Advanced clinical parameters: A complementary hydroxyurea adherence evaluation in sickle cell anemia treatment.

OBJECTIVES: Given the complex pathology of sickle cell anemia (SCA) and low adherence to hydroxyurea (HU) treatment, there is a need to seek parameters that identify recent changes in patient status. The advanced clinical parameters (ACPs) allow an early analysis of hematopoiesis. We aimed to draw the demographic profile of non-adherent SCA patients and to verify the use of ACPs as a measure of HU treatment adherence. METHOD: In a cross-sectional study, we divided 83 SCA subjects treated with HU into Children (<12 years old) and adolescents/adults (≥12 years old). Their hemogram with the ACPs, electronic medical charts and pharmacy claim data were analyzed. RESULTS: Non-adherent ≥12 years old patients had significantly increased WBC, absolute neutrophil, lymphocyte, monocyte, and basophil counts, RBC, RET, RDW, and PLT, and significantly decreased MCV and MCH. Subjects in the adolescent/adult group with IG† ≥0.035 cells/mm3 had the RR for non-adherence increased by 4.6 times (p = .014), and the systemic immune inflammation index (SII) of non-adherent patients was also significantly higher (p = .042). CONCLUSION: IG† presents clinical utility in early identification of non-adherence to HU, especially when combined with other parameters, suggesting the evaluation of ACPs in laboratory routine, as they can be easily implemented.

Population-based prevalence surveys during the Covid-19 pandemic: A systematic review.

Population-based prevalence surveys of Covid-19 contribute to establish the burden of infection, the role of asymptomatic and mild infections in transmission, and allow more precise decisions about reopen policies. We performed a systematic review to evaluate qualitative aspects of these studies, assessing their reliability and compiling practices that can influence the methodological quality. We searched MEDLINE, EMBASE, bioRxiv and medRxiv, and included cross-sectional studies using molecular and/or serological tests to estimate the prevalence of Covid-19 in the general population. Survey quality was assessed using the Joanna Briggs Institute Critical Appraisal Checklist for Prevalence Studies. A correspondence analysis correlated methodological parameters of each study to identify patterns related to higher, intermediate and lower risks of bias. The available data described 37 surveys from 19 countries. The majority were from Europe and America, used antibody testing, and reached highly heterogeneous sample sizes and prevalence estimates. Minority communities were disproportionately affected by Covid-19. Important risk of bias was detected in four domains: sample size, data analysis with sufficient coverage, measurements in standard way and response rate. The correspondence analysis showed few consistent patterns for high risk of bias. Intermediate risk of bias was related to American and European studies, municipal and regional initiatives, blood samples and prevalence >1%. Low risk of bias was related to Asian studies, nationwide initiatives, reverse-transcriptase polymerase chain reaction tests and prevalence <1%. We identified methodological standards applied worldwide in Covid-19 prevalence surveys, which may assist researchers with the planning, execution and reporting of future population-based surveys.

Systemic redox imbalance in severe COVID-19 patients.

The aim of this study was to evaluate the systemic redox state and inflammatory markers in intensive care unit (ICU) or non-ICU severe COVID-19 patients during the hospitalization period. Blood samples were collected at hospital admission (T1) (Controls and COVID-19 patients), 5-7 days after admission (T2: 5-7 days after hospital admission), and at the discharge time from the hospital (T3: 0-72 h before leaving hospital or death) to analyze systemic oxidative stress markers and inflammatory variables. The reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) were analyzed in peripheral granulocytes and monocytes. THP-1 human monocytic cell line was incubated with plasma from non-ICU and ICU COVID-19 patients and cell viability and apoptosis rate were analyzed. Higher total antioxidant capacity, protein oxidation, lipid peroxidation, and IL-6 at hospital admission were identified in both non-ICU and ICU COVID-19 patients. ICU COVID-19 patients presented increased C-reactive protein, ROS levels, and protein oxidation over hospitalization period compared to non-ICU patients, despite increased antioxidant status. Granulocytes and monocytes of non-ICU and ICU COVID-19 patients presented lower MMP and higher ROS production compared to the healthy controls, with the highest values found in ICU COVID-19 group. Finally, the incubation of THP-1 cells with plasma acquired from ICU COVID-19 patients at T3 hospitalization period decreased cell viability and apoptosis rate. In conclusion, disturbance in redox state is a hallmark of severe COVID-19 and is associated with cell damage and death.

