Case Report of Leprosy in Central Florida, USA, 2022.

Florida, USA, has witnessed an increased incidence of leprosy cases lacking traditional risk factors. Those trends, in addition to decreasing diagnoses in foreign-born persons, contribute to rising evidence that leprosy has become endemic in the southeastern United States. Travel to Florida should be considered when conducting leprosy contact tracing in any state.

Evidence for Extensive Duplication and Subfunctionalization of FCRL6 in Armadillo (Dasypus novemcinctus).

The control of infections by the vertebrate adaptive immune system requires careful modulation to optimize defense and minimize harm to the host. The Fc receptor-like (FCRL) genes encode immunoregulatory molecules homologous to the receptors for the Fc portion of immunoglobulin (FCR). To date, nine different genes (FCRL1-6, FCRLA, FCRLB and FCRLS) have been identified in mammalian organisms. FCRL6 is located at a separate chromosomal position from the FCRL1-5 locus, has conserved synteny in mammals and is situated between the SLAMF8 and DUSP23 genes. Here, we show that this three gene block underwent repeated duplication in Dasypus novemcinctus (nine-banded armadillo) resulting in six FCRL6 copies, of which five appear functional. Among 21 mammalian genomes analyzed, this expansion was unique to D. novemcinctus. Ig-like domains that derive from the five clustered FCRL6 functional gene copies show high structural conservation and sequence identity. However, the presence of multiple non-synonymous amino acid changes that would diversify individual receptor function has led to the hypothesis that FCRL6 endured subfunctionalization during evolution in D. novemcinctus. Interestingly, D. novemcinctus is noteworthy for its natural resistance to the Mycobacterium leprae pathogen that causes leprosy. Because FCRL6 is chiefly expressed by cytotoxic T and NK cells, which are important in cellular defense responses against M. leprae, we speculate that FCRL6 subfunctionalization could be relevant for the adaptation of D. novemcinctus to leprosy. These findings highlight the species-specific diversification of FCRL family members and the genetic complexity underlying evolving multigene families critical for modulating adaptive immune protection.

Mycobacterium leprae in Armadillo Tissues from Museum Collections, United States.

We examined armadillos from museum collections in the United States using molecular assays to detect leprosy-causing bacilli. We found Mycobacterium leprae bacilli in samples from the United States, Bolivia, and Paraguay; prevalence was 14.8% in nine-banded armadillos. US isolates belonged to subtype 3I-2, suggesting long-term circulation of this genotype.

Microbiota in promoting liver regeneration.

Extensive work has revealed well-coordinated mechanisms that underlie liver regeneration, albeit with a focus on intrinsic interactions within hepatic cells. Here, Hess et al.1 demonstrate that Mycobacterium leprae infection can induce liver growth of nine-banded armadillo without obvious side effects.

The Armadillo as a Model for Leprosy Nerve Function Impairment: Preventative and Therapeutic Interventions.

Mycobacterium leprae infection of peripheral nerves and the subsequent nerve function impairment (NFI), especially in response to reactional episodes, are hallmarks of leprosy. Improved treatments for M. leprae-induced nerve injury are needed, as most if not all of the disability and stigma associated with leprosy arises from the direct or indirect effects of NFI. Nine-banded armadillos (Dasypus novemcinctus), like humans, exhibit the full clinical spectrum of leprosy and extensive involvement of the peripheral nerves. In this study, state-of-the-art technology was used to compare nerve function between uninfected and M. leprae-infected armadillos. Motor nerve conduction velocity (MNCV) and compound muscle action potential (cMAP), which measure changes in the rate of impulse conduction velocity and amplitude, revealed a progression of impairment that was directly correlated with the duration of M. leprae infection and enabled development of an objective nerve impairment scoring system. Ultrasonography accompanied by color Doppler imaging detected enlargement of the M. leprae-infected nerves and increased vascularity, possibly due to inflammation. Assessment of epidermal nerve fiber density (ENFD), which shows a length-dependent innervation in armadillos that is similar to humans, identified small fiber degeneration early after M. leprae infection. Staining for neuromuscular junction (NMJ) integrity, which is an indicator of signal transduction efficiency into skeletal muscle, discerned a markedly lower number and structural integrity of NMJ in M. leprae-infected armadillo footpads. These tools for assessing nerve injury were used to monitor the effects of intervention therapy. Two potential neuro-protective drugs, ethoxyquin (EQ) and 4-aminopyridine (4-AP), were tested for their ability to ameliorate peripheral nerve injury in M. leprae-infected armadillos. 4-AP treatment improved MNCV, cMAP, and EFND compared to untreated animals, while EQ had less effect. These results support the armadillo as a model for M. leprae-induced peripheral nerve injury that can provide insights toward the understanding of NFI progression and contribute to the preclinical investigation of the safety and efficacy of neuro-preventive and neuro-therapeutic interventions for leprosy.

