Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is an important public health problem, occurring mainly in Latin America. The disease has a major social and economical effect, negatively impacting the life of the infected individuals, and bringing great costs to public health. An early and accurate diagnosis is essential for administration of early treatment. In addition, prognostic tests may aid disease management, decreasing hospitalization costs. However, the serological diagnostic scenario for CD still faces several challenges, making the development of new diagnostic kits a pressing matter. Facing this scenario, several researchers have expanded efforts in developing and testing new antigens, such as recombinant proteins and recombinant multiepitope proteins, with promising results. These recombinant antigens offer several advantages, such as improved sensitivity and specificity, in addition to facilitated scaling. Also, it has been possible to observe a rising number of studies using ELISA and point-of-care platforms, employing these antigens in the past few years. Among them, recombinant proteins were the most applied antigens, demonstrating great capacity to discriminate between positive and negative samples. Although fewer in number, recombinant multiepitope proteins also demonstrated an improved diagnostic performance. Indeed, a great number of studies employing these antigens showed sensitivity and specificity values above 90%, greatly impacting diagnostic accuracy. Nevertheless, despite the good results found, it is still possible to observe some bottlenecks in the development of new antigens, such as the scarcity of tests with sera from the acute phase and the variability of results in different geographic areas. In this sense, aiming to contribute to control and health programs, the continuous search for a more accurate serological diagnosis is essential, both for the acute and chronic phases of the disease.

We previously demonstrated that a prime-boost regime with an infectious bronchitis virus (IBV) Massachusetts (Mass)-type vaccine and recombinant LaSota virus (rLS) coexpressing IBV Arkansas (Ark)-type trimeric spike ectodomain (Se) and granulocyte macrophage colony stimulating factor (GMCSF) enhances heterologous protection against virulent Ark challenge. This study evaluates protection against Ark-type challenge conferred by administering the rLS/ArkSe.GMCSF and the attenuated Mass viruses mixed in the same vial as a combined vaccine. Chickens were vaccinated at day of hatch and challenged at 21 days of age with virulent Ark. Protection conferred by vaccination was assessed by respiratory signs, tracheal virus isolation as well as IBV RNA quantitation, and tracheal histomorphometry. Protection conferred by the combined vaccine was compared to protection induced by a commercial attenuated ArkDPI (Delmarva Poultry Industry) vaccine as well as by the attenuated Mass vaccine alone. Vaccination with the combined vaccine (rLS/ArkSe.GMCSF + Mass) as well as Mass alone provided significantly less protection against Ark challenge compared to the control using attenuated live ArkDPI vaccine. Only ArkDPI-vaccinated chickens exhibited \"sterilizing immunity,\" i.e., no virus isolated from ≥10% of chickens after challenge. Chickens vaccinated with the combined vaccine rLS/ArkSe.GMCSF + Mass showed significantly less tracheal damage than birds vaccinated with the attenuated Mass vaccine alone. In addition, the combined vaccine also resulted in less virus isolation from the trachea. We concluded that the combined vaccine containing the recombinant virus and the attenuated Mass enhanced the cross-protective ability of the attenuated Mass vaccine against heterologous challenge.
Nota de investigación- Protección cruzada conferida por una vacuna combinada que contiene el virus atenuado de la bronquitis infecciosa serotipo Massachusetts y un virus de Newcastle LaSota recombinante que expresa la proteína de la espícula del serotipo Arkansas. Previamente se demostró que un esquema de refuerzo con una vacuna tipo Massachusetts (Mass) del virus de la bronquitis infecciosa (IBV) y con un virus de Newcastle LaSota recombinante (rLS) que co-expresa el ectodominio de pico trimérico del serotipo Arkansas (Ark) del virus de la bronquitis infecciosa (Se) y el factor de estimulación de colonias de granulocitos y macrófagos (GMCSF) (virus rLS/ArkSe.GMCSF) mejora la protección heteróloga contra el desafío con serotipo Arkansas virulento. Este estudio evaluó la protección contra el desafío con el serotipo Arkansas conferida por la administración del virus recombinante rLS/ArkSe.GMCSF y el virus Massachussets atenuado mezclados en el mismo vial como una vacuna combinada. Los pollos se vacunaron el día de la eclosión y se desafiaron a los 21 días de edad con el serotipo Arkansas virulento. La protección conferida por la vacunación se evaluó mediante signos respiratorios, el aislamiento del virus traqueal, cuantificación del ARN del virus de bronquitis y por histomorfometría traqueal. La protección conferida por la vacuna combinada se comparó con la protección inducida por una vacuna comercial atenuada ArkDPI (Delmarva Poultry Industry), y con la vacuna atenuada Massachussets por si sola. La vacunación con la vacuna combinada (rLS/ArkSe.GMCSF + Mass), así como con Massachussets sola, proporcionó una protección significativamente menor contra la exposición con Arkansas en comparación con el control que utilizó la vacuna ArkDPI viva atenuada. Solo los pollos vacunados con ArkDPI exhibieron “inmunidad esterilizante”, es decir, que no se aisló ningún virus en ≥10 % de los pollos después del desafío. Los pollos vacunados con la vacuna combinada rLS/ArkSe.GMCSF + Mass mostraron significativamente menos daño traqueal que las aves vacunadas con la vacuna Massachusetts atenuada sola. Además, la vacuna combinada también resultó en un menor aislamiento del virus de la tráquea. Se concluye que la vacuna combinada que contiene el virus recombinante y el serotipo Massachussets atenuado mejoró la capacidad de protección cruzada de la vacuna de Massachusetts atenuada contra el desafío heterólogo.

