Impact of Cerebrospinal Fluid Leukocyte Infiltration and Neuroimmmune Mediators on Survival with HIV-Associated Cryptococcal Meningitis.

Introduction: Cryptococcal meningitis remains a prominent cause of death in persons with advanced HIV disease. CSF leukocyte infiltration predicts survival at 18 weeks; however, how CSF immune response relates to CSF leukocyte infiltration is unknown. Methods: We enrolled 401 adults with HIV-associated cryptococcal meningitis in Uganda who received amphotericin and fluconazole induction therapy. We assessed the association of CSF leukocytes, chemokine, and cytokine responses with 18-week survival. Results: Participants with CSF leukocytes ≥50/µL, had higher probability 68% (52/77) of 18-week survival compared with 52% (151/292) 18-week survival in those with ≤50 cells/µL (Hazard Ratio=1.63, 95% confidence intervals 1.14-2.23; p=0.008). Survival was also associated with higher expression of T helper (Th)-1, Th17 cytokines, and immune regulatory elements. CSF levels of Programmed Death-1 Ligand, CXCL10, and Interleukin (IL)-2 independently predicted survival. In multivariate analysis, CSF leukocytes were inversely associated with CSF fungal burden and positively associated with CSF protein, interferon-gamma (IFN-γ), IL-17A, tumor necrosis factor (TNF)-α, and peripheral blood CD4+ and CD8+ T cells expression. Conclusion: 18-week survival after diagnosis of cryptococcal meningitis was associated with higher CSF leukocytes at baseline with greater T helper 1 (IFN-γ, IL-2 and TNF-α cytokines), T helper 17 (IL-17A cytokine) and CXCR3+ T cell (CXCL10 chemokine) responses. These results highlight the interdependent contribution of soluble and cellular immune responses in predicting survival with HIV-associated cryptococcal meningitis.

Production of recombinant humanized monoclonal anti-human neutrophil antigen (HNA) antibodies with potential applicability as standard antibodies.

BACKGROUND: Antibodies against human neutrophil antigen (HNA) are involved in the pathogenesis of neonatal alloimmune neutropenia, autoimmune neutropenia, and transfusion-related acute lung injury. The present methods for anti-HNA antibody identification strongly depend on the presence of standard antisera with known allo/isospecificities. Here, we aimed to produce recombinant humanized antibodies to HNA from available mouse monoclonal antibodies (MoAbs). STUDY DESIGN AND METHODS: RNAs were extracted from available hybridoma cells producing mouse anti-HNA antibodies recognizing HNA-1a (TAG-1), -1b (TAG-2), -2 (TAG-4), and FcγRIIIb, and the cDNA was synthesized. Recombinant fragments consisting of the variable regions of the H and L chains of the mouse MoAb ligated to the constant region of human IgG were incorporated into an expression vector and transfected into CHO cells. Antibody specificity of the selected humanized monoclonal antibodies was confirmed, and tested by the participants of the ISBT Granulocyte Immunobiology Working Party (GIWP) workshop 2020. RESULTS: GIFT results confirmed the specific reactivity of TAGH-1 to -4, except for a cross-reactivity of TAGH-2 with HNA-1a/a neutrophils, only in flow-cytometry. MAIGA results showed clear specificity of all humanized antibodies, but the selection of the appropriate capture monoclonal antibody was essential for the test. The results of the ISBT GIWP showed high concordance among the labs. CONCLUSIONS: These are the first humanized monoclonal antibodies to HNA-1 and HNA-2 antigens produced and they will be important standard reagents for laboratories testing for neutrophil antibodies. We plan to have these humanized MoAbs available through WHO.

Risk factors for T-cell lymphopenia in frequent platelet donors: The BEST collaborative study.