Chromoblastomycosis-Leprosy Co-Infection in Central West Brazil. Presentation of Three Cases and Literature Review.

Chromoblastomycosis and leprosy are chronic diseases with high prevalence in tropical and subtropical regions. Brazil is one of the countries with the highest incidence and prevalence for both diseases, however, reports of co-infections are scarce. The aim of this study was to describe three cases of chromoblastomycosis-leprosy co-infection in patients from Mato Grosso state, Brazil. A review of chromoblastomycosis-leprosy co-infection was performed of English, Portuguese and Spanish publications in LILACS, SciELO, PubMed and Web of Science databases using the descriptors (chromoblastomycosis OR cromoblastomicose OR cromoblastomicosis) AND (leprosy OR hanseníase OR lepra), without time period delimitation. Nineteen cases were included, 16 cases were published in 11 articles, plus the three cases reported in the current study. Most reported coninfection cases came from Brazil. Majority of the patients were male with a mean age of 52.2 years. Farmer was the main occupational activity reported. In 12 patients, the clinical signs and symptoms of leprosy started first. No contacts with patients affected by leprosy, armadillos or history of injuries at the anatomical site of chromoblastomycosis lesions were reported. Five leprosy patients who received steroid treatment for leprosy reactions or neuropathies, were diagnosed with chromoblastomycosis during immunosuppressive therapy. Four cases (21.1%) were reported among the elderly patients. Co-infections in patients with chromoblastomycosis or leprosy are uncommon, but the possibility should always be considered, especially if the patient is undergoing immunosuppressive treatment or is elder.

The Presence of Mycobacterium leprae in Wild Rodents.

Leprosy is a chronic infection caused by Mycobacterium leprae. There is a lack of data regarding environmental reservoirs, which may represent a serious public health problem in Brazil, especially in the state of Pará, which occupies the fourth position in incidence of cases in the country. Previous studies report evidence of infection occurring among armadillos, mangabei monkeys, and chimpanzees. In the present study, wild animals were captured and tested for the presence of anti-PGL-1 antibodies and M. leprae DNA. Fieldwork was carried out from October to November of 2016 in the cities of Curionópolis and Canaã dos Carajás, southeast of Pará state. Small and medium-sized wild animals were captured using appropriate traps. A total of 15 animals were captured. Sera and viscera fragments were collected and tested by ELISA and PCR methods. The presence of M. leprae DNA was confirmed by sequencing of specific gyrase gene in three animals of two different species, including one Necromys lasiurus (liver sample) and two Proechimys roberti (kidney and liver samples). This unprecedented finding suggests that species other than those previously reported are responsible for maintaining M. leprae in nature.

Mycobacterium leprae induces Schwann cell proliferation and migration in a denervated milieu following intracutaneous excision axotomy in nine-banded armadillos.

