Cell Wall Integrity Pathway Involved in Morphogenesis, Virulence and Antifungal Susceptibility in Cryptococcus neoformans

Due to its location, the fungal cell wall is the compartment that allows the interaction with the environment and/or the host, playing an important role during infection as well as in different biological functions such as cell morphology, cell permeability and protection against stress. All these processes involve the activation of signaling pathways within the cell. The cell wall integrity (CWI) pathway is the main route responsible for maintaining the functionality and proper structure of the cell wall. This pathway is highly conserved in the fungal kingdom and has been extensively characterized in Saccharomyces cerevisiae. However, there are still many unknown aspects of this pathway in the pathogenic fungi, such as Cryptococcus neoformans. This yeast is of particular interest because it is found in the environment, but can also behave as pathogen in multiple organisms, including vertebrates and invertebrates, so it has to adapt to multiple factors to survive in multiple niches. In this review, we summarize the components of the CWI pathway in C. neoformans as well as its involvement in different aspects such as virulence factors, morphological changes, and its role as target for antifungal therapies among others.This work was supported by Grant PID2020-114546RB-100 from the Spanish Ministry for Science and Innovation. De Oliveira H.C. was supported by Inova Fiocruz/Fundação Oswaldo Cruz. Rossi SA is funded by postdoctoral fellowship from Fundacão de Amparo á pesquisa do Estado de São Paulo (reference FAPESP-BEPE 2020/09919-0). García-Barbazán Iwas supported by a FPI fellowship (reference PRE2018-083436). Trevijano-Contador N is funded by a “Ayudas de Atracción de Talento Investigador” Contract of Community of Madrid (reference 2019-T2/BMD-14926).S

Negative immunodiffusion test results obtained with sera of paracoccidioidomycosis patients may be related to low-avidity immunoglobulin G2 antibodies directed against carbohydrate epitopes

Immunodiffusion (ID) is the serologic test most frequently used for the diagnosis and posttherapy follow-up of patients with paracoccidioidomycosis (PCM). the ID test is highly specific (100%), but its sensitivity is relatively low (90%), leading to false-negative results. the aim of this study was to determine the profiles of antibodies in sera from patients with proven PCM and with negative results in the ID test (IDneg) versus positive results in the ID test (IDpos). We analyzed 46 sera from patients with active PCM for total immunoglobulin G (IgG) and IgG subclass responses to Paracoccidioides brasiliensis gp43 antigen (treated or not treated with sodium metaperiodate) by enzyme-linked immunosorbent assay and immunoblotting. Immunoblotting showed that both IDneg and IDpos sera recognized predominantly the gp43 fraction of the P. brasiliensis antigen used in the ID test. IDneg sera contain low-avidity antibodies, low levels of specific IgG (total) and IgGI, and high levels of IgG2 compared with IDpos sera. the antibodies present in IDpos sera were predominantly directed against carbohydrate epitopes, since treatment with sodium metaperiodate resulted in a significant decrease in antibody reactivity. These data suggest that the lack of reactivity of sera from PCM patients in the ID test may be related to the production of low-avidity IgG2 antibodies directed against carbohydrate epitopes.UNICAMP, Fac Med Sci, Dept Clin Pathol, BR-13083970 Campinas, SP, BrazilUNIFESP, Discipline Cellular Biol, São Paulo, SP, BrazilUNIFESP, Discipline Cellular Biol, São Paulo, SP, BrazilWeb of Scienc

Produção de próteses dentárias no SUS em idosos brasileiros e impacto da pandemia covid-19

