Relationship of Late Lactation Milk Somatic Cell Count and Cathelicidin with Intramammary Infection in Small Ruminants

Late lactation is a critical moment for making mastitis management decisions, but in small ruminants the reliability of diagnostic tests is typically lower at this stage. We evaluated somatic cell counts (SCC) and cathelicidins (CATH) in late lactation sheep and goat milk for their relationship with intramammary infections (IMI), as diagnosed by bacteriological culture (BC). A total of 315 sheep and 223 goat half-udder milk samples collected in the last month of lactation were included in the study. IMI prevalence was 10.79% and 15.25%, respectively, and non-aureus staphylococci were the most common finding. Taking BC as a reference, the diagnostic performance of SCC and CATH was quite different in the two species. In sheep, receiver operating characteristic (ROC) analysis produced a higher area under the curve (AUC) value for CATH than SCC (0.9041 versus 0.8829, respectively). Accordingly, CATH demonstrated a higher specificity than SCC (82.92% versus 73.67%, respectively) at comparable sensitivity (91.18%). Therefore, CATH showed a markedly superior diagnostic performance than SCC in late lactation sheep milk. In goats, AUC was <0.67 for both parameters, and CATH was less specific than SCC (61.90% versus 65.08%) at comparable sensitivity (64.71%). Therefore, both CATH and SCC performed poorly in late lactation goats. In conclusion, sheep can be screened for mastitis at the end of lactation, while goats should preferably be tested at peak lactation. In late lactation sheep, CATH should be preferred over SCC for its higher specificity, but careful cost/benefit evaluations will have to be made

Proteomic changes in the milk of water buffaloes (Bubalus bubalis) with subclinical mastitis due to intramammary infection by Staphylococcus aureus and by non-aureus staphylococci

Subclinical mastitis by Staphylococcus aureus (SAU) and by non-aureus staphylococci (NAS) is a major issue in the water buffalo. To understand its impact on milk, 6 quarter samples with >3,000,000 cells/ mL (3 SAU-positive and 3 NAS-positive) and 6 culture-negative quarter samples with <50,000 cells/ mL were investigated by shotgun proteomics and label-free quantitation. A total of 1530 proteins were identified, of which 152 were significantly changed. SAU was more impacting, with 162 vs 127 differential proteins and higher abundance changes (P < 0.0005). The 119 increased proteins had mostly structural (n = 43, 28.29%) or innate immune defence functions (n = 39, 25.66%) and included vimentin, cathelicidins, histones, S100 and neutrophil granule proteins, haptoglobin, and lysozyme. The 33 decreased proteins were mainly involved in lipid metabolism (n = 13, 59.10%) and included butyrophilin, xanthine dehydrogenase/oxidase, and lipid biosynthetic enzymes. The same biological processes were significantly affected also upon STRING analysis. Cathelicidins were the most increased family, as confirmed by western immunoblotting, with a stronger reactivity in SAU mastitis. S100A8 and haptoglobin were also validated by western immunoblotting. In conclusion, we generated a detailed buffalo milk protein dataset and defined the changes occurring in SAU and NAS mastitis, with potential for improving detection (ProteomeXchange identifier PXD012355)

Severe asthma: One disease and multiple definitions

Introduction: There is, so far, no universal definition of severe asthma. This definition usually relies on: number of exacerbations, inhaled therapy, need for oral corticosteroids, and respiratory function. The use of such parameters varies in the different definitions used. Thus, according to the parameters chosen, each patient may result in having severe asthma or not. The aim of this study was to evaluate how the choice of a specific definition of severe asthma can change the allocation of patients. Methods: Data collected from the Severe Asthma Network Italy (SANI) registry were analyzed. All the patients included were then reclassified according to the definitions of U-BIOPRED, NICE, WHO, ATS/ERS, GINA, ENFUMOSA, and TENOR. Results: 540 patients, were extracted from the SANI database. We observed that 462 (86%) met the ATS/ERS criteria as well as the GINA criteria, 259 (48%) the U-Biopred, 222 (41%) the NICE, 125 (23%) the WHO, 313 (58%) the Enfumosa, and 251 (46%) the TENOR criteria. The mean eosinophil value were similar in the ATS/ERS, U-Biopred, and Enfumosa (528, 532 and 516 cells/mcl), higher in WHO and Tenor (567 and 570 cells/mcl) and much higher in the NICE classification (624 cells/mcl). Lung function tests resulted similarly in all groups, with WHO (67%) and ATS/ERS-GINA (73%), respectively, showing the lower and upper mean FEV1 values. Conclusions: The present observations clearly evidence the heterogeneity in the distribution of patients when different definitions of severe asthma are used. However, the recent definition of severe asthma, provided by the GINA document, is similar to that indicated in 2014 by ATS/ERS, allowing mirror reclassification of the patients examined. This lack of homogeneity could complicate the access to biological therapies. The definition provided by the GINA document, which reflects what suggested by ATS/ERS, could partially overcome the problem

