Final analysis of phase II results with cemiplimab in metastatic basal cell carcinoma after hedgehog pathway inhibitors.

BACKGROUND: Metastatic basal cell carcinoma (mBCC) is a rare condition with no effective second-line treatment options. Cemiplimab is an immune checkpoint inhibitor that blocks the binding of programmed cell death-1 (PD-1) to its ligands, programmed death-ligand 1 (PD-L1) and programmed death-ligand 2 (PD-L2). Here, we present the final analysis of cemiplimab in patients with mBCC after first-line hedgehog pathway inhibitor (HHI) treatment (NCT03132636). PATIENTS AND METHODS: In this open-label, single-arm, phase II study, adults with mBCC and Eastern Cooperative Oncology Group performance status ≤1, post-HHI treatment, received cemiplimab 350 mg intravenously every 3 weeks for ≤93 weeks or until disease progression or unacceptable toxicity. The primary endpoint was objective response rate (ORR) by independent central review (ICR). Duration of response (DOR) was a key secondary endpoint. Other secondary endpoints were ORR per investigator assessment, progression-free survival (PFS), overall survival (OS), complete response rate, safety, and tolerability. RESULTS: Fifty-four patients were enrolled: 70% were male and the median age of patients was 64 [interquartile range (IQR) 57.0-73.0] years. The median duration of follow-up was 8 months (IQR 4-21 months). The ORR per ICR was 22% [95% confidence interval (CI) 12% to 36%], with 2 complete responses and 10 partial responses. Among responders, the median time to response per ICR was 3 months (IQR 2-7 months). The estimated median DOR per ICR was not reached [95% CI 10 months-not evaluable (NE)]. The disease control rate was 63% (95% CI 49% to 76%) per ICR and 70% (95% CI 56% to 82%) per investigator assessment. The median PFS per ICR was 10 months (95% CI 4-16 months); the median OS was 50 months (95% CI 28 months-NE). The most common treatment-emergent adverse events were fatigue [23 (43%)] and diarrhoea [20 (37%)]. There were no treatment-related deaths. CONCLUSIONS: Cemiplimab demonstrated clinically meaningful antitumour activity, including durable responses, and an acceptable safety profile in patients with mBCC who had disease progression on or intolerance to HHI therapy.

A randomized, controlled trial examining quarter-level somatic cell count and culture-based selective dry cow therapy against blanket dry cow therapy on early-lactation production outcomes.

The objective of this study was to determine quarters requiring antimicrobial treatment using either a benchtop somatic cell counter (S-SDCT) or culture with gram-positive selective media (C-SDCT) and compare outcomes in these cows to those receiving blanket dry cow therapy (BDCT) in a randomized, controlled trial. Two novel methods of identifying cows with intramammary infections followed by selective antimicrobial treatment were evaluated at a commercial dairy farm to determine their usefulness in decreasing antibiotic usage during the dry period without significant detrimental effects on milk quality and production. Cows (n = 840) were randomly allocated to one of 3 groups (BDCT, C-SDCT, S-SDCT) the day before dry-off and quarter-level milk samples (QLMS) were collected. The QLMS from cows in the S-SDCT group were evaluated using the cell counter and quarters were treated if somatic cell count (SCC) was ≥200,000 cells/mL, while QLMS from cows in the C-SDCT group were cultured and quarters were treated if the culture showed growth. All cows in the BDCT received antimicrobial therapy and all cows received an internal teat sealant regardless of treatment group. Outcomes measured were first and second DHIA test somatic cell count, milk production through 60 d in milk, cows leaving the farm, clinical mastitis, and bacteriologic new infections in a subset of quarters. Cows in both SDCT groups had fewer antimicrobial treatments than cows in the BDCT group as was expected, and cows in the C-SDCT group had fewer treatments than those in the S-SDCT group. Cows in both SDCT groups had higher linear score at the first DHIA test (BDCT: 1.8, S-SDCT: 2.2, C-SDCT: 2.2), however there were no other differences between groups regarding any other outcomes measured. While antimicrobial use was significantly reduced, farms should use caution in adopting the benchtop analyzer and the selective media described in this study as ways to identify infected cows for dry cow therapy as they may result in increased linear score early in lactation.

Association of time to metritis diagnosis with circulating concentration of metabolites, minerals, and haptoglobin in Jersey cows.

