Serodiagnosis of leprosy and follow-up of household contacts using a commercial rapid test containing ND-O/LID-1 antigens

Introduction: Early diagnosis of leprosy, including the diagnosis of sub-clinical disease in contacts of known cases, would be a major advance. The signs of early leprosy are often difficult to assess and a reliable diagnostic test could play an important role in identifying cases and thus reducing transmission. Methods: Subjects were recruited at Centro de Saude Jardim Guanabara, Rondonopolis, Mato Grosso State, Brazil. Sera were obtained from 174 recently diagnosed leprosy patients, 409 household contacts, 53 endemic controls and 12 patients with active TB. 186 of the household contacts were re-tested on subsequent visits. Both the NDO-LID rapid test (Orange Life, Rio de Janeiro, Brazil) and an anti-PGL-1 ELISA test were used on all samples for comparison. Results: LID-1 was positive in 25 of 125 PB cases (20%), but in 41 of 49 MB cases (83.7%), while the anti-PGL-1 ELISA was positive in 8 (6.4%) and 35 (71.4%) cases, respectively. Specificity for LID-1 was 85% and for the anti-PGL-1 ELISA, 97%. Amongst household contacts, 9 of 409 (2.2%) were positive on the LID-1 test, and a further 51 (115%) were weakly positivealthough 9 cases of leprosy were subsequently identified amongst these contacts, 6 had tested negative, 2 were weakly positive and only 1 had been positive. Conclusion: The ND-O-LID-1 assay can be easily performed by the addition of serum to the assay device, and thus can be applied in resource-poor settings. The test was found to be useful for the detection of multibacillary cases, which in the long term will help to reduce transmission of the disease. This is a specific test, but it is not very sensitive for early detection of leprosy in household contacts and paucibacillary forms.Inst Lauro Souza Lime, Bauru, SP, BrazilHlth Ctr Rondonopolis MT, Rondonopolis, BrazilFed Univ Sao Paulo Unifesp, Dept Dermatol, Sao Paulo, BrazilUniv Fed Sao Paulo Unifesp, Dept Dermatol, Sao Paulo, BrazilWeb of Scienc

Acoustic analysis of vocalization and the behavioral response associated to sound production of the nine banded armadillo Dasypus novemcinctus (Mammalia, Cingulata, Dasypodidae) in an agonistic context

Although communication capabilities are displayed by many vertebrate groups, some repertoires are poorly known, such as the case of xenarthrans, particularly armadillos, for which vocalization as a source of communicating to others remains poorly understood and relies on punctual reports of sounds. Here we provide the first description of a behavioral response associated with sound emission of two subjects of Dasypus novemcinctus. Both audio and visual registration was performed to subsequent analyses of expressed behaviors and emitted calls, which accounted for 76 vocalizations from a total of eight video recordings randomly collected from 2017 to 2019. Sound is acoustically characterized by both inhale and exhale phases composed of two vocal units, and no harmonic structure was observed. Once the subjects have always produced these vocalizations while cornered and exhibiting defensive behavior against another subject/human disturbance, these vocalizations were termed as distress. Subjects produced a hiss-purr-like sound while trying to avoid contact with another by bowing or lowering their bodies, humping, or even moving elsewhere when sound production ceased. This shows that the sound repertoire of armadillos is still to be unveiled and seems to be much more complex than previously thought

High rates of undiagnosed leprosy and subclinical infection amongst school children in the Amazon Region

Leprosy in children is correlated with community-level factors, including the recent presence of disease and active foci of transmission in the community. We performed clinical and serological examinations of 1,592 randomly selected school children (SC) in a cross-sectional study of eight hyperendemic municipalities in the Brazilian Amazon Region. Sixty-three (4%) SC, with a mean age of 13.3 years (standard deviation = 2.6), were diagnosed with leprosy and 777 (48.8%) were seropositive for anti-phenolic glycolipid-I (PGL-I). Additionally, we evaluated 256 household contacts (HHCs) of the students diagnosed with leprosy; 24 (9.4%) HHC were also diagnosed with leprosy and 107 (41.8%) were seropositive. The seroprevalence of anti-PGL-I was significantly higher amongst girls, students from urban areas and students from public schools (p < 0.0001). Forty-five (71.4%) new cases detected amongst SC were classified as paucibacillary and 59 (93.6%) patients did not demonstrate any degree of physical disability at diagnosis. The results of this study suggest that there is a high rate of undiagnosed leprosy and subclinical infection amongst children in the Amazon Region. The advantages of school surveys in hyperendemic areas include identifying leprosy patients at an early stage when they show no physical disabilities, preventing the spread of the infection in the community and breaking the chain of transmission.CNPq [576425/2008-7]CNPqFAPESPAFAPESPASESPASESPAUFPAUFP

Ticks as potential vectors of Mycobacterium leprae: Use of tick cell lines to culture the bacilli and generate transgenic strains.

