Aline graduated from the University of Grande ABC, Brazil, with a Bachelor’s degree in Biological Sciences. She obtained her Master’s degree in Microbiology from the University of Sao Paulo, Brazil and PhD degree in Biotechnology from the University of Sao Paulo/Butantan Institute, Brazil. Aline has a Postdoctoral internship at the Butantan Institute (2016-2022), at the Department of Veterinary and Animal Sciences at the University of Massachusetts, Amherst, USA (2019-2020) and at the Federal University of ABC (2022). Currently, Aline is at the Vaccine Development Laboratory at the Butantan Institute, where researches antigenic fractions of Leptospira interrogans as a vaccine target for the control of leptospirosis. Aline has experience in the area of Microbiology, Immunology, Biotechnology and Molecular Biology with emphasis on the development of recombinant proteins, evaluation of host-pathogen interactions, cell culture and immunological assays with mice and hamsters.
RESEARCH, TEACHING, or OTHER INTERESTS
Immunology and Microbiology, Molecular Biology, Biotechnology
35
Scopus Publications
Scopus Publications
Functional Validation of the Proteome-Identified LIC_13056 Putative Lipoprotein of Leptospira interrogans and the Potential Role in Pathogenesis Giovanna Costa, João P. Gaspar, Aline F. Teixeira, Ana L. T. O. Nascimento International Journal of Molecular Sciences, 2026 Leptospirosis is a widespread zoonosis of human and veterinary concern. The etiological agent of the disease is the pathogenic bacteria of the genus Leptospira. Transmission typically occurs through mucosal contact and/or injured skin with the urine of infected animals or contaminated environmental sources. Understanding the biology and pathogenesis of leptospires is the main focus of our study. In this work, we characterized a novel protein encoded by the LIC_13056 gene from L. interrogans serovar Copenhageni, having an OmpA-like domain. We show that this coding sequence (CDS), previously assigned as a hypothetical protein with an unknown function, is capable of binding to the cellular receptor α8 integrin subunit, potentially contributing to kidney colonization. Additionally, the protein bound to both purified and normal human serum (NHS) plasminogen (PLG). In both conditions, PLG bound to protein was able to generate plasmin (PLA). Furthermore, rLIC_13056 interacted with the complement system components C4b, C4BP, C8 and C9. The interaction of recombinant protein to the C9 had a negative impact on C9 polymerization. Taken together, the protein LIC_13056, having an OmpA-like domain, appears to be involved in leptospiral pathogenesis via different stages of the infection process; PLA generation together with the inhibition of the membrane attack complex (MAC) may contribute to the immune evasion mechanism of Leptospira, thus facilitating the infection.
Revisiting Host-Binding Properties of LigA and LigB Recombinant Domains Henrique M. Pires, Igor R. M. Silva, Aline F. Teixeira, Ana L. T. O. Nascimento Microorganisms, 2025 Pathogenic bacteria of the genus Leptospira are the etiological agents of leptospirosis, a disease that affects humans and animals worldwide. Despite the increasing number of studies, the mechanisms of leptospiral pathogenesis remain poorly comprehended. In this study, we report various interactions of the LigA7’-13’ and LigB1’-7’ domains with host components. The LigA7’-13’ and LigB1’-7’ were cloned into the pET28a vector, and the recombinant proteins were expressed in E. coli C43 (DE3) and E. coli BL21 (DE3), respectively. Both recombinant protein domains were expressed in soluble form and purified using nickel-chelating chromatography. The rLigA7’-13’ and rLigB1’-7’ domains exhibited binding to several types of integrins, with most interactions occurring in a dose-dependent and saturable manner, consistent with the characteristics of typical receptor-ligand interactions. The recombinant domain LigA7’-13’ demonstrated affinity for the glycosaminoglycans (GAGs) chondroitin-4-sulfate, chondroitin sulfate, heparin, chondroitin sulfate B, and heparan sulfate, while no binding was detected for LigB1’-7’ with these molecules. Both rLigA7’-13’ and rLigB1’-7’ interacted with components of the terminal complement pathway and were capable of recruiting C9 from normal human serum (NHS). These interactions may inhibit the formation of polyC9, ultimately preventing the assembly of the membrane attack complex (MAC). Collectively, our data expand the repertoire of host components that interact with rLigA7’-13’ and rLigB1’-7’, opening new avenues for understanding leptospiral immune evasion and broadening the roles of these domains in bacterial virulence.
