Trends in childhood body mass index between 1936 and 2011 showed that underweight remained more common than obesity among 398 970 Danish school children.

AIM: To examine trends in all body mass index (BMI) groups in children from 1936 to 2011. METHODS: We included 197 694 girls and 201 276 boys from the Copenhagen School Health Records Register, born between 1930 and 1996, with longitudinal weight and height measurements (6-14 years). Using International Obesity Task Force criteria, BMI was classified as underweight, normal-weight, overweight and obesity. Sex- and age-specific prevalences were calculated. RESULTS: From the 1930s, the prevalence of underweight was stable until a small increase occurred from 1950 to 1970s, and thereafter it declined into the early 2000s. Using 7-year-olds as an example, underweight changed from 10% to 7% in girls and from 9% to 6% in boys during the study period. The prevalence of overweight plateaued from 1950 to 1970s and then steeply increased from 1970s onwards and in 1990-2000s 15% girls and 11% boys at 7 years had overweight. The prevalence of obesity particularly increased from 1980s onwards and in 1990-2000s 5% girls and 4% boys at 7 years had obesity. These trends slightly differed by age. CONCLUSION: Among Danish schoolchildren, the prevalence of underweight was greater than overweight until the 1980s and greater than obesity throughout the period. Thus, monitoring the prevalence of childhood underweight remains an important public health issue.

An olivine cumulate outcrop on the floor of Jezero crater, Mars.

The geological units on the floor of Jezero crater, Mars, are part of a wider regional stratigraphy of olivine-rich rocks, which extends well beyond the crater. We investigated the petrology of olivine and carbonate-bearing rocks of the Séítah formation in the floor of Jezero. Using multispectral images and x-ray fluorescence data, acquired by the Perseverance rover, we performed a petrographic analysis of the Bastide and Brac outcrops within this unit. We found that these outcrops are composed of igneous rock, moderately altered by aqueous fluid. The igneous rocks are mainly made of coarse-grained olivine, similar to some martian meteorites. We interpret them as an olivine cumulate, formed by settling and enrichment of olivine through multistage cooling of a thick magma body.

A retrospective observational study of neuromuscular monitoring practice in 30,430 cases from six Danish hospitals.

Timely application of objective neuromuscular monitoring can avoid residual neuromuscular blockade. We assessed the frequency of objective neuromuscular monitoring with acceleromyography and the last recorded train-of-four ratio in a cohort of Danish patients. We extracted data from all patients receiving general anaesthesia from November 2014 to November 2016 at six hospitals in the Zealand Region of Denmark. Acceleromyography was available in all operating rooms and data were recorded automatically. The primary outcome measure was acceleromyography use in patients receiving neuromuscular blocking agents, divided into non-depolarising agents and succinylcholine only. The dataset included 76,743 cases, of which 30,430 received a neuromuscular blocking drug. Non-depolarising drugs were used in 16,525 (54%) and succinylcholine as the sole drug in 13,905 (46%) cases. Acceleromyography was used in 14,463 (88%) patients who received a non-depolarising neuromuscular blocking drug and in 4224 (30%) receiving succinylcholine alone. Acceleromyography use varied between the departments from 58% to 99% for non-depolarising drugs and from 3% to 79% for succinylcholine alone. The median (IQR [range]) of the last recorded train-of-four ratio before tracheal extubation was 0.97 (0.90-1.06 [0.01-2.20]) when non-depolarising drugs were used, and was less than 0.9 in 22% of cases. The OR for oxygen desaturation was higher with the use of succinylcholine [2.51 (95%CI 2.33-2.70) p < 0.001] and non-depolarising drugs [2.57 (95%CI 2.32-2.84) p < 0.001] as compared with cases where no neuromuscular blockade drug was used. In conclusion, acceleromyography was almost always used in cases where non-depolarising neuromuscular blocking drugs were used, but a train-of-four ratio of 0.9 was not always achieved. Monitoring was used in less than 30% of cases where succinylcholine was the sole drug used.

No effect of oral sucrose or IV glucose during exercise in phosphorylase b kinase deficiency.

