CD8 T lymphocytes from B-1 cell-deficient mice down-regulates fungicidal activity of macrophages challenged with E. Cuniculi.

BACKGROUND: Encephalitozoon cuniculi is an opportunistic intracellular pathogen that establishes a balanced relationship with immunocompetent individuals depending on the activity of their CD8+ T cells lymphocytes. However, lower resistance to experimental infection with E. cuniculi was found in B-1 deficient mice (Xid), besides increased the number of CD8 T lymphocytes. Here, we evaluated the profile of CD8+ T lymphocytes from Balb/c wild-type (WT) or Balb/c Xid mice (with B-1 cell deficiency) on the microbicidal activity of macrophages challenged with E. cuniculi. METHODS: Naïve CD8 T lymphocytes from WT or Xid mice uninfected and primed CD8 T lymphocytes from WT or Xid mice infected with E cuniculi were co-cultured with macrophages previously challenged with E. cuniculi. We evaluated macrophages viability and microbicidal activity, and CD8 T lymphocytes viability and presence of activating molecules (CD62L, CD69, and CD107a). RESULTS: Macrophages co-cultured with naïve CD8 T lymphocytes from WT demonstrated high microbicidal activity. Naïve CD8 T lymphocytes obtained from WT mice had a higher expression of CD69 and LAMP-1-activating molecules compared to Xid CD8+ T lymphocytes. Primed CD8 T lymphocytes from Xid mice proliferated more than those from WT mice, however, when the expression of the activating molecule CD69 associated with the expression of CD62L was kept low. In conclusion, naïve CD8+ T lymphocytes from Xid mice, deficient in B-1 cells, they had reduced expression of activation molecules and cytotoxic activity.

Measures to Contain the Transmission of Hepatitis C in a Chronic Kidney Care Hospital Unit in the Triângulo Mineiro in Brazil: A Case Study.

Objectives: Hepatitis C virus elimination is complex. The objective was to analyze measures to eliminate virus transmission in a hemodialysis unit. Methods: Case study composed of multiple units of analysis. The scenario is the hemodialysis unit of a Brazilian public hospital. Population composed of health service records. Descriptive analyzes were performed and the beginning of the event was considered as the moment of increased incidence of HCV. The intentional and purposeful collection of information for understanding the event and implementing interventions. Results: The subunits of analysis were related to: clinical-epidemiological profile, active search, transmission routes, management protocol and results achieved. In August 2019, out of 45 patients, six were reactive for anti-HCV. All received treatment. Patients had exposure to contaminated medical equipment, objects or hands of professionals. Preventive measures were adopted and routine techniques were corrected. Situational Analysis Committee guided the management of the event. No new cases were detected. Conclusions: Strategies for the microelimination of the C virus in a dialysis environment are demonstrated and it shows the multidisciplinary efforts in conducting the event.

Molecular characterization of a new SARS-CoV-2 recombinant cluster XAG identified in Brazil.

Recombination events have been described in the Coronaviridae family. Since the beginning of the SARS-CoV-2 pandemic, a variable degree of selection pressure has acted upon the virus, generating new strains with increased fitness in terms of viral transmission and antibody scape. Most of the SC2 variants of concern (VOC) detected so far carry a combination of key amino acid changes and indels. Recombination may also reshuffle existing genetic profiles of distinct strains, potentially giving origin to recombinant strains with altered phenotypes. However, co-infection and recombination events are challenging to detect and require in-depth curation of assembled genomes and sequencing reds. Here, we present the molecular characterization of a new SARS-CoV-2 recombinant between BA.1.1 and BA.2.23 Omicron lineages identified in Brazil. We characterized four mutations that had not been previously described in any of the recombinants already identified worldwide and described the likely breaking points. Moreover, through phylogenetic analysis, we showed that the newly named XAG lineage groups in a highly supported monophyletic clade confirmed its common evolutionary history from parental Omicron lineages and other recombinants already described. These observations were only possible thanks to the joint effort of bioinformatics tools auxiliary in genomic surveillance and the manual curation of experienced personnel, demonstrating the importance of genetic, and bioinformatic knowledge in genomics.

