Leptospira borgptersenii and Leptospira interrogans identified in wild mammals in Rio Grande do Sul, Brazil.

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.

Isolation and molecular identification of Leptospira santarosai and Leptospira interrogans in equines from eastern Mexico.

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.

Factors associated with differential seropositivity to Leptospira interrogans and Leptospira kirschneri in a high transmission urban setting for leptospirosis in Brazil.

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.

Leptospira interrogans serogroup Pomona strains isolated from river buffaloes.

At present, little is known regarding the prevalence of buffalo leptospirosis worldwide, especially with respect to which Leptospira strains may infect this animal species. Furthermore, most investigations into this disease in buffaloes have only been performed with serological studies. In Brazil, particularly in the Amazon, buffalo production is growing and is just as important as cattle production, although few studies have been performed on buffalo compared to cattle. Thus, the aim of this study was to isolate and characterise Leptospira strains from river buffaloes raised in the Brazilian Amazon region. We collected 109 kidney samples from slaughtered buffaloes raised in the Amazon Delta region of Brazil. The samples were analysed by bacteriological culture for the isolation of leptospires, and the obtained isolates were serologically and molecularly characterised by microscopic agglutination test (MAT), DNA sequencing and multiple locus variable-number tandem repeat analysis (MLVA). Five isolates were obtained, and in serogrouping analyses, these isolates were only reactive for the Pomona serogroup, with an observed titre of 25,600. The DNA sequencing results revealed that all the isolates belonged to the species Leptospira interrogans, and the MLVA results showed that the VNTR loci 4, 7 and 10 profile of all the isolates was 4-1-10. In this study, we observed that Pomona serogroup strains circulate in buffaloes in the Amazon, showing that in Brazil, buffaloes can be affected by Leptospira strains other than the Sejroe group, which are adapted to cattle.

Unusual case of polyarthritis and hepatorenal syndrome associated with Leptospira interrogans infection in a dog: A case report.

Leptospirosis is a zoonotic disease caused by spirochetal bacterial of the genus Leptospira affecting virtually all mammals. The infection has a broad range of effects, from mild clinical manifestation to multiple organ failure, and ultimately death. A 5-months-old male unvaccinated dog was admitted to the University Veterinary Teaching Hospital presenting dullness, dehydration, jaundiced mucous, bloody diarrhea, vomiting, and hyporexia. Microscopic agglutination test (MAT) detected serological titers of 1:1.600 for serogroup Canicola. After five days of monitoring by the medical team he developed fever and swelling of carpal and tarsal joints, accompanied by functional limitation. Initial antimicrobial treatment was instituted for leptospirosis. Polyarthritis responsiveness to glucocorticoid therapy was observed through decreasing signs of inflammation of the affected joints. The diagnosis of leptospirosis was further confirmed by molecular investigation for Leptospira spp. on blood and synovial fluid samples. Amplification and sequencing of the secY partial gene characterized the infective bacterial as Leptospira interrogans. From the 7th day the respiratory condition worsened and on Day 14 the patient evolved to death, when necropsy and histological evaluation were performed. Prominent anatomopathological findings included: fibrinous polyarthritis, bronchointerstitial pneumonia, intense hepatocyte dissociation, cholestasis, and periportal multifocal hepatitis, diffuse acute tubular necrosis, and significant dystrophic mineralization in the renal parenchyma, lungs, and atrial endocardium. Here, we present a case report of systemic clinical manifestations polyarthritis associated with the presence of leptospiras in the synovial fluid. We highlight the need for richer knowledge about the different clinical manifestations of leptospirosis.

Case Report: Fatal Human Leptospirosis Caused by Leptospira interrogans Genotype ST149.