Leprosy is an infectious disease caused by Mycobacterium leprae and frequently resulting in irreversible deformities and disabilities. Ticks play an important role in infectious disease transmission due to their low host specificity, worldwide distribution, and the biological ability to support transovarial transmission of a wide spectrum of pathogens, including viruses, bacteria and protozoa. To investigate a possible role for ticks as vectors of leprosy, we assessed transovarial transmission of M. leprae in artificially-fed adult female Amblyomma sculptum ticks, and infection and growth of M. leprae in tick cell lines. Our results revealed M. leprae RNA and antigens persisting in the midgut and present in the ovaries of adult female A. sculptum at least 2 days after oral infection, and present in their progeny (eggs and larvae), which demonstrates the occurrence of transovarial transmission of this pathogen. Infected tick larvae were able to inoculate viable bacilli during blood-feeding on a rabbit. Moreover, following inoculation with M. leprae, the Ixodes scapularis embryo-derived tick cell line IDE8 supported a detectable increase in the number of bacilli for at least 20 days, presenting a doubling time of approximately 12 days. As far as we know, this is the first in vitro cellular system able to promote growth of M. leprae. Finally, we successfully transformed a clinical M. leprae isolate by inserting the reporter plasmid pCHERRY3; transformed bacteria infected and grew in IDE8 cells over a 2-month period. Taken together, our data not only support the hypothesis that ticks may have the potential to act as a reservoir and/or vector of leprosy, but also suggest the feasibility of technological development of tick cell lines as a tool for large-scale production of M. leprae bacteria, as well as describing for the first time a method for their transformation

Toxoplasma gondii and Leptospira spp. infection in free-ranging armadillos

Armadillos are primitive mammals used as food, mostly in rural areas. These animals may be sources of toxoplasmosis and leptospirosis infection for humans, but there is little information about their potential risk as reservoirs. In order to determine the prevalence of armadillos infected by Toxoplasma gondii and Leptospira spp., serum samples of 31 nine-banded armadillos (Dasypus novemcinctus), three six-banded armadillos (Euphractus sexcinctus), two naked-tailed armadillos (Cabassous tatouay) and two long-nosed armadillos (D. hybridus), captured in the mid-west region of the state of São Paulo, were analyzed for leptospirosis using the Microscopic Agglutination Test (MAT-l), and for toxoplasmosis using the Modified Agglutination Test (MAT-t). Only 4/31 (12.90%) nine-banded armadillos were positive for T gondii, while 3/31 (9.68%) nine-banded armadillos and 1/3 (33.33%) six-banded armadillos presented antibodies to Leptospira spp., demonstrating the potential risk of T gondii and Leptospira spp. transmission to humans, mainly due the habit of eating the meat of these animals in rural areas. (C) 2008 Elsevier B.V. All rights reserved.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Antimicrobial resistance among leprosy patients in Brazil : real-world data based on the national surveillance plan

Brazil ranks second among countries for new cases and first for relapse cases of leprosy worldwide. The Mycobacterium leprae Resistance Surveillance Plan was established. We aimed to present the results of a 2-year follow-up of the National Surveillance Plan in Brazil. A cross-sectional study of leprosy cases was performed to investigate antimicrobial resistance (AMR) in Brazil from October 2018 to September 2020. Molecular screening targeting genes related to dapsone (folP1), rifampin (rpoB), and ofloxacin resistance (gyrA) was performed. During the referral period, 63,520 active leprosy patients were registered in Brazil, and 1,183 fulfilled the inclusion criteria for molecular AMR investigation. In total, only 16 (1.4%) patients had genetic polymorphisms associated with AMR. Of these, 8 (50%) had cases of leprosy relapse, 7 (43.8%) had cases of suspected therapeutic failure with standard treatment, and 1 (6.2%) was a case of new leprosy presentation. M. leprae strains with AMR-associated mutations were found for all three genes screened. Isolates from two patients showed simultaneous resistance to dapsone and rifampin, indicating multidrug resistance (MDR). No significant relationship between clinical variables and the presence of AMR was identified. Our study revealed a low frequency of AMR in Brazil. Isolates were resistant mainly to dapsone, and a very low number of isolates were resistant to rifampin, the main bactericidal agent for leprosy, or presented MDR, reinforcing the importance of the standard World Health Organization multidrug therapy. The greater frequency of AMR among relapsed patients supports the need to constantly monitor this group

Leprosy survey among rural communities and wild armadillos from Amazonas state, Northern Brazil.

