PGL I expression in live bacteria allows activation of a CD206/PPARγ cross-talk that may contribute to successful Mycobacterium leprae colonization of peripheral nerves.

Mycobacterium leprae, an obligate intracellular bacillus, infects Schwann cells (SCs), leading to peripheral nerve damage, the most severe leprosy symptom. In the present study, we revisited the involvement of phenolic glycolipid I (PGL I), an abundant, private, surface M. leprae molecule, in M. leprae-SC interaction by using a recombinant strain of M. bovis BCG engineered to express this glycolipid. We demonstrate that PGL I is essential for bacterial adhesion and SC internalization. We also show that live mycobacterium-producing PGL I induces the expression of the endocytic mannose receptor (MR/CD206) in infected cells in a peroxisome proliferator-activated receptor gamma (PPARγ)-dependent manner. Of note, blocking mannose recognition decreased bacterial entry and survival, pointing to a role for this alternative recognition pathway in bacterial pathogenesis in the nerve. Moreover, an active crosstalk between CD206 and the nuclear receptor PPARγ was detected that led to the induction of lipid droplets (LDs) formation and prostaglandin E2 (PGE2), previously described as fundamental players in bacterial pathogenesis. Finally, this pathway was shown to induce IL-8 secretion. Altogether, our study provides evidence that the entry of live M. leprae through PGL I recognition modulates the SC phenotype, favoring intracellular bacterial persistence with the concomitant secretion of inflammatory mediators that may ultimately be involved in neuroinflammation.

Applicability of the Monocyte Activation Test (MAT) in the quality control of the 17DD yellow fever vaccine.

The need for alternatives to animal use in pyrogen testing has been driven by the Three Rs concept. This has resulted in the inclusion of the monocyte activation test (MAT) in the European Pharmacopoeia, 2010. However, some technical and regulatory obstacles must be overcome to ensure the effective implementation of the MAT by the industry, especially for the testing of biological products. The yellow fever (YF) vaccine (17DD-YFV) was chosen for evaluation in this study, in view of: a) the 2016-2018 outbreak of YF in Brazil; b) the increase in demand for 17DD-YFV doses; c) the complex production process with live attenuated virus; d) the presence of possible test interference factors, such as residual process components (e.g. ovalbumin); and e) the need for the investigation of other pyrogens that are not detectable by the methods prescribed in the YF vaccine monograph. The product-specific testing was carried out by using cryopreserved and fresh whole blood, and IL-6 and IL-1ß levels were used as the marker readouts. After assessing the applicability of the MAT on a 1:10 dilution of 17DD-YFV, endotoxin and non-endotoxin pyrogens were quantified in spiked batches, by using the lipopolysaccharide and lipoteichoic acid standards, respectively. The quantitative analysis demonstrated the correlation between the MAT and the Limulus amoebocyte lysate (LAL) assays, with respect to the limits of endotoxin recovery in spiked batches and the detection of no pyrogenic contamination in commercial batches of 17DD-YFV. The data demonstrated the applicability of the MAT for 17DD-YFV pyrogen testing, and as an alternative method that can contribute to biological quality control studies.

Deciphering the contribution of lipid droplets in leprosy: multifunctional organelles with roles in Mycobacterium leprae pathogenesis.

Leprosy is an infectious disease caused by Mycobacterium leprae that affects the skin and nerves, presenting a singular clinical picture. Across the leprosy spectrum, lepromatous leprosy (LL) exhibits a classical hallmark: the presence of a collection of M. leprae-infected foamy macrophages/Schwann cells characterised by their high lipid content. The significance of this foamy aspect in mycobacterial infections has garnered renewed attention in leprosy due to the recent observation that the foamy aspect represents cells enriched in lipid droplets (LD) (also known as lipid bodies). Here, we discuss the contemporary view of LD as highly regulated organelles with key functions in M. leprae persistence in the LL end of the spectrum. The modern methods of studying this ancient disease have contributed to recent findings that describe M. leprae-triggered LD biogenesis and recruitment as effective mycobacterial intracellular strategies for acquiring lipids, sheltering and/or dampening the immune response and favouring bacterial survival, likely representing a fundamental aspect of M. leprae pathogenesis. The multifaceted functions attributed to the LD in leprosy may contribute to the development of new strategies for adjunctive anti-leprosy therapies.