Our objective was to investigate the association of early metritis [EMET, diagnosed at <5 d in milk (DIM)] and late metritis (LMET, diagnosed at ≥5 DIM) with circulating concentrations of energy metabolites, minerals, and haptoglobin (Hp) throughout the first 14 d postpartum. A total of 379 purebred Jersey cows were enrolled in a prospective cohort study from a single herd in west Texas. Cows were examined for metritis using the Metricheck device (Simcro Ltd.) at 4, 7, and 10 DIM. Cows identified by farm employees as possible metritis cases were also evaluated for metritis. Blood samples were collected for analysis of concentrations of Ca, Mg, and glucose at DIM 1 through 5, 7, 10, and 14. Albumin, urea, fructosamine, free fatty acids (FFA), creatinine, and ß-hydroxybutyrate (BHB) were analyzed at DIM 3, 5, 7, 10, and 14, and Hp at DIM 1 through 5 and 7. Data were analyzed using the MIXED and PHREG procedures of SAS (SAS Institute Inc.). A series of mixed general linear models accounting for repeated measures were fitted to the data. The independent variables metritis [no metritis (NMET), EMET, and LMET], DIM of analyte assessment, and parity were forced in all models. Multivariable Cox proportional hazard models were built to assess the risk of pregnancy and culling within 150 DIM. The overall metritis incidence was 26.9% (EMET = 49; LMET = 53; NMET = 277). Average concentrations of glucose, Mg, and urea were not associated with metritis. The associations of Ca, creatinine, BHB, and fructosamine with metritis were dependent on the DIM of analyte assessment. Cows categorized as EMET and LMET had, on average, lower albumin and fructosamine compared with NMET cows. Both EMET and LMET cows had, on average, greater BHB than NMET cows. A greater FFA concentration was only observed in cows diagnosed with EMET compared with NMET cows (EMET = 0.58, LMET = 0.52, NMET = 0.48 mmol/L). Additionally, circulating Hp concentration was greater for LMET and EMET compared with NMET cows, and EMET cows had greater Hp compared with LMET cows (EMET = 1.15; LMET = 1.00; NMET = 0.84). In conclusion, several blood biomarkers were temporally associated with early versus late metritis diagnosis in postpartum Jersey cows. No meaningful differences were observed in production, reproduction, or culling between EMET and LMET cows. These results suggest that cows with EMET undergo a more severe degree of inflammation and negative energy balance compared with NMET cows.

The potential role of physical activity and a healthy diet in increasing nitric oxide during COVID-19 outbreak.

The potential role of physical activity and a healthy diet in increasing nitric oxide during COVID-19 outbreak. This manuscript presents a perspective which provide new insights about the promising role of nitric oxide on COVID-19. Demonstration that nitric oxide was an important cornerstone against viral infections, including SARS-CoV-1 in 2009. Thus, given the concern that higher NO- could improve endothelial health and might be a protection factor against COVID-19, should we critically consider non-pharmacological strategies that increase NO- bioavailability as medicine for COVID-19? From this perspective, we highlight the potential effect of physical activity and healthy diet in stimulating the increase of NO- bioavailability.

Les rôles potentiels de l'activité physique et d'une alimentation saine dans l'augmentation de l'oxyde nitrique pendant l'épidémie de COVID-19. Ce manuscrit présente une perspective qui fournit de nouvelles informations sur le rôle prometteur de l'oxyde nitrique sur la protection contre le risque de COVID-19. Dès 2009, a été évoqué le rôle de l'oxyde nitrique contre le risque d'infections virales, y compris contre la première pandémie liée au coronavirus SARS-CoV-1. Compte tenu de l'hypothèse qu'une augmentation de la production de NO− permettrait d'améliorer la santé endothéliale et pourrait être un facteur de protection contre COVID-19, la question se pose sur la promotion de stratégies non pharmacologiques qui augmentent la biodisponibilité du NO−. C'est dans cette perspective que sont, ici, discutés les effets potentiels de l'activité physique et d'une alimentation saine pour stimuler l'augmentation de la biodisponibilité du NO et la prévention contre la COVID-19.

Prolonged, low-grade inflammation in the first week of lactation: Associations with mineral, protein, and energy balance markers, and milk yield, in a clinically healthy Jersey cow cohort.

Our objectives were to perform a proof-of-concept study to assess the association of a prolonged inflammatory state (based on a continually elevated haptoglobin concentration at the end of the first week after parturition) with mineral, protein, and energy balance markers in the first 2 wk after parturition, and milk production in the first 60 d of lactation in clinically healthy cows. We conducted a cohort study in 1 herd in west Texas that was milking Jersey and Jersey-Holstein crosses. Only multiparous Jersey cows were eligible for enrollment. Cows were classified as having or not having elevated haptoglobin concentrations based on plasma concentrations evaluated on d 4 and 7 postpartum. We used median concentrations of haptoglobin in the reference population (i.e., before the exclusion of cows diagnosed with clinical diseases) as the limits for categorizing cows into 2 groups: cows with plasma haptoglobin concentrations greater than the median values on both d 4 (0.49 g/L) and 7 (0.35 g/L) had continually elevated haptoglobin (with eHp); and cows with plasma haptoglobin concentrations lower than or equal to the median values of the reference population on d 4 or 7 did not have continually elevated haptoglobin (without eHp). Next, cows with clinical diseases in the first 2 wk of the postpartum period were excluded, so that 233 cows remained for the final analyses. Evaluated outcomes were average daily milk production across the first 60 d of lactation, plasma Ca, Mg, and glucose concentrations on d 1, 2, 3, 4, 5, 7, 10, and 14 postpartum, and ß-hydroxybutyrate (BHB), nonesterified fatty acids (NEFA), fructosamine, albumin, urea, and creatinine concentrations on d 3, 5, 7, 10, and 14 postpartum. Rectal temperatures measured on d 4, 7, and 10 postpartum were also analyzed. We performed statistical analyses using linear mixed models while accounting for the repeated effect of sampling time (plasma analytes and rectal temperature models) and weekly milk test (milk production model). Cows with eHp had lower plasma Ca concentrations in the first 2 wk after calving, but no differences in Mg, BHB, NEFA, glucose, or rectal temperatures compared to cows without eHp. Cows with eHp had lower plasma fructosamine, albumin, and urea concentrations in a time-dependent manner. They also had lower milk production (2.3 kg/d less than cows without eHp in the first 60 DIM). Our study demonstrated that 25% of cows without clinical disorders in the first 2 wk after parturition had continually elevated haptoglobin concentrations at d 7 after parturition relative to d 4, suggestive of a prolonged, low-grade systemic inflammatory state. More observational studies are needed to more fully characterize the duration of prolonged postpartum subclinical inflammation in cows without clinical diseases, as well as its long-term effects, and to evaluate the use of other potential markers of systemic inflammation to describe this disorder.