Utility of a urine sediment score in hyperbilirubinemia/hyperbilirubinuria
.

AIMS: Acute and chronic kidney dysfunction is common in patients with end-stage liver disease. Differentiation between acute kidney injury (AKI) due to hepatorenal syndrome (HRS) or acute tubular necrosis (ATN) remains difficult, however urine cast scoring systems using renal tubular epithelial cells (RTECs) and granular casts (GCs) can help to identify intrinsic kidney diseases. The objective of this study was to evaluate the urine sediment profile of patients with liver disease and hyperbilirubinemia/hyperbilirubinuria and the use of a urine sediment scoring system to identify the most common score in AKI patients and high urine bilirubin concentrations. MATERIALS AND METHODS: A retrospective study in the database of a large laboratory that assists a hospital-complex in Brazil was performed. RESULTS: Urinary casts, in particular GCs, as well as RTECs were observed more frequently in patients with hyperbilirubinemia/hyperbilirubinuria, while hyaline casts were observed in patients without hyperbilirubinemia/hyperbilirubinuria. Regardless of the AKI or non-AKI condition, the relative risk for scores 2 or 3 (sediment consistent with tubular damage, with GCs and/or RTECs in different quantities) in group 4 was 3.61 times higher compared to patients in group 1. CONCLUSION: In patients with higher urinary bilirubin levels, the urine sediment had greater numbers of GCs and RTECs and higher urine sediment scores (scores 2 or 3). The presence of a larger number of urine particles (RTECs and GCs) originating in the kidneys in the groups with higher levels of urinary bilirubin suggests an association between hyperbilirubinemia/hyperbilirubinuria and tubular injury independent of AKI or non-AKI.

Bright Field Microscopy to Detect Decoy Cells Due to BK Virus Infection in the Fresh and Unstained Urine Sediment in Kidney Allograft Recipients.

BACKGROUND: BK virus (BKV) may reactivate in kidney allograft recipients ultimately leading to BKV nephropathy and graft loss. Decoy cells (DCs) are one of the early marks of BKV reactivation, and these can be detected in the urine sediment. METHODS: A cohort of 102 kidney transplant patients was followed during months 3 and 6 after the transplant procedure. Urine samples were obtained to detect the presence of DC in the fresh and unstained urine sediment under bright field microscopy (BFM), in concomitance to the determination of the amount of BK viruria by qPCR. RESULTS: Decoy cells were found in 14.7% of patients (15/102). There was a strong agreement (P < 0.001) between qualitative DC detection by two experienced analysts and by qPCR. The positive predictive value, negative predictive value, specificity, and accuracy of BFM were 80%, 75%, 97%, and 75%, respectively. Test sensitivity was 16%. The comparative method was the qPCR. CONCLUSIONS: Despite its limited sensitivity, BFM of unstained urine sediment is an easily available, fast and cheap method to identify DCs in the population of kidney allograft recipients. The diagnostic performance of BFM on the hands of less experienced analysts deserves further investigation.

Association between a frequent variant of the FTO gene and anthropometric phenotypes in Brazilian children.

BACKGROUND: Our goal was to analyze the association of the fat mass and obesity- associated (FTO) gene rs9939609 variant (T/A) with the anthropometric and dietary intake phenotypes related to obesity in Brazilian children. METHODS: We analyzed the association of this single nucleotide polymorphism (SNP) with phenotypes related to the accumulation of body mass in a cohort of 348 children followed from the time of birth until 8 years old and then replicated the main findings in an independent schoolchildren sample (n = 615). RESULTS: At the age of 4, we observed a significant association between the A/A genotype and a higher mean BMI Z-score (P = 0.036). At the age of 8, the A/A individuals still presented with a higher BMI Z-score (P = 0.011) and with marginal differences in the volume of subcutaneous fat (P = 0.048). We replicated these findings in the schoolchildren sample, which showed that those with at least one copy of the A allele presented with a higher BMI Z-score (P = 0.029) and volume of subcutaneous fat (P = 0.016). CONCLUSION: Our results indicate that this FTO variant is associated with increased body mass and subcutaneous fat in Brazilian children beginning at the age of 4.