Drug development costs can be significantly reduced if proven \"platform\" technologies are allowed to be used without having to validate their use. The most recent US Food and Drug Administration (FDA) guideline brings more clarity, as well as a greater focus on the most complex technologies that can now be used for faster drug development. The FDA has highlights the use of lipid nanoparticles (LNPs) to package and deliver mRNA vaccines, gene therapy, and short (2-20 length) synthetic nucleotides (siRNA). Additionally, monoclonal antibody cell development is targeted. The FDA provides a systematic process of requesting platform status to benefit from its advantages. It brings advanced science and rationality into regulatory steps for the FDA\'s approval of drugs and biologicals.

Continuous recombination and variation during replication could lead to rapid evolution and genetic diversity of HIV-1. Some studies had identified that it was easy to develop new recombinant strains of HIV-1 among the populations of men who have sex with men (MSM). Surveillance of genetic variants of HIV-1 in key populations was crucial for comprehending the development of regional HIV-1 epidemics. The finding was reported the identification of two new unique recombinant forms (URF 20110561 and 21110743) from individuals infected with HIV-1 in Tongzhou, Beijing in 2020-2022. Sequences of near full-length genome (NFLG) were amplified, then identification of amplification products used phylogenetic analyses. The result showed that CRF01_AE was the main backbone of 20110561 and 21110743. In the gag region of the virus, 20110561 was inserted two fragments from CRF07_BC, while in the pol and tat regions of the virus, 21110743 was inserted four fragments from CRF07_BC. The CRF01_AE parental origin in the genomes of the two URFs was derived from the CRF01_AE Cluster 4. In the phylogenetic tree, the CRF07_BC parental origin of 20110561 clustered with 07BC_N and the CRF07_BC parental origin of 21110743 clustered with 07BC_O. In summary, the prevalence of novel second-generation URFs of HIV-1 was monitored in Tongzhou, Beijing. The emergence of the novel CRF01_AE/CRF07_BC recombination demonstrated that there was a great significance of continuous monitoring of new URFs in MSM populations to prevent and control the spreading of new HIV-1 URFs.