Leptospirosis is an important zoonotic disease with worldwide distribution and nonspecific clinical manifestation. We report a case of fatal leptospirosis in a previously healthy woman with a causative agent. A young adult Indian woman was brought in dead to the forensic department. Ten days before, she developed fever, dizziness with headache, myalgia, diarrhea, and vomiting. Routine inquest and autopsy were performed on the deceased, revealing hemorrhagic lungs with extensive intra-alveolar hemorrhages, pale liver with dissociation and separation of hepatocyte plates, and edematous brain with histiocyte and lymphocyte infiltration in the parenchyma and meninges. Heart tissue depicts myocarditis and pericarditis inflammatory changes. Cerebrospinal fluid (CSF) was turbid in appearance with mildly elevated leukocytes, predominantly lymphocytes. Real-time PCR targeting lipL32 gene of pathogenic Leptospira was detected in the blood, CSF, brain, kidney, heart, and liver. The genetic profile of the causative agent was ST149 (multi-locus sequence typing Scheme 3). This study illustrates the usefulness of Leptospira PCR assay in postmortem diagnosis and addresses the need for further surveillance to identify the epidemiological link of the disease.

Serological study of Leptospira interrogans serovar Copenhageni and L. borgpetersenii serovars Tarassovi and Ballum in beef cattle, sheep and deer in New Zealand.

AIMS: To estimate animal-level seroprevalence of Leptospira interrogans serovar Copenhageni and L. borgpetersenii serovars Ballum and Tarassovi, in beef cattle, sheep and deer on New Zealand farms, and herd/flock-level seroprevalence of any serovar when existing same-sera data for serovars Hardjobovis and Pomona were included, and to determine associations between risk factors and animal-level seroprevalence. METHODS: Banked sera from sheep (n = 82), beef (n = 54) and deer (n = 62) herds/flocks (n = 3,878 animals) from seven regions were analysed using the microscopic agglutination test. Titres of ≥48 were designated positive. Herds/flocks were considered positive if either ≥1, ≥2 or ≥3 animals were positive. Existing same-sera data for serovars Hardjobovis and Pomona were included to establish farm-level any-serovar seropositivity. Factors associated with serological status were analysed using generalised estimating equations. RESULTS: Animal-level seroprevalence for serovars Ballum, Copenhageni, and Tarassovi, respectively, was 13.7 (95% CI = 11.7-16.0)%, 12.6 (95% CI = 10.6-14.7)% and 18.0 (95% CI = 15.7-20.5)% for beef cattle, 10.5 (95% CI = 9.0-12.1)%, 16.7 (95% CI = 14.9-18.6)% and 14.0 (95% CI = 12.4-15.8)% for sheep and 6.6 (95% CI = 5.3-8.2)%, 15.5 (95% CI = 13.5-17.7)% and 3.6 (95% CI = 2.7-4.8)% for deer, respectively. Herd/flock-level seroprevalence for Ballum was 86.6, 52.4 and 39.0% for sheep, 85.2, 52.7 and 33.3% for beef cattle and 50.8, 27.9 and 21.3% for deer at definitions ≥1, ≥2 and ≥3 seropositive animals per species, respectively. For Copenhageni, corresponding data were 95.1, 73.2 and 56.1% for sheep, 68.5, 48.2 and 29.6% for beef cattle and 73.8, 57.4 and 41.0% for deer, and for Tarassovi, 80.5, 59.7 and 45.1% for sheep, 83.3, 68.5 and 61.1% for beef cattle, and 42.6, 16.4 and 4.9% for deer. Seropositivity to all serovars was observed from all regions, with some differences in seroprevalence observed between species and regions, but not between islands. Combining with Hardjobovis and Pomona data, herd/flock-level seropositivity for all animal species and all five Leptospira serovars was 100% at definition ≥1 animal positive, and 97.5 and 96.3% for sheep flocks, 87.8 and 97.8% for beef cattle herds, and 89.3 and 75% for deer herds at ≥2 and ≥3 animals positive, respectively. CONCLUSIONS: Seropositivity to serovars Ballum, Copenhageni and Tarassovi is common in sheep, beef cattle and deer New Zealand and most, or all farms have ≥1 livestock species seropositive to ≥1 serovar. CLINICAL RELEVANCE: Serovars Ballum, Tarassovi and Copenhageni should be considered when clinical or subclinical signs of leptospirosis are observed in sheep, beef cattle or deer. Livestock sector workers are potentially at risk of exposure.

