Evolution of Tick Vaccinology Highlights Changes in Paradigms in This Research Area.

Ticks and tick-borne diseases affect human and animal health worldwide. Although some tick-protective antigens have been identified and characterized, further research is needed for the development and application of effective anti-tick vaccines, which currently are unavailable for human protection. To study the trends and gaps in anti-tick vaccine approaches, herein we used scientometric analysis to evaluate several aspects of tick vaccinology. Co-authorship and citations networks pointed out two main research fronts, one focused on the laboratory protocols driving the recognition of candidate antigens and the other devoted to field experiments of protection against ticks. The analysis demonstrated the prominence of research in European countries on the topic. The scientometric approach allowed the recognition of isolated teams working casually on the topic, the lack of cooperation between middle- and low-income countries, and the need for sustained and integrated research. Most important, we identified a considerable lack of new candidates for vaccine development, as well as the participation of African and Asian countries. These results provide significant insights obtained from bibliographical analysis, suggest the strength and weaknesses in this field of research, and highlight new directions to advance in the development of effective vaccines for the control of tick infestations and tick-borne pathogens.

Co-Immunization Efficacy of Recombinant Antigens against Rhipicephalus microplus and Hyalomma anatolicumTick Infestations.

The immunoprophylactic management of ticks is the most effective option to control tick infestations and counter spread the acaricide resistance problem worldwide. Several researchers reported an inconsistent efficacy of the single antigen-based immunization of hosts against different tick species. In the present study, to develop a multi-target immunization protocol, proteins from Rhipicephalus microplus BM86 and Hyalomma anatolicum subolesin (SUB) and tropomyosin (TPM) were targeted to evaluate the cross-protective potential. The sequence identities of the BM86, SUB, and TPM coding genes amongst Indian tick isolates of targeted species were 95.6-99.8%, 98.7-99.6%, and 98.9-99.9%, respectively, while at the predicted amino acid level, the identities were 93.2 to 99.5, 97.6 to 99.4, and 98.2 to 99.3%. The targeted genes were expressed in the eukaryotic expression system, pKLAC2-Kluyveromyces lactis, and 100 µg each of purified recombinant protein (Bm86-89 kDa, SUB-21 kDa, and TPM-36 kDa) mixed with adjuvant was injected individually through the intramuscular route at different sites of the body on days 0, 30, and 60 to immunize cross-bred cattle. Post-immunization, a statistically significant (p < 0.001) antibody response (IgG, IgG1, and IgG2) in comparison to the control, starting from 15 to 140 days, against each antigen was recorded. Following multi-antigen immunization, the animals were challenged twice with the larvae of R. microplus and H. anatolicum and theadults of H. anatolicum, and a significant vaccine efficacy of 87.2% and 86.2% against H. anatolicum larvae and adults, respectively, and 86.7% against R. microplus was obtained. The current study provides significant support to develop a multi-antigen vaccine against cattle tick species.

Challenges at the host-arthropod-coronavirus interface and COVID-19: a One Health approach.

Background: The world faces the challenge posed by the interaction between hosts and Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) with potential role for arthropod vectors, and the effect of SARS-CoV-2 variants on acquired immunity, vaccine efficacy and coronavirus disease-19 (COVID-19) pandemic control. Proposal: The characterization of the role played by animal hosts and host-virus interactions is essential to address this challenge. Zoonotic (animal-to-human) and reverse zoonotic (human-to-animal) routes may be involved in virus transmission with a possible still unconfirmed role for arthropod vectors. Herein we propose to consider the risks posed by the possible role of arthropod vectors in COVID-19 and that immunity against SARS-CoV-2 may increase the risk for zoonotic virus transmission. These risks should be considered when evaluating vaccine efficacy and monitoring animal SARS-CoV-2 variants. Conclusion: Virus surveillance, epidemiology, sequencing and evaluation of susceptibility to antibodies and other protective immune mechanisms from vaccinated individuals should be improved. A One Health approach such as the one applied by our group SaBio is necessary for a more effective control of COVID-19 and prevention of future pandemics.

Application of highly sensitive saturation labeling to the analysis of differential protein expression in infected ticks from limited samples.