Early Emergence and Dispersal of Delta SARS-CoV-2 Lineage AY.99.2 in Brazil.

The year of 2021 was marked by the emergence and dispersal of a number of SARS-CoV-2 lineages, resulting in the "third wave" of COVID-19 in several countries despite the level of vaccine coverage. Soon after the first confirmed cases of COVID-19 by the Delta variant in Brazil, at least seven Delta sub-lineages emerged, including the globally spread AY.101 and AY.99.2. In this study we performed a detailed analysis of the COVID-19 scenario in Brazil from April to December 2021 by using data collected by the largest private medical diagnostic company in Latin America (Dasa), and SARS-CoV-2 genomic sequences generated by its SARS-CoV-2 genomic surveillance project (GENOV). For phylogenetic and Bayesian analysis, SARS-CoV-2 genomes available at GISAID public database were also retrieved. We confirmed that the Brazilian AY.99.2 and AY.101 were the most prevalent lineages during this period, overpassing the Gamma variant in July/August. We also estimated that AY.99.2 likely emerged a few weeks after the entry of the B.1.617.2 in the country, at some point between late April and May and rapidly spread to other countries. Despite no increased fitness described for the AY.99.2 lineage, a rapid shift in the composition of Delta SARS-CoV-2 lineages prevalence in Brazil took place. Understanding the reasons leading the AY.99.2 to become the dominant lineage in the country is important to understand the process of lineage competitions that may inform future control measures.

SARS-CoV-2 BA.1 and BA.2 coinfection detected by genomic surveillance in Brazil, January 2022.

In January 2022, our genomic surveillance network identified a SARS-CoV-2 BA.1 and BA.2 coinfection in a sample from a patient residing in Brazil. Our results suggest that the true number of SARS-CoV-2 coinfections remains largely underestimated.

SARS-CoV-2 saliva testing is a useful tool for Covid-19 diagnosis.

SARS-CoV-2 is the etiologic agent of coronavirus disease 2019 (COVID-19) and is mainly detected by RT-PCR methods from upper respiratory specimens, as recommended by the World Health Organization. Oro/nasopharyngeal swabbing can be discomfortable to the patients, requires trained healthcare personnel and may generate aerosol, increasing the risk of nosocomial infections. In this study, we describe two SARS-CoV-2 RNA extraction-free single RT-PCR protocols on saliva samples and compared the results with the paired oro/nasopharyngeal swab specimens from 400 patients. The two saliva protocols demonstrated a substantial agreement when compared to the oro/nasopharyngeal swab protocol. Moreover, the positivity rate of saliva protocols increased according to the disease period. The 95 % limit of detection of one of the therefore implemented saliva protocol was determined as 9441 copies/mL. Our results support the conclusion that RNA extraction-free RT-PCR using self-collected saliva specimens is an alternative to nasopharyngeal swabs, especially in the early phase of symptom onset.

DNA Recovery Using Ethanol-Based Liquid Medium from FTA Card-Stored Samples for HPV Detection.