HIV-1 is characterized by remarkable genetic diversity resulting from its high replication rate, error-prone reverse transcriptase enzyme and recombination events. In Uganda, HIV-1 subtype diversity is mostly dominated by subtypes A, D, and A1/D Unique Recombinant Forms (URFs). In this study, deep sequences of HIV from patients with known antiretroviral therapy (ART) status were analyzed to determine the subtypes and to identify drug-resistance mutations circulating in the study population. Of the 187 participant samples processed for next-generation sequencing (NGS), 137 (73%) were successfully classified. The majority of HIV-1 strains were classified as subtype A (75, 55%), D (43, 31%), with other subtypes including C (3, 2%), A1/D (9, 7%) and CRF10_CD (1, <1%). Recombinant analysis of nine complete A1/D HIV genomes identified novel recombination patterns described herein. Furthermore, we report for the first time in Uganda, an HIV-1 CRF10_CD strain from a fisherfolk in a Lake Victoria Island fishing community.

The single variable domains of camelid heavy-chain only antibodies, known as nanobodies, have taken a long journey since their discovery in 1989 until the first nanobody-based drug\'s entrance to the market in 2022. On account of their unique properties, nanobodies have been successfully used for diagnosis and therapy against various diseases or conditions. Although research on the application of recombinant antibodies has focused on human medicine, the development of nanobodies has paved the way for incorporating recombinant antibody production in favour of veterinary medicine. Currently, despite many efforts in developing these biomolecules with diversified applications, significant opportunities exist for exploiting these highly versatile and cost-effective antibodies in veterinary medicine. The present study attempts to identify existing gaps and shed light on paths for future research by presenting an updated review on camelid nanobodies with potential applications in veterinary medicine.

Potyvirus genomes are expressed as polyproteins that are autocatalytically cleaved to produce 10 to 12 multifunctional proteins, among which P1 is the most variable. It has long been hypothesized that P1 plays role(s) in host adaptation and host specificity. We tested this hypothesis using two phylogenetically distinct potyviruses: soybean mosaic virus (SMV), with a narrow host range, and clover yellow vein virus (ClYVV), with a broader host range. When the full-length P1 cistron of SMV-N was replaced with P1 from ClYVV-No.30, the chimera systemically infected only SMV-N-permissive hosts. Hence, there were no changes in the host range or host specificity of the chimeric viruses. Despite sharing only 20.3% amino acid sequence identity, predicted molecular models of P1 proteins from SMV-N and ClYVV-No.30 showed analogous topologies. These observations suggest that P1 of ClYVV-No.30 can functionally replace P1 of SMV-N. However, the P1 proteins of these two potyviruses are not determinants of host specificity and host range.

BACKGROUND: The variations in sequence, three-dimensional structure, and post-translational modifications (PTMs) of human serum albumin (HSA) are crucial for its physiological functions. This study aims to analyze and compare the disparities in PTMs between HSA derived from human plasma and genetically recombinant sources for clinical treatments in China.
METHODS: Six distinct PTMs, namely acetylation, succinylation, crotonylation, phosphorylation, beta-hydroxybutyrylation, and lactylation, were identified using pan-specific antibodies via Western blot analysis. The samples, comprising human plasma-derived HSA (pHSA) from six different manufacturers and recombinant HSA (rHSA) expressed in yeast and Oryza sativa, underwent detection for various types of PTMs. Additionally, a 4D label-free quantitative proteomic analysis was performed to identify N-glycosylation and the aforementioned PTMs in both pHSA and rHSA samples. This analysis aimed to discern disparities in modification sites and levels.
RESULTS: Through Western blot analysis, all six pHSA and two rHSA samples displayed positive bands for albumin (66.5 kDa) across the six PTMs. Subsequent analysis using 4D label-free quantitative proteomics revealed 25 (29) acetylated, 30 (32) succinylated, 41 (50) malonylated, 15 (23) phosphorylated, 36 (30) beta-hydroxybutyrylated, and 27 (34) lactylated modification sites in pHSA and rHSA samples, with no N-glycosylation modification sites detected. The analysis identified 1 acetylation (ALB_K160), 2 beta-hydroxybutyrylation (ALB_K569, ALB_K426), and 3 crotonylation (ALB_K264, ALB_K581, ALB_K560) specific modification sites in pHSA, as well as 3 crotonylation (ALB_K560, ALB_K562, ALB_K75), 1 succinylation (ALB_K490), and 23 phosphorylation specific modification sites in rHSA. In pHSA (rHSA), 2 (6) acetylation, 10 (12) succinylation, 0 (9) crotonylation, 1 (9) phosphorylation, 6 (0) beta-hydroxybutyrylation, and 0 (7) lactylation specific modification sites were found. Moreover, in the shared modification sites between pHSA and rHSA, pHSA exhibited up-regulation of amberylation (16:1) and beta-hydroxybutyrylation (12:2) in more sites, and up-regulation of acetylation (7:11), crotonylation (2:11), phosphorylation (1:8), and lactylation (1:14) in fewer sites compared to rHSA.
CONCLUSIONS: In clinical practice, both pHSA and rHSA utilized in China commonly display acetylation, succinylation, crotonylation, phosphorylation, beta-hydroxybutyrylation, and lactylation. Notably, there exist distinctions in the site characteristics and modification levels of these alterations between pHSA and rHSA. Further experimental inquiries are imperative to delve into the implications of these disparities in PTMs on the biological functionality, effectiveness, and safety of pHSA and rHSA.

