Identification and characterization of a polyomavirus in the thornback skate (Raja clavata).

Members of the family Polyomaviridae have a circular double-stranded DNA genome that have been identified in various hosts ranging from mammals to arachnids. Here we report the identification and analysis of a complete genome sequence of a novel polyomavirus, Raja clavata polyomavirus (RcPyV1), from a cartilaginous fish, the thornback skate (Raja clavata). The genome sequence was determined using a metagenomics approach with an aim to provide baseline viral data in cartilaginous fish in different ecosystems. The RcPyV1 genome (4,195 nucleotides) had typical organization of polyomavirus, including early antigens (small T; Large T) encoded on one strand and late viral proteins (VP1; VP2) on the complementary strand. Maximum-likelihood phylogenetic analysis of the large T-antigen revealed that RcPyV1 clusters with a polyomavirus obtained from another cartilaginous fish, the guitarfish polyomavirus 1 (GfPyV1). These two share ~ 56% pairwise identity in LT and VP1 protein sequences. These analyses support the hypothesis that cartilaginous fishes have a specific lineage of polyomaviruses.

Mastectomy or Margin Re-excision? A Nomogram for Close/Positive Margins After Lumpectomy for DCIS.

BACKGROUND: Anatomic extent of ductal carcinoma in situ (DCIS) may be uncertain in spite of clinical, pathologic, and imaging data. Consequently close/positive margins are common with lumpectomy for DCIS and often lead to a challenge in deciding whether to perform a re-excision or mastectomy. PATIENTS AND METHODS: From a single health system, we identified cases of lumpectomy for DCIS with close/positive margins who underwent re-excision for the purpose of constructing a nomogram. In total, 289 patients were available for analysis. The patients were randomly divided into two sets allocating 70% to the modeling and 30% to the validation set. A multivariable logistic regression model was used to estimate the probability of overall positive margin status using multiple clinicopathologic predictors. Nomogram validation included internal tenfold cross-validation, internal bootstrap validation, and external validation for which a concordance index was calculated to assess the external validity. RESULTS: Significant predictors of persistent positive margins from regression modeling included necrosis at diagnosis (non-comedo or comedo); DCIS not associated with calcifications on core biopsy; high-grade DCIS; progesterone receptor positivity; and number of positive margins at initial surgery. When subjected to internal validation, the nomogram achieved an uncorrected concordance index of 0.7332, a tenfold cross-validation concordance index of 0.6795, and a bootstrap-corrected concordance index of 0.6881. External validation yielded an estimated concordance index of 0.7095. CONCLUSION: Using clinical and pathologic variables from initial diagnosis and surgery for DCIS, this nomogram predicts persistent positive margins with margin re-excision, and may be a valuable tool in surgical decision-making.

Biological and clinical significance of dysplastic hematopoiesis in patients with newly diagnosed multiple myeloma.

Risk of developing myelodysplastic syndrome (MDS) is significantly increased in both multiple myeloma (MM) and monoclonal gammopathy of undetermined significance, suggesting that it is therapy independent. However, the incidence and sequelae of dysplastic hematopoiesis at diagnosis are unknown. Here, we used multidimensional flow cytometry (MFC) to prospectively screen for the presence of MDS-associated phenotypic alterations (MDS-PA) in the bone marrow of 285 patients with MM enrolled in the PETHEMA/GEM2012MENOS65 trial (#NCT01916252). We investigated the clinical significance of monocytic MDS-PA in a larger series of 1252 patients enrolled in 4 PETHEMA/GEM protocols. At diagnosis, 33 (11.6%) of 285 cases displayed MDS-PA. Bulk and single-cell-targeted sequencing of MDS recurrently mutated genes in CD34+ progenitors (and dysplastic lineages) from 67 patients revealed clonal hematopoiesis in 13 (50%) of 26 cases with MDS-PA vs 9 (22%) of 41 without MDS-PA; TET2 and NRAS were the most frequently mutated genes. Dynamics of MDS-PA at diagnosis and after autologous transplant were evaluated in 86 of 285 patients and showed that in most cases (69 of 86 [80%]), MDS-PA either persisted or remained absent in patients with or without MDS-PA at diagnosis, respectively. Noteworthy, MDS-associated mutations infrequently emerged after high-dose therapy. Based on MFC profiling, patients with MDS-PA have altered hematopoiesis and T regulatory cell distribution in the tumor microenvironment. Importantly, the presence of monocytic MDS-PA at diagnosis anticipated greater risk of hematologic toxicity and was independently associated with inferior progression-free survival (hazard ratio, 1.5; P = .02) and overall survival (hazard ratio, 1.7; P = .01). This study reveals the biological and clinical significance of dysplastic hematopoiesis in newly diagnosed MM, which can be screened with moderate sensitivity using cost-effective MFC.