The VapBC-4 Characterization Indicates It Is a Bona Fide Toxin-Antitoxin Module of Leptospira interrogans: Initial Evidence for a Role in Bacterial Adaptation Bruna Oliveira Pigatto Azevedo, Deborah Kohn Damiano, Aline Florencio Teixeira, Ana Lucia Tabet Oller Nascimento, Luis Guilherme Virgilio Fernandes, et al. Microorganisms, 2025 Toxin-antitoxin (TA) systems are one of the bacterial adaptation mechanisms to adverse conditions. Leptospira interrogans serovar Copenhageni contains nine putative TA systems. To date, only VapBC-3 and VapBC-1 have been experimentally characterized and considered functional modules. This study shows that the VapBC-4 module is a novel bona fide TA system constituted by VapB-4 antitoxin and VapC-4 toxin. Overexpression of the recombinant toxin in Escherichia coli resulted in growth inhibition, which was rescued by co-expression of the VapB-4 antitoxin. The toxin-antitoxin binding capability, essential to TA functionality, was demonstrated by dot blot assay in vitro, while the pull-down assay indicates that the toxin and antitoxin interact in vivo. In addition, we confirmed that VapC-4 is a PIN domain endoribonuclease capable of degrading viral MS2 substrate. The transcriptional studies suggest that vapC-4 may be involved in the virulence and adaptability of L. interrogans serovar Copenhageni for adverse environmental conditions. Taken together, these results show that the VapBC-4 module is functional and can be considered a bona fide module.
Evaluation of binding activities of a putative lipoprotein LIC_13355 of Leptospira spp. Igor R. M. Silva, Maria B. Takahashi, Aline F. Teixeira, Ana L. T. O. Nascimento FEBS Open Bio, 2025 Pathogenic Leptospira is the etiological cause of the zoonotic life‐threatening infection called leptospirosis. The disease is spread worldwide with higher risk in tropical regions. Although leptospirosis represents a burden to the health of humans and animals, the pathogenic mechanisms of Leptospira infection are yet to be clarified. Leptospirosis infection is multifactorial, involving functionally redundant proteins with the capability to invade, disseminate, and escape the host's immune response. In this work, we describe a putative lipoprotein encoded by the gene LIC_13355, genome annotated as hypothetical of unknown function. The coding sequence is conserved among pathogenic Leptospira spp. with high percentage of coverage and identity. The recombinant protein, rLIC_13355, was expressed in Escherichia coli host system in its insoluble form. The circular dichroism spectrum of the refolded protein showed it containing a mixture of secondary structures. rLIC_13355 interacts with extracellular matrix (ECM) component laminin and binds plasminogen (PLG), generating plasmin (PLA), thus possibly participating during the adhesion and dissemination processes. The rLIC_13355 has the ability to interact with Ea.hy926 and HMEC‐1 endothelial cells either in monolayer or suspension. The binding of rLIC_13355 with monolayer cells is dose‐dependent on protein concentration. Taken together, our data suggest that this is presumably an adhesion lipoprotein that may play diverse roles in host–Leptospira interactions by mediating the interaction with host components and with endothelial cell.