BACKGROUND: Severe T-cell lymphopenia of uncertain clinical significance has been observed in frequent apheresis platelet donors. Two commonly used plateletpheresis instruments are the Trima Accel, which uses a leukoreduction system (LRS) chamber to trap leukocytes and the Fenwal Amicus, which does not use an LRS chamber. STUDY DESIGN AND METHODS: We performed an international, multicenter, observational study comparing T-cell populations in frequent platelet donors collected exclusively using the Trima instrument (n = 131) or the Amicus instrument (n = 77). Age- and sex-matched whole blood donors (n = 126) served as controls. RESULTS: CD4+ T-cell counts <200 cells/µL were found in 9.9% of frequent Trima (LRS+) platelet donors, 4.4% of frequent Amicus (LRS-) platelet donors, and 0 whole blood donors (p < .0001). CD4+ T-cell counts <200 cells/µL were only seen in platelet donors with ≥200 lifetime donations. In multivariable analysis, age, lifetime donations, and instrument (Trima vs. Amicus) were independent risk factors for lymphopenia. In 40 Trima platelet donors, a plasma rinseback procedure was routinely performed following platelet collections. No Trima platelet donors receiving plasma rinseback had a CD4+ T-cell count <200 cells/µL versus 13/91 Trima platelet donors not receiving plasma rinseback (p = .01). DISCUSSION: Recurrent bulk lymphocyte removal appears to contribute to the development of T-cell lymphopenia in frequent, long-term platelet donors. Lymphopenia is more common when an LRS chamber is used during platelet collection but can occur without an LRS chamber. Blood centers using LRS chambers can mitigate donor lymphopenia by performing plasma rinseback.

Elevated Levels of Neutrophil-to Monocyte Ratio Are Associated with the Initiation of Paroxysmal Documented Atrial Fibrillation in the First Two Months after Heart Transplantation: A Uni-Institutional Retrospective Study.

BACKGROUND: Heart transplantation represents the treatment for patients with end-stage heart failure (HF) being symptomatic despite optimal medical therapy. We investigated the role of NMR (neutrophil-to-monocyte ratio), NLR (neutrophil-to-lymphocyte ratio), NPR (neutrophil-to-platelet ratio), NWR (neutrophil-to-white cells ratio), MLR (monocyte-to-lymphocyte ratio), PLR (platelet-to-lymphocyte ratio), MWR (neutrophil-to-white cells ratio), and LWR (lymphocyte-to-white cells ratio) at the same cut-off values previously studied, to predict complications after heart transplant within 2 months after surgery. METHODS: From May 2014 to January 2021, was included 38 patients in our study from the Cardiovascular and Transplant Emergency Institute of Târgu Mureș. RESULTS: Preoperative NMR > 8.9 (OR: 70.71, 95% CI: 3.39-1473.64; p = 0.006) was a risk factor for the apparition of post-operative paroxysmal atrial fibrillation (Afib). In contrast, preoperative MWR > 0.09 (OR: 0.04, 95% CI: 0.003-0.58; p = 0.0182) represented a protective factor against AFib, but being the risk of complications of any cause (OR: 14.74, 95% CI: 1.05-206.59, p = 0.0458). CONCLUSION: Preoperative elevated levels of NMR were associated with the apparition of documented AFib, with high levels of MWR as a protective factor. High MWR was a risk factor in developing complications of any cause in the first 2 months after heart transplantation.

Differential Activity of Human Leukocyte Extract on Systemic Immune Response and Cyst Growth in Mice with Echinococcus Multilocularis Infection After Oral, Subcutaneous and Intraperitoneal Routes of Administration.

Alveolar echinococcosis (AE) caused by the larval stage of Echinococcus multilocularis is serious parasitic diseases associated with the host´s immunosuppression. The effects of human non-immune dialyzable leukocyte extract (DLE) on immune cells in blood and spleen and parasitic cysts weight in Balb/c mice after oral (PO), subcutaneous (SC) and intraperitoneal administration (IP) were compared. The reduction in cysts weight (p < 0.01) was recorded after PO route, whereas moderate reduction was found after SC and IP routes. The elevation of lymphoid populations in blood and spleen was found after PO administration (p < 0.01) in parallel with reduced myeloid population. Infection-elicited decline in B220+B cells was partially abolished by PO route, but DLE routes did not influence the CD3+ T cells. The proportions of CD3+CD4+Th lymphocytes were moderately upregulated, whereas CD3+CD8+Tc populations were reduced after all DLE routes (p < 0.01). PO administration increased CD11b+MHCIIhigh blood monocytes, CD11b-SigleF+ cell, but not CD11b+Si-glecF+ eosinophils in the blood, stimulated after SC and IP routes. DLE induced downregulation of NO production by LPS-stimulated adherent splenocytes ex vivo. Con A-triggered T lymphocyte proliferation was associated with the elevated IFN-γ production and transcription factor Tbet mRNA expression. The alleviation of Th2 (IL-4) and Treg (TGF-ß) cytokine production by lymphocytes ex vivo paralleled with downregulation of gene transcription for cytokines, GATA and FoxP3. Reduction of myeloid cells with suppressive activity was found. The SC and IP routes affected partially the cysts weights, diminished significantly gene transcription, NO levels and Th2 and Treg cytokines production. Results showed that PO route of DLE administration was the most effective in ameliorating immunosuppression via stimulation of Th1 type, reducing Th2 and Treg type of immunity and CD3+CD8+Tc lymphocytes in the blood and spleens during E. multilocularis infection in mice.

