Mitochondrial variants of complex I genes associated with leprosy clinical subtypes.

Leprosy is a chronic bacterial infection mainly caused by Mycobacterium leprae that primarily affects skin and peripheral nerves. Due to its ability to absorb carbon from the host cell, the bacillus became dependent on energy production, mainly through oxidative phosphorylation. In fact, variations in genes of Complex I of oxidative phosphorylation encoded by mtDNA have been associated with several diseases in humans, including bacterial infections, which are possible influencers in the host response to leprosy. Here, we investigated the presence of variants in the mtDNA genes encoding Complex I regarding leprosy, as well as the analysis of their pathogenicity in the studied cohort. We found an association of 74 mitochondrial variants with either of the polar forms, Pole T (Borderline Tuberculoid) or Pole L (Borderline Lepromatous and Lepromatous) of leprosy. Notably, six variants were exclusively found in both clinical poles of leprosy, including m.4158A>G and m.4248T>C in MT-ND1, m.13650C>A, m.13674T>C, m.12705C>T and m.13263A>G in MT-ND5, of which there are no previous reports in the global literature. Our observations reveal a substantial number of mutations among different groups of leprosy, highlighting a diverse range of consequences associated with mutations in genes across these groups. Furthermore, we suggest that the six specific variants exclusively identified in the case group could potentially play a crucial role in leprosy susceptibility and its clinical differentiation. These variants are believed to contribute to the instability and dysregulation of oxidative phosphorylation during the infection, further emphasizing their significance.

Whole mitogenome sequencing uncovers a relation between mitochondrial heteroplasmy and leprosy severity.

BACKGROUND: In recent years, the mitochondria/immune system interaction has been proposed, so that variants of mitochondrial genome and levels of heteroplasmy might deregulate important metabolic processes in fighting infections, such as leprosy. METHODS: We sequenced the whole mitochondrial genome to investigate variants and heteroplasmy levels, considering patients with different clinical forms of leprosy and household contacts. After sequencing, a specific pipeline was used for preparation and bioinformatics analysis to select heteroplasmic variants. RESULTS: We found 116 variants in at least two of the subtypes of the case group (Borderline Tuberculoid, Borderline Lepromatous, Lepromatous), suggesting a possible clinical significance to these variants. Notably, 15 variants were exclusively found in these three clinical forms, of which five variants stand out for being missense (m.3791T > C in MT-ND1, m.5317C > A in MT-ND2, m.8545G > A in MT-ATP8, m.9044T > C in MT-ATP6 and m.15837T > C in MT-CYB). In addition, we found 26 variants shared only by leprosy poles, of which two are characterized as missense (m.4248T > C in MT-ND1 and m.8027G > A in MT-CO2). CONCLUSION: We found a significant number of variants and heteroplasmy levels in the leprosy patients from our cohort, as well as six genes that may influence leprosy susceptibility, suggesting for the first time that the mitogenome might be involved with the leprosy process, distinction of clinical forms and severity. Thus, future studies are needed to help understand the genetic consequences of these variants.

Specialized active leprosy search strategies in an endemic area of the Brazilian Amazon identifies a hypermutated Mycobacterium leprae strain causing primary drug resistance.

Introduction: Leprosy, an infectious disease caused by Mycobacterium leprae, remains a public health concern in endemic countries, particularly in Brazil. In this study, we conducted an active surveillance campaign in the hyperendemic city of Castanhal in the northeastern part of the state of Pará using clinical signs and symptoms combined with serological and molecular tools to diagnose new cases and to identify drug resistance of circulating M. leprae strains and their distribution in the community. Methods: During an active surveillance of one week, we enrolled 318 individuals using three different strategies to enroll subjects for this study: (i) an active survey of previously treated cases from 2006 to 2016 found in the Brazil National Notifiable Disease Information System database (n = 23) and their healthy household contacts (HHC) (n = 57); (ii) an active survey of school children (SC) from two primary public schools in low-income neighborhoods (n = 178), followed by visits to the houses of these newly diagnosed SC (n = 7) to examine their HHC (n = 34) where we diagnosed additional new cases (n = 6); (iii) and those people who spontaneously presented themselves to our team or the local health center with clinical signs and/or symptoms of leprosy (n = 6) with subsequent follow-up of their HHC when the case was confirmed (n = 20) where we diagnosed two additional cases (n = 2). Individuals received a dermato-neurological examination, 5 ml of peripheral blood was collected to assess the anti-PGL-I titer by ELISA and intradermal earlobe skin scrapings were taken from HHC and cases for amplification of the M. leprae RLEP region by qPCR. Results: Anti-PGL-I positivity was highest in the new leprosy case group (52%) followed by the treated group (40.9%), HHC (40%) and lowest in SC (24.6%). RLEP qPCR from SSS was performed on 124 individuals, 22 in treated cases, 24 in newly diagnosed leprosy cases, and 78 in HHC. We detected 29.0% (36/124) positivity overall in this sample set. The positivity in treated cases was 31.8% (7/22), while in newly diagnosed leprosy cases the number of positives were higher, 45.8% (11/23) and lower in HHC at 23.7% (18/76). Whole genome sequencing of M. leprae from biopsies of three infected individuals from one extended family revealed a hypermutated M. leprae strain in an unusual case of primary drug resistance while the other two strains were drug sensitive. Discussion: This study represents the extent of leprosy in an active surveillance campaign during a single week in the city of Castanhal, a city that we have previously surveyed several times during the past ten years. Our results indicate the continuing high transmission of leprosy that includes fairly high rates of new cases detected in children indicating recent spread by multiple foci of infection in the community. An unusual case of a hypermutated M. leprae strain in a case of primary drug resistance was discovered. It also revealed a high hidden prevalence of overt disease and subclinical infection that remains a challenge for correct clinical diagnosis by signs and symptoms that may be aided using adjunct laboratory tests, such as RLEP qPCR and anti-PGL-I serology.

