Breaking Barriers: A Future Perspective on Glioblastoma Therapy with mRNA-Based Immunotherapies and Oncolytic Viruses.

The use of mRNA-based immunotherapies that leverage the genomes of oncolytic viruses holds significant promise in addressing glioblastoma (GBM), an exceptionally aggressive neurological tumor. We explore the significance of mRNA-based platforms in the area of immunotherapy, introducing an innovative approach to mitigate the risks associated with the use of live viruses in cancer treatment. The ability to customize oncolytic virus genome sequences enables researchers to precisely target specific cancer cells, either through viral genome segments containing structural proteins or through a combination of regions with oncolytic potential. This strategy may enhance treatment effectiveness while minimizing unintended impacts on non-cancerous cells. A notable case highlighted here pertains to advanced findings regarding the application of the Zika virus (ZIKV) in GBM treatment. ZIKV, a member of the family Flaviviridae, shows oncolytic properties against GBM, opening novel therapeutic avenues. We explore intensive investigations of glioblastoma stem cells, recognized as key drivers in GBM initiation, progression, and resistance to therapy. However, a comprehensive elucidation of ZIKV's underlying mechanisms is imperative to pave the way for ZIKV-based clinical trials targeting GBM patients. This investigation into harnessing the potential of oncolytic-virus genomes for mRNA-based immunotherapies underscores its noteworthy implications, potentially paving the way for a paradigm shift in cancer treatment strategies.

Viral load: We need a new look at an old problem?

The concept of viral load was introduced in the 1980s to measure the amount of viral genetic material in a person's blood, primarily for human immunodeficiency virus (HIV). It has since become crucial for monitoring HIV infection progression and assessing the efficacy of antiretroviral therapy. However, during the coronavirus disease 2019 pandemic, the term "viral load" became widely popularized, not only for the scientific community but for the general population. Viral load plays a critical role in both clinical patient management and research, providing valuable insights for antiviral treatment strategies, vaccination efforts, and epidemiological control measures. As measuring viral load is so important, why don't researchers discuss the best way to do it? Is it simply acceptable to use raw Ct values? Relying solely on Ct values for viral load estimation can be problematic due to several reasons. First, Ct values can vary between different quantitative polymerase chain reaction assays, platforms, and laboratories, making it difficult to compare data across studies. Second, Ct values do not directly measure the quantity of viral particles in a sample and they can be influenced by various factors such as initial viral load, sample quality, and assay sensitivity. Moreover, variations in viral RNA extraction and reverse-transcription steps can further impact the accuracy of viral load estimation, emphasizing the need for careful interpretation of Ct values in viral load assessment. Interestingly, we did not observe scientific articles addressing different strategies to quantify viral load. The absence of standardized and validated methods impedes the implementation of viral load monitoring in clinical management. The variability in cell quantities within samples and the variation in viral particle numbers within infected cells further challenge accurate viral load measurement and interpretation. To advance the field and improve patient outcomes, there is an urgent need for the development and validation of tailored, standardized methods for precise viral load quantification.

Molecular survey of potentially pathogenic microorganisms in ticks collected from coatis (Nasua nasua) in Iguaçu National Park, Atlantic Forest biome, southern Brazil.

Human contact with wild animals in synanthropic habits is often mediated by arthropod vectors such as ticks. This is an important method of spreading infectious agents that pose a risk to human health. Thus, this study aimed to molecularly detect Ehrlichia spp., Anaplasma spp., Borrelia spp., and protozoa of the order Piroplasmida in ticks collected from coatis of Iguaçu National Park (PNI), Paraná, Brazil. This study involved 553 ticks DNA, including Amblyomma spp. larvae, Haemaphysalis juxtakochi nymphs, Amblyomma brasiliense, Amblyomma coelebs, and adults of Amblyomma ovale. The DNA extracted from each sample was subjected to polymerase chain reaction (PCR) targeting the genes 23S rRNA for the Anaplasmataceae family, 16S rRNA for Anaplasma spp., dsb for Ehrlichia spp., flaB, 16S rRNA, hpt, and glpQ for Borrelia spp., and 18S rRNA for Piroplasmid protozoans. DNA from Anaplasma sp. was detected in ticks of the species A. coelebs (4/553); Borrelia sp. DNA was detected in A. coelebs (3/553), A. ovale (1/553), and Amblyomma larvae (1/553); and Theileria sp. was detected in A. coelebs (2/553). All tested samples were negative for Ehrlichia spp. Our study constitutes the newest report in South America of these microorganisms, which remain poorly studied.