Normal Values for Cerebrospinal Fluid in Neonates: A Systematic Review.

BACKGROUND: The diagnosis of neonatal meningitis often rests on microscopic and biochemical findings in the cerebrospinal fluid (CSF). There is ongoing uncertainty about age-related normal values for CSF findings in neonates, and many previous studies have included infants in whom antibiotics were administered before lumbar puncture or in whom viral meningitis was not excluded. METHODS: A systematic search was done using MEDLINE and EMBASE to identify original studies which investigated CSF normal values in either healthy neonates or febrile neonates in whom bacterial and viral meningitis were reliably excluded. RESULTS: We identified seven studies investigating 270 term and 96 preterm neonates. There were minimal differences between preterm and term neonates in the CSF white blood cell (WBC) count and glucose concentration. In contrast, the CSF neutrophil count and protein concentration were influenced by gestational and chronological age. In the four studies that reported individual patient data, in 95% of cases the CSF WBC count was <12 cells/µL in preterm and <10 cells/µL in term neonates, the neutrophil count was <16 and 8 cells/µL, and the protein concentration was <210 and 110 mg/dL, respectively. CONCLUSION: The normal range for CSF parameters in neonates is different to that in older infants, and some parameters are influenced by gestational and chronological age. CSF parameters alone are not sufficiently reliable to exclude meningitis.

Platelet and White Cell Reactivity to Top-Load Intravenous Perfluorocarbon Infusion in Healthy Sheep.

BACKGROUND: Perfluorocarbon emulsions (PFCs) are intravenous artificial oxygen carriers with enhanced gas solubility. As lipid micelle nanoparticle emulsions, PFCs may have a class effect that causes degrees of thrombocytopenia. Understanding the extent of the platelet effects, including mechanism and potential inflammation after PFC infusion, is important for safe human trials. METHODS: Normal sheep (Dorper) were infused with 5 mL/kg of Oxygent (w/v 60% PFC) or Perftoran (w/v 20% PFC). Controls received 6% Hetastarch or were naive. Blood samples were analyzed from baseline, time 0 (the end of infusion), 3 and 24 hours, and 4 and 7 days. Platelet count, plateletcrit, mean platelet volume, platelet distribution width, and CD-62p (a platelet activation-dependent membrane protein) were measured. Neutrophils, monocytes, and total white blood cell counts were analyzed. RESULTS: In these inflammatory cell lines, there were no consistent changes or cellular activation after PFC infusion. A decrease (<10% from baseline and naive controls) in platelet count was seen on day 4 after Oxygent infusion (3 g/kg), which recovered by day 7. No platelet effect was seen in Perftoran (1 g/kg). Plateletcrit, mean platelet volume, and platelet distribution width did not change significantly at any time point among the groups. CD-62p, ADP, and collagen aggregometry showed no significant change in platelet function. CONCLUSION: There was no evidence of overall reduction in platelet number, or any correlation with the change in platelet activation or inhibition. Therefore, the risk of increased thrombosis/bleeding after PFC intravenous infusion is low in this non-trauma sheep model.

The utility of a monocyte monolayer assay in the assessment of intravenous immunoglobulin-associated hemolysis.

