The benefits from treatment with antivenom sera are indubitable. However, the mechanism for toxin neutralization has not been completely elucidated. A mixture of anti-bothropic and anti-crotalic horse antivenom has been reported to be more effective in neutralizing the effects of Bothrops jararacussu snake venom than anti-bothropic antivenom alone. This study determined which regions in the three PLA₂s from B. jararacussu snake venom are bound by antibodies in tetravalent anti-bothropic and monovalent anti-crotalic commercial horse antivenom. Mapping experiments of BthTX-I, BthTX-II and BthA-I using two small libraries of 69 peptides each revealed six major IgG-binding epitopes that were recognized by both anti-bothropic and anti-crotalic horse antivenom. Two epitopes in BthTX-I were only recognized by the anti-bothropic horse antivenom, while anti-crotalic horse antivenom recognized four unique epitopes across the three PLA₂s. Our studies suggest that the harmful activities of the PLA₂s present in the venom of B. jararacussu are neutralized by the combinatorial treatment with both antivenom sera through their complementary binding sites, which provides a wide coverage on the PLA₂s. This is the first peptide microarray of PLA₂s from B. jararacussu snake venom to survey the performance of commercial horse antiophidic antivenom. Regions recognized by the protective antivenom sera are prime candidates for improved venom cocktails or a chimeric protein encoding the multiple epitopes to immunize animals as well as for designing future synthetic vaccines.

The increasing detection of infections of Trypanosoma cruzi, the etiological agent of Chagas disease, in non-endemic regions beyond Latin America has risen to be a major public health issue. With an impact in the millions of people, current treatments rely on antiquated drugs that produce severe side effects and are considered nearly ineffective for the chronic phase. The minimal progress in the development of new drugs highlights the need for advances in basic research on crucial biochemical pathways in T. cruzi to identify new targets. Here, we report on the T. cruzi presenilin-like transmembrane aspartyl enzyme, a protease of the aspartic class in a unique phylogenetic subgroup with T. vivax separate from protozoans. Computational analyses suggests it contains 9 transmembrane domains and an active site with the characteristic PALP motif of the A22 family. Multiple linear B-cell epitopes were identified by SPOT synthesis analysis with Chagasic patient sera. Two were chosen to generate rabbit antisera, whose signal was primarily localized to the flagellar pocket, intracellular vesicles and endoplasmic reticulum in parasites by whole cell immunofluorescence. The results suggest that the parasitic presenilin-like enzyme could have a role in the secretory pathway and serve as a biomarker for infections.

Background: Health care-associated infections (HAIs) are a significant public health problem worldwide, favoring multidrug-resistant (MDR) microorganisms. The SARS-CoV-2 infection was negatively associated with the increase in antimicrobial resistance, and the ESKAPE group had the most significant impact on HAIs. The study evaluated the bactericidal effect of a high concentration of O3 gas on some reference and ESKAPE bacteria. Material and Methods: Four standard strains and four clinical or environmental MDR strains were exposed to elevated ozone doses at different concentrations and times. Bacterial inactivation (growth and cultivability) was investigated using colony counts and resazurin as metabolic indicators. Scanning electron microscopy (SEM) was performed. Results: The culture exposure to a high level of O3 inhibited the growth of all bacterial strains tested with a statistically significant reduction in colony count compared to the control group. The cell viability of S. aureus (MRSA) (99.6%) and P. aeruginosa (XDR) (29.2%) was reduced considerably, and SEM showed damage to bacteria after O3 treatment Conclusion: The impact of HAIs can be easily dampened by the widespread use of ozone in ICUs. This product usually degrades into molecular oxygen and has a low toxicity compared to other sanitization products. However, high doses of ozone were able to interfere with the growth of all strains studied, evidencing that ozone-based decontamination approaches may represent the future of hospital cleaning methods.

Peptide microarrays have become increasingly more affordable in recent years with the SPOT technique being one of the most frequently used methods for synthesis and screening of peptides in arrays. Here, a protocol is presented for the identification of the amino acid sites involved in the conversion of human IgG to IgE response during the passive administration of therapeutic, anti-snake venom sera. Similarly, the minimal region of both the IgG and IgE binding epitopes, important for its interaction with ligand, were identified. As the ratio of concentrations for IgG to IgE in human serum is 1:10,000, also presented is a reproductive protocol of chemiluminescence-scanning for the detection of both immunoglobulins.