The role of leptospiral proteins in immune evasion and inflammatory response stimulation in HEK293T cell monolayers Igor R. M. Silva, Aline F. Teixeira, Ana L. T. O. Nascimento Frontiers in Immunology, 2025 Pathogenic Leptospira spp. are the causative agents of leptospirosis, a significant zoonotic disease that has emerged as a crucial public health concern. This study aims to evaluate the interactions of two L. interrogans proteins, LIC_10499 and LIC_12339, with host components as well as with endothelial and epithelial cells. The coding sequences (CDSs) for LIC_10499 and LIC_12339 were cloned, expressed in Escherichia coli , and successfully purified from inclusion bodies. Both recombinant proteins demonstrated interactions with fibronectin, fibrinogen, and plasminogen (PLG). Notably, these proteins were capable of sequestering PLG from normal human serum (NHS). In the presence of an activator, the bound PLG is converted to plasmin (PLA), a broad-spectrum protease involved in pathogen invasion and immune evasion. Additionally, LIC_10499 and LIC_12339 were found to bind to complement system regulators, including factor H and C4b-binding protein, as well as to components C7, C8, and C9. We observed that the formation of C9 complexes was inhibited in the presence of recombinant proteins, and a higher survival rate of E. coli was noted when the proteins were incubated with NHS. The protein rLIC_10499 was able to bind to both monolayer and suspension cells of HMEC, Ea.hy926, and HEK293T, whereas rLIC_12339 only bound to HEK293T suspension cells. A significant production of IFN-γ was detected after 24 h when HEK293T epithelial cells were incubated with rLIC_10499, while a modest production of IL-6 and IL-8 was observed. No cytokine production occurred when HEK293T cells were stimulated with rLIC_12339. Collectively, these findings suggest that these proteins play a role in leptospiral immune evasion and have the potential to induce an inflammatory response in host cell monolayers.
The Toxin of VapBC-1 Toxin-Antitoxin Module from Leptospira interrogans Is a Ribonuclease That Does Not Arrest Bacterial Growth but Affects Cell Viability Deborah K. Damiano, Bruna O. P. Azevedo, George S. C. Fernandes, Aline F. Teixeira, Viviane M. Gonçalves, et al. Microorganisms, 2024 Bacterial ubiquitous Toxin-Antitoxin (TA) systems are considered to be important survival mechanisms during stress conditions. In regular environmental conditions, the antitoxin blocks the toxin, whereas during imbalanced conditions, the antitoxin concentration decreases, exposing the bacteria cell to a range of toxic events. The most evident consequence of this disequilibrium is cell growth arrest, which is the reason why TAs are generally described as active in the function of bacterial growth kinetics. Virulence-associated proteins B and C (VapBC) are a family of type II TA system, in which VapC is predicted to display the toxic ribonuclease activity while VapB counteracts this activity. Previously, using in silico data, we designated four VapBC TA modules in Leptospira interrogans serovar Copenhageni, the main etiological agent of human leptospirosis in Brazil. The present study aimed to obtain the proteins and functionally characterize the VapBC-1 module. The expression of the toxin gene vapC in E. coli did not decrease the cell growth rate in broth culture, as was expected to happen within active TA modules. However, interestingly, when the expression of the toxin was compared to that of the complexed toxin and antitoxin, cell viability was strongly affected, with a decrease of three orders of magnitude in colony forming unity (CFU). The assumption of the affinity between the toxin and the antitoxin was confirmed in vivo through the observation of their co-purification from cultivation of E. coli co-expressing vapB-vapC genes. RNAse activity assays showed that VapC-1 cleaves MS2 RNA and ribosomal RNA from L. interrogans. Our results indicate that the VapBC-1 module is a potentially functional TA system acting on targets that involve specific functions. It is very important to emphasize that the common attribution of the functionality of TA modules cannot be defined based merely on their ability to inhibit bacterial growth in a liquid medium.
Leucine-rich repeat proteins of Leptospira interrogans that interact to host glycosaminoglycans and integrins Bruno B. Foltran, Aline F. Teixeira, Eliete C. Romero, Luis G. V. Fernandes, Ana L. T. O. Nascimento Frontiers in Microbiology, 2024 Pathogenic spirochaetes of the genus Leptospira are the etiological agents of leptospirosis, a zoonotic infection worldwide. The disease is considered an emerging and re-emerging threat due to global warming, followed by heavy rainfall and flooding when outbreaks of leptospirosis occur. Adhesion to host tissues is mediated by surface/extracellular proteins expressed by pathogens during infection. Leucine-rich repeat (LRR) domain-containing proteins seem to be important for the virulence of pathogenic Leptospira and their role has been recently examined. Here, we report the characterization of two LRR-proteins encoded by LIC11051 and LIC11505. They present 7 and 17 LRR motifs, respectively. LIC11051 was found mainly in the P1 subclade, whereas LIC11505 was identified with higher identity in subclade P1, but was also found in subclades P2, S1, and S2. The recombinant proteins were recognized by antibodies in leptospirosis serum samples, suggesting their expression during infection. rLIC11505 contains a broad spectrum of ligands, including GAG and integrin receptors, whereas rLIC11051 showed limited binding activity. The attachment of proteins to ligands was specific, dose-dependent, and saturable. Compared to their role in adhesion, both proteins were shown to be secreted, with the ability to reassociate with the bacteria. Taken together, our data suggested that LIC11051 and LIC11505 participate in leptospiral pathogenesis. To the best of our knowledge, this is the first report showing leptospiral LRR-proteins exhibiting GAG and integrin receptor-binding properties.