Extra-renal bovine leptospirosis: Molecular characterization of the Leptospira interrogans Sejroe serogroup on the uterus of non-pregnant cows.

Bovine genital leptospirosis is a chronic disease that causes reproductive disorders such as abortions, stillbirths, and estrus repetition, as well as economic losses. Despite clinical signs related to reproductive failure, the majority of studies have focused on the detection of Leptospira spp. in the urine, while few have considered the reproductive tract. Consequently, the aim of the present study was to investigate the uterus as an important extra-renal site of leptospiral infection in cows. A total of 42 non-pregnant cows were studied at a slaughterhouse. Blood samples and uterine fragments were collected for serology and molecular analysis, respectively. Concerning serologic results, 20.5 % presented as reactive, all of them against the Sejroe serogroup. Regarding lipL32 PCR, 26.2 % (11/42) of samples were positive for pathogenic Leptospira sp. Sequencing the secY gene short region enabled nine strains to be characterized, all of which were L. interrogans, with high identity (98.8 %-99.8 %) with serovar Hardjo. The use of molecular tools substantially improved the sensitivity of Leptospira sp. detection at species level and demonstrated that the uterus is an important site of bovine leptospiral infection. The findings of the present study reinforce our understanding that leptospiral uterine infection are associated to members of the Sejroe serogroup.

Leptospira interrogans in wild Boa constrictor snakes from Northeast Brazil peri­urban rainforest fragments.

Leptospirosis, a disease that occurs worldwide, especially in tropical regions, is caused by bacteria of the genus Leptospira and affects mammals, amphibians, and reptiles. Boa constrictor snakes are commonly found in Atlantic rainforest fragments in peri­urban areas, which indicates a greater possibility of the contact of these animals with humans residing there. Therefore, the aim of this work was to detect Leptospira spp infection through molecular assays in wild B. constrictor snakes rescued in peri­urban areas and verify seroreactivity, by the microscopic agglutination test (MAT), as well as the most common serogroups. Among the 46 samples tested, 7 (15.21%) were positive according to PCR and confirmed as Leptospira interrogans through secY gene sequencing. In MAT, 37 (80.43%) of the 46 samples were classified as reactive. Panama was the serogroup with the highest occurrence. The results showed the presence of Leptospira spp DNA in asymptomatic snakes rescued in rainforest fragments located in peri­urban areas and support further investigations on the influence of these animals in the epidemiology of leptospirosis in tropical peri­urban areas.

Novel genotypes of Leptospira interrogans serogroup Sejroe isolated from human patients in Okinawa Prefecture, Japan.

Leptospirosis is a zoonotic disease caused by pathogenic spirochetes of Leptospira species. It is a public health issue in the tropics, including Okinawa, the southernmost prefecture of Japan. This study reports the first isolation of L. interrogans serogroup Sejroe from two human patients in Japan, and describes its molecular characterization using multilocus sequence typing (MLST) and multiple-locus variable-number tandem repeat analysis (MLVA). MLST on the two isolates, 168036 and 178129, showed that pfkB in 178129 is a novel allele, and that both isolates constitute novel sequence types (STs); ST286 for 168036 and ST287 for 178129. A minimum spanning tree based on seven alleles of L. interrogans indicates that both isolates are genetically close, but are distinct from known L. interrogans serogroup Sejroe strains. MLVA using 11 loci demonstrated that seven of the 11 loci were identical between the two isolates, whereas the identity between the isolates and the seven reference strains of L. interrogans serogroup Sejroe was zero to three loci. These results indicate that the isolates investigated in this study have novel genotypes, and are genetically closest to each other among the known L. interrogans serogroup Sejroe strains.

M16-Type Metallopeptidases Are Involved in Virulence for Invasiveness and Diffusion of Leptospira interrogans and Transmission of Leptospirosis.