INTRODUCTION: Alternative methods of dry storage and transportation may be a viable alternative to the use of liquid storage medium for cervical samples, especially for screening programs in places with few resources. OBJECTIVE: The objective of this study is to verify the viability and efficacy of human papillomavirus DNA (HPV-DNA) detection in cervical cell samples collected and stored on a Flinders Technology Associates (FTA) card (Whatman Indicating FTA® Elute Micro Card) and subsequently recovered in ethanol-based liquid medium and to compare the results to those obtained using samples stored directly in ethanol-based liquid medium. STUDY DESIGN: Thirty-four women submitted to ETZ (excision of the transformation zone of the cervix) were included in this study. Before ETZ, 2 samples of exfoliated cervical cells were collected from each woman by a doctor and stored in ethanol-based liquid medium and on an FTA card. DNA recovery from FTA samples was performed using ethanol-based liquid medium. Detection of HPV-DNA in the samples was performed using the Cobas® 4800 HPV Test Platform. RESULTS AND CONCLUSIONS: The HPV-DNA detection positivity rates were 70.6% for the samples collected directly in liquid medium and 64.7% for the samples stored on the FTA card, with high detection accuracy in the DNA samples recovered from the FTA card (area under the curve = 0.958; 95% confidence interval = 0.890-1.000). The concordance between the results obtained using the 2 storage media was 94.1% (Kappa = 0.866). These preliminary results suggest that collection of cervical material on an FTA card may be an alternative to storage in liquid medium since the liquid medium has some limitations. In addition, DNA recovery from the card using ethanol-based liquid medium streamlines the workflow in the laboratory and reduces the cost associated with reagents, thereby facilitating access to the HPV test in places with few resources and potentially improving cervical cancer screening.

Local Transmission of SARS-CoV-2 Lineage B.1.1.7, Brazil, December 2020.

In December 2020, research surveillance detected the B.1.1.7 lineage of severe acute respiratory syndrome coronavirus 2 in São Paulo, Brazil. Rapid genomic sequencing and phylogenetic analysis revealed 2 distinct introductions of the lineage. One patient reported no international travel. There may be more infections with this lineage in Brazil than reported.

Low-cost Gent type sampler constructed for urban atmospheric aerosol sampling.

The importance of studying the atmospheric pollution due to its effects on human health and other ecosystems, the inexistence of national production of equipment for air sample collection, and the high cost of the imported equipment (especially in developing countries) led the authors of the present work to construct a low-cost Gent type sampler. The construction of the sampler was carried out by combining low-cost materials with good mechanical strength (such as nylon 6.0), hydraulic piping PVC, and the use of a 3D printer. The innovation of the present work is the employment of a 3D printer using ABS polymer to create the grids that cannot be machined. In addition to the sampler, the system is composed of a vacuum pump, a gas meter, and a rotameter. The total cost of the sampling system amounted at about 1200 USD, and the cost of the manufactured Gent type sampler did not reach 100 USD. The results obtained while using this set for sampling atmospheric aerosol for a period of 11 months were compared with the mass concentration of PM10 obtained from the official environmental company, CETESB of São Paulo State, Brazil, showing good correlation with those from CETESB - which confirmed its effectiveness and suitability for use. The low cost, easy operation, and versatility of the built Gent type sampler enable its use for scientific and academic purposes. The equipment can be useful in environmental monitoring networks, in low-income regions, and as an instrument for environmental education used in universities.

HPV Vaccine: Updates and Highlights.

HPV is the most common sexually transmitted biological agent and is the cause of many conditions in men and women, including precancer lesions and cancer. Three prophylactic HPV vaccines targeting high-risk HPV types are available in many countries worldwide: 2-, 4- and 9-valent vaccines. All the 3 vaccines use recombinant DNA technology and are prepared from the purified L1 protein that self-assembles to form HPV type-specific empty shells. This non-systematic review aims to summarize the HPV epidemiology and the vaccine development to review the landmark trials of HPV vaccine, to present to most remarkable results from clinical trials and the real world, and to stress the challenges and the barriers for HPV vaccine implementation.

Establishment, molecular and biological characterization of HCB-514: a novel human cervical cancer cell line.