The role of immune subtyping in glioma mRNA vaccine development.

Studies on the development of mRNA vaccines for central nervous system tumors have used gene expression profiles, clinical data and RNA sequencing from sources such as The Cancer Genome Atlas and Chinese Glioma Genome Atlas to identify effective antigens. These studies revealed several immune subtypes of glioma, each one linked to unique prognoses and genetic/immune-modulatory changes. Potential antigens include ARPC1B, BRCA2, COL6A1, ITGB3, IDH1, LILRB2, TP53 and KDR, among others. Patients with immune-active and immune-suppressive phenotypes were found to respond better to mRNA vaccines. While these findings indicate the potential of mRNA vaccines in cancer therapy, further research is required to optimize administration and adjuvant selection, and precisely identify target antigens.

Scientists study special vaccines for hard-to-treat brain tumors. They looked at things, such as information about patients and the small parts of cells that make up the tumor, to find ways to help. They found that brain tumors can make our body's defenses act differently. They also found some possible targets and unique defense patterns that are special to each patient when fighting these tumors. Patients with these special defenses and good targets might respond better to treatment with vaccines. This is exciting because it means that in the future, we might have treatments made for each person. But we still need to do more research to figure out how to get these vaccines to the tumor, so this research gives us hope that we can find better treatments and more choices for people with brain cancer. If we keep researching, we might find even better treatments in the future.

Borrelia theileri in Bovine in the northern and southeastern regions of Brazil.

The present study aimed to describe the occurrence of Borrelia spp. in cattle in the states of Minas Gerais and Pará in southeastern and northern Brazil, respectively. Bovine whole blood samples were examined by blood smear and polymerase chain reaction (PCR) to detect the flagellin B (flaB) gene of Borrelia spp. Frequencies of positive animals for Borrelia spp. were 1.52% (2/132) in the municipality of Unaí, Minas Gerais, and 14.2% (2/7) in the municipality of Marabá, Pará. Subsequent genetic sequencing confirmed that the detected spirochetes close to the species B. theileri. In both locations, the animals positive for B. theileri were also highly infested by Rhipicephalus microplus ticks. Despite the low frequency of Borrelia spp., the occurrence of this spirochete indicates that further studies are needed to determine the consequences in cattle herds.

miR-383-5p, miR-181a-5p, and miR-181b-5p as Predictors of Response to First-Generation Somatostatin Receptor Ligands in Acromegaly.

Acromegaly is a chronic systemic disease caused in the vast majority of cases by growth hormone (GH)-secreting adenoma, with surgery being the first-line treatment. When a cure is not attained with surgery, first-generation somatostatin receptor ligands (fg-SRLs) are the most common medication prescribed. Predictors of response to fg-SRLs have been studied; however, they cannot fully predict the response to fg-SRL. MicroRNAs are small RNAs, the main role of which is messenger RNA (mRNA) post-transcriptional regulation. This study aimed to identify the microRNAs involved in resistance to treatment with fg-SRLs in acromegaly. Ten patients with acromegaly undergoing treatment with fg-SRLs were selected to undergo miRNA sequencing: five controlled and five uncontrolled with treatment. Bioinformatic analysis was performed to detect differentially expressed miRNAs. Then, the same 10 samples were used for validation by qPCR and an additional 22 samples were analyzed, totaling 32 samples. e We found 59 differentially expressed miRNAs in the first analysis. miR-181a-5p and miR-181b-5p were downregulated, and miR-383-5p was upregulated in the uncontrolled group. Receiver operating characteristic (ROC) curve analysis of miR-383-5p showed an NPV of 84.3% and a PPV of 84.5%. In summary, miR-181a-5p, miR-181b-5p, and miR-383-5p are biomarkers of response to fg-SRLs, and they can be used individually or included in prediction models as tools to guide clinical decisions.