There is evidence that in southern US, leprosy is a zoonosis infecting wild Dasypus novemcinctus armadillos but the extent of this finding is unknown. This ecological study investigated leprosy in rural communities and in wild armadillos from the Brazilian Amazon. The study area was the Mamiá Lake of Coari municipality, Amazonas State, Northern region, a hyper endemic leprosy area where residents live on subsistence farming, fishing and armadillo hunting and its meat intake are frequent. The leprosy survey was conducted in sixteen communities by a visiting team of specialists. Local partakers provided wild armadillos to investigate M. leprae infection. Volunteers had complete dermato-neurological examination by a dermatologist with expertise in leprosy diagnosis, suspect skin lesions were biopsied for histopathology (Hematoxylin-eosin/HE, Fite-Faraco/FF staining); slit skin smears were collected. Armadillos' tissue fragments (skins, spleens, livers, lymph nodes, adrenal glands, others) were prepared for histopathology (HE/FF) and for M. leprae repetitive element-RLEP-qPCR. Among 176 volunteers, six new indeterminate leprosy cases were identified (incidence = 3.4%). Suspect skin sections and slit skin smears were negative for bacilli. Twelve wild D. novemcinctus were investigated (48 specimens/96 slides) and histopathological features of M. leprae infection were not found, except for one skin presenting unspecific inflammatory infiltrate suggestive of indeterminate leprosy. Possible traumatic neuroma, granuloma with epithelioid and Langhans cells, foreign-body granuloma were also identified. Granulomatous/non-granulomatous dermatitides were periodic-acid-Schiff/PAS negative for fungus. M. leprae-RLEP-qPCR was negative in all armadillos' tissues; no bacillus was found in histopathology. Our survey in rural communities confirmed the high endemicity for leprosy while one armadillo was compatible with paucibacillary M. leprae infection. At least in the highly endemic rural area of Coari, in the Brazilian Amazon region where infectious sources from untreated multibacillary leprosy are abundant, M. leprae infected armadillos may not represent a major source of infection nor a significant public health concern

Autophagy Is an Innate Mechanism Associated with Leprosy Polarization

Leprosy is a chronic infectious disease that may present different clinical forms according to the immune response of the host. Levels of IFN-gamma are significantly raised in paucibacillary tuberculoid (T-lep) when compared with multibacillary lepromatous (L-lep) patients. IFN-gamma primes macrophages for inflammatory activation and induces the autophagy antimicrobial mechanism. The involvement of autophagy in the immune response against Mycobacterium leprae remains unexplored. Here, we demonstrated by different autophagic assays that LC3-positive autophagosomes were predominantly observed in T-lep when compared with L-lep lesions and skin-derived macrophages. Accumulation of the autophagic receptors SQSTM1/p62 and NBR1, expression of lysosomal antimicrobial peptides and colocalization analysis of autolysosomes revealed an impairment of the autophagic flux in L-lep cells, which was restored by IFN-gamma or rapamycin treatment. Autophagy PCR array gene-expression analysis revealed a significantly upregulation of autophagy genes (BECN1, GPSM3, ATG14, APOL1, and TPR) in T-lep cells. Furthermore, an upregulation of autophagy genes (TPR, GFI1B and GNAI3) as well as LC3 levels was observed in cells of L-lep patients that developed type 1 reaction (T1R) episodes, an acute inflammatory condition associated with increased IFN-gamma levels. Finally, we observed increased BCL2 expression in L-lep cells that could be responsible for the blockage of BECN1-mediated autophagy. In addition, in vitro studies demonstrated that dead, but not live M. leprae can induce autophagy in primary and lineage human monocytes, and that live mycobacteria can reduce the autophagy activation triggered by dead mycobacteria, suggesting that M. leprae may hamper the autophagic machinery as an immune escape mechanism. Together, these results indicate that autophagy is an important innate mechanism associated with the M. leprae control in skin macrophages