Cervical cancer is the fourth most common cancer in women. Although cure rates are high for early stage disease, clinical outcomes for advanced, metastatic, or recurrent disease remain poor. To change this panorama, a deeper understanding of cervical cancer biology and novel study models are needed. Immortalized human cancer cell lines such as HeLa constitute crucial scientific tools, but there are few other cervical cancer cell lines available, limiting our understanding of a disease known for its molecular heterogeneity. This study aimed to establish novel cervical cancer cell lines derived from Brazilian patients. We successfully established one (HCB-514) out of 35 cervical tumors biopsied. We confirmed the phenotype of HCB-514 by verifying its' epithelial and tumor origin through cytokeratins, EpCAM and p16 staining. It was also HPV-16 positive. Whole-exome sequencing (WES) showed relevant somatic mutations in several genes including BRCA2, TGFBR1 and IRX2. A copy number variation (CNV) analysis by nanostring and WES revealed amplification of genes mainly related to kinases proteins involved in proliferation, migration and cell differentiation, such as EGFR, PIK3CA, and MAPK7. Overexpression of EGFR was confirmed by phospho RTK-array and validated by western blot analysis. Furthermore, the HCB-514 cell line was sensitive to cisplatin. In summary, this novel Brazilian cervical cancer cell line exhibits relevant key molecular features and constitutes a new biological model for pre-clinical studies.

Is virtual reality effective in improving the motor performance of children with developmental coordination disorder? A systematic review.

INTRODUCTION: Although virtual reality (VR) is an increasingly popular approach within studies that propose interventions for children with developmental coordination disorder (DCD), evidence on the effectiveness of VR remains debatable. The aim of this study was to synthesize evidence on the effectiveness of VR interventions for motor performance improvement in children with DCD. EVIDENCE ACQUISITION: Searches were conducted in the MEDLINE/PubMed, Scopus, Web of Science and ERIC databases to select studies published from 1 January 2006 to 30 November 2017. Two independent reviewers performed the primary study selection based on titles, abstracts and full-text reading; this selection included randomized controlled trials (RCTs) that applied VR interventions to children with DCD and assessed outcomes related to motor performance. The methodological quality of the studies included in the search was assessed through the PEDro scale. PRISMA guidelines and Cochrane recommendations for systematic reviews were followed. The effect size of each intervention was calculated to allow for the interpretation of clinical effects, and the body of evidence was synthesised through the GRADE approach. EVIDENCE SYNTHESIS: A total of 2160 publications were retrieved; by the end of the selection process, twelve RCTs had been included. Of these twelve, seven were classified as having high methodological quality. Only three studies satisfied the homogeneity conditions to be assessed through the GRADE system, which showed a low level of evidence in favor of VR for improving the motor performance of children with DCD. CONCLUSIONS: Not enough evidence currently exists to support or refute the use of VR over non-VR interventions for improving motor performance in children with DCD. Despite the potential for improving the motor performance of DCD children, the absence of specific protocols prevents formal recommendations of VR for these children. Future studies should consider VR protocols that are more specific regarding the tasks, features and target motor skills to be developed by DCD children. In addition, comparisons of similar groups at baseline, the concealment of allocation and the blinding of assessors are internal validity aspects which deserve researchers' attention.

Adaptation of Alpha-Papillomavirus over Millennia.

Papillomaviruses (PVs) are a group of small DNA viruses that, with around 350 million years of evolution, acquired the capacity of infecting a broad range of vertebrates, including humans. To date, more than 300 PV types have been isolated. Viruses that have a long common evolutionary history with their host typically cause unapparent infections. However, in some Alpha-PV infections, lesions become apparent and may cause benign proliferative disorders or even malignant proliferative lesions of the cervix, vulva, vagina, anus, penis, and oropharynx. The incongruence observed between the topology of the phylogenetic tree of Alpha-PVs and that of their hosts suggests that virus-host codivergence is not the only evolutionary force that has driven the progression of PVs. The integration of the precursors of E5, E6, and E7 on the genome of the ancestral Alpha-PV was important and made the colonization of new niches and the emergence of carcinogenic types possible.

Loss of Raf kinase inhibitor protein expression is associated with human papillomavirus 16 infection in anal tumors.