Survey of bacterial and protozoan agents in ticks and fleas found on wild animals in the state of Rio de Janeiro, Brazil.

This study evaluates the presence of bacterial and protozoan agents in ticks and fleas found on wild animals in the state of Rio de Janeiro, Brazil. These ectoparasites were collected on mammal species Hydrochoerus hydrochaeris, Tapirus terrestris, Dicotyles tajacu, Didelphis aurita, Cuniculus paca, Cerdocyon thous, and Coendou prehensilis, and on the terrestrial bird Dromaius novaehollandiae. Ticks and fleas were identified morphologically using specific taxonomic keys. A total of 396 ticks and 54 fleas were tested via polymerase chain reaction (PCR) for the presence of Rickettsia spp., Borrelia spp., microorganisms of the order Piroplasmida and Anaplasmataceae family. This total is distributed among nine tick species of the genus Amblyomma and one flea species. Rickettsia bellii was detected in Amblyomma dubitatum and Amblyomma pacae; Rickettsia sp. strain AL was found in Amblyomma longirostre; Rickettsia parkeri strain Atlantic rainforest was found in Amblyomma ovale; and "Candidatus Rickettsia senegalensis" and Rickettsia felis were detected in Ctenocephalides felis felis. Wolbachia sp. was detected in C. f. felis, and Borrelia sp. was detected in Amblyomma calcaratum (here named Borrelia sp. strain Acalc110). All tested samples were negative for Ehrlichia spp. and microorganisms of the Piroplasmida order. This study detected a new bacterial strain, Borrelia sp. strain Acalc 110 (which is genetically close to B. miyamotoi and B. venezuelensis) and the Rickettsia sp. strain 19P, which is 100% similar to "Ca. R. senegalensis", a bacterium recently discovered and now being reported for the first time in Brazil.

Characterization of sporadic somatotropinomas with high GIP receptor expression.

PURPOSE: To analyze the expression of glucose-dependent insulinotropic polypeptide receptor (GIPR) in somatotropinomas specimens and compare clinical, biochemical, radiological, therapeutic, molecular, and pathological data among those who overexpressed (GIPR +) and those who did not overexpress (GIPR - ) GIPR. METHODS: Clinical, biochemical, radiological, molecular, and pathological data were collected. GNAS1 sequencing was performed with the Sanger method. Protein expression of somatostatin receptor subtypes 2 and 5 and CAM 5.2 were analyzed by immunohistochemistry. Quantitative real-time PCR was performed to analyze the mRNA expression of GIPR with the TaqMan® method. Positive expression was considered when the fold change (FC) was above 17.2 (GIPR +). RESULTS: A total of 74 patients (54% female) were included. Eighteen tumors (24%) were GIPR + . Gsp mutation was detected in 30 tumors (40%). GIPR + tumors were more frequently densely granulated adenomas (83% vs 47%, p = 0.028). There was no difference in clinical, biochemical, radiological, therapeutic (surgical cure or response to medical therapy), or other pathological features between GIPR + and GIPR -  tumors. Twenty-eight out of 56 (50%) GIPR -  tumors harbored a gsp mutation, whereas two out of 18 (11%) GIPR + tumors harbored a gsp mutation (p = 0.005). CONCLUSION: We described, for the first time, that GIPR + and gsp mutations are not mutually exclusive, but gsp mutations are less common in GIPR + tumors. GIPR + and GIPR -  tumors have similar clinical, biochemical, radiological, therapeutic, and pathological features, with the exception of a high frequency of densely granulated adenomas among GIPR + tumors.

Detection of Rickettsia spp. in ring-tailed coatis (Nasua nasua) and ticks of the Iguaçu National Park, Brazilian Atlantic Rainforest.