BACKGROUND: Ticks are vectors of pathogens that affect human and animal health worldwide. Proteomics and genomics studies of infected ticks are required to understand tick-pathogen interactions and identify potential vaccine antigens to control pathogen transmission. One of the limitations for proteomics research in ticks is the amount of protein that can be obtained from these organisms. In the work reported here, individual naturally-infected and uninfected Rhipicephalus spp. ticks were processed using a method that permits simultaneous extraction of DNA, RNA and proteins. This approach allowed using DNA to determine pathogen infection, protein for proteomics studies and RNA to characterize mRNA levels for some of the differentially expressed proteins. Differential protein expression in response to natural infection with different pathogens was characterized by two-dimensional (2-D) differential in gel electrophoresis (DIGE) saturation labeling in combination with mass spectrometry analysis. To our knowledge, this is the first report of the application of DIGE saturation labeling to study tick proteins. RESULTS: Questing and feeding Rhipicephalus spp. adult ticks were collected in 27 farms located in different Sicilian regions. From 300 collected ticks, only 16 were found to be infected: R. sanguineus with Rickettsia conorii and Ehrlichia canis; R. bursa with Theileria annulata; and R. turanicus with Anaplasma ovis. The proteomic analysis conducted from a limited amount of proteins allowed the identification of host, pathogen and tick proteins differentially expressed as a consequence of infection. CONCLUSION: These results showed that DIGE saturation labeling is a powerful technology for proteomics studies in small number of ticks and provided new information about the effect of pathogen infection in ticks.

A Novel Combined Scientific and Artistic Approach for the Advanced Characterization of Interactomes: The Akirin/Subolesin Model.

The main objective of this study was to propose a novel methodology to approach challenges in molecular biology. Akirin/Subolesin (AKR/SUB) are vaccine protective antigens and are a model for the study of the interactome due to its conserved function in the regulation of different biological processes such as immunity and development throughout the metazoan. Herein, three visual artists and a music professor collaborated with scientists for the functional characterization of the AKR2 interactome in the regulation of the NF-κB pathway in human placenta cells. The results served as a methodological proof-of-concept to advance this research area. The results showed new perspectives on unexplored characteristics of AKR2 with functional implications. These results included protein dimerization, the physical interactions with different proteins simultaneously to regulate various biological processes defined by cell type-specific AKR-protein interactions, and how these interactions positively or negatively regulate the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway in a biological context-dependent manner. These results suggested that AKR2-interacting proteins might constitute suitable secondary transcription factors for cell- and stimulus-specific regulation of NF-κB. Musical perspective supported AKR/SUB evolutionary conservation in different species and provided new mechanistic insights into the AKR2 interactome. The combined scientific and artistic perspectives resulted in a multidisciplinary approach, advancing our knowledge on AKR/SUB interactome, and provided new insights into the function of AKR2-protein interactions in the regulation of the NF-κB pathway. Additionally, herein we proposed an algorithm for quantum vaccinomics by focusing on the model proteins AKR/SUB.

Tick and Host Derived Compounds Detected in the Cement Complex Substance.

Ticks are obligate hematophagous arthropods and vectors of pathogens affecting human and animal health worldwide. Cement is a complex protein polymerization substance secreted by ticks with antimicrobial properties and a possible role in host attachment, sealing the feeding lesion, facilitating feeding and pathogen transmission, and protection from host immune and inflammatory responses. The biochemical properties of tick cement during feeding have not been fully characterized. In this study, we characterized the proteome of Rhipicephalus microplus salivary glands (sialome) and cement (cementome) together with their physicochemical properties at different adult female parasitic stages. The results showed the combination of tick and host derived proteins and other biomolecules such as α-Gal in cement composition, which varied during the feeding process. We propose that these compounds may synergize in cement formation, solidification and maintenance to facilitate attachment, feeding, interference with host immune response and detachment. These results advanced our knowledge of the complex tick cement composition and suggested that tick and host derived compounds modulate cement properties throughout tick feeding.

Evaluation of baits for oral vaccination of European wild boar piglets.

The objective of this study was to develop and evaluate new baits for the oral delivery of vaccine preparations to 2-4 month-old wild boar piglets. Baits were prepared using a matrix composed of wild boar feed, wheat flour, paraffin, sacarose and cinnamon-truffle powder attractant with polyethylene capsules dipped into the matrix to introduce vaccine formulation. Physical stability studies demonstrated that baits were stable for at least three days at temperatures as high as 42 degrees C. Recombinant Escherichia coli expressing the membrane-displayed BM95-MSP1a fusion protein were used to test bacterial viability in the baits and the antibody response in orally immunized wild boar. The E. coli viability was not significantly affected after bait incubation at 25 and 37 degrees C for 96h. Bait acceptance studies using artificial feeders in the field showed that baits were accepted by 2-3month-old animals, the preferred age for vaccination. Orally immunized wild boar piglets excreted recombinant E. coli in the feces and developed antibody titers to recombinant BM95-MSP1a protein, thus confirming that vaccine composition was released and reached the wild boar gastrointestinal track. The results of these experiments support the use of these baits for oral delivery of vaccine formulations to 2-4month-old wild boar piglets.