Immunogenomic identification and characterization of granulocytic myeloid-derived suppressor cells in multiple myeloma.

Granulocytic myeloid-derived suppressor cells (G-MDSCs) promote tumor growth and immunosuppression in multiple myeloma (MM). However, their phenotype is not well established for accurate monitoring or clinical translation. We aimed to provide the phenotypic profile of G-MDSCs based on their prognostic significance in MM, immunosuppressive potential, and molecular program. The preestablished phenotype of G-MDSCs was evaluated in bone marrow samples from controls and MM patients using multidimensional flow cytometry; surprisingly, we found that CD11b+CD14-CD15+CD33+HLADR- cells overlapped with common eosinophils and neutrophils, which were not expanded in MM patients. Therefore, we relied on automated clustering to unbiasedly identify all granulocytic subsets in the tumor microenvironment: basophils, eosinophils, and immature, intermediate, and mature neutrophils. In a series of 267 newly diagnosed MM patients (GEM2012MENOS65 trial), only the frequency of mature neutrophils at diagnosis was significantly associated with patient outcome, and a high mature neutrophil/T-cell ratio resulted in inferior progression-free survival (P < .001). Upon fluorescence-activated cell sorting of each neutrophil subset, T-cell proliferation decreased in the presence of mature neutrophils (0.5-fold; P = .016), and the cytotoxic potential of T cells engaged by a BCMA×CD3-bispecific antibody increased notably with the depletion of mature neutrophils (fourfold; P = .0007). Most interestingly, RNA sequencing of the 3 subsets revealed that G-MDSC-related genes were specifically upregulated in mature neutrophils from MM patients vs controls because of differential chromatin accessibility. Taken together, our results establish a correlation between the clinical significance, immunosuppressive potential, and transcriptional network of well-defined neutrophil subsets, providing for the first time a set of optimal markers (CD11b/CD13/CD16) for accurate monitoring of G-MDSCs in MM.

Growth and maturity of the lesser-spotted dogfish Scyliorhinus canicula (Linnaeus, 1758) in the southern Portuguese continental coast.

Growth and reproductive parameters were estimated for Scyliorhinus canicula from the south-western Portuguese coast. The sample consisted of 148 specimens with total length ranging from 187 to 580 mm (82 males and 66 females). Maximum ages assigned to males and females were 12 and 13 years, respectively. Linf and k were estimated as 63.6 cm and 0.16 year-1 for males and 63.2 and 0.15 year-1 for females. Length and age at first maturity were estimated as 42.6 cm and 6 years for males and 44.5 cm and 7 years for females.

Immunologic characterization of COVID-19 patients with hematological cancer.

Not available.

Diabetes Risk Assessment in Latinas: Effectiveness of a Brief Diabetes Risk Questionnaire for Detecting Prediabetes in a Community-Based Sample.