Ω3-Polyunsaturated fatty acids prevent lipoperoxidation, modulate antioxidant enzymes, and reduce lipid content but do not alter glycogen metabolism in the livers of diabetic rats fed on a high fat thermolyzed diet.

Ω3-Polyunsaturated fatty acids (Ω3-PUFAs) are known to act as hypolipidaemics, but the literature is unclear about the effects that Ω3-PUFAs have on oxidative stress in obese and diabetic patients. In this study, our aim was to investigate the effects of Ω3-PUFAs on oxidative stress, including antioxidant enzyme activity and hepatic lipid and glycogen metabolism in the livers of diabetic and non-diabetic rats fed on a high fat thermolyzed diet. Rats were divided into six groups: (1) the control group (C), (2) the control diabetic group (D), (3) the high fat thermolyzed diet group (HFTD), which were fed a diet that was enriched in fat that was heated for 60 min at 180°C, (4) the high fat thermolyzed diet diabetic group (D + HFTD), (5) the high fat thermolyzed diet + Ω3 polyunsaturated fatty acid group (HFTD + Ω3), and (6) the high fat thermolyzed diet + Ω3 polyunsaturated fatty acid diabetic group (D + HFTD + Ω3). The most important finding of this study was that Ω3-PUFAs are able to reduce triglycerides, non-esterified fatty acid, lipoperoxidation levels, advanced glycation end products, SOD/CAT enzymatic ratio, and CAT immunocontent and increase SOD2 levels in the livers of diabetic rats fed with a HFTD. However, Ω3-PUFAs did not alter the observed levels of protein damage, blood glucose, or glycogen metabolism in the liver. These findings suggest that Ω3-PUFAs may represent an important auxiliary adjuvant in combating some diseases like diabetes mellitus, insulin resistance, and non-alcoholic fatty liver disease.

Acute kidney injury associated with rhabdomyolysis in a patient with COVID-19.

Rhabdomyolysis is defined as the breakdown of skeletal muscle leading to the release of muscle contents into the extracellular fluid. Patients with rhabdomyolysis can be asymptomatic or have myalgia symptoms, weakness, myoglobinuria with dark urine, significant electrolyte imbalance, and acute kidney injury. Here we describe a case on acute kidney injury associated to rhabdomyolysis in a patient with COVID-19.

Comparative Genomics and Characterization of SARS-CoV-2 P.1 (Gamma) Variant of Concern From Amazonas, Brazil.

BACKGROUND: P.1 lineage (Gamma) was first described in the State of Amazonas, northern Brazil, in the end of 2020, and has emerged as a very important variant of concern (VOC) of SARS-CoV-2 worldwide. P.1 has been linked to increased infectivity, higher mortality, and immune evasion, leading to reinfections and potentially reduced efficacy of vaccines and neutralizing antibodies. METHODS: The samples of 276 patients from the State of Amazonas were sent to a central referral laboratory for sequencing by gold standard techniques, through Illumina MiSeq platform. Both global and regional phylogenetic analyses of the successfully sequenced genomes were conducted through maximum likelihood method. Multiple alignments were obtained including previously obtained unique human SARS-CoV-2 sequences. The evolutionary histories of spike and non-structural proteins from ORF1a of northern genomes were described and their molecular evolution was analyzed for detection of positive (FUBAR, FEL, and MEME) and negative (FEL and SLAC) selective pressures. To further evaluate the possible pathways of evolution leading to the emergence of P.1, we performed specific analysis for copy-choice recombination events. A global phylogenomic analysis with subsampled P.1 and B.1.1.28 genomes was applied to evaluate the relationship among samples. RESULTS: Forty-four samples from the State of Amazonas were successfully sequenced and confirmed as P.1 (Gamma) lineage. In addition to previously described P.1 characteristic mutations, we find evidence of continuous diversification of SARS-CoV-2, as rare and previously unseen P.1 mutations were detected in spike and non-structural protein from ORF1a. No evidence of recombination was found. Several sites were demonstrated to be under positive and negative selection, with various mutations identified mostly in P.1 lineage. According to the Pango assignment, phylogenomic analyses indicate all samples as belonging to the P.1 lineage. CONCLUSION: P.1 has shown continuous evolution after its emergence. The lack of clear evidence for recombination and the positive selection demonstrated for several sites suggest that this lineage emergence resulted mainly from strong evolutionary forces and progressive accumulation of a favorable signature set of mutations.