OBJECTIVE: D escribe t he t rends i n t he p roduction o f d ental p rosthetics b y t he U nified Health System (SUS) in older people aged 60 years or older in Brazil and country regions from 2010 to 2019 and the impact of the covid-19 pandemic on the expected production for 2020 and 2021.METHODS: A time series study using secondary data from the SUS database (Datasus-Tabnet) and the Instituto Brasileiro de Geografia e Estatística (Brazilian Institute of Geography and Statistics - IBGE) from 2010 to 2021. Age-standardized rates were calculated for Brazil and regions for each year analyzed. Generalized linear regressions estimated production trends using the Prais-Winstein estimation method.RESULTS: A growth trend occurred in the standardized production rate of complete denturesand other prosthetics per 100,000 inhabitants in Brazil and all country regions. The increasein the production of complete dentures was higher in the Northeast region (50.3%/year) and lower in the North region (19.1%/year). Trends in the production of other prosthetics were higher in the Southeast region (120.7%/year) and lower in the North region (24.5%/year). The output of prosthetics for both groups decreased in the pandemic years. In 2020, the relative difference ranged from -36.4% (North) to -61.7% (Northeast) for producing complete dentures and from -17.9% (North) to -68.4%(Northeast) for other prosthetics. In 2021, standardized rates and total production increased compared to the previous year. However, compared with expected values, the differences were close to those in 2020.CONCLUSION: Policies aimed at producing complete dentures and other prosthetics have been increasing. However, production remains far from the population’s needs, and there is no equity in providing services. The covid-19 pandemic negatively impacted the production of dental prosthetics by SUS.OBJETIVO: Descrever as tendências na produção de próteses odontológicas pelo Sistema Único de Saúde (SUS) em idosos de 60 anos ou mais no Brasil e nas regiões do país de 2010 a 2019, e o impacto da pandemia de covid-19 na produção esperada para 2020 e 2021. MÉTODOS: Estudo de séries temporais que utilizou dados secundários provenientes do banco de dados do SUS (Datasus–Tabnet), e do Instituto Brasileiro de Geografia e Estatística, no período de 2010 a 2021. Calcularam-se as taxas padronizadas por idade para o Brasil e regiões de cada ano analisado. Regressões lineares generalizadas, pelo método de estimação de Prais-Winstein, estimaram as tendências das produções. RESULTADOS: Houve tendência de crescimento na taxa padronizada de produção das próteses totais e das demais próteses para cada 100 mil habitantes, no Brasil e em todas as regiões do país. O aumento na produção de próteses totais foi maior na região Nordeste (50,3%/ano) e menor na região Norte (19,1%/ano). As tendências na produção das demais próteses foram maiores na região Sudeste (120,7%/ano) e menores na região Norte (24,5%/ano). A produção de próteses para ambos os grupos diminuiu nos anos de pandemia. Em 2020, a diferença relativa variou de -36,4% (Norte) até -61,7% (Nordeste) para a produção de próteses totais e de -17,9% (Norte) até -68,4% (Nordeste) para as demais próteses. Em 2021, as taxas padronizadas e a produção absoluta aumentaram quando comparadas ao ano anterior, todavia, em relação aos valores esperados, as diferenças foram próximas às encontradas em 2020. CONCLUSÃO: Políticas voltadas para a produção de próteses totais e das demais próteses vêm tomando forças e crescendo. Todavia, a produção se mantém distante da necessidade da população e não há equidade na oferta dos serviços. A pandemia de covid-19 impactou negativamente na produção de próteses dentárias pelo SUS