Nine-banded armadillos develop peripheral neuropathy after experimental Mycobacterium leprae infection that recapitulates human disease. We used an intracutaneous excision axotomy model to assess the effect of infection duration by M. leprae on axonal sprouting and Schwan cell density. 34 armadillos (17 naïve and 17 M. leprae-infected) underwent 3 mm skin biopsies to create an intracutaneous excision axotomy followed by a concentric 4-mm overlapping biopsy 3 and 12-months post M. leprae inoculation. A traditional distal leg biopsy was obtained at 15mo for intraepidermal nerve fiber (IENF) density. Serial skin sections were immunostained against a axons (PGP9.5, GAP43), and Schwann cells (p75, s100) to visualize regenerating nerves. Regenerative axons and proliferation of Schwann cells was measured and the rate of growth at each time point was assessed. Increasing anti-PGL antibody titers and intraneural M. leprae confirmed infection. 15mo following infection, there was evidence of axon loss with reduced distal leg IENF versus naïve armadillos, p < 0.05. This was associated with an increase in Schwann cell density (11,062 ± 2905 vs. 7561 ± 2715 cells/mm3, p < 0.01). Following excisional biopsy epidermal reinnervation increased monotonically at 30, 60 and 90 days; the regeneration rate was highest at 30 days, and decreased at 60 and 90 days. The reinnervation rate was highest among animals infected for 3mo vs those infected for 12mo or naïve animals (mean ± SD, 27.8 ± 7.2 vs.16.2 ± 5.8vs. 15.3 ± 6.5 mm/mm3, p < 0.05). The infected armadillos displayed a sustained Schwann cell proliferation across axotomy time points and duration of infection (3mo:182 ± 26, 12mo: 256 ± 126, naive: 139 ± 49 cells/day, p < 0.05). M. leprae infection is associated with sustained Schwann cell proliferation and distal limb nerve fiber loss. Rates of epidermal reinnervation were highest 3mo after infection and normalized by 12 mo of infection. We postulate that excess Schwann cell proliferation is the main pathogenic process and is deleterious to sensory axons. There is a compensatory initial increase in regeneration rates that may be an attempt to compensate for the injury, but it is not sustained and eventually followed by axon loss. Aberrant Schwann cell proliferation may be a novel therapeutic target to interrupt the pathogenic cascade of M. leprae.

Mycobacterium leprae Infection in a Wild Nine-Banded Armadillo, Nuevo León, Mexico.

Nine-banded armadillos (Dasypus novemcinctus) are naturally infected with Mycobacterium leprae and are implicated in the zoonotic transmission of leprosy in the United States. In Mexico, the existence of such a reservoir remains to be characterized. We describe a wild armadillo infected by M. leprae in the state of Nuevo León, Mexico.

TISSUE AND GASTROINTESTINAL PARASITES OF COLONIZING NINE-BANDED ARMADILLOS AT THE EDGE OF THEIR NORTHERN RANGE.

Originally endemic to South America, the nine-banded armadillo (Dasypus novemcinctus) has recently expanded its range northward to Illinois. With this range expansion comes concern regarding potential incoming pathogens; our research, conducted during 2012-2020, consisted of screening armadillos for the presence of helminths, Trypanosoma cruzi, and Mycobacterium leprae. We screened for the presence of T. cruzi and M. leprae, 2 pathogens known to infect humans, using polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. All 80 samples screened for T. cruzi and all 25 samples screened for M. leprae were negative. No parasite specific to the nine-banded armadillo, such as Aspidodera sogandaresi, was detected. This lack of infection is consistent with the idea that animals may be isolated from their common parasites during periods of range expansion. Lack of infection by T. cruzi in an endemic area suggests that these mammals may not be exposed to the infective stages at this early phase of their colonization. Presently, the armadillo has become established in Illinois, yet they have not introduced their parasites into the area. Our study represents the first effort to document the parasitological record of the expanding armadillo within 30 yr of their initial record in Illinois and the American Midwest. This helminthological record of armadillos in Illinois sets the timeline to observe the establishment of A. sogandaresi in the Midwest.

Leprosy: what is new.

Leprosy, also known as Hansen's disease, is an age-old chronic granulomatous infection characterized by prominent cutaneous and neurologic findings. Long known to be caused by Mycobacterium leprae, a new etiologic species was identified and linked in 2008, Mycobacterium lepromatosis. The BCG vaccine with highly variable efficacy may soon be replaced by the first leprosy-specific subunit vaccine LepVax, which has recently moved forward in human trials. Recent evidence supporting theories of zoonotic transmission from armadillos and the less-discussed Eurasian red squirrels has emerged. Knowledge on genetic polymorphisms that may increase leprosy susceptibility, such as the newly uncovered mitochondrial ribosomal protein S5 (MRPS5) polymorphism in the Chinese population, has provided a fresh perspective and direction. Further, we will delineate the latest information on leprosy, including the possible effects of leprosy coinfection with COVID-19, HIV, and HTLV-1, and the shift to newer leprosy therapies and treatment regimens.