Transmission dynamics of SARS-CoV-2 variants in the Brazilian state of Pará.

Introduction: After three years since the beginning of the pandemic, the new coronavirus continues to raise several questions regarding its infectious process and host response. Several mutations occurred in different regions of the SARS-CoV-2 genome, such as in the spike gene, causing the emergence of variants of concern and interest (VOCs and VOIs), of which some present higher transmissibility and virulence, especially among patients with previous comorbidities. It is essential to understand its spread dynamics to prevent and control new biological threats that may occur in the future. In this population_based retrospective observational study, we generated data and used public databases to understand SARS-CoV-2 dynamics. Methods: We sequenced 1,003 SARS-CoV-2 genomes from naso-oropharyngeal swabs and saliva samples from Pará from May 2020 to October 2022. To gather epidemiological data from Brazil and the world, we used FIOCRUZ and GISAID databases. Results: Regarding our samples, 496 (49.45%) were derived from female participants and 507 (50.55%) from male participants, and the average age was 43 years old. The Gamma variant presented the highest number of cases, with 290 (28.91%) cases, followed by delta with 53 (5.28%). Moreover, we found seven (0.69%) Omicron cases and 651 (64.9%) non-VOC cases. A significant association was observed between sex and the clinical condition (female, p = 8.65e-08; male, p = 0.008961) and age (p = 3.6e-10). Discussion: Although gamma had been officially identified only in December 2020/January 2021, we identified a gamma case from Belém (capital of Pará State) dated May 2020 and three other cases in October 2020. This indicates that this variant was circulating in the North region of Brazil several months before its formal identification and that Gamma demonstrated its actual transmission capacity only at the end of 2020. Furthermore, the public data analysis showed that SARS-CoV-2 dispersion dynamics differed in Brazil as Gamma played an important role here, while most other countries reported a new infection caused by the Delta variant. The genetic and epidemiological information of this study reinforces the relevance of having a robust genomic surveillance service that allows better management of the pandemic and that provides efficient solutions to possible new disease-causing agents.

Leprosy among children in an area without primary health care coverage in Caratateua Island, Brazilian Amazon.

Introduction: The detection of leprosy in children is an important epidemiological marker of the disease, indicating the community's early exposure to Mycobacterium leprae and active transmission of the infection. Methods: In order to detect new cases among children by combining clinical evaluation and laboratory tests, we conducted an active case finding among individuals under 15 years old on Caratateua Island, located in the city of Belém, in the Pará state, an endemic region in the Amazon. Dermato-neurological examination, collection of 5 mL of peripheral blood for IgM anti-PGL-I antibody titration, and intradermal scraping for bacilloscopy and amplification of the specific RLEP region by qPCR were performed. Results: Out of the 56 examined children, 28/56 (50%) new cases were identified. At the time of evaluation, 38/56 (67.8%) children presented one or more clinical alterations. Seropositivity was detected in 7/27 (25.9%) new cases and 5/24 (20.8%) undiagnosed children. DNA amplification of Mycobacterium leprae was observed in 23/28 (82.1%) of new cases and in 5/26 (19.2%) of non-cases. Out of the total cases, 11/28 (39.2%) were exclusively diagnosed by clinical evaluation performed during the active case finding. Seventeen new cases (60.8%) were detected considering the clinical alterations found in addition to positive results for qPCR. In this group, 3/17 (17.6%) qPCR-positive children presented significant clinical changes 5.5 months after the first evaluation. Discussion: Our research detected a number of cases 5.6 times higher compared to the total number of pediatric cases recorded throughout the year 2021 in the municipality of Belém, which shows a critical scenario of underdiagnosing of leprosy among children under 15 years old in the region. We propose the use of qPCR technique to identify new cases among children with oligosymptomatic or early disease in endemic areas, in addition to the training of Primary Health Care professionals and the implementation of the Family Health Strategy coverage in the visited area.

Enzymatic isolation of Lacazia loboi cells from skin lesions of lobomycosis.

Lacazia loboi is the etiologic agent of Jorge Lobo's disease, a cutaneous and subcutaneous mycosis endemic to Latin America tropical regions and characterized by chronic nodular or keloidal lesions which develop after traumatic events. A new method for the extraction of L. loboi yeast cells from biopsies of lobomycosis skin lesions is presented. The method is based on the proteolytic action of the enzyme dispase which is known for its action against fibronectin and collagen type IV. Fungal identification was based on histological examination of the biological material and molecular analysis based on 18S ribosomal sequences. Observations under optic and fluorescence microscopy proved the efficacy of enzymatic isolation of the lobomycosis etiologic agent, as well as identifying the organism's main parasitic characteristics. Molecular phylogenetic analysis corroborated the histological examination and indicated L. loboi relationship with other members of the Onygenales. Use of dispase proved to be ideal for the isolation of L. loboi from human biopsies, shows promise as an important tool for improving biological studies of this peculiar fungus.

Isolation of Fonsecaea pedrosoi from thorns of Mimosa pudica, a probable natural source of chromoblastomycosis.

We report the isolation of Fonsecaea pedrosoi from thorns of the plant Mimosa pudica L. at the place of infection identified by one of our patients. Clinical diagnosis of chromoblastomycosis was established by direct microscopic examination and cultures from the patient's lesion. The same species was isolated from the patient and from the plant. Scanning electron microscopy of the surface of the thorns showed the characteristic conidial arrangement of F. pedrosoi. These data indicate that M. pudica could be a natural source of infection for the fungus F. pedrosoi.