Canine Fetal Peritonitis

Background: The occurrence of neonatal diseases and neonatal deaths in canines are considered higher in the first days of life of individuals and bacterial diseases are considered the second most common cause. Inflammation of the mesothelial lining of the abdominal cavity is referred peritonitis, which can be classified as primary or secondary and there is no description on the occurrence of canine fetal peritonitis in the veterinary literature. The objective of this study is to describe a case of canine fetus peritonitis, presenting abnormal sonographic findings of the fetus and abnormalities identified after birth that were not yet described in veterinary literature.Case: A female Poodle, six years of age, considered healthy and without clinical complications during the gestational period, was referred for sonographic evaluation at the end of the pregnancy, in order to evaluate of the fetal viability and estimate the gestational period. By ultrasound examination, two canine fetuses were identified and based on its morphogenesis and biparietal diameter, the estimated gestational age was approximately 58 days. On the individual examination of each of the fetus by ultrasound, it was found that one of them had normal sonographic findings and on the second canine fetus it was found an increase of the intra-abdominal echogenicity of tissues, loss of the sonographic definition of the intestinal loops and presence of laminar anechoic areas between the interfaces of organs, consistent with abdominal effusion. Parturition occurred the following day of the ultrasound examination no complications and with the birth of two canine fetuses. About six hours after birth, one of the neonates showed dyspnea, hypothermia, areas of ecchymosis, suffusion in the ventral abdominal region, icteric and pale mucous membrane and the presence of abdominal fluid and came to death about 30 min after. Cytological analysis of the sample revealed exuberant amount of erythrocytes, fibrin and leukocytes. Culture revealed infection by Staphylococccus aureus and Enterococcus sp. Presence of bloody fluid in the abdominal cavity was observed at necropsy, with yellowish parietal surface of the thoracic and abdominal cavities and petechiae on the lungs, heart, stomach surfaces and intestinal mucous membrane. Microscopically, the lungs showed extensive areas of parenchymal consolidation, vascular congestion and intense inflammatory infiltrate in the alveolar wall; the liver was characterized by vascular congestion, areas of degeneration of hepatocytes and presence of calcified foci, necrosis and inflammatory infiltrates. Associating the results and findings of the exams was possible to conclude the diagnostic of fetal peritonitis.Discussion: The visualization of abnormal findings in affected fetus by a gestational ultrasound exam makes this report inedited and relevant for veterinary obstetrics. This demonstrates the importance of ultrasound exam in pregnant bitches for early detection of maternal fetal diseases. Presence of abdominal fluid and increased mesenteric ochogenicity associated with loss of the intestinal loop definition corroborates with the clinical suspicion of peritonitis in fetus. Findings on the anatomopathological examination, cytology and culture allowed conclusion of the cause of death. Bacterial forms in the cytological smear supported the diagnostic suspicion of peritonitis. Gestational ultrasound exam performed in this report was essential for proper conduct of the pregnant bitch and viability of the fetuses, suggesting that the detected fetal abnormalities showed predictive value in the viability of diseased individual. This case report contributes with the description of canine fetal peritonitis

Minilungs from Human Embryonic Stem Cells to Study the Interaction of Streptococcus pneumoniae with the Respiratory Tract

The new generation of organoids derived from human pluripotent stem cells holds a promising strategy for modeling host-bacteria interaction studies. Organoids recapitulate the composition, diversity of cell types, and, to some extent, the functional features of the native organ. We generated lung bud organoids derived from human embryonic stem cells to study the interaction of Streptococcus pneumoniae (pneumococcus) with the alveolar epithelium. Invasive pneumococcal disease is an important health problem that may occur as a result of the spread of pneumococcus from the lower respiratory tract to sterile sites. We show here an efficient experimental approach to model the main events of the pneumococcal infection that occur in the human lung, exploring bacterial adherence to the epithelium and internalization and triggering an innate response that includes the interaction with surfactant and the expression of representative cytokines and chemokines. Thus, this model, based on human minilungs, can be used to study pneumococcal virulence factors and the pathogenesis of different serotypes, and it will allow therapeutic interventions in a reliable human context. Importance: Streptococcus pneumoniae is responsible for high morbidity and mortalities rates worldwide, affecting mainly children and adults older than 65 years. Pneumococcus is also the most common etiologic agent of bacterial pneumonia and nonepidemic meningitis, and it is a frequent cause of bacterial sepsis. Although the introduction of pneumococcal vaccines has decreased the burden of pneumococcal disease, the rise of antibiotic-resistant strains and nonvaccine types by serotype replacement is worrisome. To study the biology of pneumococcus and to establish a reliable human model for pneumococcal pathogenesis, we generated human minilungs from embryonic stem cells. The results show that these organoids can be used to model some events occurring during the interaction of pneumococcus with the lung, such as adherence, internalization, and the initial alveolar innate response. This model also represents a great alternative for studying virulence factors involved in pneumonia, drug screening, and other therapeutic interventions.This work was supported by grant PI19CIII/00003 from the ISCIII to A.Z. and grantsPID2020-119298RB-I00 and PID2020-114546RB-I00 from the Spanish Ministry of Scienceand Innovation (MICINN) to J.Y. and O.Z., respectively. M.P.D.L. and J.M.R.-C. received grantsupport from AESI (PI20CIII/0029), the Spanish Association Against Cancer (AECC,CGB14143035THOM), and CIBERONC (group CB16/12/00273). S.A.R. was granted apostdoctoral fellowship by the Fundacão de Amparo á pesquisa do Estado do SãoPaulo (reference FAPESP-BEBE 2020/09919-0).S