The value of the biomarkers cathelicidin, milk amyloid A, and haptoglobin to diagnose and classify clinical and subclinical mastitis

Timely and objective diagnosis and classification of mastitis is crucial to ensure adequate management and therapeutic decisions. Analyzing specific biomarkers in milk could be advantageous compared with subjec-tive or semiquantitative criteria, such as palpation of the udder in clinical mastitis cases or evaluation of somatic cell count using cow side tests (e.g., California Mastitis Test) in subclinical mastitis quarters. The ob-jective of this study was to investigate the diagnostic value of 3 biomarkers; cathelicidin, milk amyloid A, and haptoglobin for the diagnosis of subclinical and clinical mastitis. Furthermore, the suitability of these biomarkers to differentiate between mild, moderate, and severe clinical mastitis and the influence of differ-ent pathogens on biomarker levels was tested. A total of 67 healthy cows, 119 cows with subclinical mastitis, and 212 cows with clinical mastitis were enrolled in the study. Although cathelicidin, haptoglobin, and milk amyloid A were measured in all samples from healthy cows and those with subclinical mastitis, haptoglobin, and cathelicidin results were only available from 121 out of 212 cows with clinical mastitis. Milk amyloid A was measured in all samples. In cows with clinical mastitis, the mastitic quarter and a second healthy quarter serv-ing as a healthy in-cow control quarter were sampled. It was possible to differentiate between healthy quarters, quarters with subclinical mastitis, and quarters with clinical mastitis using all 3 biomarkers. Concerning cathelicidin, thresholds were 0.000 [sensitivity (Se) = 0.83, specificity (Sp) = 0.97] and 0.053 (Se = 0.98, Sp = 0.99) for normalized optical density at 450 nm (NOD450) for differentiating between healthy quarters and quarters with subclinical or clinical mastitis, re-spectively. Thresholds of 1.28 \u3bcg/mL (Se = 0.65, Sp = 0.76) and 1.81 \u3bcg/mL (Se = 0.77, Sp = 0.83) for milk amyloid A and 3.65 \u3bcg/mL (Se = 0.92, Sp = 0.94) and 5.40 \u3bcg/mL mL (Se = 0.96, Sp = 0.99) for hap-toglobin were calculated, respectively. Healthy in-cow control quarters from cows with CM showed elevated milk amyloid A and haptoglobin levels compared with healthy quarters from healthy cows. Only the level of milk amyloid A was higher in severe clinical mastitis cases compared with mild ones. In contrast to clini-cal mastitis, cathelicidin and haptoglobin in subclini-cal mastitis quarters were significantly influenced by different bacteriological results. The measurement of cathelicidin, milk amyloid A, and haptoglobin in milk proved to be a reliable method to detect quarters with subclinical or clinical mastiti