BACKGROUND: Leptospirosis is a global zoonotic infectious disease caused by Leptospira interrogans. The pathogen rapidly invades into hosts and diffuses from bloodstream into internal organs and excretes from urine to cause transmission of leptospirosis. However, the mechanism of leptospiral invasiveness remains poorly understood. METHODS: Proteolytic activity of M16-type metallopeptidases (Lep-MP1/2/3) of L. interrogans was determined by spectrophotometry. Expression and secretion of Lep-MP1/2/3 during infection of cells were detected by quantitative reverse-transcription polymerase chain reaction, Western blot assay, and confocal microscopy. Deletion and complementation mutants of the genes encoding Lep-MP1/2/3 were generated to determine the roles of Lep-MP1/2/3 in invasiveness using transwell assay and virulence in hamsters. RESULTS: Leptospira interrogans but not saprophytic Leptospira biflexa strains were detectable for Lep-MP-1/2/3-encoding genes. rLep-MP1/2/3 hydrolyzed extracellular matrix proteins, but rLep-MP1/3 displayed stronger proteolysis than rLep-MP2, with 123.179/340.136 µmol/L Km and 0.154/0.159 s-1 Kcat values. Expression, secretion and translocation of Lep-MP1/2/3 during infection of cells were increased. ΔMP1/3 but not ΔMP2 mutant presented attenuated transmigration through cell monolayers, decreased leptospiral loading in the blood, lungs, liver, kidneys, and urine, and 10/13-fold decreased 50% lethal dose and milder histopathologic injury in hamsters. CONCLUSIONS: Lep-MP1 and 3 are involved in virulence of L. interrogans in invasion into hosts and diffusion in vivo, and transmission of leptospirosis.

Leptospirosis in Northern Sea Otters (Enhydra lutris kenyoni) from Washington, USA.

We diagnosed leptospirosis in six northern sea otters (Enhydra lutris kenyoni) that stranded on beaches in Washington State, US in 2002. Significant gross findings included cyanotic oral mucous membranes, renal swelling, congestion or pale streaks on the cut surface of renal lobules, hematuria, dehydration, lymphadenopathy, pulmonary congestion, and rarely adrenal hemorrhage and congestion. Histopathology showed lymphoplasmacytic tubulointerstitial nephritis with intraluminal spirochetes and immunoreactivity to leptospiral antigens in the renal tubules and interstitium. A quantitative polymerase chain reaction (qPCR) using kidney or urine for the leptospiral lipL32 gene was positive with cycle threshold values indicative of abundant or moderate amounts of nucleic acid. A microscopic agglutination test showed the highest serum antibody titer to serovar Pomona and positive titers to serovars Autumnalis, Bratislava, Hebdomadis, Grippotyphosa, Icterohaemorrhagiae, Pyrogenes, Ballum, Canicola, and Hardjo. Although antibodies to Leptospira interrogans have been previously detected in sea otters, this report describes the pathology of leptospirosis in diseased free-ranging sea otters.

A canine vaccine against Leptospira serovars Icterohaemorrhagiae, Canicola and Grippotyphosa provides cross protection against Leptospira serovar Copenhageni.

Efficacy of the Leptospira components of multivalent vaccine DAPPi-L was previously demonstrated against virulent challenge with three serovars of Leptospira interrogans (Canicola, Icterohaemorrhagiae and Grippotyphosa) carried out 14 days after primary vaccination. In this study we demonstrate that this vaccine provides, two weeks after vaccination, an additional protection (prevention of mortality, clinical signs, renal infection, bacterial excretion, renal carriage and renal lesions) against fatal leptospirosis due to Leptospira interrogans serovar Copenhageni (serovar of major medical importance).

Development of antibody ELISA specific of Leptospira interrogans serovar Grippotyphosa, Canicola, and Icterohaemorrhagiae to monitor vaccine immunogenicity.