Treatment and Evaluation Advances in Leprosy Neuropathy.

Neuropathy and related disabilities are the major medical consequences of leprosy, which remains a global medical concern. Despite major advances in understanding the mechanisms of M. leprae entry into peripheral nerves, most aspects of the pathogenesis of leprosy neuropathy remain poorly understood. Sensory loss is characteristic of leprosy, but neuropathic pain is sometimes observed. Effective anti-microbial therapy is available, but neuropathy remains a problem especially if diagnosis and treatment are delayed. Currently there is intense interest in post-exposure prophylaxis with single-dose rifampin in endemic areas, as well as with enhanced prophylactic regimens in some situations. Some degree of nerve involvement is seen in all cases and neuritis may occur in the absence of leprosy reactions, but acute neuritis commonly accompanies both Type 1 and Type 2 leprosy reactions and may be difficult to manage. A variety of established as well as new methods for the early diagnosis and assessment of leprosy neuropathy are reviewed. Corticosteroids offer the primary treatment for neuritis and for subclinical neuropathy in leprosy, but success is limited if nerve function impairment is present at the time of diagnosis. A candidate vaccine has shown apparent benefit in preventing nerve injury in the armadillo model. The development of new therapeutics for leprosy neuropathy is greatly needed.

Mycobacterium leprae Infection in Ticks and Tick-Derived Cells.

Leprosy is a zoonosis in the southern United States involving humans and wild armadillos. The majority of patients presenting with zoonotic strains of Mycobacterium leprae note extensive outdoor activity but only rarely report any history of direct contact with wild armadillos. Whether M. leprae is transmitted to new vertebrate hosts through the environment independently or with the aid of other organisms, e.g., arthropod vectors, is a fundamental question in leprosy transmission. The objectives of this study were to assess the potential for ticks to transmit M. leprae and to test if viable M. leprae can be maintained in tick-derived cells. To evaluate tick transmission, nymphal Amblyomma maculatum ticks were injected with isolated M. leprae. Infection and transmission were assessed by qPCR. Ticks infected as nymphs harbored M. leprae through vertical transmission events (nymph to adult and adult to progeny); and, horizontal transmission of M. leprae to a vertebrate host was observed. Mycobacterium leprae DNA was detected in multiple tick life cycle stages. Likewise, freshly isolated M. leprae (Thai-53) was used to infect a tick-derived cell line, and enumeration and bacterial viability were assessed at individual time points for up to 49 days. Evaluations of the viability of long-term cultured M. leprae (Thai-53 and Br4923) were also assessed in a mouse model. Tick-derived cells were able to maintain viable M. leprae over the 49-day course of infection and M. leprae remained infectious within tick cells for at least 300 days. The results of this study suggest that ticks themselves might serve as a vector for the transmission of M. leprae and that tick cells are suitable for maintenance of viable M. leprae for an extended period of time.

Detection and Monitoring of Mycobacterium leprae Infection in Nine Banded Armadillos (Dasypus novemcinctus) Using a Quantitative Rapid Test.