New insights into the putative role of leucine-rich repeat proteins of Leptospira interrogans and their participation in host cell invasion: an in silico analysis Bruno B. Foltran, João P. Gaspar, Igor R. M. Silva, Henrique M. Pires, Fernanda B. Andrade, et al. Frontiers in Cellular and Infection Microbiology, 2024 Pathogenic Leptospira are spirochetes that cause leptospirosis, a worldwide zoonotic disease. Leptospirosis affects humans and animals, with approximately 1 million human infections and 60,000 deaths per year. The diversity of leptospiral strains and serovars allied to the fact that pathogenesis is not yet fully understood, make the development of an effective vaccine against leptospirosis a challenge. Outer membrane and secreted proteins are considered potential antigens since they play a vital role in mediating interactions with host molecules. Several domains or motifs have been reported to participate in the leptospiral infection process. Among them, leucine-rich repeat (LRR) proteins have been highlighted as attractive multipurpose proteins, exhibiting a broad spectrum of ligands and having a putative role in bacterial pathogenesis. Indeed, genome annotation of leptospiral species pointed out that LRR proteins are predominant in pathogenic strains, a feature that corroborates this hypothesis. A few LRR proteins of L. santarosai, L. borgpetersenii and L. interrogans have been studied and their possible role in virulence was proposed. Yet, a mechanistic and broad investigation of LRR proteins was not fully performed. In this review, a comprehensive in silico analysis of 21 LRR proteins of L. interrogans was performed in relation to structure, function, dynamics and virulent potential that will contribute to understanding the key role of these domains in the underlying mechanisms of leptospiral infection.
LipL41 and LigA/LigB Gene Silencing on a LipL32 Knockout Leptospira interrogans Reveals the Impact of Multiple Mutations on Virulence Luis Guilherme V. Fernandes, Bruno B. Foltran, Aline F. Teixeira, Ana Lucia Tabet Oller Nascimento Pathogens, 2023 Leptospirosis is a global zoonosis caused by pathogenic bacteria of the genus Leptospira. The application of the CRISPR/Cas9 system has facilitated the generation of mutants and subsequent evaluation of phenotypes. Since DNA breaks induced by RNA-guided Cas9 nuclease are lethal to Leptospira, different methodologies were implemented to overcome this limitation. Initially, CRISPR interference (CRISPRi) was employed to create knockdown mutants, utilizing a catalytically inactive Cas9 (dCas9). Subsequently, the co-expression of CRISPR/Cas9 and a DNA repair system from Mycobacterium smegmatis enabled the generation of scarless knockout mutants. We eliminated plasmids from the lipL32 knockout L. interrogans strain and further achieved multiple gene mutations via gene silencing in this knockout background. Strains lacking both LipL41 and LipL32 and LigA, LigB, and LipL32, were evaluated. The absence of proteins LipL32 and LipL41 had no effect on leptospiral virulence. On the other hand, mutants lacking LigA, LigB, and LipL32 were unable to cause acute disease. The expanded apparatus for genetic manipulation of pathogenic leptospires via the CRISPR/Cas9 system has allowed the evaluation of multiple mutations upon leptospiral virulence. This work shows that LipL32 and LipL41 are not required for acute disease and consolidates LigA and LigB proteins as virulence factors.