BACKGROUND: Hemolysis following the administration of intravenous immunoglobulin (IVIG) is an important adverse event (AE). While the monocyte monolayer assay (MMA) has been used to predict in vivo hemolysis when serologically incompatible blood may be transfused, it has also been shown to correlate with IVIG-associated hemolysis. In this study, the MMA was examined for its utility in assessing the risk of hemolysis after IVIG. STUDY DESIGN AND METHODS: Forty-two non-blood group O patients receiving high-dose IVIG (≥2 g/kg) were examined using an autologous and allogeneic MMA. Hemolysis was defined by a drop in hemoglobin of ≥1 g/L, a positive direct antiglobulin test (DAT) and eluate, and a decrease in haptoglobin or increase in lactate dehydrogenase and/or reticulocytes. RESULTS: Forty-two patients provided 50 assessable postinfusion samples, with hemolysis observed in 20 (40%) of cases. Autologous MMA using post-IVIG red blood cells significantly correlated with clinical outcomes when compared to allogeneic MMA (P = .0320 vs .5806, t test). No significant difference in receiver operating characteristics was observed when comparing autologous MMA testing against DAT for the diagnosis of IVIG-associated hemolysis. However, when using samples collected 5 to 10 days after receipt of high-dose IVIG, the autologous MMA had higher sensitivity than the DAT. CONCLUSION: MMA testing with autologous monocytes collected 5 to 10 days after receipt of high-dose IVIG can be used for the diagnosis of IVIG-associated hemolysis and may be of particular value in cases in which the Day 5 to 10 DAT is negative.

Full blood count and white cell differential count reference ranges obtained from a healthy urban South African population residing in the Western Cape of South Africa.

BACKGROUND: Research has suggested that individuals of African descent have lower white cell and neutrophil counts than Caucasians. These differences could lead to incorrect clinical decisions, and therefore, ethnic-specific reference ranges are required. The Western Cape region of South Africa is uniquely diverse, comprising Caucasian, Mixed Ancestry and those of African descent. The aim of this study was to compare the full blood count and differential counts across the three major ethnic groups residing in this area and to propose appropriate RIs. METHODS: The study formed part of the international project led by the Committee on Reference Intervals and Decision Limits (C-RIDL), and therefore, the strict guidelines laid out by the committee were followed. Full blood count and differential counts were performed on a Beckman Coulter ACT 5 diff AL analyser within 2-4 hours of collection and were reported as mean (standard deviation), 2.5th and 97.5th percentiles. Comparisons were analysed using Spss v25 and Statistica v13, and a P value of < 0.05 was considered significant. RESULTS: Reference ranges for Caucasian and Mixed Ancestry individuals were similar while white cell (P = 0.016), monocyte (P < 0.001), neutrophil (P = 0.034) and red cell indices were significantly different amongst the three population groups. There were however no statistical and clinical significant differences between the eosinophil, lymphocyte, red cell and platelet counts across the three groups. CONCLUSION: In conclusion, subjects of Mixed Ancestry, in this region, have similar reference intervals to those of European descent, while lower white cell and neutrophil counts in Africans have been confirmed.

Identification and characterization of ORF19.1725, a novel gene contributing to the white cell pheromone response and virulence-associated functions in Candida albicans.

An epigenetic transition between white cells and opaque cells influences several properties of Candida albicans biology, including cellular morphology, biofilm formation, virulence, and sexual mating. In particular, these two cell types exhibit marked differences in their ability to undergo sex. A previous study identified the transcriptional regulator of pheromone response in both the white and opaque states as Cph1 because deletion of this gene abolished both pheromone-induced cell adhesion in white cells and sexual mating in opaque cells. To further explore how these cell types exhibit distinct biological outputs upon pheromone stimulation, we selected five Cph1-regulated genes with significant expression during the pheromone response in the white state but not the opaque state. These phase-specific pheromone-induced genes are ORF19.1539, ORF19.1725, ORF19.2430, ORF19.2691 and ORF19.5557. Deletion of each gene revealed that orf19.1539Δ, orf19.1725Δ, orf19.2430Δ and orf19.5557Δ showed significant decreases in pheromone-stimulated cell adhesion in the white state but retained normal mating competency in the opaque state, indicating that a particular role in white cell pheromone response is mediated by these four genes. Interestingly, the defects of orf19.1725Δ in pheromone-stimulated cell adhesion also abolished conventional biofilms and hyphal growth. Zebrafish egg infection assays further demonstrated that ORF19.1725 is involved in cell adhesion, penetration and virulence. Overall, four Cph1-regulated downstream targets were identified in the regulation of white cell pheromone response. We also clarified the roles of C. albicans ORF19.1725 in cell adhesion, hyphal growth, biofilm formation and virulence.