Nerve agents pose significant threats to civilian and military populations. The reactivation of acetylcholinesterase (AChE) is critical in treating acute poisoning, but there is still lacking broad-spectrum reactivators, which presents a big challenge. Therefore, insights gained from the reactivation kinetic analysis and molecular docking are essential for understanding the behavior of reactivators towards intoxicated AChE. In this research, we present a systematic determination of the reactivation kinetics of three V agents-inhibited four human ChEs [(AChE and butyrylcholinesterase (BChE)) from either native or recombinant resources, namely, red blood cell (RBC) AChE, rhAChE, hBChE, rhBChE) reactivated by five standard oximes. We unveiled the effect of native and recombinant ChEs on the reactivation kinetics of V agents ex vitro, where the reactivation kinetics characteristic of Vs-inhibited BChE was reported for the first time. In terms of the inhibition type, all of the five oxime reactivators exhibited noncompetitive inhibition. The inhibition potency of these reactivators would not lead to the difference in the reactivation kinetics between native and recombinant ChE. Despite the significant differences between the native and recombinant ChEs observed in the inhibition, aging, and spontaneous reactivation kinetics, the reactivation kinetics of V agent-inhibited ChEs by oximes were less differentiated, which were supported by the ligand docking results. We also found differences in the reactivation efficiency between five reactivators and the phosphorylated enzyme, and molecular dynamic simulations can further explain from the perspectives of conformational stability, hydrogen bonding, binding free energies, and amino acid contributions. By Poisson-Boltzmann surface area (MM-PBSA) calculations, the total binding free energy trends aligned well with the experimental kr2 values.

Adaptive immunity to viral infections requires time to neutralize and clear viruses to resolve infection. Fast growing and pathogenic viruses are quickly established, are highly transmissible and cause significant disease burden making it difficult to mount effective responses, thereby prolonging infection. Antibody-based passive immunotherapies can provide initial protection during acute infection, assist in mounting an adaptive immune response, or provide protection for those who are immune suppressed or immune deficient. Historically, plasma-derived antibodies have demonstrated some success in treating diseases caused by viral pathogens; nonetheless, limitations in access to product and antibody titer reduce success of this treatment modality. Monoclonal antibodies (mAbs) have proven an effective alternative, as it is possible to manufacture highly potent and specific mAbs against viral targets on an industrial scale. As a result, innovative technologies to discover, engineer and manufacture specific and potent antibodies have become an essential part of the first line of treatment in pathogenic viral infections. However, a mAb targeting a specific epitope will allow escape variants to outgrow, causing new variant strains to become dominant and resistant to treatment with that mAb. Methods to mitigate escape have included combining mAbs into cocktails, creating bi-specific or antibody drug conjugates but these strategies have also been challenged by the potential development of escape mutations. New technologies in developing antibodies made as recombinant polyclonal drugs can integrate the strength of poly-specific antibody responses to prevent mutational escape, while also incorporating antibody engineering to prevent antibody dependent enhancement and direct adaptive immune responses.