Evaluation of milk cathelicidin for detection of dairy sheep mastitis

Mastitis due to intramammary infections is one of the most detrimental diseases in dairy sheep farming, representing a major cause of reduced milk productions and quality losses. In particular, subclinical mastitis presents significant detection and control problems, and the availability of tools enabling its timely, sensitive, and specific detection is therefore crucial. We have previously demonstrated that cathelicidins, small proteins implicated in the innate immune defense of the host, are specifically released in milk of mastitic animals by both epithelial cells and neutrophils. Here, we describe the development of an ELISA for milk cathelicidin and assess its value against somatic cell counts (SCC) and bacteriological culture for detection of ewe mastitis. Evaluation of the cathelicidin ELISA was carried out on 705 half-udder milk samples from 3 sheep flocks enrolled in a project for improvement of mammary health. Cathelicidin was detected in 35.3% of milk samples (249/705), and its amount increased with rising SCC values. The cathelicidin-negative (n\ua0=\ua0456) and cathelicidin-positive (n\ua0=\ua0249) sample groups showed a clear separation in relation to SCC, with median values of 149,500 and 3,300,000 cells/mL, respectively. Upon bacteriological culture, 20.6% (145/705) of the milk samples showed microbial growth, with coagulase-negative staphylococci being by far the most frequent finding. A significant proportion of all bacteriologically positive milk samples were positive for cathelicidin (110/145, 75.9%). Given the lack of a reliable gold standard for defining the true disease status, sensitivity (Se) and specificity (Sp) of the cathelicidin ELISA were assessed by latent class analysis against 2 SCC thresholds and against bacteriological culture results. At an SCC threshold of 500,000 cells/mL, Se and Sp were 92.3 and 92.3% for cathelicidin ELISA, 89.0 and 94.9% for SCC, and 39.4 and 93.6% for bacteriological culture, respectively. At an SCC threshold of 1,000,000 cells/mL, Se and Sp were 93.3 and 91.9% for cathelicidin ELISA, 80.0 and 97.1% for SCC, and 39.4 and 93.5% for bacteriology, respectively. In view of the results obtained in this study, the measurement of cathelicidin in milk by ELISA can provide added Se while maintaining a high Sp and may therefore improve detection of subclinical mastitis