Wild animals are of considerable importance in the ecology of infectious agents, as they can function as hosts and even as possible vectors. In this study, DNA from Rickettsia spp. was detected on ticks and fragments of skin collected from wild coatis with synanthropic habits in the Iguaçu National Park (INP) in the state of Paraná in southern Brazil. Testing was carried out on a total of 566 ticks, comprising Amblyomma spp. larvae, nymphs of Haemaphysalis juxtakochi, Amblyomma brasiliense, Amblyomma coelebs, and adults of Amblyomma ovale. The samples were tested by polymerase chain reaction (PCR) by amplifying htrA, gltA, ompA, and ompB gene fragments to detect Rickettsia spp. A fragment of each positive sample was sequenced in both directions, submitted to Genbank for a homology search, and also used for phylogenetic analyses. Samples of A. coelebs (1.90%, 8/420), A. ovale (13%, 6/45), and ring-tailed coati skin (1%, 1/75) amplified Rickettsia spp. DNA. Through sequencing, Rickettsia bellii was observed in A. ovale, Rickettsia amblyommatis in A. coelebs, while Rickettsia rhipicephali was detected in the skin samples. Wild ring-tailed coatis with synanthropic habits in the INP and their ticks are infected by Rickettsia spp., and associations with new hosts have been described.

Orthohantavirus Survey in Indigenous Lands in a Savannah-Like Biome, Brazil.

In Brazil, the first confirmed cases of hantavirus cardiopulmonary syndrome in Indigenous populations occurred in 2001. The purpose of this study was to determine the seroprevalence of orthohantavirus infections in the Utiariti Indigenous land located in the southeastern region of the Brazilian Amazon. In December 2014 and 2015, a survey was conducted using an enzyme-linked immunosorbent assay in nine villages belonging to the Haliti-Paresí Indigenous communities. A total of 301 participants were enrolled in the study. Of the two study cohorts, the one from 2014 showed a prevalence of 12.4%, whereas the one from 2015 had a serum prevalence of 13.4%. Analysis of the paired samples of 110 Indigenous people who participated in both stages of the study enabled identification of four individuals who had seroconverted during the study period. Identifying the circulation of orthohantaviruses in the Utiariti Indigenous land highlights a serious public health problem in viral expansion and highlights the need to implement preventive measures appropriate to the sociocultural reality of these communities.

Effect of Convalescent Plasma in Critically Ill Patients With COVID-19: An Observational Study.

Background: Convalescent plasma is a potential therapeutic option for critically ill patients with coronavirus disease 19 (COVID-19), yet its efficacy remains to be determined. The aim was to investigate the effects of convalescent plasma (CP) in critically ill patients with COVID-19. Methods: This was a single-center prospective observational study conducted in Rio de Janeiro, Brazil, from March 17th to May 30th, with final follow-up on June 30th. We included 113 laboratory-confirmed COVID-19 patients with respiratory failure. Primary outcomes were time to clinical improvement and survival within 28 days. Secondary outcomes included behavior of biomarkers and viral loads. Kaplan-Meier analyses and Cox proportional-hazards regression using propensity score with inverse-probability weighing were performed. Results: 41 patients received CP and 72 received standard of care (SOC). Median age was 61 years (IQR 48-68), disease duration was 10 days (IQR 6-13), and 86% were mechanically ventilated. At least 29 out of 41CP-recipients had baseline IgG titers ≥ 1:1,080. Clinical improvement within 28 days occurred in 19 (46%) CP-treated patients, as compared to 23 (32%) in the SOC group [adjusted hazard ratio (aHR) 0.91 (0.49-1.69)]. There was no significant change in 28-day mortality (CP 49% vs. SOC 56%; aHR 0.90 [0.52-1.57]). Biomarker assessment revealed reduced inflammatory activity and increased lymphocyte count after CP. Conclusions: In this study, CP was not associated with clinical improvement or increase in 28-day survival. However, our study may have been underpowered and included patients with high IgG titers and life-threatening disease. Clinical Trial Registration: The study protocol was retrospectively registered at the Brazilian Registry of Clinical Trials (ReBEC) with the identification RBR-4vm3yy (http://www.ensaiosclinicos.gov.br).

Misinterpretation of viral load in COVID-19 clinical outcomes.