Dehydration and Hemodynamic Changes as Causa Mortis Associated with Trichuris vulpis in a Dog

Background: Trichuris vulpis, a species that belongs to Trichuris and shows a cosmopolitan distribution, parasitizes the gastrointestinal system of dogs causing trichuriasis. The infection occurs owing to ingestion of larval eggs and subsequent fixation of their adult form in the large intestine of the host. The objective of this paper is to report the case of a Border Collie dog that arrived at the Veterinary Pathology Laboratory (LPV) of the Federal University of Jataí (UFJ) after exhibiting signs of intense dehydration and sudden death caused by severe T. vulpis infection, and to elucidate the macroscopic and microscopic histopathological correlations observed during necropsy.Case: A 7-year-old male Border Collie dog was referred for routine necroscopic examination on suspicion of death from intoxication. In the history, there were reports of bone ingestion, restlessness, and polydipsia for three days, followed by sudden death. Although the animal presented an adequate body state during the necroscopic procedure, enophthalmia and pale oral and ocular mucosa were observed, which are characteristic signs of severe dehydration and anemia. On opening the abdominal cavity, the visceral serosa were found to be stained and severely dry. Greenish mucous content was observed throughout the intestine, and in the large intestine, a moderate amount of mucus associated with high parasitic infestation by elongated parasites attached to the mucosa was identified. The parasites were harvested, stored in airtight vials containing 70% alcohol, processed, and subsequently identified as T. vulpis.Discussion: The necroscopic findings associated with the histopathology were compatible with T. vulpis infestation at high parasite intensity, with severe destruction of the intestinal mucosa and inability of water absorption, resulting in severe dehydration. In cases of parasitism, villous atrophy with crypt hypertrophy occurs, a fact observed in the present case. Possibly crypt hypertrophy occurs prior to villous atrophy, and occurs independently of previous lesions of the absorptive compartment. This results in poor absorption. Advanced loss of gastrointestinal tract fluids can usually be extensive and lead to progressive dehydration with loss of isotonic and hypertonic fluids, which can also be observed in the observation of blood components, severely accelerating changes such as hypovolemia and posterior hemoconcentration. Hypovolemia from severe dehydration also decreases renal perfusion and reduces the rate of glomerular filtration. Hypovolemia also leads to decreased blood pumping capacity, which may lead to heart failure and pulmonary circulatory disorders with notable effects on gas exchange, which may cause hypoxemia and possibly lethal metabolic acidosis. The necropsy performed on the animal found a severe dehydration that could be sustained mainly owing to signs of hypovolemia, associated with a series of pathophysiological events with the massive presence of parasites identified as T. vulpis. The damage caused to the intestinal mucosa by the oral stylet, the movement, and toxins of the parasites led to a severe condition of villosities destruction and tissue necrosis, leading to a large loss of the absorption function of nutrients in the intestines and especially of water, resulting in a condition of severe dehydration. This imbalance of system functioning also alters cardiac function because of increased blood viscosity generating possible lardaceous clots, which in turn are indicative of anemia. This cycle of deleterious changes can result in hypovolemic shock and consequent sudden death