Leprosy is an infectious disease caused by Mycobacterium leprae with tropism for skin and peripheral nerves. Incessant transmission in endemic areas is still impeding elimination of leprosy. Although detection of M. leprae infection remains a challenge in asymptomatic individuals, the presence of antibodies specific for phenolglycolipid-I (PGL-I) correlate with bacterial load. Therefore, serosurveillance utilizing field-friendly tests detecting anti-PGL-I antibodies, can be applied to identify those who may transmit bacteria and to study (reduction of) M. leprae transmission. However, serology based on antibody detection cannot discriminate between past and present M. leprae infection in humans, nor can it detect individuals carrying low bacillary loads. In humans, anti-PGL-I IgM levels are long-lasting and usually detected in more individuals than anti-PGL-I IgG levels. Inherent to the characteristically long incubation time of leprosy, IgM/IgG relations (antibody kinetics) in leprosy patients and infected individuals are not completely clear. To investigate the antibody response directly after infection, we have measured antibody levels by ELISA, in longitudinal samples of experimentally M. leprae infected, susceptible nine-banded armadillos (Dasypus novemcinctus). In addition, we assessed the user- and field-friendly, low-cost lateral flow assay (LFA) utilizing upconverting reporter particles (UCP), developed for quantitative detection of human anti-PGL-I IgM (UCP-LFA), to detect treatment- or vaccination-induced changes in viable bacterial load. Our results show that serum levels of anti-PGL-I IgM, and to a lesser extent IgG, significantly increase soon after experimental M. leprae infection in armadillos. In view of leprosy phenotypes in armadillos, this animal model can provide useful insight into antibody kinetics in early infection in the various spectral forms of human leprosy. The UCP-LFA for quantitative detection of anti-PGL-I IgM allows monitoring the efficacy of vaccination and rifampin-treatment in the armadillo leprosy model, thereby providing a convenient tool to evaluate the effects of drugs and vaccines and new diagnostics.

Leprosy in wild chimpanzees.

Humans are considered as the main host for Mycobacterium leprae1, the aetiological agent of leprosy, but spillover has occurred to other mammals that are now maintenance hosts, such as nine-banded armadillos and red squirrels2,3. Although naturally acquired leprosy has also been described in captive nonhuman primates4-7, the exact origins of infection remain unclear. Here we describe leprosy-like lesions in two wild populations of western chimpanzees (Pan troglodytes verus) in Cantanhez National Park, Guinea-Bissau and Taï National Park, Côte d'Ivoire, West Africa. Longitudinal monitoring of both populations revealed the progression of disease symptoms compatible with advanced leprosy. Screening of faecal and necropsy samples confirmed the presence of M. leprae as the causative agent at each site and phylogenomic comparisons with other strains from humans and other animals show that the chimpanzee strains belong to different and rare genotypes (4N/O and 2F). These findings suggest that M. leprae may be circulating in more wild animals than suspected, either as a result of exposure to humans or other unknown environmental sources.

Mycobacterium lepromatosis as a Second Agent of Hansen's Disease.

Mycobacterium lepromatosis was identified as a new species and second causal agent of Hansen's disease (HD, or leprosy) in 2008, 150years after the disease was first attributed to Mycobacterium leprae. M. lepromatosis has been implicated in a small number of HD cases, and clinical aspects of HD caused by M. lepromatosis are poorly characterized. HD is a recognized zoonosis through transmission of M. leprae from armadillos, but the role of M. lepromatosis as a zoonotic agent of HD is unknown. M. lepromatosis was initially associated with diffuse lepromatous leprosy, but subsequent case reports and surveys have linked it to other forms of HD. HD caused by M. lepromatosis has been reported from three endemic countries: Brazil, Myanmar, and Philippines, and three non-endemic countries: Mexico, Malaysia, and United States. Contact with armadillos in Mexico was mentioned in 2/21 M. lepromatosis HD case reports since 2008. M. lepromatosis in animals has been investigated only in non-endemic countries, in squirrels and chipmunks in Europe, white-throated woodrats in Mexico, and armadillos in the United States. To date, there have only been a small number of positive findings in Eurasian red squirrels in Britain and Ireland. A single study of environmental samples found no M. lepromatosis in soil from a Scottish red squirrel habitat. Future studies must focus on endemic countries to determine the true proportion of HD cases caused by M. lepromatosis, and whether viable M. lepromatosis occurs in non-human sources.

Culturing Mycobacteria.