This case report investigated exercise metabolism and the effect of oral sucrose and intravenous glucose supplementation in a 30-year-old, mildly affected man with muscle phosphorylase b kinase (PHK) deficiency caused by a novel c.586G>A mutation in the PHKA1 gene. Only 12 patients with PHK deficiency have been reported and it is unclear to what extent patients exhibit symptoms during exercise. Carbohydrate and fat metabolism were measured during 30 min of exercise at ∼ 70% of peak oxidative capacity using stabile isotope technique and signaling proteins and enzymes in the energy pathway were analyzed by Western blot. Results were compared to four healthy subjects. These studies show that neither oral nor intravenous glucose improved exercise tolerance in this patient with PHK deficiency. Despite Western blots indicated affected metabolism on protein level, systemic substrate turnover studies showed that carbohydrate and fatty acid oxidations were normal.

A novel non-invasive method for measuring knee joint laxity in four dof: In vitro proof-of-concept and validation.

Knee joint laxity or instability is a common problem that may have detrimental consequences for patients. Unfortunately, assessment of knee joint laxity is limited by current methodologies resulting in suboptimal diagnostics and treatment. This paper presents a novel method for accurately measuring non-invasive knee joint laxity in four degrees-of-freedom (DOF). An arthrometer, combining a parallel manipulator and a six-axis force/moment sensor, was developed to be used in combination with a low-dose biplanar x-ray system and 3D image data to reconstruct tibiofemoral position and orientation of laxity measurements. As proof-of-concept, four cadaveric knees were tested in the device. Each cadaveric knee was mounted in the device at approximately 30° of flexion and twelve monoplanar anteroposterior, mediolateral and internal/external load cases were applied. Additionally, four biplanar load cases were applied, consisting of different combinations of anteroposterior and internal/external loads. The arthrometer was limited to four DOF to address the specific measurements. For validation purposes, the pose reconstructions of tibia and femur were compared with pose reconstructions of bone pin marker frames mounted on each bone. The measurements from the arthrometer in terms of translation and rotations displayed comparable values to what have previously been presented in the literature. Furthermore, the measurements revealed coupled motions in multiple planes, demonstrating the importance of multi DOF laxity measurements. The validation displayed an average mean difference for translations of 0.08 mm and an average limit of agreement between -1.64 mm and 1.80 mm. The average mean difference for rotations was 0.10° and the limit of agreement was between -0.85° and 1.05°. The presented method eliminates several limitations present in current methods and may prove a valuable tool for assessing knee joint laxity.

The impact of luteal serum progesterone levels on live birth rates-a prospective study of 602 IVF/ICSI cycles.