OBJECTIVE: Numerous validated questionnaires use self-reported data to quantify individuals' risk of having diabetes or developing it in the future. Evaluations of these tools have primarily used nationally representative data, limiting their application in clinical and community settings. This analysis tested the effectiveness of the American Diabetes Association (ADA) risk questionnaire for identifying prediabetes in a community-based sample of Latinas. METHODS: Data were collected using the ADA risk questionnaire and assessing A1C. Among 204 participants without diabetes, we examined the association between individual characteristics and glycemic status. We then calculated the performance characteristics (sensitivity, specificity, positive predictive value [PPV], and negative predictive value [NPV]) of the ADA risk questionnaire for detecting prediabetes, using A1C results as the gold standard to define the outcome. RESULTS: All participants were women of self-reported Hispanic/Latino ethnicity. Their mean ADA risk score was 5.6 ± 1.6. Latinas who had prediabetes were older, with significantly higher rates of hypertension and a higher ADA risk score than those without prediabetes. At a risk score ≥5-the threshold for high risk set by the ADA-the questionnaire had the following test performance characteristics: sensitivity 77.8%, specificity 41.7%, PPV 76.2%, and NPV 43.9%. CONCLUSION: The ADA risk questionnaire demonstrates reasonable performance for identifying prediabetes in a community-based sample of Latinas. Our data may guide other groups' use of this tool in the same target population. Future research should examine the effectiveness of this questionnaire for recruiting diverse populations into diabetes prevention programs. In addition, unique diabetes risk assessment tools for specific target populations are needed and may outperform questionnaires developed using nationally representative data.

Trial of labour after caesarean section: Two-year analysis at a Portuguese centre.

We conducted a retrospective study aimed at determining variables associated with a higher success rate for vaginal delivery after caesarean section, and assessing the impact of induction of labour. Secondarily, we aimed to describe our vaginal delivery and uterine rupture rates with the use of a controlled-release dinoprostone vaginal insert for cervical ripening. Of 292 women who met the inclusion criteria, induction of labour occurred in 48% (94% with dinoprostone). There was a non-significant difference between the vaginal delivery rate of spontaneous labour (57%) and induction of labour (33%), after adjusting for confounding variables. The success rate was influenced by a Bishop score ≥6, previous vaginal delivery and previous caesarean for dystocia or failed induction. There was only one case of uterine rupture, which was associated with dinoprostone use (overall rate 0.34%, 0.77% for dinoprostone). Impact statement Trial of labour after caesarean section is considered an alternative to elective repeat caesarean. Both present associated benefits and risks, the most fearsome of which is uterine rupture during labour (0.78% in term pregnancies). Induction is also possible but carries a higher risk of uterine rupture and lower success rate for vaginal birth. Prostaglandins have been of particular concern due to a higher risk of uterine scar rupture, estimated at 2% for dinoprostone; however, its use as a controlled-release vaginal insert has been under-reported. Our study confirms the reported impact of previous vaginal delivery, previous caesarean indication and Bishop score at admission on success rate for vaginal birth after caesarean. We were unable to prove a lower success rate for induction of labour after adjusting for other variables. Despite our study limitations, we report on the use of a controlled-release vaginal insert with 10mg of dinoprostone in 130 women with a uterine rupture rate of 0.77%, lower than previously reported and similar to the overall rate estimated for term pregnancies. This dinoprostone formulation may be safer than previously reported but larger studies, and preferably randomised controlled trials, are needed to confirm these findings.

Should Patients with Trichorhinophalangeal Syndrome be Tested for Growth Hormone Deficiency?

Type 1 Trichorhinophalangeal syndrome (TRPS) is characterized by typical facial and skeletal abnormalities. These patients frequently exhibit short stature; however, only one case with growth hormone (GH) deficiency can be found in the literature. Our patient is a 10-year-old girl with two novel nonsense pathogenic mutations in the TRPS1 gene, both in heterozygosity: c. 1198C>T (p. Gln400X) and c.2086C>T (p. Arg696X). She has an additional GH deficiency. The patient is short in stature, with a growth velocity of 1.5 cm per year (SDS - 4.07), a bone age of 4.5 years, and she shows no response to the GH stimulation tests. According to a previous report of an identical case, catch-up growth will occur after beginning GH treatment. We believe that GH stimulation tests should be performed on patients with TRPS1 exhibiting a growth velocity below the normal range expected for their age and sex. If the result is subnormal, then GH therapy should be attempted.

Identification of potential essential fish habitats for skates based on fishers' knowledge.