Severe asthma: One disease and multiple definitions

123noopenIntroduction: There is, so far, no universal definition of severe asthma. This definition usually relies on: number of exacerbations, inhaled therapy, need for oral corticosteroids, and respiratory function. The use of such parameters varies in the different definitions used. Thus, according to the parameters chosen, each patient may result in having severe asthma or not. The aim of this study was to evaluate how the choice of a specific definition of severe asthma can change the allocation of patients. Methods: Data collected from the Severe Asthma Network Italy (SANI) registry were analyzed. All the patients included were then reclassified according to the definitions of U-BIOPRED, NICE, WHO, ATS/ERS, GINA, ENFUMOSA, and TENOR. Results: 540 patients, were extracted from the SANI database. We observed that 462 (86%) met the ATS/ERS criteria as well as the GINA criteria, 259 (48%) the U-Biopred, 222 (41%) the NICE, 125 (23%) the WHO, 313 (58%) the Enfumosa, and 251 (46%) the TENOR criteria. The mean eosinophil value were similar in the ATS/ERS, U-Biopred, and Enfumosa (528, 532 and 516 cells/mcl), higher in WHO and Tenor (567 and 570 cells/mcl) and much higher in the NICE classification (624 cells/mcl). Lung function tests resulted similarly in all groups, with WHO (67%) and ATS/ERS-GINA (73%), respectively, showing the lower and upper mean FEV1 values. Conclusions: The present observations clearly evidence the heterogeneity in the distribution of patients when different definitions of severe asthma are used. However, the recent definition of severe asthma, provided by the GINA document, is similar to that indicated in 2014 by ATS/ERS, allowing mirror reclassification of the patients examined. This lack of homogeneity could complicate the access to biological therapies. The definition provided by the GINA document, which reflects what suggested by ATS/ERS, could partially overcome the problem.restrictedopenBagnasco D.; Paggiaro P.; Latorre M.; Folli C.; Testino E.; Bassi A.; Milanese M.; Heffler E.; Manfredi A.; Riccio A.M.; De Ferrari L.; Blasi F.; Canevari R.F.; Canonica G.W.; Passalacqua G.; Guarnieri G.; Patella V.; Maria Pia F.B.; Carpagnano G.E.; Colle A.D.; Scioscia G.; Gerolamo P.; Puggioni F.; Racca F.; Favero E.; Iannacone S.; Savi E.; Montagni M.; Camiciottoli G.; Allegrini C.; Lombardi C.; Spadaro G.; Detoraki C.; Menzella F.; Galeone C.; Ruggiero P.; Yacoub M.R.; Berti A.; Scichilone N.; Durante C.; Costantino M.T.; Roncallo C.; Braschi M.; D'Adda A.; Ridolo E.; Triggiani M.; Parente R.; Maria D.A.; Verrillo M.V.; Rolla G.; Brussino L.; Frazzetto A.V.; Cristina Z.M.; Lilli M.; Crimi N.; Bonavia M.; Corsico A.G.; Grosso A.; Del Giacco S.; Deidda M.; Ricciardi L.; Isola S.; Cicero F.; Amato G.; Vita F.; Spanevello A.; Pignatti P.; Cherubino F.; Visca D.; Massimo Ricciardolo F.L.; Anna Carriero V.M.; Bertolini F.; Santus P.; Barlassina R.; Airoldi A.; Guida G.; Eleonora N.; Aruanno A.; Rizzi A.; Caruso C.; Colantuono S.; Senna G.; Caminati M.; Arcolaci A.; Vianello A.; Bianchi F.C.; Marchi M.R.; Centanni S.; Luraschi S.; Ruggeri S.; Rinaldo R.; Parazzini E.; Calabrese C.; Flora M.; Cosmi L.; Di Pietro L.; Maggi E.; Pini L.; Macchia L.; Di Bona D.; Richeldi L.; Condoluci C.; Fuso L.; Bonini M.; Farsi A.; Carli G.; Montuschi P.; Santini G.; Conte M.E.; Turchet E.; Barbetta C.; Mazza F.; D'Alo S.; Pucci S.; Caiaffa M.F.; Minenna E.; D'Elia L.; Pasculli C.; Viviano V.; Tarsia P.; Rolo J.; Di Proietto M.; Lo Cicero S.Bagnasco, D.; Paggiaro, P.; Latorre, M.; Folli, C.; Testino, E.; Bassi, A.; Milanese, M.; Heffler, E.; Manfredi, A.; Riccio, A. M.; De Ferrari, L.; Blasi, F.; Canevari, R. F.; Canonica, G. W.; Passalacqua, G.; Guarnieri, G.; Patella, V.; Maria Pia, F. B.; Carpagnano, G. E.; Colle, A. D.; Scioscia, G.; Gerolamo, P.; Puggioni, F.; Racca, F.; Favero, E.; Iannacone, S.; Savi, E.; Montagni, M.; Camiciottoli, G.; Allegrini, C.; Lombardi, C.; Spadaro, G.; Detoraki, C.; Menzella, F.; Galeone, C.; Ruggiero, P.; Yacoub, M. R.; Berti, A.; Scichilone, N.; Durante, C.; Costantino, M. T.; Roncallo, C.; Braschi, M.; D'Adda, A.; Ridolo, E.; Triggiani, M.; Parente, R.; Maria, D. A.; Verrillo, M. V.; Rolla, G.; Brussino, L.; Frazzetto, A. V.; Cristina, Z. M.; Lilli, M.; Crimi, N.; Bonavia, M.; Corsico, A. G.; Grosso, A.; Del Giacco, S.; Deidda, M.; Ricciardi, L.; Isola, S.; Cicero, F.; Amato, G.; Vita, F.; Spanevello, A.; Pignatti, P.; Cherubino, F.; Visca, D.; Massimo Ricciardolo, F. L.; Anna Carriero, V. M.; Bertolini, F.; Santus, P.; Barlassina, R.; Airoldi, A.; Guida, G.; Eleonora, N.; Aruanno, A.; Rizzi, A.; Caruso, C.; Colantuono, S.; Senna, G.; Caminati, M.; Arcolaci, A.; Vianello, A.; Bianchi, F. C.; Marchi, M. R.; Centanni, S.; Luraschi, S.; Ruggeri, S.; Rinaldo, R.; Parazzini, E.; Calabrese, C.; Flora, M.; Cosmi, L.; Di Pietro, L.; Maggi, E.; Pini, L.; Macchia, L.; Di Bona, D.; Richeldi, L.; Condoluci, C.; Fuso, L.; Bonini, M.; Farsi, A.; Carli, G.; Montuschi, P.; Santini, G.; Conte, M. E.; Turchet, E.; Barbetta, C.; Mazza, F.; D'Alo, S.; Pucci, S.; Caiaffa, M. F.; Minenna, E.; D'Elia, L.; Pasculli, C.; Viviano, V.; Tarsia, P.; Rolo, J.; Di Proietto, M.; Lo Cicero, S

ALICE upgrades during the LHC Long Shutdown 2

International audienceA Large Ion Collider Experiment (ALICE) has been conceived and constructed as a heavy-ion experiment at the LHC. During LHC Runs 1 and 2, it has produced a wide range of physics results using all collision systems available at the LHC. In order to best exploit new physics opportunities opening up with the upgraded LHC and new detector technologies, the experiment has undergone a major upgrade during the LHC Long Shutdown 2 (2019–2022). This comprises the move to continuous readout, the complete overhaul of core detectors, as well as a new online event processing farm with a redesigned online-offline software framework. These improvements will allow to record Pb-Pb collisions at rates up to 50 kHz, while ensuring sensitivity for signals without a triggerable signature