Similar health benefits of endurance and high-intensity interval training in obese children

Purpose: To compare two modalities of exercise training (i.e., Endurance Training [ET] and High-Intensity Interval Training [HIT]) on health-related parameters in obese children aged between 8 and 12 years. Methods: Thirty obese children were randomly allocated into either the ET or HIT group. The ET group performed a 30 to 60-minute continuous exercise at 80% of the peak heart rate (HR). The HIT group training performed 3 to 6 sets of 60-s sprint at 100% of the peak velocity interspersed by a 3-min active recovery period at 50% of the exercise velocity. HIT sessions last similar to 70% less than ET sessions. At baseline and after 12 weeks of intervention, aerobic fitness, body composition and metabolic parameters were assessed. Results: Both the absolute (ET: 26.0%; HIT: 19.0%) and the relative VO2 peak (ET: 13.1%; HIT: 14.6%) were significantly increased in both groups after the intervention. Additionally, the total time of exercise (ET: 19.5%; HIT: 16.4%) and the peak velocity during the maximal graded cardiorespiratory test (ET: 16.9%; HIT: 13.4%) were significantly improved across interventions. Insulinemia (ET: 29.4%; HIT: 30.5%) and HOMA-index (ET: 42.8%; HIT: 37.0%) were significantly lower for both groups at POST when compared to PRE. Body mass was significantly reduced in the HIT (2.6%), but not in the ET group (1.2%). A significant reduction in BMI was observed for both groups after the intervention (ET: 3.0%; HIT: 5.0%). The responsiveness analysis revealed a very similar pattern of the most responsive variables among groups. Conclusion: HIT and ET were equally effective in improving important health related parameters in obese youth.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho Nacional de Pesquisa e Desenvolvimento (CNPq)Conselho Nacional de Pesquisa e Desenvolvimento (CNPq)CAPES (Coordenacao de Aperfeicoamento de Pessoal de Ensino Superior)Coordenacao de Aperfeicoamento de Pessoal de Ensino Superior (CAPES

Identification of Off-Patent Compounds That Present Antifungal Activity Against the Emerging Fungal Pathogen Candida auris

Candida auris is an emerging fungal pathogen of great concern among the scientific community because it is causing an increasing number of hospital outbreaks of difficult management worldwide. In addition, isolates from this species frequently present reduced susceptibility to azole and echinocandin drugs. For this reason, it is necessary to develop new antifungal strategies to better control the disease caused by this yeast. In this work, we screened drugs from the Prestwick chemical library, which contains 1,280 off-patent compounds that are already approved by the Food and Drug Administration, with the aim of identifying molecules with antifungal activity against C. auris. In an initial screening, we looked for drugs that inhibited the growth of three different C. auris strains and found 27 of them which it did so. Ten active compounds were selected to test the susceptibility profile by using the EUCAST protocol. Antifungal activity was confirmed for seven drugs with MICs ranging from 0.5 to 64 mg/L. Some of these drugs were also tested in combination with voriconazole and anidulafungin at sub-inhibitory concentrations. Our results suggest synergistic interactions between suloctidil and voriconazole with fractional inhibitory concentration index (FICI) values of 0.11 to 0.5 and between ebselen and anidulafungin (FICI, 0.12 to 0.44). Our findings indicate that drug repurposing could be a viable alternative to managing infections by C. auris

The frequency of metal enrichment of cool helium-atmosphere white dwarfs using the DESI early data release