A metaproteomics approach reveals changes in mandibular lymph node microbiota of wild boar naturally exposed to an increasing trend of Mycobacterium tuberculosis complex infection.

Constraints in the characterization of microbiota community that circulates in the host have limited the extent of co-infection studies in natural populations. In this study, we used a metaproteomics approach to characterize the mandibular lymph nodes microbiota of wild boar (Sus scrofa) naturally exposed to an increasing trend of Mycobacterium tuberculosis complex (MTC) infection. Our results showed a reduction in microbiota diversity and changes in the composition, structure and functionality of the microbiota community associated with an increase in tuberculosis prevalence, from 45% in 2002/06 to 83% in 2009/12. These temporal changes were accompanied by an increase in the relative abundance of Babesia, Theileria and Pestivirus genera and a decrease in the Ascogregarina and Chlorella. A positive association was also evidenced between the prevalence of tuberculosis and the presence of microbial proteins responsible for carbohydrate transport and metabolism. Our findings suggest MTC-host-microbiota interactions at the population level, which may occur in order to ensure sufficient metabolic resources for MTC survival, growth and transmission. We strongly recommend the use of metaproteomics when studying microbiota communities in wildlife populations, for which traditional diagnostic techniques are limited and in which new organisms with a pathogenic potential for domestic animals and humans may appear.

A vaccinology Approach to the Identification and Characterization of Dermanyssus Gallinae Candidate Protective Antigens for the Control of Poultry Red Mite Infestations.

The poultry red mite (PRM), Dermanyssus gallinae, is a hematophagous ectoparasite considered as the major pest in the egg-laying industry. Its pesticide-based control is only partially successful and requires the development of new control interventions such as vaccines. In this study, we follow a vaccinology approach to identify PRM candidate protective antigens. Based on proteomic data from fed and unfed nymph and adult mites, we selected a novel PRM protein, calumenin (Deg-CALU), which is tested as a vaccine candidate on an on-hen trial. Rhipicephalus microplus Subolesin (Rhm-SUB) was chosen as a positive control. Deg-CALU and Rhm-SUB reduced the mite oviposition by 35 and 44%, respectively. These results support Deg-CALU and Rhm-SUB as candidate protective antigens for the PRM control.

Repercusiones hemodinamicas y pulmonares de la embolia aerea en posición sentado / Haemodynamic and respiratory repercussions of venous air embolization in the sitting position

La embolia aérea puede ocurrir cada vez que exista una gradiente entre la altura de cavidades cardiacas derechas y el lugar donde se realiza la intervención. Este último debe estar a mayor altura. La entrada de aire al sistema venoso es un riesgo significativo de la posición sentado empleada en procedimiento neuroquirúrgicos. El propósito de este trabajo es estudiar nuestra incidencia de embolia aérea y sus repercusiones cardiopulmonares. Fueron estudiados 50 pacientes operados en posición sentada en forma consecutiva. El diagnóstico de embolia aérea se formuló sólo cuando se aspiró aire através del catéter central. Nueve enfermos (18%) presentaron una embolia aérea. La monitorización de estos pacientes con Doppler, presión venosa central, presión arterial directa, electrocardiografía continua, gases arteriales pre y postembolia permitió evidenciar que en el 55% de las embolias hay hipotensión arterial significativa con presiones arteriales medias bajo 50 mmHg. Tres de los pacientes con embolia aérea presentaron extrasistolia ventricular frecuente y dos de ellos tuvieron un alza mayor de un 50% en la presión venosa central. 44% de los pacientes que tuvieron una embolia aérea presentaron un alza en la PaCO2 y 33% una caída en la PaO2 inmediatamente después del episodio embólico. Ningún paciente presentó problemas pulmonares clínicamente significativos en el postoperatorio. En esta serie no hubo mortalidad. La embolia aérea es un cuadro grave. El Doppler se mostró como el monitor de mayor sensibilidad para el diagnóstico y nos parece indispensable contar con un catéter central para un tratamiento adecuado. La posición sentado por los riesgos que conlleva debe reservarse exclusivamente para cirugía que no pueda realizarse en otra posición