Understanding of spatio-temporal patterns of sensitive fish species such as skates (Rajidae) is essential for implementation of conservation measures. With insufficient survey data available for these species in Portuguese Continental waters, this study shows that fishery-dependent data associated with fishers' knowledge can be used to identify potential Essential Fish Habitats (EFH) for seven skate species. Sites with similar geomorphology were associated with the occurrence of juveniles and/or adults of the same group of species. For example, sites deeper than 100 m with soft sediment include predominantly adults of Raja clavata, and are the habitat for egg deposition of this species. Raja undulata and R. microocellata are the more coastal species, preferring sand or gravel habitats, while coastal areas with rocks and sand seabed are potential nursery areas for R. brachyura, R. montagui and R. clavata. The main output of this study is the identification of preferential fishing sites enclosing potential EFH for some species, associated with egg-laying and nursery grounds. The location of these areas will be considered for future seasonal closures, and studies will be conducted to evaluate the biological and socio-economic impacts of such measures. As in the past, fishermen will collaborate in the process of evaluating those impacts, since they have practical and applied knowledge that is extremely valuable for evaluating the advantages and disadvantages of such closures. In conclusion, this study is a first contribution to the understanding and identification of EFH for skate species, associated with nursery and egg deposition sites, with direct application to management.

HIV Associated Lung Cancer: Unique Clinicopathologic Features and Immune Biomarkers Impacting Lung Cancer Screening and Management.

OBJECTIVES: Lung cancer contributes significantly to morbidity and mortality in people with HIV (PWH). We study the clinicopathologic characteristics and immune microenvironment in HIV associated lung cancer. MATERIAL AND METHODS: Clinicopathological characteristics including immunotherapy outcomes were collected for 174 PWH diagnosed with lung cancer. Immunohistochemical staining for PD-L1, CD4, and CD8 was performed. RESULTS: At diagnosis, patients with HIV associated lung cancer were significantly younger (56.9 vs. 69 years, P < .0001) and more frequently had advanced disease (70% vs. 53%, P = .01). The majority were African American (60% vs. 42%, P < .0001) and were smoking at the time of diagnosis or smoked in the past (98% vs. 86%, P = .0001). Only 10% of HIV associated lung cancer was diagnosed through the screening program. The median CD4+ lymphocyte count was 334 cells/µL, 31% had a CD4 ≤200 cells/µL and 63% of the cohort was virally suppressed. HIV associated non-small-cell lung cancer(NSCLC) was characterized by limited PD-L1 expression compared to the HIV negative cohort, 64% vs. 31% had TPS <1%, and 20% vs. 34% had TPS≥50%, respectively (P = .04). Higher CD8+ TILs were detected in PD-L1-high tumors (P < .0001). 50% of patients achieved disease control in the metastatic setting with the use of immunotherapy, and there were no new safety signals in 19 PWH treated with immunotherapy. CONCLUSION: Lung cancer in PWH demonstrates unique features highlighting the need for a specialized screening program. Despite low PD-L1 expression, immunotherapy is well tolerated with reasonable disease control. Altered immune system in lung cancer pathogenesis in PWH should be further investigated.

Immune dysfunction prior to and during vaccination in multiple myeloma: a case study based on COVID-19.

Infection is the leading cause of death in multiple myeloma (MM). However, the cellular composition associated with immune dysfunction is not defined. We analyzed immune profiles in the peripheral blood of patients with MM (n = 28) and B-cell chronic lymphoproliferative disorders (n = 53) vs. health care practitioners (n = 96), using multidimensional and computational flow cytometry. MM patients displayed altered distribution of most cell types (41/56, 73%), particularly within the B-cell (17/17) and T-cell (20/30) compartments. Using COVID-19 as a case study, we compared the immune response to vaccination based on 64,304 data points generated from the analysis of 1099 longitudinal samples. MM patients showed limited B-cell expansion linked to lower anti-RBD and anti-S antibody titers after the first two doses and booster. The percentages of B cells and CD4+ T cells in the blood, as well as the absolute counts of B cells and dendritic cells, predicted vaccine immunogenicity at different time points. In contrast with the humoral response, the percentage and antigen-dependent differentiation of SARS-CoV-2-specific CD8+ T cells was not altered in MM patients. Taken together, this study defined the cellular composition associated with immune dysfunction in MM and provided biomarkers such as the B-cell percentage and absolute count to individualize vaccination calendars.