STUDY QUESTION: Is the chance of a live birth following IVF treatment and fresh embryo transfer affected by early and mid-luteal serum progesterone (P4) levels? SUMMARY ANSWER: Low as well as high serum P4 levels in the early and mid-luteal phase reduce the chance of a live birth following IVF treatment with fresh embryo transfer. WHAT IS KNOWN ALREADY: Data from non-human studies and studies of frozen-thawed embryo transfer cycles indicate that low as well as high P4 levels during the mid-luteal phase decrease the chance of pregnancy. The altered P4 pattern may disrupt the endometrial maturation leading to asynchrony between embryonic development and endometrial receptivity, thereby, compromising implantation and early development of pregnancy. STUDY DESIGN, SIZE, DURATION: Prospective multicenter cohort study of 602 women undergoing IVF treatment. Patients were recruited from four Danish public Fertility Centers from May 2014 to June 2017. The study population was unselected, thus, representing a normal everyday patient cohort. Patients were treated in a long GnRH-agonist protocol or a GnRH-antagonist protocol and triggered for final oocyte maturation with either hCG or a GnRH-agonist. The same vaginal luteal support regimen was applied in all patients. PARTICIPANTS/MATERIALS, SETTING, METHODS: Serum P4 levels from the early or mid-luteal phase were correlated to positive hCG and live birth rates (delivery > gestational week 20). Patients were divided into four P4 groups based on raw data of P4 serum levels and reproductive outcomes during early luteal phase (P4<60 nmol/l, P4 60-100 nmol/l, P4 101-400 nmol/l and P4>400 nmol/l) and during mid-luteal phase (P4<150 nmol/l, P4 150-250 nmol/l, P4 251-400 nmol/l and P4>400 nmol/l). MAIN RESULTS AND THE ROLE OF CHANCE: The optimal chance of pregnancy was achieved with serum P4 levels of 60-100 nmol/l in the early luteal phase whereas the optimal P4 level during the mid-luteal phase was 150-250 nmol/l. Below, but most distinctly above these levels, the chance of pregnancy was consistently reduced. With an early luteal P4 level of 60-100 nmol/l, the chance of a positive hCG-test was 73%, 95% CI: [59, 84] following cleavage stage embryo transfer. In contrast, with P4 levels >400 nmol/l, the chance of a positive hCG-test was significantly reduced to 35%, 95% CI: [17, 57], thus, an absolute risk difference of -38%, P = 0.01. A similar negative association between early luteal P4 and live birth rate was found, although it did not reach statistical significance. During the mid-luteal phase, a P4 level of 150-250 nmol/l resulted in an optimal chance of live birth: 54%, 95% CI: [37, 70] compared to 38%, 95% CI: [20, 60] with a P4 level >400 nmol/l, thus, an absolute risk difference of -16%, P = 0.14. All estimates were adjusted for maternal age, maternal BMI, study site, final follicle count and late follicular P4 levels. LIMITATIONS, REASONS FOR CAUTION: This study is the first to explore the possible upper and lower thresholds for luteal P4 following IVF treatment and fresh embryo transfer, and the optimal P4 ranges found in this study should be corroborated in future clinical trials. Furthermore, the P4 thresholds in this study only apply to fresh IVF cycles, using vaginal luteal phase support, as the optimal P4 level in cycles using intramuscular P4 may be different. WIDER IMPLICATIONS OF THE FINDINGS: Future studies are necessary to explore whether additional exogenous luteal P4 supplementation in the low P4 group could increase the chance of a live birth following fresh embryo transfer, and whether patients with luteal P4 levels >400 nmol/l would benefit from segmentation followed by subsequent transfer in frozen/thawed cycles. TRIAL REGISTRATION NUMBER: NCT02129998 (Clinicaltrials.gov). STUDY FUNDING/COMPETING INTEREST(S): L.H.T. received an unrestricted grant from Ferring Pharmaceuticals, Denmark, to support this study. P.H. received unrestricted research grants from MSD, Merck, Gedeon Richter and Ferring Pharmaceuticals outside of this work as well as honoraria for lectures from MSD, Merck and Gedeon Richter outside of this work. U.K. received honoraria for lectures from MSD and Ferring Pharmaceuticals outside of this work. C.A. received unrestricted research grants from MSD, IBSA, and Ferring Pharmaceuticals outside of this work as well as honoraria for lectures from MSD and IBSA. H.O.E. and B.B.P. received an unrestricted research grant from Gedeon Richter outside of this work. K.E., L.B., D.P. and B.H. have no conflict of interest. Furthermore, grants from 'The Health Research Fund of Central Denmark Region', 'The Research Foundation of the Hospital of Central Jutland', 'The Research Foundation of A.P. Møller', 'The Research Foundation of Aase & Ejnar Danielsen', 'The Research Foundation of Dagmar Marshall', 'The Research Foundation of Dir. Jacob Madsen & Hustru Olga Madsen', 'The Research Foundation of Fam. Hede Nielsen' and 'The Danish Medical Research Grant' supported conducting this study. The providers of funding were neither involved in the conduction of the study nor in the writing of the scientific report.

Sensory distribution of the lateral femoral cutaneous nerve block - a randomised, blinded trial.

BACKGROUND: The lateral femoral cutaneous nerve (LFCN) block may be used for post-operative pain management in patients undergoing total hip arthroplasty. The aim of this trial was to investigate the sensory coverage of the posterior and the lateral incision lines and the involvement of the femoral nerve after an LFCN block. METHODS: The study was a randomised, blinded trial in 20 healthy volunteers. All subjects received a bilateral LFCN block randomised to 8 ml ropivacaine on the right side and 8 ml isotonic saline on the left side, or vice versa. An orthopaedic surgeon depicted the incision lines (invisible to the investigators) prior to block performance. The distribution of the blocked area and the coverage of the incision lines were assessed with temperature discrimination and pinprick test before unblinding the incision lines. Pain during tonic heat stimulation and involvement of the femoral nerve by measuring quadriceps strength were assessed. RESULTS: The mean difference in block coverage of the posterior (primary outcome) and the lateral incision lines tested with temperature discrimination were 5.8% (95% CI: -2.2 to 14.0%, P = 0.146) and 18.9% (95% CI: 6.5-31.4%, P = 0.005), respectively, comparing the active with the placebo side. A varying anatomic distribution area was observed. No clinically significant differences for experimental pain and quadriceps muscle strength were found. The block failure rate was 15%. CONCLUSION: An LFCN block consisting of 8 ml 0.75% ropivacaine had limited coverage of the posterior and lateral incision lines.