Phenotypic switch: The enigmatic white-gray-opaque transition system of Candida albicans.

Candida albicans represents the most common commensal and opportunistic fungal pathogen colonizing humans. As a member of the normal microflora, it is present on the skin and the mucous membranes of the upper respiratory tract, gastrointestinal tract and female genital tracts. It is therefore not transmitted. It lies in wait for a change in some aspect of the host physiology that normally suppress growth and invasiveness through an enigmatic phenomenon called Phenotypic Switch System or White-Opaque Transition. This system involves reversible and heritable switching between alternative cellular phenotypes. White-opaque switching in Candida albicans was first discovered in 1987. This was initially identified in strain WO-1. Switching has been demonstrated to occur at sites of infection and to occur between recurrent episodes of infection in select cases esp. AIDS and diabetes.

Efecto de la Suplementación con Semilla de Canavalia ensiformis sobre Parámetros Sanguíneos de Ovinos Tropicales con Infecciones Parasitarias Gastrointestinales / Effect of Supplementation with Canavalia ensiformis Seeds on Blood Variables of Parasitised Tropical Lambs

Con el objetivo de evaluar el efecto de la suplementación con semilla molida de Canavalia ensiformis sobre la hematología y la química sanguínea, se distribuyeron al azar 21 corderas West African, balanceadas por peso y carga parasitaria gastrointestinal en tres grupos homogéneos (n=7). Los animales pastorearon 6 h/d, luego de la suplementación con uno de los tres tratamientos: T (sin canavalia), C (2,5g de canavalia/kg PV) y C+ (5g de canavalia/kg PV). Las variables sanguíneas se determinaron a través de muestreos bisemanales y se analizaron a través de un ANAVAR. Los resultados indicaron que C+ causó anemia debido a los valores menores de hematocrito (25,6%) en comparación con T (28,3%) y C (26,2%) (P=0,0003); y con valores de hemoglobina menores en C+ (8,8 g/dL), en comparación con T (9,8 g/dL) y C (9,15 g/dL) (P=0,0006). Los valores de alanina transaminasa, glucosa y colesterol, de todos los animales, se encontraron dentro de los rangos referenciales. En conclusión, la suplementación con 5 g/kg PV de semillas molidas de Canavalia ensiformis, puede provocar signos de anemia en corderas tropicales en crecimiento. En menor proporción (2,5 g/kg PV) no afecta la hematología ni la química sanguínea.

The purpose of this investigation was to evaluate the effect of supplementation with Canavalia ensiformis ground seeds on hematology and blood chemistry of West African lambs. For that, a total of 21 lambs (grouped by similar weights and parasite load) were randomly divided into three homogeneous groups of seven animals each (n=7). The animals grazed 6 hours/day, after supplementation with one of the following three treatments: T (not supplemented with canavalia); C (supplemented with 2.5g canavalia/kg BW); and C+ (supplemented with canavalia 5g/kg BW). Blood variables were determined through sampling biweekly and analyzed through ANOVA. The results of the investigation showed that the C+ group caused anemia, as reflected by a lower hematocrit value (25.6%), when compared to T (28.3%) and C (26.2%); (P=0.0003). Furthermore, hemoglobin levels were also lower for C+ (8.8 g /dL), as compared to T (9.8 g /dL) and C (9.15 g / dL); (P=0.0006). Alanine transaminase, glucose and cholesterol values of all animals were within reference ranges for the species. In conclusion, supplementation with a higher amount of canavalia seeds (5g/kg BW), can cause signs of anemia in growing tropical lambs. In contrast, a lesser proportion of this substance (2.5g/Kg BW) did not affect hematology and blood chemistry.