Role of innate immunological/inflammatory pathways in myelodysplastic syndromes and AML: a narrative review.

Dysregulation of the innate immune system and inflammatory-related pathways has been implicated in hematopoietic defects in the bone marrow microenvironment and associated with aging, clonal hematopoiesis, myelodysplastic syndromes (MDS), and acute myeloid leukemia (AML). As the innate immune system and its pathway regulators have been implicated in the pathogenesis of MDS/AML, novel approaches targeting these pathways have shown promising results. Variability in expression of Toll like receptors (TLRs), abnormal levels of MyD88 and subsequent activation of NF-κß, dysregulated IL1-receptor associated kinases (IRAK), alterations in TGF-ß and SMAD signaling, high levels of S100A8/A9 have all been implicated in pathogenesis of MDS/AML. In this review we not only discuss the interplay of various innate immune pathways in MDS pathogenesis but also focus on potential therapeutic targets from recent clinical trials including the use of monoclonal antibodies and small molecule inhibitors against these pathways.

Preclinical models for prediction of immunotherapy outcomes and immune evasion mechanisms in genetically heterogeneous multiple myeloma.

The historical lack of preclinical models reflecting the genetic heterogeneity of multiple myeloma (MM) hampers the advance of therapeutic discoveries. To circumvent this limitation, we screened mice engineered to carry eight MM lesions (NF-κB, KRAS, MYC, TP53, BCL2, cyclin D1, MMSET/NSD2 and c-MAF) combinatorially activated in B lymphocytes following T cell-driven immunization. Fifteen genetically diverse models developed bone marrow (BM) tumors fulfilling MM pathogenesis. Integrative analyses of ∼500 mice and ∼1,000 patients revealed a common MAPK-MYC genetic pathway that accelerated time to progression from precursor states across genetically heterogeneous MM. MYC-dependent time to progression conditioned immune evasion mechanisms that remodeled the BM microenvironment differently. Rapid MYC-driven progressors exhibited a high number of activated/exhausted CD8+ T cells with reduced immunosuppressive regulatory T (Treg) cells, while late MYC acquisition in slow progressors was associated with lower CD8+ T cell infiltration and more abundant Treg cells. Single-cell transcriptomics and functional assays defined a high ratio of CD8+ T cells versus Treg cells as a predictor of response to immune checkpoint blockade (ICB). In clinical series, high CD8+ T/Treg cell ratios underlie early progression in untreated smoldering MM, and correlated with early relapse in newly diagnosed patients with MM under Len/Dex therapy. In ICB-refractory MM models, increasing CD8+ T cell cytotoxicity or depleting Treg cells reversed immunotherapy resistance and yielded prolonged MM control. Our experimental models enable the correlation of MM genetic and immunological traits with preclinical therapy responses, which may inform the next-generation immunotherapy trials.

FlowCT for the analysis of large immunophenotypic data sets and biomarker discovery in cancer immunology.

Large-scale immune monitoring is becoming routinely used in clinical trials to identify determinants of treatment responsiveness, particularly to immunotherapies. Flow cytometry remains one of the most versatile and high throughput approaches for single-cell analysis; however, manual interpretation of multidimensional data poses a challenge when attempting to capture full cellular diversity and provide reproducible results. We present FlowCT, a semi-automated workspace empowered to analyze large data sets. It includes pre-processing, normalization, multiple dimensionality reduction techniques, automated clustering, and predictive modeling tools. As a proof of concept, we used FlowCT to compare the T-cell compartment in bone marrow (BM) with peripheral blood (PB) from patients with smoldering multiple myeloma (SMM), identify minimally invasive immune biomarkers of progression from smoldering to active MM, define prognostic T-cell subsets in the BM of patients with active MM after treatment intensification, and assess the longitudinal effect of maintenance therapy in BM T cells. A total of 354 samples were analyzed and immune signatures predictive of malignant transformation were identified in 150 patients with SMM (hazard ratio [HR], 1.7; P < .001). We also determined progression-free survival (HR, 4.09; P < .0001) and overall survival (HR, 3.12; P = .047) in 100 patients with active MM. New data also emerged about stem cell memory T cells, the concordance between immune profiles in BM and PB, and the immunomodulatory effect of maintenance therapy. FlowCT is a new open-source computational approach that can be readily implemented by research laboratories to perform quality control, analyze high-dimensional data, unveil cellular diversity, and objectively identify biomarkers in large immune monitoring studies. These trials were registered at www.clinicaltrials.gov as #NCT01916252 and #NCT02406144.