The antibody response after primary vaccination and annual revaccination with a multivalent DAPPi-L vaccine was assessed respectively in SPF dogs and in client owned dogs against the Grippotyphosa (Lg), Canicola (Lc) and Icterohaemorrhagiae (Li) Leptospira serovars. To overcome limitations of the microscopic agglutination test (MAT), we developed serovar-specific and sensitive blocking ELISA assays. Serovar-specific antibodies against Lg, Lc and Li were detected in 100%, 45% and 91% of dogs, respectively, after the first dose of vaccine, and in 100% of dogs for all serovars after the second dose. In addition, mean ELISA antibody titers increased 14 days after annual revaccination with most dogs remaining ELISA antibody positive against Lg (85.3%), Lc (90%) and Li (100%). Parallel testing of sera from the annual revaccination study in the MAT and ELISA assays resulted in an overall agreement of 72%, 67%, 77% of samples for Lg, Lc and Li serovars, respectively. More sera tested positive by ELISA than by MAT, suggesting that the ELISA assay is more sensitive than the MAT. These three new antibody-based assays are the first suitable and reliable ELISA assays for the assessment of the canine antibody response following vaccination and an attractive alternative to the MAT.

Presence of Antibodies Against Leptospira interrogans Serovar hardjo in Serum Samples from Cattle in Ukraine.

The article presents data on serological studies of 573 sera samples of cattle that were collected from the farms affected by leptospirosis in different regions of Ukraine in the period of 2014-2015. Samples were investigated by the microscopic agglutination test (MAT), which was conducted within eight serological groups of Leptospira and nine serovars: Sejroe (serovars polonica and hardjo), Hebdomadis (serovar kabura), Tarassovi (serovar tarassovi), Pomona (serovar pomona), Grippotyphosa (serovar grippotyphosa), Canicola (serovar canicola), Icterohaemorrhagiae (serovar copenhageni), and Australis (serovar bratislava). The circulation of L. interrogans serovar hardjo among cattle has been observed in all 11 regions of Ukraine investigated within 25.8-60.0% of the leptospirosis-positive serum samples in these regions. Antibodies in the cattle sera against serovar hardjo (serogroup Sejroe) were detected in 139 of the 370 cows reacting positively in MAT. Overall, they were detected in 24.3% animals out of the total of 573 cows investigated. These are the preliminary results, however, in our opinion, they should allow to include the serovar hardjo in a standard panel of strains for MAT in Ukraine.The article presents data on serological studies of 573 sera samples of cattle that were collected from the farms affected by leptospirosis in different regions of Ukraine in the period of 2014­2015. Samples were investigated by the microscopic agglutination test (MAT), which was conducted within eight serological groups of Leptospira and nine serovars: Sejroe (serovars polonica and hardjo), Hebdomadis (serovar kabura), Tarassovi (serovar tarassovi), Pomona (serovar pomona), Grippotyphosa (serovar grippotyphosa), Canicola (serovar canicola), Icterohaemorrhagiae (serovar copenhageni), and Australis (serovar bratislava). The circulation of L. interrogans serovar hardjo among cattle has been observed in all 11 regions of Ukraine investigated within 25.8­60.0% of the leptospirosis-positive serum samples in these regions. Antibodies in the cattle sera against serovar hardjo (serogroup Sejroe) were detected in 139 of the 370 cows reacting positively in MAT. Overall, they were detected in 24.3% animals out of the total of 573 cows investigated. These are the preliminary results, however, in our opinion, they should allow to include the serovar hardjo in a standard panel of strains for MAT in Ukraine.

A Novel Genotype of Leptospira interrogans Recovered from Leptospirosis Outbreak Samples from Southern Thailand.

We performed Leptospira culture analysis of 76 clinical samples collected from animals and of six soil samples for the investigation of a leptospirosis outbreak in southern Thailand in 2017. Leptospires were recovered from a kidney sample (a fatal canine leptospirosis case) and from all the soil samples. Next, 16S rRNA sequence analysis demonstrated that the clinical isolate was closely related to the pathogenic L. interrogans, whereas the soil isolates represented different species, including pathogenic L. ellisii, intermediate L. wolffii, and nonpathogenic L. yanagawae. Multilocus sequence typing identified an isolate of L. interrogans as a novel sequence type (ST263), suggesting that the causative agent of the canine leptospirosis in the southern Thailand outbreak has a unique genetic profile.

Severe Myasthenic Manifestation of Leptospirosis Associated with New Sequence Type of Leptospira interrogans.

We report the rapid development of a myasthenic crisis as the first-time manifestation of myasthenia gravis. The symptoms developed in the course of acute leptospirosis associated with a new sequence type of Leptospira interrogans. Antibiotic treatment led to rapid amelioration of myasthenia.