Knowledge of viral load is essential to formulate strategies for antiviral treatment, vaccination, and epidemiological control of COVID-19. Moreover, identification of patients with high viral loads can also be useful to understand risk factors such as age, comorbidities, severity of symptoms and hypoxia, to decide on the need for hospitalization. Several ongoing studies are analyzing viral load in different types of samples and evaluating its relationship with clinical outcomes and viral transmission pathways. However, in a great number of emerging studies, cycle threshold (Ct) values alone are often used as viral load indicators, which may be a mistake. In this study, we compared tracheal aspirate with nasopharyngeal swab samples obtained from critically ill COVID-19 patients and here we report how the raw Ct can lead to misinterpretation of results. Furthermore, based on analysis of nasopharyngeal swab samples we propose a method to reduce evaluation errors that could occur from using raw Ct data. Based on these findings, we show the impact that normalization of Ct values has on interpretation of SARS-CoV-2 viral load from different biological samples.

Telomerase expression in clinically non-functioning pituitary adenomas.

PURPOSE: Non-functioning pituitary adenomas (NFPA) are benign tumors, however, some are agressive. We aimed to assess if human telomerase reverse transcriptase (hTERT) is present in NFPA and if it can be used as a marker of aggressiveness and proliferation. METHODS: Consecutive patients operated for NFPA whose fresh frozen tumors were available were included. We analyzed tumor's aggressiveness (based on radiological progression) and proliferation (based on Ki-67), as well as hTERT mRNA by quantitative real-time polymerase chain reaction (RT-qPCR). RESULTS: We included 109 samples from 86 patients followed for a median period of 60 months (5-120 months). Aggressive tumors were present in 66% cases and proliferative tumors in 47.7%. Seven (6.4%) samples expressed hTERT: 3 (42.8%) had aggressive and proliferative tumors, 2 (28.6%) only exhibited aggressiveness and the remaining 2 (28.6%) only proliferation. From the aggressive and proliferative tumors, 14% and 16%, respectively, expressed hTERT. From the non-aggressive and non-proliferative tumors, 9% and 6%, respectively, expressed hTERT. CONCLUSION: hTERT expression is present in a minority of NFPA and does not seem to be related to aggressiveness or proliferation in NFPA.

Genetic diversity of Anaplasma marginale in calves with anaplasmosis on farms in Minas Gerais, Brazil.

The objective of this study was to identify Anaplasma marginale strains in dairy heifers from farms with a history of anaplasmosis in the northwest region of the State of Minas Gerais, Brazil. Among the examined animals of the four farms, the overall prevalence total of A. marginale was 55.7 % for gene msp5 and 36.7 % for blood smear. Thirty DNA samples (from 24 asymptomatic and six symptomatic animals) positive for A. marginale msp1α were sequenced to study genotype and strain diversity. The majority (28/30) were the E genotype, followed by C (1/30) and G (1/30). Thirteen different strains were found: α-ß-F-F-F (nine animals), 13-27-27 (three animals), τ-27-18 (three animals), α-ß-ß- BRA1-31 (three animals), α- 22-1318 (three animals), 80-F-F- F-F (three animals), and α -22-13-13, α-ß-ß-Г, M-φ-φ-φ-φ-F, 42-25- 25-31, Q-Q-Q-M, B-Q-B-Q-B-M, and 16-17-F-F (one animal each). A new structure repeated in tandem was described and named BRA 1 (TDSSSASGVLSQSGQASTSSQLG). The α-ß-F-F-F strain was present in all animals with acute anaplasmosis and in three animals asymptomatic. Thus, although 13 strains were observed in the animals evaluated, only the α-ß-F-F-F strain was identified during occurrence of acute disease and mortality, we suggest that this strain has important pathogenicity for calves in northeastern Minas Gerais.

MicroRNAs and Mammarenaviruses: Modulating Cellular Metabolism.

Mammarenaviruses are a diverse genus of emerging viruses that include several causative agents of severe viral hemorrhagic fevers with high mortality in humans. Although these viruses share many similarities, important differences with regard to pathogenicity, type of immune response, and molecular mechanisms during virus infection are different between and within New World and Old World viral infections. Viruses rely exclusively on the host cellular machinery to translate their genome, and therefore to replicate and propagate. miRNAs are the crucial factor in diverse biological processes such as antiviral defense, oncogenesis, and cell development. The viral infection can exert a profound impact on the cellular miRNA expression profile, and numerous RNA viruses have been reported to interact directly with cellular miRNAs and/or to use these miRNAs to augment their replication potential. Our present study indicates that mammarenavirus infection induces metabolic reprogramming of host cells, probably manipulating cellular microRNAs. A number of metabolic pathways, including valine, leucine, and isoleucine biosynthesis, d-Glutamine and d-glutamate metabolism, thiamine metabolism, and pools of several amino acids were impacted by the predicted miRNAs that would no longer regulate these pathways. A deeper understanding of mechanisms by which mammarenaviruses handle these signaling pathways is critical for understanding the virus/host interactions and potential diagnostic and therapeutic targets, through the inhibition of specific pathologic metabolic pathways.