Short communication: Associations of serum biomarkers of stress and inflammation measured at arrival with health, mortality, and growth of calves transported within the first 4 days of life.

The objective of this study was to assess the association between serum concentrations of haptoglobin (Hp), cortisol, and l-lactate following transport of dairy heifer calves within their first week of life with health, mortality, and growth during the preweaning period. Blood samples were collected at arrival and included a cohort of 168 Jersey and Jersey-cross calves from 8 different sources located in Minnesota. All calves were raised at a single facility in New Mexico. Incidence of respiratory disease (BRD) and mortality during the preweaning period (60 d of life) were extracted from the farm's software database. Individual body weight was measured at birth and at weaning to estimate average daily gain. No simple linear correlations were found between the biomarkers. The incidence of BRD during the preweaning period was 7.7%. While cortisol and l-lactate serum concentrations were not associated with BRD, Hp was negatively associated with BRD. Receiving operating characteristic curve analysis was performed to determine the optimal threshold value for Hp [threshold ≤0.63 µmol/L (63 µg/mL), area under the curve = 0.65]. A Cox's proportional hazards model revealed that calves with Hp concentration ≤0.63 µmol/L were more likely to be diagnosed with BRD (hazard ratio = 5.0, 95% confidence interval = 1.3-19.0). A mixed linear model showed that calves with Hp concentration ≤0.63 µmol/L tended to have lower average daily gain (454.4 vs. 479.9 g/d) during the preweaning period than calves with Hp >0.63 µmol/L at arrival. Overall mortality of the cohort was 3.5%, and Hp was not associated with mortality. Although circulating concentrations of l-lactate and cortisol measured at arrival were not associated with BRD incidence during the preweaning period of heifer calves transported within the first 4 d of life, calves with serum Hp concentration >0.63 µmol/L were less likely to be diagnosed with BRD and tended to grow more than calves with Hp ≤0.63 µmol/L. Further research is needed to replicate these results in a larger cohort and to better understand the possible influence of greater inflammatory status at arrival on health and growth of calves transported to a calf-raising facility.

Association of blood calcium concentration in the first 3 days after parturition and energy balance metabolites at day 3 in milk with disease and production outcomes in multiparous Jersey cows.

Research exploring specific associations of markers of negative energy balance and Ca in postpartum Jersey cows with lactation performance is lacking. Our objectives were to evaluate the associations of total Ca concentration (tCa) measured at 1 through 3 d in milk (DIM) and free fatty acids (FFA), ß-hydroxybutyrate (BHB), and glucose measured at 3 DIM with (1) the risk of multiparous Jersey cows being diagnosed with early-lactation diseases and culling, (2) milk production in the first 9 wk of lactation, and (3) the risk of pregnancy in the first 150 DIM. A cohort study was performed in 1 dairy herd in Texas. Multivariable Poisson regression models were built to evaluate the association of the analytes of interest with the risks of early-lactation diseases and culling in the first 60 DIM (i.e., binary outcomes). Linear mixed models were used to evaluate the association of the analytes of interest with milk production within the first 9 wk of lactation, and a Cox proportional hazard model was built to assess the risk of pregnancy within 150 DIM. A total of 380 cows were used in the final analyses. Total Ca measured at 1 through 3 DIM was not associated with the risk of metritis. Cows with increased FFA and BHB had an increased risk of being diagnosed with metritis and clinical mastitis, respectively. Increased concentrations of glucose and FFA and decreased tCa at 3 DIM were associated with an increased risk of culling. Reduced tCa concentrations at 1 DIM (≤1.84 mmol/L) and 2 DIM (≤2.04 mmol/L) were associated with increased milk production across the first 9 wk of lactation compared with tCa concentrations above those thresholds. Total Ca was not associated with milk production when assessed at 3 DIM, whereas increased FFA (≥0.37 mmol/L) and decreased glucose (≤2.96 mmol/L) at 3 DIM were associated with increased milk production. None of the metabolites measured were associated with the risk of pregnancy in the first 150 DIM. Our results demonstrate that tCa concentration assessed in the first 3 DIM show temporary associations with milk production and culling in multiparous Jersey cows. Although increased concentration of FFA assessed at 3 DIM was associated with greater milk yield, it was a detrimental factor for the risk of metritis. This study attempted to better elucidate the relationship of tCa, FFA, BHB, and glucose assessed in early postpartum with health and performance of Jersey cows. Based on this study, assessments performed at 3 DIM using tCa concentration ≤1.99 mmol/L for increased risk of early-lactation culling and FFA ≥0.43 mmol/L for increased risk of metritis could be used as starting points. More studies evaluating the dynamics of energy balance markers and tCa in postpartum Jersey cows using a greater number of herds are needed to better inform dairy consultants on critical levels for exacerbated postpartum negative energy balance and subclinical hypocalcemia for the Jersey breed.

Association of transient, persistent, or delayed subclinical hypocalcemia with early lactation disease, removal, and milk yield in Holstein cows.