Characterization of CRISPR-Cas systems in Leptospira reveals potential application of CRISPR in genotyping of Leptospira interrogans.

Leptospirosis is a zoonotic disease caused by pathogenic Leptospira. However, understanding of the pathogenic mechanism of Leptospira is still elusive due to the limited number of genetic tools available for this microorganism. Currently, the reason for the genetic inaccessibility of Leptospira is still unknown. It is well known that as an acquired immunity of bacteria, Clustered Regularly Interspaced Short Palindromic Repeat-CRISPR-associated gene (CRISPR-Cas) systems can help bacteria against invading mobile genetic elements. In this study, the occurrence and diversity of CRISPR-Cas systems in 41 genomes of Leptospira strains were investigated. Three subtypes (subtype I-B, subtype I-C and subtype I-E) of CRISPR-Cas systems were identified in both pathogenic and intermediate Leptospira species but not in saprophytic species. Noteworthy, the majority of pathogenic species harbor two different types of CRISPR-Cas systems (subtype I-B and subtype I-E). Furthermore, Cas2 protein of subtype I-C in L. interrogans exhibited a metal-dependent DNase activity in a nonspecific manner. CRISPR spacers in subtype I-B are highly conserved within the same serovars and hypervariable across different serovars of L. interrogans. Based on the subtype I-B CRISPR arrays, the serotypes of different L. interrogans strains were easily identified. Investigation of the origin of CRISPR spacers showed that 192 spacers (23.5%) matched to mobile genetic elements, indicating CRISPR-Cas systems may play an important role in the defense of foreign invading DNA.

Genetic diversity of Leptospira interrogans circulating isolates and vaccine strains in China from 1954-2014.

Leptospirosis is one of the most important but neglected, infectious tropical diseases worldwide. Leptospira interrogans is now recognized as a leading cause of the disease. Little is known of the genetic diversity and phylogenetic characteristics of L. interrogans within China. To better understand the transmission and genetic diversity of L. interrogans populations, we characterized 271 isolates and seven vaccine strains from China during 1954-2014 using multilocus variable-number tandem repeat analysis (MLVA). 110 different L. interrogans MLVA profiles (MTs) were identified, of which five were predominant, reflecting a high level of genetic diversity in L. interrogans population in China. Different from that of circulating isolates, seven vaccine strains have different MT, of which some are phylogenetically away from the circulating isolates. The results showed that Icterohaemorrhagiae, Hebdomadis, and Canicola ranked as the top three serogroups among L. interrogans strains tested. The cluster analysis demonstrate the clonal links between rodent and human isolates, suggesting the rodent species played a key role in the transmission of leptospirosis to humans, and contributed to the circulation of the pathogen in humans. Taken together, these findings should provide insight into a better knowledge of the epidemiology and molecular evolution of L. interrogans in China. Furthermore, the results should facilitate the selection of candidate vaccine strains in the future.

Analysis of sequencing strategies and tools for taxonomic annotation: Defining standards for progressive metagenomics.

Metagenomics research has recently thrived due to DNA sequencing technologies improvement, driving the emergence of new analysis tools and the growth of taxonomic databases. However, there is no all-purpose strategy that can guarantee the best result for a given project and there are several combinations of software, parameters and databases that can be tested. Therefore, we performed an impartial comparison, using statistical measures of classification for eight bioinformatic tools and four taxonomic databases, defining a benchmark framework to evaluate each tool in a standardized context. Using in silico simulated data for 16S rRNA amplicons and whole metagenome shotgun data, we compared the results from different software and database combinations to detect biases related to algorithms or database annotation. Using our benchmark framework, researchers can define cut-off values to evaluate the expected error rate and coverage for their results, regardless the score used by each software. A quick guide to select the best tool, all datasets and scripts to reproduce our results and benchmark any new method are available at https://github.com/Ales-ibt/Metagenomic-benchmark . Finally, we stress out the importance of gold standards, database curation and manual inspection of taxonomic profiling results, for a better and more accurate microbial diversity description.