There has been an increase in the incidence of anal cancer in the past two decades, with squamous cell carcinoma (SCC) being the most frequent histological type identified. Among the risk factors, high-risk human papillomavirus (HPV) infection is the most pervasive. Raf kinase inhibitor protein (RKIP) is expressed in a number of normal human tissues and previous studies have demonstrated the prognostic value of the loss of RKIP expression in several gastrointestinal tumors. Therefore, the present study aimed to evaluate the clinical implications of RKIP expression in a series of neoplastic lesions of the anal canal. The resected tumors of 48 patients [8 high-grade intraepithelial lesions (HSILs), 14 adenocarcinomas and 26 squamous cell carcinomas (SCCs)] were immunohistochemically evaluated for RKIP expression, and the results were correlated with clinicopathological data. The results identified a decreased 5-year overall survival rate in patients with adenocarcinoma (40.8%) compared with patients with SCC (76.7%), and a decreased 5-year disease-free survival rate in patients at clinical stages III/IV (37.3 vs. 62.5 and 82.6% for clinical stages 0 and I/II, respectively). Low RKIP expression was revealed in 62.5% of HSILs, 88.5% of SCCs and 100.0% of the adenocarcinomas. High RKIP expression was associated with patient ethnicity (37.5% in non-Caucasians vs. 7.5% in Caucasians) and patient age (33.3% in younger patients vs. 0.0% in older patients). Finally, high RKIP expression was correlated with HPV16 infection status (40% in HPV- vs. 5.3% in HPV+ patients). A correlation was identified between high RKIP expression and lesions with a generally improved prognosis, such as those diagnosed in younger patients, in situ lesions and lesions of lower clinical grades; there was also a negative correlation between high RKIP expression and HPV16 positivity in patients.

Self-sampling coupled to the detection of HPV 16 and 18 E6 protein: A promising option for detection of cervical malignancies in remote areas.

OBJECTIVE: To evaluate both the performance and acceptability of a method coupling self-sampling with detection of cervical malignancy via elevated HPV 16 and 18 E6 oncoproteins (OncoE6™ Cervical Test) in remote areas in Brazil. METHODS: Women living in rural villages in proximity to Coari city, Amazonas, Brazil were invited to participate in a cervical cancer screening study. 412 subjects were enrolled; there were no refusals. In addition to E6 protein detection, DNA was extracted from the brushes and evaluated for HPV genotypes by PCR (PGMY09/11), followed by typing by the Papillocheck™ if positive. Subjects who were found to be positive for OncoE6 or HPV-DNA were referred for colposcopy. RESULTS: For 110 subjects (27%) this was the first cervical cancer exam. Overall the HPV-DNA prevalence was 19.1% (n = 79); 1.4% (n = 6) were positive by the OncoE6 Test. Fifty-six women attended the invitation for colposcopy where nine had an abnormal cervix and were subsequently biopsied. Histopathological analysis revealed 2 CIN3, 2 carcinomas and 5 CIN1. OncoE6 called two out of the three HPV 16 or 18 associated CIN3+ lesions. CONCLUSIONS: The findings suggest that self-administered sample collection in combination with OncoE6 Test is feasible in this population. This could enable expanded screening coverage while ensuring a high specificity which is imperative given the remote geographic location, since women bearing abnormal test results would necessitate travel and logistical burden to access colposcopy and treatment.

Organic amaranth starch: A study of its technological properties after heat-moisture treatment.

Organic amaranth starch (Amaranthus caudatus) was studied after heat-moisture treatment (HMT) using different moisture contents and different times. The starch extracted by the aqueous method presented low lipid and protein content. After HMT, an increase in the thermal stability was identified. The onset and peak temperatures were higher with an increase in moisture content and the times used in the modification. The gelatinisation enthalpy varied due to the heterogeneity of the crystals formed after the structural reorganisation caused by HMT. The relative crystallinity was lower for the physically modified starches. An increase in the pasting temperature was accompanied by a decrease in the viscosity, setback and breakdown, which were proportional to the moisture and time used. The morphology of the HMT-modified samples was not altered; however, agglomerations were noted. Low levels of dispersion homogeneity and suspension stability were observed for the modified samples due to the strong presence of agglomerates.