Our objectives were to evaluate the association of subclinical hypocalcemia (SCH) dynamics with the risk of early lactation disease, removal, and milk production. We conducted a prospective observational cohort study in 407 Holstein cows in 2 dairy herds in New York. Cows were stratified by parity group (144 primiparous, 263 multiparous) and classified into 1 of 4 groups based on postpartum plasma Ca concentrations previously associated with improved milk production or increased risk of disease: normocalcemic (NC; primiparous [Ca] >2.15 mmol/L at 1 and 2 d in milk, n = 67; multiparous [Ca] >1.77 at 1 d in milk and 2.20 mmol/L at 4 d in milk, n = 109); transient SCH (tSCH; primiparous [Ca] ≤2.15 at 1 d in milk and >2.15 mmol/L at 2 d in milk, n = 25; multiparous [Ca] ≤1.77 at 1 d in milk and >2.20 mmol/L at 4 d in milk, n = 50); persistent SCH (pSCH; primiparous [Ca] ≤2.15 mmol/L at 1 and 2 d in milk, n = 33; multiparous [Ca] ≤1.77 at 1 d in milk and ≤2.20 mmol/L at 4 d in milk, n = 34); or delayed SCH (dSCH; primiparous [Ca] >2.15 at 1 d in milk and ≤2.15 mmol/L at 2 d in milk, n = 19; multiparous [Ca] >1.77 at 1 d in milk and ≤2.20 mmol/L at 4 d in milk, n = 70). Evaluated outcomes were development of an adverse event [hyperketonemia (blood ß-hydroxybutyrate concentration ≥1.2 mmol/L at 3, 5, 7, or 10 d in milk), metritis, displaced abomasum, or herd removal in the first 60 d in milk] and average milk yield per day across the first 10 wk of lactation. Multivariable Poisson regression was used to analyze the adverse event outcome and generalized linear mixed models for milk yield analysis. Primiparous cows with tSCH were no more likely to have an adverse event than NC cows [risk ratio = 1.3; 95% confidence interval (CI) = 0.5 to 3.2], whereas multiparous cows tended to have a higher risk for an adverse event than NC cows (risk ratio = 1.4; 95% CI = 0.9 to 2.1). However, pSCH cows were 4.1 (95% CI = 2.1 to 7.9, primiparous) and 1.8 (95% CI = 1.2 to 2.7, multiparous) times more likely, and dSCH cows 3.2 (95% CI = 1.5 to 7.0, primiparous) and 1.9 (95% CI = 1.3 to 2.6, multiparous) times more likely, to have an adverse event than NC cows. Primiparous and multiparous cows with tSCH made more milk per day than NC, pSCH, or dSCH cows across the first 10 wk of lactation. Primiparous cows averaged 28.5 ± 0.7, 31.9 ± 1.1, 29.7 ± 0.9, and 28.7 ± 1.2 kg/d, and multiparous cows averaged 44.6 ± 0.7, 49.1 ± 1.1, 46.4 ± 1.3, and 41.4 ± 0.9 kg/d for NC, tSCH, pSCH, and dSCH cows, respectively. Our results suggest that cows with tSCH adapt well to early lactation, develop fewer disease or removal events than pSCH or dSCH cows, and make more milk than NC, pSCH, or dSCH cows. Cows with pSCH or dSCH, regardless of parity group, are at an increased risk for early lactation disease or removal events.

Technical note: Effect of delayed analysis of cooled lithium-heparinized whole blood on the stability of ionized calcium, ionized magnesium, sodium, potassium, chloride, glucose, and lactate in samples from dairy cows.

The objectives of this study were to describe the stability of bovine whole-blood electrolytes, glucose, and lactate in samples collected in lithium heparin tubes and stored in thermoconductive modules immersed in ice water. A total of 99 Jersey cows (40 first-parity, 18 second-parity, and 41 third-parity or greater cows) from a commercial dairy farm in West Texas were enrolled between June and July 2018. Blood was collected from the jugular vein using a 60-mL polypropylene syringe and equally distributed into 5 spray-dried evacuated lithium heparin tubes. Baseline samples were analyzed within 90 s of collection using a benchtop blood gas analyzer. The remaining 4 tubes were stored in a thermoconductive, passive-temperature-regulating module inside a cooler with ice water. At 30 min and 2, 4, and 8 h post-collection, samples were removed from the temperature-regulating module, gently inverted for 10 s, and analyzed. Repeated-measures models were built to evaluate the effect of time on the stability of ionized Ca (iCa), ionized Mg (iMg), Na, K, Cl, glucose, and lactate. Most of the analytes investigated remained stable up to 8 h under ice water storage conditions before analysis, including iCa, iMg, Cl, glucose, and lactate. However, Na and K were significantly affected by delayed analysis: Na remained stable up to 4 h post-collection, but K was not stable starting at 2 h post-collection. The results of this study are useful in helping future researchers and consultants to recognize acceptable time delays between whole blood collection and processing or analysis for electrolytes, glucose, and lactate.

Technical note: Effect of storage time and temperature on total calcium concentrations in bovine blood.