CD200 expression in hematopoietic neoplasms: Beyond a marker for diagnosis of B-cell neoplasms.

CD200 (OX-2) is expressed in myeloid cells, B cells, subsets of T cells and on other normal and neoplastic non-hematopoietic cells. It interacts with CD200R and has a suppressive effect on T cells immune mediated response. We aimed to review CD200 expression and its role in the immune evasion of non-B cell hematopoietic neoplasms. In acute myeloid leukemia, CD200 seems to be related to the worst outcome, even in diseases of good prognosis, possibly due to an immunosuppressive effect. In plasma cell myeloma studies, while some have associated CD200 expression with worst prognosis possibly due to its suppressive effect on monocyte and T cell-mediated immune response, in others CD200 appeared to be a marker of a better outcome, or even showed no impact in event-free survival (EFS). Few studies have evaluated CD200 expression in T cell neoplasms; however, it appears to be a good immunophenotypic marker for angioimmunoblastic T cell lymphoma. In conclusion, CD200 appears to be involved in the immune evasion of malignant cells, which could affect the survival of these patients.

Neutrophils phagocytosing fungal hyphae in urinary sediment.

The Phagocytosis of fungal structures by neutrophils is a well-documented function of these immune cells. However, neutrophil phagocytosis of hyphal structures in the urine sediment is not usually observed during routine sample evaluation. This is a case of hyphal phagocytosis by neutrophils in the urine of a kidney allograft recipient patient.

Survey on reporting of epithelial cells in urine sediment as part of external quality assessment programs in Brazilian laboratories.

INTRODUCTION: Epithelial cells (ECs) are structures regularly observed during urine microscopy analysis. The correct identification of EC subtypes can be useful since renal tubular epithelial cells (RTECs) are clinically relevant. We investigate the urinary ECs report and the judgement of its clinical importance by Brazilian laboratories. MATERIALS AND METHODS: A survey with four questions was made available to participants of the Urinalysis External Quality Assessment Program (EQAP) from Controllab. Laboratories composed 3 groups: (1) differentiating ECs subtypes: "squamous", "transitional" and "RTECs"; (2) differentiating ECs subtypes: "squamous" or "non-squamous" cells; (3) without ECs subtype identification. Participants did not necessarily answer to all questions and the answers were evaluated both within the same laboratory's category and within different categories of laboratories. RESULTS: A total of 1336 (94%) laboratories answered the survey; Group 1, 119/140 (85%) reported that ECs differentiation is important to the physician and 62% want to be evaluated by EQAP, while in Group 3, 455/1110 (41%) reported it is useful to them, however only 25% want be evaluated by EQAP. Group 2 laboratories 37/51 (73%) reported that the information is important, but only 13/52 (25%) are interested in an EQAP with differentiation of the 3 ECs subtypes. CONCLUSION: Most of the laboratories do not differentiate ECs in the three subtypes, despite the clinical importance of RTECs. Education of laboratory staff about the clinical significance of urinary particles should be considered a key priority.

Predominance of the SARS-CoV-2 Lineage P.1 and Its Sublineage P.1.2 in Patients from the Metropolitan Region of Porto Alegre, Southern Brazil in March 2021.

Almost a year after the COVID-19 pandemic had begun, new lineages (B.1.1.7, B.1.351, P.1, and B.1.617.2) associated with enhanced transmissibility, immunity evasion, and mortality were identified in the United Kingdom, South Africa, and Brazil. The previous most prevalent lineages in the state of Rio Grande do Sul (RS, Southern Brazil), B.1.1.28 and B.1.1.33, were rapidly replaced by P.1 and P.2, two B.1.1.28-derived lineages harboring the E484K mutation. To perform a genomic characterization from the metropolitan region of Porto Alegre, we sequenced viral samples to: (i) identify the prevalence of SARS-CoV-2 lineages in the region, the state, and bordering countries/regions; (ii) characterize the mutation spectra; (iii) hypothesize viral dispersal routes by using phylogenetic and phylogeographic approaches. We found that 96.4% of the samples belonged to the P.1 lineage and approximately 20% of them were assigned as the novel P.1.2, a P.1-derived sublineage harboring signature substitutions recently described in other Brazilian states and foreign countries. Moreover, sequences from this study were allocated in distinct branches of the P.1 phylogeny, suggesting multiple introductions in RS and placing this state as a potential diffusion core of P.1-derived clades and the emergence of P.1.2. It is uncertain whether the emergence of P.1.2 and other P.1 clades is related to clinical or epidemiological consequences. However, the clear signs of molecular diversity from the recently introduced P.1 warrant further genomic surveillance.