There is an overwhelming evidence that white dwarfs host planetary systems; revealed by the presence, disruption, and accretion of planetary bodies. A lower limit on the frequency of white dwarfs that host planetary material has been estimated to be ≃ 25–50 per cent; inferred from the ongoing or recent accretion of metals on to both hydrogen-atmosphere and warm helium-atmosphere white dwarfs. Now with the unbiased sample of white dwarfs observed by the Dark Energy Spectroscopic Instrument (DESI) survey in their Early Data Release (EDR), we have determined the frequency of metal enrichment around cool-helium atmosphere white dwarfs as 21 ± 3 per cent using a sample of 234 systems. This value is in good agreement with values determined from previous studies. With the current samples we cannot distinguish whether the frequency of planetary accretion varies with system age or host-star mass, but the DESI data release 1 will contain roughly an order of magnitude more white dwarfs than DESI EDR and will allow these parameters to be investigated

Dolutegravir twice-daily dosing in children with HIV-associated tuberculosis: a pharmacokinetic and safety study within the open-label, multicentre, randomised, non-inferiority ODYSSEY trial

Background: Children with HIV-associated tuberculosis (TB) have few antiretroviral therapy (ART) options. We aimed to evaluate the safety and pharmacokinetics of dolutegravir twice-daily dosing in children receiving rifampicin for HIV-associated TB. Methods: We nested a two-period, fixed-order pharmacokinetic substudy within the open-label, multicentre, randomised, controlled, non-inferiority ODYSSEY trial at research centres in South Africa, Uganda, and Zimbabwe. Children (aged 4 weeks to <18 years) with HIV-associated TB who were receiving rifampicin and twice-daily dolutegravir were eligible for inclusion. We did a 12-h pharmacokinetic profile on rifampicin and twice-daily dolutegravir and a 24-h profile on once-daily dolutegravir. Geometric mean ratios for trough plasma concentration (Ctrough), area under the plasma concentration time curve from 0 h to 24 h after dosing (AUC0–24 h), and maximum plasma concentration (Cmax) were used to compare dolutegravir concentrations between substudy days. We assessed rifampicin Cmax on the first substudy day. All children within ODYSSEY with HIV-associated TB who received rifampicin and twice-daily dolutegravir were included in the safety analysis. We described adverse events reported from starting twice-daily dolutegravir to 30 days after returning to once-daily dolutegravir. This trial is registered with ClinicalTrials.gov (NCT02259127), EudraCT (2014–002632-14), and the ISRCTN registry (ISRCTN91737921). Findings: Between Sept 20, 2016, and June 28, 2021, 37 children with HIV-associated TB (median age 11·9 years [range 0·4–17·6], 19 [51%] were female and 18 [49%] were male, 36 [97%] in Africa and one [3%] in Thailand) received rifampicin with twice-daily dolutegravir and were included in the safety analysis. 20 (54%) of 37 children enrolled in the pharmacokinetic substudy, 14 of whom contributed at least one evaluable pharmacokinetic curve for dolutegravir, including 12 who had within-participant comparisons. Geometric mean ratios for rifampicin and twice-daily dolutegravir versus once-daily dolutegravir were 1·51 (90% CI 1·08–2·11) for Ctrough, 1·23 (0·99–1·53) for AUC0–24 h, and 0·94 (0·76–1·16) for Cmax. Individual dolutegravir Ctrough concentrations were higher than the 90% effective concentration (ie, 0·32 mg/L) in all children receiving rifampicin and twice-daily dolutegravir. Of 18 children with evaluable rifampicin concentrations, 15 (83%) had a Cmax of less than the optimal target concentration of 8 mg/L. Rifampicin geometric mean Cmax was 5·1 mg/L (coefficient of variation 71%). During a median follow-up of 31 weeks (IQR 30–40), 15 grade 3 or higher adverse events occurred among 11 (30%) of 37 children, ten serious adverse events occurred among eight (22%) children, including two deaths (one tuberculosis-related death, one death due to traumatic injury); no adverse events, including deaths, were considered related to dolutegravir. Interpretation: Twice-daily dolutegravir was shown to be safe and sufficient to overcome the rifampicin enzyme-inducing effect in children, and could provide a practical ART option for children with HIV-associated TB