Our objectives were to examine the stability of (1) serum and plasma total calcium (tCa) after delayed separation of bovine whole blood stored at 4°C, and (2) frozen serum and plasma tCa stored at -80°C. Whole blood was collected from 19 cows into vacutainer tubes (Becton Dickinson, Franklin Lakes, NJ) containing no additives (serum) or lithium heparin (plasma). Baseline tCa concentrations (0-h tCa) were determined on serum and plasma harvested within 2 h of blood collection. To evaluate the effect of prolonged whole blood storage at 4°C, serum and plasma were harvested from samples after 6, 24, 48, 72 h, and 7 and 14 d of storage. For evaluation of the stability of frozen serum and plasma stored at -80°C, samples were stored for 7 d and 1, 3, 6, and 12 mo. The median 0-h tCa concentration for serum and plasma was 2.25 mmol/L (range: 1.58 to 2.60 mmol/L) and 2.28 mmol/L (range: 1.60 to 2.65 mmol/L), respectively. The average difference in tCa from 0-h samples ranged from -0.02 to 0.03 mmol/L for all samples stored at either 4 or -80°C. The median within-cow variability for whole blood samples stored at 4°C was 1.1 and 1.0% for serum and plasma, respectively, and for serum and plasma stored at -80°C was 1.1 and 1.3%, respectively. When whole blood was stored at 4°C, no differences in serum or plasma tCa concentrations were observed across the evaluated time points as compared with 0-h concentrations. Similarly, frozen serum and plasma stored at -80°C remained stable up to 12 mo. Therefore, our findings show that whole blood samples may be stored for at least 14 d at 4°C in vacutainer tubes containing no additives or lithium heparin without biologically important changes in tCa concentrations beyond expected analytical variation. Additionally, serum or plasma may be stored at -80°C for up to 12 mo with no effect on tCa concentrations.

Effects of oral calcium bolus supplementation on intracellular polymorphonuclear leukocyte calcium levels and functionality in primiparous and multiparous dairy cows.

The objectives of this study were (1) to characterize Ca levels and polymorphonuclear leukocyte (PMN) function in primiparous and multiparous animals following oral Ca bolus supplementation, and (2) to determine differential responses of boluses containing a lower dose of Ca than traditionally used in primiparous animals on Ca levels and PMN function. Jersey × Holstein crossbred animals (n = 104) were enrolled within 24 h of parturition. All animals were blocked by time relative to calving and randomly assigned to treatment. The Ca boluses were composed of a mixture of Ca chloride, Ca sulfate, and Ca propionate. For objective 1, animals were assigned to control (CON; no Ca supplementation), or a series of 2 Ca boluses given 24 h apart for a total of 50 g of Ca. Objective 2 treatments included control (CON; no Ca supplementation), a series of 2 Ca boluses given 24 h apart containing 50 g of Ca, or a series of 2 Ca boluses given 24 h apart containing 25 g of Ca. Blood samples were collected on d 1 (<24 h), 2, 3, 5, and 7 relative to parturition. Total serum Ca, serum haptoglobin, PMN intracellular Ca, PMN intracellular Ca after stimulation with an environmental Escherichia coli, PMN L-selectin surface expression, and PMN phagocytic and oxidative burst activities were analyzed. For objective 1 a tendency was detected for a treatment difference on basal intracellular PMN Ca and a treatment difference on E. coli-stimulated intracellular PMN Ca. We detected a parity × DIM effect for PMN oxidative burst intensity. However, no other interactions or parity effects on other functional PMN variables were detectable. In primiparous animals, we found a treatment difference for E. coli-stimulated intracellular PMN Ca among animals given 50 g of Ca but no treatment difference on basal intracellular PMN Ca. The 50 g of Ca treatment increased both PMN phagocytosis and oxidative burst intensities. Supplementing animals with 50 g of oral Ca increased intracellular PMN Ca and influenced PMN function.

Eurytrema coelomaticum: updated morphology of adult worms using advanced microscopy experiments.

Eurytrema coelomaticum is a digenean flatworm of ruminants that is the causative agent of eurytrematosis, a disease of veterinary health concern. Although modern techniques of morphological analysis have provided new insights about the morphology and anatomy of parasitic helminths, most studies on E. coelomaticum adults are based on conventional light microscopy. In the present study, a combined approach using brightfield, fluorescence, confocal and scanning electron microscopies (SEMs), together with the cryofracture technique, have updated morphological data on E. coelomaticum recovered from cattle in Rio de Janeiro State, Brazil. Light microscopy confirmed the presence of several structures present in the current description, such as suckers, pharynx, oesophagus, intestinal bifurcation and the cirrus-sac. Fluorescence stereomicroscopy revealed for the first time the cubic crystal protein inclusions in the forebody, which were further detailed by confocal and SEMs. Confocal microscopy provided detailed information of the muscular architecture associated with the attachment structures (suckers), digestive system (pharynx and oesophagus), egg-forming complex (ovary, Mehlis' gland and Laurer's canal) and male reproductive system, which are similar to those found in other digenean flukes. SEM images of cryofractured parasites showed mucus and developing eggs within uterine loops. It was demonstrated that the combination of advanced tools generated complementary information, confirming the importance of experimental morphology in parasitology. Therefore, the knowledge of the adult structural organization of E. coelomaticum was improved and this work has contributed to propose new morphological criteria to evaluate the effects of antiparasitic drugs on flukes of medical and veterinary importance.

Predicting the effectiveness of different mulching techniques in reducing post-fire runoff and erosion at plot scale with the RUSLE, MMF and PESERA models.