Mayaro virus (MAYV), which causes mayaro fever, is endemic to limited regions of South America that may expand due to the possible involvement of Aedes spp. mosquitoes in its transmission. Its effective control will require the accurate identification of infected individuals, which has been restricted to nucleic acid-based tests due to similarities with other emerging members of the Alphavirus genus of the Togaviridae family; both in structure and clinical symptoms. Serological tests have a more significant potential to expand testing at a reasonable cost, and their performance primarily reflects that of the antigen utilized to capture pathogen-specific antibodies. Here, we describe the assembly of a synthetic gene encoding multiple copies of antigenic determinants mapped from the nsP1, nsP2, E1, and E2 proteins of MAYV that readily expressed as a stable chimeric protein in bacteria. Its serological performance as the target in ELISAs revealed a high accuracy for detecting anti-MAYV IgM antibodies. No cross-reactivity was observed with serum from seropositive individuals for dengue, chikungunya, yellow fever, Zika, and other infectious diseases as well as healthy individuals. Our data suggest that this bioengineered antigen could be used to develop high-performance serological tests for MAYV infections.

Mayaro virus produces a nonspecific, sublethal disease in man with symptoms that are often confused with dengue. The symptoms of arthralgia, often associated with these viral infections, can cause an incapacitating disability. To date, outbreaks have been localized and sporadic within the Pan-Amazonia forest since its first isolation in 1954 (Trinidad and Tobago). The available literature is diverse, scarce and dispersed. Mayaro virus is an alphavirus, which is phylogenetically related to the Semliki forest antigenic complex. In the New World, Mayaro and the related UNA viruses are the only members of this complex that have been isolated. The genome of Mayaro consists of single-stranded RNA with a positive charge and a length of 12 kb that can be subdivided into genomic and subgenomic regions that encode nonstructural and structural proteins, respectively. Mayaro has shown great plasticity in hosts for vertebrate infections, but high specificity in invertebrates towards the family Culicidae (mosquitoes). Risk factors for infection are forested areas in northern South America and the rainy season. Two genotypes of MAYV have been identified, L (restricted to Belterra, Brazil) and D (widely distributed in the Pan-Amazonia). The enzootic cycle is similar to the jungle cycle of yellow fever, which involves Haemagogus mosquitoes and monkeys as reservoirs. Of concern is the potential spread of the virus by the involvement of other secondary vectors and hosts such as Aedes aegypti, Aedes albopictus and Aedes scapularis that have been shown experimentally to efficiently transmit the virus. Together with the observed high viremia levels of infected individuals, a significant risk exists for an emerging disease in urban, rural and peridomestic locations close to enzootic foci of Mayaro virus.

Serine proteases are involved in several biochemical processes that are essential for the biology of pathogens, including Leishmania sp. Considering their importance, an interest in serine proteases for vaccine development against leishmaniasis has been raised. As targets, these enzymes have demonstrated a dual role in a vaccine against leishmaniasis, both protective and a counter-protective, depending on the conditions that they are evaluated. In this work, serine proteases or inhibitors of them that have been used as components of vaccines to Leishmania sp. are presented, aiming to disseminate the knowledge gained about these proteases and their potential in potential vaccine against leishmaniasis.

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome virus 2 (SARS-CoV-2), has led to a global crisis that included collapsing healthcare systems and shut-down communities, producing considerable economic burden. Despite the number of effective vaccines quickly implemented, the emergence of new variants is a primary concern. The scientific community undertook a rapid response to better study this new virus. However, critical questions about viral protein-protein interactions and mechanisms of its physiopathology are still unclear. Although severe COVID-19 was associated with hematological dysfunctions, scarce experimental data were produced about iron dysmetabolism and the viral proteins’ possible interaction with hemoglobin (Hb) chains. This work demonstrates the binding of SARS-CoV-2 proteins to hemin and Hb using a multimethodological approach. In silico analysis indicated binding motifs between a cavity in the viral nucleoprotein and hemoglobin’s porphyrin coordination region. Different hemin binding capacities of mock and SARS-CoV-2-infected culture extracts were noticed using gel electrophoresis and TMB staining. Hemin-binding proteins were isolated from SARS-CoV-2-infected cells by affinity chromatography and identified by shotgun proteomics, indicating that structural (nucleoprotein, spike, and membrane protein) and non-structural (Nsp3 and Nsp7) viral proteins interact with hemin. In vitro analyses of virus adsorption to host cells and viral replication studies in Vero cells demonstrated inhibitory activities - at different levels - by hemin, protoporphyrin IX (PpIX) Hb. Strikingly, free Hb at 1mM suppressed viral replication (99 %), and its interaction with SARS-CoV-2 was localized to the RBD region of the Spike protein. The findings showed clear evidence of new avenues to disrupt viral replication and understand virus physiopathology that warrants further investigation.