Increased LPS levels coexist with systemic inflammation and result in monocyte activation in severe COVID-19 patients.

Mucosal barrier alterations may play a role in the pathogenesis of several diseases, including COVID-19. In this study we evaluate the association between bacterial translocation markers and systemic inflammation at the earliest time-point after hospitalization and at the last 72 h of hospitalization in survivors and non-survivors COVID-19 patients. Sixty-six SARS-CoV-2 RT-PCR positive patients and nine non-COVID-19 pneumonia controls were admitted in this study. Blood samples were collected at hospital admission (T1) (Controls and COVID-19 patients) and 0-72 h before hospital discharge (T2, alive or dead) to analyze systemic cytokines and chemokines, lipopolysaccharide (LPS) concentrations and soluble CD14 (sCD14) levels. THP-1 human monocytic cell line was incubated with plasma from survivors and non-survivors COVID-19 patients and their phenotype, activation status, TLR4, and chemokine receptors were analyzed by flow cytometry. COVID-19 patients presented higher IL-6, IFN-γ, TNF-α, TGF-ß1, CCL2/MCP-1, CCL4/MIP-1ß, and CCL5/RANTES levels than controls. Moreover, LPS and sCD14 were higher at hospital admission in SARS-CoV-2-infected patients. Non-survivors COVID-19 patients had increased LPS levels concomitant with higher IL-6, TNF-α, CCL2/MCP-1, and CCL5/RANTES levels at T2. Increased expression of CD16 and CCR5 were identified in THP-1 cells incubated with the plasma of survivor patients obtained at T2. The incubation of THP-1 with T2 plasma of non-survivors COVID-19 leads to higher TLR4, CCR2, CCR5, CCR7, and CD69 expression. In conclusion, the coexistence of increased microbial translocation and hyperinflammation in patients with severe COVID-19 may lead to higher monocyte activation, which may be associated with worsening outcomes, such as death.

Urine Sediment Findings and the Immune Response to Pathologies in Fungal Urinary Tract Infections Caused by Candida spp.

Fungi are pathogenic agents that can also cause disseminated infections involving the kidneys. Besides Candida, other agents like Cryptococcus spp. can cause urinary tract infection (UTI), as well as other non-yeast fungi, especially among immunocompromised patients. The detection and identification of fungi in urine samples (by microscopy and culture) plays an essential role in the diagnosis of fungal UTI. However, variable cutoff definitions and unreliable culture techniques may skew analysis of the incidence and outcome of candiduria. The sediment analysis plays a key role in the identification of fungal UTI because both yeasts and pseudohyphae are easily identified and can be used as a clinical sign of fungal UTI but should not be overinterpreted. Indeed, urine markers of the immune response (leukocytes), urine barriers of tissue protection (epithelial cells), and urine markers of kidney disease (urinary casts) can be found in urine samples. This work explores the manifestations associated with the fungal UTI from the urinalysis perspective, namely the urinary findings and clinical picture of patients with fungal UTI caused by Candida spp., aspects associated with the immune response, and the future perspectives of urinalysis in the diagnosis of this clinical condition.

Effects of undernutrition on glutamatergic parameters in the cerebral cortex of young rats.

Perinatal undernutrition impairs maturational events in the development of the brain, resulting in a variety of brain dysfunctions, which affect cognitive functions. This study investigated the effects of pre- and post-natal undernutrition (diet: 8% protein; control group: 25% protein) on some glutamatergic and behavioral parameters of 21-day-old rats. In the cerebral cortex, undernutrition reduced the Na-independent [(3)H]Glutamate binding in cellular membranes and [(3)H]Glutamate vesicular uptake, without affecting the [(3)H]Glutamate uptake by slices preparation. Behavioral parameters were affected, showing a strong amnesic effect both in the short- and long-term memory of inhibitory avoidance tasks, and a significant reduction in the number of crossings in an open field. The effects of perinatal undernutrition in 21-day-old rats, which alter some glutamatergic parameters may be related to the impairment of memory in certain behavioral tasks.