inflammatory cells composed mainly of macrophages and lymphocytes. It remains unclear whether Leptospira directly infects the muscle and how the infiltrating inflammatory cells are involved in muscle fiber destruction. This study evaluated the relationship between histopathological changes and leptospiral localization in the muscle tissue of a hamster model. The influence of macrophages in skeletal muscle injury was also investigated, using selective depletion of macrophages by administration of liposomal clodronate. Hamsters infected subcutaneously with Leptospira interrogans serovar Manilae strain UP-MMC-SM showed myositis of the thighs adjacent to the inoculated area beginning at 6 days post-infection. The myositis was non-purulent and showed sporadic degeneration and necrosis of muscle fibers. The degeneration of myofibers was accompanied by aggregations of macrophages. Immunofluorescence staining revealed leptospires surrounding the damaged muscle fibers. Subcutaneous injection of formalin-killed Leptospira or intraperitoneal injection of live Leptospira caused no myositis in hamster thighs. Liposomal clodronate treatment in infected hamsters reduced macrophage infiltration in muscle tissue without impacting bacterial clearance. Muscle necrosis was still observed in the infected hamsters treated with liposomal clodronate, and there was no significant change in serum creatine kinase levels compared to those in animals treated with liposomes alone. Our findings suggest that leptospiral invasion of muscle tissue from an inoculation site leads to the destruction of muscle fibers and causes non-purulent myositis, whereas the infiltrating macrophages contribute less to muscle destruction.
Assuntos
Leptospira interrogans , Leptospira , Leptospirose , Miosite , Cricetinae , Humanos , Animais , Ácido Clodrônico , Leptospirose/microbiologia , Músculo Esquelético/patologia , NecroseRESUMO
Bacterial motility is often a crucial virulence factor for pathogenic species. A common approach to study bacterial motility is fluorescent labeling, which allows detection of individual bacterial cells in a population or in host tissues. However, the use of fluorescent labeling can be hampered by protein expression stability and/or interference with bacterial physiology. Here, we apply machine learning to microscopic image analysis for label-free motion tracking of the zoonotic bacterium Leptospira interrogans on cultured animal cells. We use various leptospiral strains isolated from a human patient or animals, as well as mutant strains. Strains associated with severe disease, and mutant strains lacking outer membrane proteins (OMPs), tend to display fast mobility and reduced adherence on cultured kidney cells. Our method does not require fluorescent labeling or genetic manipulation, and thus could be applied to study motility of many other bacterial species.
Assuntos
Leptospira interrogans , Leptospira , Leptospirose , Spirochaeta , Animais , Humanos , Spirochaetales , Leptospirose/genética , Leptospirose/microbiologia , Leptospirose/patologia , Leptospira interrogans/genética , Proteínas de Membrana/metabolismoRESUMO
Introduction: A high incidence of human leptospirosis is recorded on Mayotte, an oceanic island located in southwestern Indian Ocean, but the severity of the disease appears relatively mild in terms of mortality rate and admission to the intensive care unit. It has been proposed that mild leptospirosis may result from a limited virulence of some of the occurring Leptospira species to which the population is exposed. Methods: Clinical and biological data of patients admitted to the Centre Hospitalier de Mayotte were collected and the infecting Leptospira species were determined through molecular typing. Results: Leptospira interrogans was detected in the minority of admitted patients but most of these patients suffered from severe forms, with 50% admitted to intensive care unit and suffering from organ failures. Nineteen percent of patients infected with Leptospira borgpetersenii were admitted to the intensive care, with 13% displaying organ failures, and one patient died. Leptospira mayottensis was found in 28% of the patients and not a single severe case was observed. Discussion: The distribution of Leptospira species in patients was not different from that reported 10-15 years ago and bacterial genotypes were very closely related to those previously reported. These results highlight the importance of the diversity of pathogenic Leptospira circulating on Mayotte island and are in keeping with distinct outcome of the disease depending on the infecting Leptospira. Altogether, presented data support that the infecting Leptospira species is an important driver of disease severity in humans.