Circulating Tumor and Immune Cells for Minimally Invasive Risk Stratification of Smoldering Multiple Myeloma.

PURPOSE: Early intervention in smoldering multiple myeloma (SMM) requires optimal risk stratification to avoid under- and overtreatment. We hypothesized that replacing bone marrow (BM) plasma cells (PC) for circulating tumor cells (CTC), and adding immune biomarkers in peripheral blood (PB) for the identification of patients at risk of progression due to lost immune surveillance, could improve the International Myeloma Working Group 20/2/20 model. EXPERIMENTAL DESIGN: We report the outcomes of 150 patients with SMM enrolled in the iMMunocell study, in which serial assessment of tumor and immune cells in PB was performed every 6 months for a period of 3 years since enrollment. RESULTS: Patients with >0.015% versus ≤0.015% CTCs at baseline had a median time-to-progression of 17 months versus not reached (HR, 4.9; P < 0.001). Presence of >20% BM PCs had no prognostic value in a multivariate analysis that included serum free light-chain ratio >20, >2 g/dL M-protein, and >0.015% CTCs. The 20/2/20 and 20/2/0.015 models yielded similar risk stratification (C-index of 0.76 and 0.78). The combination of the 20/2/0.015 model with an immune risk score based on the percentages of SLAN+ and SLAN- nonclassical monocytes, CD69+HLADR+ cytotoxic NK cells, and CD4+CXCR3+ stem central memory T cells, allowed patient' stratification into low, intermediate-low, intermediate-high, and high-risk disease with 0%, 20%, 39%, and 73% rates of progression at 2 years. CONCLUSIONS: This study showed that CTCs outperform BM PCs for assessing tumor burden. Additional analysis in larger series are needed to define a consensus cutoff of CTCs for minimally invasive stratification of SMM.

Primary and Secondary Hypogonadism in Male Persons with Diabetes Mellitus.

AIMS: To characterize hypogonadism in male persons with diabetes mellitus. Patients and Methods. 184 consecutive male persons with diabetes were studied. Besides the usual care, total testosterone (TT), estradiol (E2), FSH, and LH were measured in the last appointment and in 40 patients, also in the next two appointments. Statistical analysis compared groups and explored factors for TT and LH levels. RESULTS: TT levels were stable and highly correlated (r > 0.750, p < 0.001) over a 6-12-month period. 20% of the patients presented secondary hypogonadism (SH) and 18% presented primary hypogonadism (PH). SH was inversely related to HbA1 (partial r (rp) = 0.229, p < 0.005), while PH was directly related to age (r = 0.356, p < 0.001). TT levels were reduced independently by metformin (364 ± 160 vs. 431 ± 242 ng/dL, t = 2.241, p < 0.05) and statins (359 ± 156 vs. 424 ± 230 ng/dl, t = 2.224, p < 0.05). TT levels were inversely related to microvascular disease (rp = -0.169, p < 0.05). Discussion. TT levels were stable over time and hypogonadism was common. SH, generally clinically, is related to the diabetic state, while PH, generally subclinically, is an age-dependent process unrelated to diabetes. Low TT levels were related to older age, poor metabolic control, metformin and statins use, and microvascular disease.

Immune biomarkers to predict SARS-CoV-2 vaccine effectiveness in patients with hematological malignancies.