Wildfires have become a recurrent threat for many Mediterranean forest ecosystems. The characteristics of the Mediterranean climate, with its warm and dry summers and mild and wet winters, make this a region prone to wildfire occurrence as well as to post-fire soil erosion. This threat is expected to be aggravated in the future due to climate change and land management practices and planning. The wide recognition of wildfires as a driver for runoff and erosion in burnt forest areas has created a strong demand for model-based tools for predicting the post-fire hydrological and erosion response and, in particular, for predicting the effectiveness of post-fire management operations to mitigate these responses. In this study, the effectiveness of two post-fire treatments (hydromulch and natural pine needle mulch) in reducing post-fire runoff and soil erosion was evaluated against control conditions (i.e. untreated conditions), at different spatial scales. The main objective of this study was to use field data to evaluate the ability of different erosion models: (i) empirical (RUSLE), (ii) semi-empirical (MMF), and (iii) physically-based (PESERA), to predict the hydrological and erosive response as well as the effectiveness of different mulching techniques in fire-affected areas. The results of this study showed that all three models were reasonably able to reproduce the hydrological and erosive processes occurring in burned forest areas. In addition, it was demonstrated that the models can be calibrated at a small spatial scale (0.5 m2) but provide accurate results at greater spatial scales (10 m2). From this work, the RUSLE model seems to be ideal for fast and simple applications (i.e. prioritization of areas-at-risk) mainly due to its simplicity and reduced data requirements. On the other hand, the more complex MMF and PESERA models would be valuable as a base of a possible tool for assessing the risk of water contamination in fire-affected water bodies and for testing different land management scenarios.

Epidemiology of subclinical hypocalcemia in early-lactation Holstein dairy cows: The temporal associations of plasma calcium concentration in the first 4 days in milk with disease and milk production.

The objectives of this study were to characterize the epidemiology of subclinical hypocalcemia (SCH) in Holstein dairy cows by assessing the temporal associations of plasma Ca concentrations in the first 4 d in milk (DIM) with the risk of cows being diagnosed with metritis or displaced abomasum (or both), and milk production across the first 15 wk of lactation. A prospective cohort study was conducted in 2 dairy herds in New York State, in which cows had a blood sample collected daily for the first 4 DIM. A total of 396 Holstein cows (137 primiparous and 259 multiparous) were enrolled. Multivariable Poisson regression models were built to evaluate the associations of plasma Ca concentration at each of the 4 d following parturition with the risk of primiparous cows being diagnosed with metritis and multiparous cows being diagnosed with metritis, displaced abomasum, or both. Similarly, generalized linear mixed models were built to evaluate the associations of plasma Ca concentration with milk production across the first 15 wk of lactation. Plasma Ca concentration was assessed on a continuous scale in all models; dichotomization and SCH classification only occurred in the final models if the Ca concentration variable was meaningful by creating an optimized threshold based on receiver operating characteristic curve analysis. Plasma Ca concentration assessed at 1 DIM was not associated with the risk of metritis in primiparous cows, but an association was observed at 2, 3, and 4 DIM (critical thresholds were plasma Ca concentration ≤2.15, 2.10, and 2.15 mmol/L, respectively). Plasma Ca concentration was associated with the risk of metritis or displaced abomasum diagnosis (or both) for 2nd parity animals at 2 DIM (threshold ≤1.97 mmol/L), and at 4 DIM for 3rd and greater lactations (threshold ≤2.20 mmol/L). Reduced plasma Ca concentration was associated with higher milk production when assessed at 1 DIM in primiparous and multiparous cows, and lower milk production when assessed at 4 DIM in multiparous cows only. For primiparous cows, plasma Ca concentration was not associated with lower milk production at any of the DIM assessed. In conclusion, assessments of SCH at the individual cow level must take into account the DIM of Ca concentration measurement and parity of the cow, as the epidemiology of the disorder was demonstrated to be highly dependent on these variables. This study advances the knowledge of the epidemiology of SCH and better establishes thresholds for optimizing SCH diagnosis.

Method comparison and validation of a prototype device for measurement of ionized calcium concentrations cow-side against a point-of-care instrument and a benchtop blood-gas analyzer reference method.

The objective of this study was to assess an optimized ion-selective electrode Ca-module prototype as a potential cow-side device for ionized Ca (iCa) measurements in bovine blood. A linearity experiment showed no deviation from linearity over a range of iCa concentrations compared with a commercial point-of-care (POC) device commonly used in the field (POCVS; VetScan i-STAT, Abaxis North America, Union City, CA) and a laboratory gold standard benchtop blood-gas analyzer [reference analyzer (RA); ABL-800 FLEX, Radiometer Medical, Copenhagen, Denmark]. Coefficient of variation on 3 samples with high, within-range, and low iCa concentrations ranged from 1.0 to 3.9% for the prototype. A follow-up validation experiment was performed, in which our objectives were to (1) assess the performance of the prototype cow-side against the POCVS (farm gold-standard) using fresh non-anticoagulated whole-blood samples; (2) assess the performance of the prototype and the POCVS against the RA in a diagnostic laboratory using blood collected in a heparin-balanced syringe; and (3) assess the agreement of the prototype and POCVS on-farm (fresh non-anticoagulated whole blood) against the RA on heparin-balanced blood. Finally, sensitivity and specificity of the results obtained by the prototype and the POCVS cow-side compared with the results obtained by the laboratory RA using 3 different iCa cut points for classification of subclinical hypocalcemia were calculated. A total of 101 periparturient Holstein cows from 3 dairy farms in New York State were used for the second experiment. Ionized Ca results from the prototype cow-side were, on average, 0.06 mmol/L higher than the POCVS. With heparin-balanced samples under laboratory conditions, the prototype and POCVS measured an average 0.04 mmol/L higher and lower, respectively, compared with the RA. Results from the prototype and POCVS cow-side were 0.01 mmol/L higher and 0.05 mmol/L lower, respectively, compared with results from the laboratory RA on heparinized blood. Sensitivity and specificity for the prototype and the POCVS under farm conditions at 3 potential subclinical hypocalcemia cut points were 100 and ≥93.5%, respectively. This novel ion-selective electrode Ca-module could become a rapid low-cost tool for assessing iCa cow-side, while qualitatively allowing classification of subclinical hypocalcemia on-farm.