Utility of inflammatory markers in predicting the aetiology of pneumonia in children.

We aimed to investigate the diagnostic value of applying cut-off levels of inflammatory markers and to develop a prediction model for differentiation between bacterial and viral infections in paediatric community-acquired pneumonia based on C-reactive protein (CRP), neutrophil, and white cell counts (WCC). Amongst 401 children, those with bacterial pneumonia were older than those with viral pneumonia (P<0.001). Compared to viral, bacterial infections had a higher median CRP level (P<0.001), whereas WCC and neutrophil count were not different. Bacterial infections were associated with higher CRP >80 mg/L than viral infections (P=0.001), but levels <20 mg/L were not discriminatory (P=0.254). Receiver operating characteristic curve of the model for differentiating bacterial from viral pneumonia based on age, CRP, and neutrophil count produced area under the curve of 0.894 with 75.7% sensitivity and 89.4% specificity. This aetiological discriminant prediction model is a potentially useful tool in clinical management and epidemiological studies of paediatric pneumonia.

Effect of increased glucose levels on short-term outcome in hypertensive spontaneous intracerebral hemorrhage.

OBJECTIVE: Spontaneous intracerebral hemorrhage (ICH) can be a devastating event. Increased glucose levels in the plasma may be related to poor outcomes; however, the precise association remains unclear. METHODS: We retrospectively assessed 116 patients with hypertensive ICH. Glucose level in the plasma was assessed at days 0, 1, and 3. Outcome variables were mortality within 7 and 30days and the National Institutes of Health Stroke Scale (NIHSS) score at day 14 after ICH onset. RESULTS: Twenty deaths had occurred by day 7, and the 30-day mortality rate was 31.9%. Hyperglycemia at day 0 was significantly more common in patients who died within 7days or 30days. Hyperglycemia at day 1 was more common in patients with an NIHSS score >15 on admission and at day 14. No differences in glucose levels were found between diabetic and non-diabetic patients. Among non-diabetic patients, higher glucose levels were related to poorer outcomes (death or an NIHSS score >15). In multivariate analysis, glucose levels >140mg/dL at day 1 were related to the 30-day mortality (hazard ratio=2.65; 95% confidence interval [CI]=1.15-6.12, p=0.02), and glucose levels >160mg/dL at day 1 were associated with an NIHSS score >15 at day 14 (odds ratio=3.08; 95% CI=0.9-10.5, p=0.07). White blood cell counts were directly associated with poorer outcomes and significantly correlated to glucose levels. CONCLUSION: Initially increased glucose levels and increased levels within 24h of ICH onset were related to poorer outcomes. Altered glucose metabolism may be due to inflammatory cell activation. Further studies are needed to clarify the association between immune activation and glucose metabolism after ICH onset.

Physical examination instead of laboratory tests for most infants born to mothers colonized with group B Streptococcus: support for the Centers for Disease Control and Prevention's 2010 recommendations.

OBJECTIVE: To compare 2 approaches in the management of neonates at risk for group B Streptococcus early-onset sepsis: laboratory tests plus standardized physical examination and standardized physical examination alone. STUDY DESIGN: Prospective, sequential study over 2 consecutive 12-month periods, carried out in the maternity hospitals of the region Friuli-Venezia Giulia (north-eastern Italy). All term infants were included (7628 in the first period, 7611 in the second). In the first period, complete blood count and blood culture were required for all infants at risk, followed by a 48-hour period of observation with a standardized physical examination. In the second period, only standardized physical examination was performed. Study outcomes were: (1) number of neonates treated with antibiotics; and (2) time between onset of signs of possible sepsis and beginning of treatment. RESULTS: There was no difference between the 2 periods in the rate of maternal colonization (19.7% vs 19.8%, P = .8), or in other risk factors. The interval between onset of signs of sepsis and starting of antibiotics was not different in the 2 periods. Significantly fewer infants were treated with antibiotics in the second period (0.5% vs 1.2%, P < .001). CONCLUSIONS: Laboratory tests together with standardized physical examination seem to offer no advantage over standardized physical examination alone; the latter was associated with fewer antibiotic treatments. Our results are in agreement with the Center for Disease Control and Prevention's 2010 recommendations.