Building upon the foundational research of Robert Koch, who demonstrated the ability to grow Mycobacterium tuberculosis for the first time in 1882 using media made of coagulated bovine serum, microbiologists have continued to develop new and more efficient ways to grow mycobacteria. Presently, all known mycobacterial species can be grown in the laboratory using either axenic culture techniques or in vivo passage in laboratory animals. This chapter provides conventional protocols to grow mycobacteria for diagnostic purposes directly from clinical specimens, as well as in research laboratories for scientific purposes. Detailed protocols used for production of M. tuberculosis in large scale (under normoxic and hypoxic conditions) in bioreactors and for production of obligate intracellular pathogens such as Mycobacterium leprae and "Mycobacterium lepromatosis" using athymic nude mice and armadillos are provided.

Mycobacterium leprae transmission characteristics during the declining stages of leprosy incidence: A systematic review.

BACKGROUND: Leprosy is an infectious disease caused by Mycobacterium leprae. As incidence begins to decline, the characteristics of new cases shifts away from those observed in highly endemic areas, revealing potentially important insights into possible ongoing sources of transmission. We aimed to investigate whether transmission is driven mainly by undiagnosed and untreated new leprosy cases in the community, or by incompletely treated or relapsing cases. METHODOLOGY/PRINCIPAL FINDINGS: A literature search of major electronic databases was conducted in January, 2020 with 134 articles retained out of a total 4318 records identified (PROSPERO ID: CRD42020178923). We presented quantitative data from leprosy case records with supporting evidence describing the decline in incidence across several contexts. BCG vaccination, active case finding, adherence to multidrug therapy and continued surveillance following treatment were the main strategies shared by countries who achieved a substantial reduction in incidence. From 3950 leprosy case records collected across 22 low endemic countries, 48.3% were suspected to be imported, originating from transmission outside of the country. Most cases were multibacillary (64.4%) and regularly confirmed through skin biopsy, with 122 cases of suspected relapse from previous leprosy treatment. Family history was reported in 18.7% of cases, while other suspected sources included travel to high endemic areas and direct contact with armadillos. None of the countries included in the analysis reported a distinct increase in leprosy incidence in recent years. CONCLUSIONS/SIGNIFICANCE: Together with socioeconomic improvement over time, several successful leprosy control programmes have been implemented in recent decades that led to a substantial decline in incidence. Most cases described in these contexts were multibacillary and numerous cases of suspected relapse were reported. Despite these observations, there was no indication that these cases led to a rise in new secondary cases, suggesting that they do not represent a large ongoing source of human-to-human transmission.

IL-1R1-Dependent Signals Improve Control of Cytosolic Virulent Mycobacteria In Vivo.

Mycobacterium tuberculosis infections claim more than a million lives each year, and better treatments or vaccines are required. A crucial pathogenicity factor is translocation from phagolysosomes to the cytosol upon phagocytosis by macrophages. Translocation from the phagolysosome to the cytosol is an ESX-1-dependent process, as previously shown in vitro Here, we show that in vivo, mycobacteria also translocate to the cytosol but mainly when host immunity is compromised. We observed only low numbers of cytosolic bacilli in mice, armadillos, zebrafish, and patient material infected with M. tuberculosis, M. marinum, or M. leprae In contrast, when innate or adaptive immunity was compromised, as in severe combined immunodeficiency (SCID) or interleukin-1 receptor 1 (IL-1R1)-deficient mice, significant numbers of cytosolic M. tuberculosis bacilli were detected in the lungs of infected mice. Taken together, in vivo, translocation to the cytosol of M. tuberculosis is controlled by adaptive immune responses as well as IL-1R1-mediated signals.IMPORTANCE For decades, Mycobacterium tuberculosis has been one of the deadliest pathogens known. Despite infecting approximately one-third of the human population, no effective treatment or vaccine is available. A crucial pathogenicity factor is subcellular localization, as M. tuberculosis can translocate from phagolysosome to the cytosol in macrophages. The situation in vivo is more complicated. In this study, we establish that high-level cytosolic escape of mycobacteria can indeed occur in vivo but mainly when host resistance is compromised. The IL-1 pathway is crucial for the control of the number of cytosolic mycobacteria. The establishment that immune signals result in the clearance of cells containing cytosolic mycobacteria connects two important fields, cell biology and immunology, which is vital for the understanding of the pathology of M. tuberculosis.