Association of immediate postpartum plasma calcium concentration with early-lactation clinical diseases, culling, reproduction, and milk production in Holstein cows.

The objective of this study was to evaluate the association of postpartum plasma Ca concentration with early-lactation disease outcomes, culling within 60 d in milk, pregnancy to first service, and milk production. A total of 1,453 cows from 5 commercial dairy farms in New York State were enrolled in a prospective cohort study from February to November 2015. Blood samples were collected within 12 h of parturition, and plasma was submitted to a diagnostic laboratory for total Ca measurement. Early-lactation disease, reproductive performance, and milk production from Dairy Herd Improvement Association (DHIA) test-day data were compiled from each farm's management software. Multivariable Poisson regression models were built to evaluate the association of plasma Ca with the risks of retained placenta (RP), metritis, displaced abomasum (DA), clinical mastitis, culling within 60 d in milk, and pregnancy to first service. Repeated-measures ANOVA were used to evaluate the association of Ca at parturition with milk production across the first 9 DHIA tests. Herd was considered a random effect in all models. Primiparous cows were modeled separately from multiparous cows if differential responses were observed. Calcium was not associated with the risk of RP, metritis, clinical mastitis, or pregnancy to first service in primiparous or multiparous cows. For multiparous cows only, higher Ca concentration tended to be associated with increased culling within the first 60 d in milk. Multiparous cows with Ca ≤1.85 mmol/L had an increased risk of being diagnosed with a DA compared with cows with Ca >1.85 mmol/L. For the milk production models, Ca was not associated with the amount of milk produced within the first 9 DHIA tests in primiparous cows; however, multiparous cows with Ca ≤1.95 mmol/L produced, on average, 1.1 kg more milk per day across the 9 DHIA tests than their multiparous counterparts with Ca >1.95 mmol/L. Our results indicate that plasma Ca concentration measured within 12 h of parturition is a poor predictor of early-lactation health outcomes. Reduced Ca concentration in the immediate postpartum period was associated with higher milk production in multiparous cows. From these results, we caution that studies attempting to categorize subclinical hypocalcemia based on a single sample in the immediate postpartum period could misclassify the disorder.

Differential effects of a single dose of oral calcium based on postpartum plasma calcium concentration in Holstein cows.

Our objectives were to determine (1) the effect of a single dose of an oral Ca bolus within 24 h after parturition on plasma Ca concentration, (2) the response of primiparous (PP) and multiparous (MP) cows to this supplementation strategy, and (3) differential responses based on plasma Ca at enrollment. For objective 1, cows from 1 commercial dairy in New York State were enrolled within 19 h after parturition (mean ± standard deviation = 8.3 ± 5.3 h) and randomized within parity group (first, second, and ≥third) to control [CON (n = 25); no placebo] or a single dose bolus treatment [BOL (n = 25); 3 oral Ca boluses supplying 54 to 64 g of Ca]. Plasma Ca was measured repeatedly between 1 and 24 h following treatment. For objectives 2 and 3, cows on 6 commercial farms in New York State were assigned to treatment as for objective 1 (CON, n = 1,973; BOL, n = 1,976). Herd records for health, reproduction, and Dairy Herd Improvement Association test day milk production were collected. Mixed effect multivariable models were developed using repeated measures ANOVA, Poisson regression, or proportional hazard models. Objective 2 analyses considered treatment with periparturient risk factors, whereas objective 3 analyses also considered Ca status. No difference was observed for plasma Ca between 1 and 24 h after treatment. Primiparous cows assigned to BOL calving at >712 d old had decreased risk of one or more health disorders [≤30 d in milk; risk ratio (RR) = 0.65, 95% confidence interval (CI) = 0.51 to 0.84] and those with body condition score >3.5 responded to BOL with increased milk production (CON = 31.7 ± 1.1, BOL = 35.1 ± 1.1 kg/d), as did those with days carried calf >277 (CON = 31.9 ± 1.0, BOL = 34.7 ± 1.0 kg/d). Reduced risk of one or more health disorders was observed in parity ≥3 (RR = 0.85, 95% CI = 0.81 to 0.89) and MP cows with body condition score >3.5 (retained placenta; RR = 0.70, 95% CI = 0.58 to 0.84) or that were lame (displaced abomasum; RR = 0.49, 95% CI = 0.32 to 0.75). Differential responses for PP cows by Ca status were minimal. For MP cows with low plasma Ca, BOL decreased risk of additional Ca treatment (≤1.8 mmol/L; RR = 0.57, 95% CI = 0.40 to 0.80) as well as risk of one or more health disorders (≤2.15 mmol/L; RR = 0.90, 95% CI = 0.85 to 0.95). Supplementation with a single oral dose of Ca could be targeted to periparturient risk groups for improved health. Calcium status did not differentiate responses of PP cows, but MP cows with low Ca at parturition had improved health status when supplemented.

Factors Associated with Contralateral Deep Venous Thrombosis after Iliocaval Venous Stenting.