Abdominopelvic Tuberculosis with a Frozen Section Analysis Consistent with Ovarian Cancer.

Pelvic tuberculosis is a type of extrapulmonary tuberculosis. The disease is accompanied by clinical and laboratory findings which may be unspecific and present aspects of other diseases, including gynecological malignancies. In this report, the authors presented a case of pelvic tuberculosis associated with peritoneal tuberculosis in a young woman exhibiting imaging and tumor markers consistent with ovarian neoplasm. An intraoperative frozen section analysis detected atypical cells that were suggestive of ovarian borderline or malignant epithelial neoplasia. The pathological analysis showed granulomatous inflammation in the right ovary and fallopian tube with a pattern of mycobacteriosis that was consistent with the presence of mycobacteria morphologically compatible with Mycobacterium tuberculosis. The patient had a complete remission after the use of antituberculosis drugs.

Characterization of topoisomerase II α and minichromosome maintenance protein 2 expression in anal carcinoma.

The present study aimed to ascertain the significance of topoisomerase II α (TOP2A) and minichromosome maintenance protein (MCM) 2 expression in anal carcinoma. A total of 75 anal lesions were retrieved from the files of the Department of Pathology of Barretos Cancer Hospital (Barretos, Brazil) in order to verify the human papillomavirus (HPV) statuses of these lesions and characterize the immunohistochemical expression levels of TOP2A and MCM2 in anal carcinoma, as these are important markers for cervical HPV-induced lesions; their expression was also compared with respect to p16 and Ki-67. The vast majority of the cases tested positive for HPV16 (84%); 1 case tested positive for both HPV16 and HPV18. Positive HPV16 status was more frequent in early stages than in advanced stages (P=0.008). Positive immunohistochemical reactivity for MCM2 and TOP2A protein was observed in 71.6 and 100% of cases, respectively. Positive reactivity for p16 was significantly associated (P=0.001) with histological grade, and was more commonly expressed in squamous cell carcinoma than adenocarcinomas. HPV16 was strongly associated with positive p16 protein expression (76.6%). However, the high expression of Ki-67 combined with the high expression of p16 was predominantly observed in Stage III-IV cases. MCM2, TOP2A, p16 and Ki-67 exhibited intense positive staining in the anal lesions, indicating that these markers were significantly and constantly expressed in anal carcinoma.

Transmission between Archaic and Modern Human Ancestors during the Evolution of the Oncogenic Human Papillomavirus 16.

Every human suffers through life a number of papillomaviruses (PVs) infections, most of them asymptomatic. A notable exception are persistent infections by Human papillomavirus 16 (HPV16), the most oncogenic infectious agent for humans and responsible for most infection-driven anogenital cancers. Oncogenic potential is not homogeneous among HPV16 lineages, and genetic variation within HPV16 exhibits some geographic structure. However, an in-depth analysis of the HPV16 evolutionary history was still wanting. We have analyzed extant HPV16 diversity and compared the evolutionary and phylogeographical patterns of humans and of HPV16. We show that codivergence with modern humans explains at most 30% of the present viral geographical distribution. The most explanatory scenario suggests that ancestral HPV16 already infected ancestral human populations and that viral lineages co-diverged with the hosts in parallel with the split between archaic Neanderthal-Denisovans and ancestral modern human populations, generating the ancestral HPV16A and HPV16BCD viral lineages, respectively. We propose that after out-of-Africa migration of modern human ancestors, sexual transmission between human populations introduced HPV16A into modern human ancestor populations. We hypothesize that differential coevolution of HPV16 lineages with different but closely related ancestral human populations and subsequent host-switch events in parallel with introgression of archaic alleles into the genomes of modern human ancestors may be largely responsible for the present-day differential prevalence and association with cancers for HPV16 variants.