Mycobacterium leprae-induced Insulin-like Growth Factor I attenuates antimicrobial mechanisms, promoting bacterial survival in macrophages.

Mycobacterium leprae (ML), the etiologic agent of leprosy, can subvert macrophage antimicrobial activity by mechanisms that remain only partially understood. In the present study, the participation of hormone insulin-like growth factor I (IGF-I) in this phenomenum was investigated. Macrophages from the dermal lesions of the disseminated multibacillary lepromatous form (LL) of leprosy expressed higher levels of IGF-I than those from the self-limited paucibacillary tuberculoid form (BT). Higher levels of IGF-I secretion by ML-infected macrophages were confirmed in ex vivo and in vitro studies. Of note, the dampening of IGF-I signaling reverted the capacity of ML-infected human and murine macrophages to produce antimicrobial molecules and promoted bacterial killing. Moreover, IGF-I was shown to inhibit the JAK/STAT1-dependent signaling pathways triggered by both mycobacteria and IFN-γ most probably through its capacity to induce the suppressor of cytokine signaling-3 (SOCS3). Finally, these in vitro findings were corroborated by in vivo observations in which higher SOCS3 expression and lower phosphorylation of STAT1 levels were found in LL versus BT dermal lesions. Altogether, our data strongly suggest that IGF-I contributes to the maintenance of a functional program in infected macrophages that suits ML persistence in the host, reinforcing a key role for IGF-I in leprosy pathogenesis.

Deciphering the contribution of lipid droplets in leprosy: multifunctional organelles with roles in Mycobacterium leprae pathogenesis

Leprosy is an infectious disease caused by Mycobacterium leprae that affects the skin and nerves, presenting a singular clinical picture. Across the leprosy spectrum, lepromatous leprosy (LL) exhibits a classical hallmark: the presence of a collection of M. leprae-infected foamy macrophages/Schwann cells characterised by their high lipid content. The significance of this foamy aspect in mycobacterial infections has garnered renewed attention in leprosy due to the recent observation that the foamy aspect represents cells enriched in lipid droplets (LD) (also known as lipid bodies). Here, we discuss the contemporary view of LD as highly regulated organelles with key functions in M. leprae persistence in the LL end of the spectrum. The modern methods of studying this ancient disease have contributed to recent findings that describe M. leprae-triggered LD biogenesis and recruitment as effective mycobacterial intracellular strategies for acquiring lipids, sheltering and/or dampening the immune response and favouring bacterial survival, likely representing a fundamental aspect of M. leprae pathogenesis. The multifaceted functions attributed to the LD in leprosy may contribute to the development of new strategies for adjunctive anti-leprosy therapies.

Protection against cutaneous leishmaniasis by intranasal vaccination with lipophosphoglycan.

We previously showed the opposing effect of systemic and mucosal vaccination with whole Leishmania amazonensis antigen (LaAg). Here, the role played by lipophosphoglycan (LPG) as the key disease-promoting component of intramuscular (i.m.) LaAg and its usefulness as a defined intranasal vaccine was investigated in murine cutaneous leishmaniasis. BALB/c mice were twice vaccinated by the i.m. route with 25mug of intact LaAg or with LaAg that was pretreated with anti-LPG 3A1-La monoclonal antibody, prior to infection with L. amazonensis. LPG neutralization rendered the otherwise disease-promoting LaAg antigen protective, as observed by the smaller lesion sizes and reduced parasite burden. The increased resistance was accompanied by a markedly lower antigen-driven TGF-beta and IL-10 responses in the lesion-draining lymph nodes, concomitant with significantly higher IFN-gamma production. To test for intranasal efficacy, 10 microg of affinity-purified LPG and its parental LaAg were twice instilled in the nostrils prior to L. amazonensis infection. In both cases, similarly slower lesion growth and lower parasite burden were found that was associated with increased IFN-gamma and IL-10 responses in the lesion-draining lymph nodes. These results support a role for LPG in the dual route-related effect of LaAg and shows its strong potential as a defined needle-free and adjuvant-free vaccine for cutaneous leishmaniasis.

Isolation and characterization of azotobacter strains from brazilian soils

Thirty-eight Azotobacter spp strains isolated from Brasilian soils were characterized for the foollowing properties : utilization of different carbon sources, diffusible pigment production, melanin production, sodium-dependent growth, siderophore production, streptomycin resistance and antibiotic-like substances production. Four strains were identified : one as A. armeniacus and three as A. paspali.