BACKGROUND: The majority of iliac venous obstructions occur on the left side, and endovascular therapy has become the first line treatment for this condition. A left common iliac venous stent will protrude into the inferior vena cava (IVC) to some extent, thereby covering the contralateral common iliac vein (CIV) outflow. This may increase the risk of thrombosis of the contralateral iliac vein. The aim of this paper was to determine the rate of, and factors associated with, contralateral lower limb venous thrombosis after stenting, and to evaluate the results of salvage revascularisation. METHODS: A total of 376 patients (102 from UCH, Galway, Ireland, 2008-16, and 274 from, CHU Nord, Marseille, France, 2000-15) with symptomatic acute or chronic left iliocaval venous obstruction were retrospectively evaluated. Either duplex ultrasound scanning (DUS) or computed tomographic venography (CTV) was used for pre- and post-operative imaging. Data were collected from the PACS system (IMPAX, Agfa, BE) of the Radiology Department, UCH, Galway, and from the electronic medical records of Vascular Surgery department, CHU Nord, Marseille. RESULTS: The median age of stented patients was 46 (range 15-86 years), 80% were female (301/376). Following left CIV stent placement, 10 patients later presented with a right (contralateral) iliac deep venous thrombosis (DVT) resulting in a cumulative incidence of contralateral DVT of 4% according to Kaplan-Meier analysis. Acute DVT (p=.001), non-compliance with the prescribed 6 months anticoagulation (p = 0.05), pre-operative contralateral internal iliac vein (IIV) thrombosis (p = 0.001), and pre-existing IVC filter placement (p = 0.003) were all statistically significantly associated with contralateral DVT. All patients with symptomatic contralateral iliac DVT underwent clot removal in the acute phase. The primary patency of these limbs was 100% at 3 years. CONCLUSION: Stent placement across the iliocaval confluence from the left CIV is associated with a low but definite rate of contralateral iliac vein thrombosis. Acute DVT, pre-operative contralateral IIV thrombosis, pre-existing IVC filters, and anticoagulation non-compliance are significant risk factors.

Risk factors associated with postpartum subclinical hypocalcemia in dairy cows.

Our study objectives were to evaluate the association of prepartum plasma Mg concentrations with subclinical hypocalcemia (SCH) classification at parturition and to evaluate the association of other cow-level risk factors with SCH classification at calving or at 2 d in milk (DIM). A total of 301 animals from 2 dairy herds located in New York were enrolled in a cohort study. Blood samples were collected at approximately 1 wk before the expected calving date, within 4 h of calving, and at 2 DIM. Prepartum samples had plasma macromineral concentrations (Ca, K, Mg, P), albumin, and ß-hydroxybutyrate analyzed. Samples collected at calving were analyzed for Ca only, and samples from 2 DIM had macromineral and albumin concentrations determined. Postpartum SCH was defined as Ca concentrations ≤2.1 mmol/L. The prevalence of SCH at calving was 2, 40, and 66% for first, second, and third or greater parities, respectively. Only 4% of cows could be classified with prepartum subclinical hypomagnesemia (Mg concentrations <0.8 mmol/L), which did not provide enough power to appropriately determine the association of plasma Mg with postpartum Ca concentrations and its effect on SCH classification. Multiparous cows with Ca concentrations ≤2.4 mmol/L in the prepartum period and third or greater parity cows had a higher risk of being categorized as SCH at calving [relative risk (RR) = 1.4 and 1.7, respectively]. The risk of SCH at 2 DIM was associated with the interaction of Ca status at calving and lameness score. Nonlame cows with Ca concentrations ≤2.1 mmol/L (RR = 3.2) and normocalcemic lame cows at parturition (RR = 3.4) were more likely to be SCH at 2 DIM compared with nonlame normocalcemic cows. In conclusion, we identified a prepartum Ca cut-point for identification of cows that are more likely to be classified as SCH at calving. Different risk factors were associated with SCH depending on the timing of diagnosis relative to parturition.

A novel microfluidic platform for size and deformability based separation and the subsequent molecular characterization of viable circulating tumor cells.

Circulating tumor cells (CTCs) were introduced as biomarkers more than 10 years ago, but capture of viable CTCs at high purity from peripheral blood of cancer patients is still a major technical challenge. Here, we report a novel microfluidic platform designed for marker independent capture of CTCs. The Parsortix™ cell separation system provides size and deformability-based enrichment with automated staining for cell identification, and subsequent recovery (harvesting) of cells from the device. Using the Parsortix™ system, average cell capture inside the device ranged between 42% and 70%. Subsequent harvest of cells from the device ranged between 54% and 69% of cells captured. Most importantly, 99% of the isolated tumor cells were viable after processing in spiking experiments as well as after harvesting from patient samples and still functional for downstream molecular analysis as demonstrated by mRNA characterization and array-based comparative genomic hybridization. Analyzing clinical blood samples from metastatic (n = 20) and nonmetastatic (n = 6) cancer patients in parallel with CellSearch(®) system, we found that there was no statistically significant difference between the quantitative behavior of the two systems in this set of twenty six paired separations. In conclusion, the epitope independent Parsortix™ system enables the isolation of viable CTCs at a very high purity. Using this system, viable tumor cells are easily accessible and ready for molecular and functional analysis. The system's ability for enumeration and molecular characterization of EpCAM-negative CTCs will help to broaden research into the mechanisms of cancer as well as facilitating the use of CTCs as "liquid biopsies."