There is evidence of reduced SARS-CoV-2 vaccine effectiveness in patients with hematological malignancies. We hypothesized that tumor and treatment-related immunosuppression can be depicted in peripheral blood, and that immune profiling prior to vaccination can help predict immunogenicity. We performed a comprehensive immunological characterization of 83 hematological patients before vaccination and measured IgM, IgG, and IgA antibody response to four viral antigens at day +7 after second-dose COVID-19 vaccination using multidimensional and computational flow cytometry. Health care practitioners of similar age were the control group (n = 102). Forty-four out of 59 immune cell types were significantly altered in patients; those with monoclonal gammopathies showed greater immunosuppression than patients with B-cell disorders and Hodgkin lymphoma. Immune dysregulation emerged before treatment, peaked while on-therapy, and did not return to normalcy after stopping treatment. We identified an immunotype that was significantly associated with poor antibody response and uncovered that the frequency of neutrophils, classical monocytes, CD4, and CD8 effector memory CD127low T cells, as well as naive CD21+ and IgM+D+ memory B cells, were independently associated with immunogenicity. Thus, we provide novel immune biomarkers to predict COVID-19 vaccine effectiveness in hematological patients, which are complementary to treatment-related factors and may help tailoring possible vaccine boosters.

Immunological Biomarkers of Fatal COVID-19: A Study of 868 Patients.

Information on the immunopathobiology of coronavirus disease 2019 (COVID-19) is rapidly increasing; however, there remains a need to identify immune features predictive of fatal outcome. This large-scale study characterized immune responses to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection using multidimensional flow cytometry, with the aim of identifying high-risk immune biomarkers. Holistic and unbiased analyses of 17 immune cell-types were conducted on 1,075 peripheral blood samples obtained from 868 COVID-19 patients and on samples from 24 patients presenting with non-SARS-CoV-2 infections and 36 healthy donors. Immune profiles of COVID-19 patients were significantly different from those of age-matched healthy donors but generally similar to those of patients with non-SARS-CoV-2 infections. Unsupervised clustering analysis revealed three immunotypes during SARS-CoV-2 infection; immunotype 1 (14% of patients) was characterized by significantly lower percentages of all immune cell-types except neutrophils and circulating plasma cells, and was significantly associated with severe disease. Reduced B-cell percentage was most strongly associated with risk of death. On multivariate analysis incorporating age and comorbidities, B-cell and non-classical monocyte percentages were independent prognostic factors for survival in training (n=513) and validation (n=355) cohorts. Therefore, reduced percentages of B-cells and non-classical monocytes are high-risk immune biomarkers for risk-stratification of COVID-19 patients.

Pediatric preseptal and orbital cellulitis: A 10-year experience.

OBJECTIVE: Characterize clinical features, epidemiology and treatment of hospitalized pediatric cases of preseptal and orbital cellulitis. METHODS: Retrospective study of children/adolescents admitted to a central hospital with preseptal and orbital cellulitis between 2007 and 2017. RESULTS: A total of 122 cases were included, 80.3% with preseptal cellulitis and 19.7% with orbital cellulitis. Patients had a median age of 5 years. Sinusitis was the most common predisposing factor (40.2%), followed by dental abscess (20.4%) in preseptal cellulitis and by external ocular infections (12.5%) in orbital cellulitis. Sinusitis (p < 0.001) was associated with orbital cellulitis, whereas patients with dental abscess (p = 0.007) and trauma (p = 0.040) were most likely to have preseptal cellulitis. Fever, photophobia, ocular pain, painful eye movements, proptosis, rhinorrhea and vison impairment were related with orbital cellulitis. Leukocytosis was present in 34.4% of patients, and associated with orbital cellulitis (p = 0.001). Nearly half of patients (49.2%) had a CT-scan performed. Systemic corticosteroids were used in 19.7%. Complications occurred in 13 patients. Imaging revaluation through CT was performed in 6.6%, with no patients showing deterioration; 1.6% of patients required surgery. CONCLUSIONS: Identification of orbital involvement signs suggested orbital cellulitis. We emphasize the impact of dental abscess as a predisposing factor for preseptal cellulitis. Repeated imaging had no impact on treatment or outcome. A high percentage of patients was treated with steroids despite their controversial use.