Investigation of Bartonella spp. in brazilian mammals with emphasis on rodents and bats from the Atlantic Forest.

The Bartonella species are zoonotic agents that infect mammals and are transmitted by arthropod vectors. Approximately 18 distinct genotypes cause diseases in humans, and may be spread by both domestic and wild animals. In Brazil, Bartonella genotypes have been identified in several species of wild mammals, and in the present study, we analyzed samples from non-human primates (marmosets), marsupials, rodents, and bats, and compared them with the genotypes described in mammals from Brazil, to examine the distribution of Bartonella genotypes in two impacted areas of Rio de Janeiro state, in southeastern Brazil. We used polymerase chain reaction (PCR) methods to detect the Bartonella DNA using partial sequences of the gltA, ftsZ, and groEL genes. We generated Bayesian inference and maximum likelihood trees to characterize the positive PCR samples and infer the phylogenetic relationships of the genotypes. A total of 276 animals were captured, including 110 bats, 91 rodents, 38 marsupials, and 37 marmosets. The DNA of Bartonella was amplified from tissue samples collected from 12 (4.34%) of the animals, including eight rodents - Akodon cursor (5/44) and Nectomys squamipes (3/27) - and four bats, Artibeus lituratus (3/58) and Carollia perspicillata (1/15). We identified Bartonella genotypes closely related to those described in previous studies, as well as new genotypes in both the rodent and the bat samples. Considering the high diversity of the Bartonella genotypes and hosts identified in the present study, further research is needed to better understand the relationships between the different Bartonella genotypes and their vectors and host species. The presence of Bartonella in the wild rodents and bats from the study area indicates that the local human populations may be at risk of infection by Bartonella due to the spillover of these strains from the wild environment to domestic and peri-domestic environments.

Cyclin A in nonfunctioning pituitary adenomas.

PURPOSE: Assess cyclin A in nonfunctioning pituitary adenomas (NFPA) and compare its expression in non-invasive and non-proliferative tumors with invasive and proliferative tumors (12× higher risk of recurrence). METHODS: Quantitative real time polymerase chain reaction to analyze cyclin A using normal pituitary gland as reference. Fold change (FC) > 1 was considered as increased. Tumor invasion was based on Knosp criteria (grades 3-4 considered invasive) and proliferation on the presence of at least two of three criteria: Ki-67 ≥ 3%; mitoses > 2/10; positive p53. Both groups were compared with Mann-Whitney test considering p value < 0.05 as statistically significant. RESULTS: Thirty-one patients with NFPA were included. Tumors were mainly of gonadotrophic origin (74.2%), followed by corticotrophic (19.4%) and lactotrophic (3.2%) origins and null-cell adenomas (3.2%). Median tumor diameter was 3.5 cm (1.8-8.0) and Ki-67 was 3.0% (0.3-11%). Sixteen patients had tumors classified as non-invasive and non-proliferative and 15 as invasive and proliferative. Median FC was 0.31 in all tumors (0.13-1.94). Cyclin A was not related to invasion or proliferation (FC 0.41 in non-invasive and non-proliferative tumors and FC 0.30 in invasive and proliferative tumors; p = 0.968). Four (12.9%) patients had tumors that exhibited increased cyclin A [median FC of 1.04 (1.02-1.94)]-three of gonadotrophic origin and one null-cell adenoma, with two tumors classified as non-invasive and non-proliferative and two tumors classified as invasive and proliferative. Median tumor diameter in these samples was 3.4 cm (2.4-3.6) and Ki-67 was 5.1% (2-11%). CONCLUSIONS: Cyclin A was increased in a minority of NFPA and does not seem to be related to invasion or proliferation.