Gamma-Hydroxybutyrate (GHB) Content in Hair Samples Correlates Negatively with Age in Succinic Semialdehyde Dehydrogenase Deficiency.

Gamma-hydroxybutyrate (GHB) is a drug of abuse, an approved therapeutic for narcolepsy, an agent employed for facilitation of sexual assault, as well as a biomarker of succinic semialdehyde dehydrogenase deficiency (SSADHD). Our laboratory seeks to identify surrogate biomarkers in SSADHD that can shed light on the developmental course of this neurometabolic disease. Since GHB may be quantified in hair as a potential surrogate to identify victims of drug-related assault, we have opted to examine its level in SSADHD. We quantified GHB in hair derived from ten patients with SSADHD, and documented a significant negative age correlation. These findings are consistent with recent results in patient biological fluids, including plasma and red blood cells. These findings may provide additional insight into the developmental course of SSADHD (Jansen et al., J Inherit Metab Dis 39:795-800, 2016).

Lateral femoral cutaneous nerve block after total hip arthroplasty: a randomised trial.

BACKGROUND: Peripheral regional nerve blocks are commonly used for pain management after lower extremity surgery, but motor blockade can be a significant concern. The lateral femoral cutaneous nerve (LFCN) is a purely sensory nerve from the lumbar plexus. We hypothesised that an LFCN block would reduce movement-related pain after total hip arthroplasty (THA) in patients with moderate-to-severe pain. METHODS: Sixty patients with visual analogue scale (VAS) score > 40 mm during 30-degree active flexion of the hip on either the first or second postoperative day after THA were included in this prospective, randomised, blinded, placebo-controlled trial. Group A received an LFCN block with 8 ml of 0.75% ropivacaine followed after 45 min by an additional LFCN block with 8 ml of saline. Group B received an LFCN block with 8 ml of saline followed after 45 min by an additional LFCN block with 8 ml of 0.75% ropivacaine. RESULTS: We found a difference of 17 mm (95% CI, 4-31 mm; P < 0.02) in VAS pain score during 30-degree flexion of the hip 45 min after the first block (primary outcome) in favour of group A. No other significant difference between groups regarding pain during mobilisation and at rest was found. The overall non-responder rate (< 15 mm pain reduction) was 42%. CONCLUSIONS: LFCN block reduced movement-related pain in patients with moderate-to-severe pain after THA. The substantial non-responder rate limits recommendations of this block as part of a standard analgesic treatment regimen.

IGF-I at 9 and 36 months of age ­ relations with body composition and diet at 3 years ­ the SKOT cohort.

OBJECTIVE: High infancy levels of insulin-like growth factor-I (IGF-I) have been associated with increased linear growth and fat-free mass (FFM) but also with risk of obesity. This paper examines how IGF-I at 9 and 36 months relates to diet and body composition. DESIGN: Healthy term infants from the prospective cohort study, SKOT, were examined at 9 and 36 months with anthropometry, bioelectrical impedance (36 months), 7-day food records and blood analysis of IGF-I and IGFBP-3 by chemiluminescent immunometric assay. RESULTS: IGF-I at 36 months (n = 229) was positively correlated with 9 months values and values were considerably higher in girls (43%). Children breastfed at 9 months had lower IGF-I concentrations at 9 months but reached the same IGF-I concentrations at 36 months as infants not breastfed at 9 months. IGF-I at 36 months was positively associated with height, weight, BMI, predicted FFM and FFM index (FFM/height (kg/m2)). Although there also was a positive association with predicted fat mass (FM) there was no association with FM index (FM/height (kg/m2)). Further, a negative association with skin fold thickness was observed. A change in IGF-I from 9­36 months was positively related to FFM and FFM index but not BMI, FM and FM index. No associations were seen between IGF-I and current intake of milk, meat or protein energy percentage, but both fat and saturated fat energy percentage were negatively associated with IGF-I. CONCLUSION: IGF-I concentrations were positively associated with growth but not with adiposity at this age. However, the higher tempo of growth may influence age at adiposity rebound and thereby later risk of obesity. Milk and protein intake at 36 months did not influence IGF-I but there was a negative association with intake of fat and saturated fat. The implications of this finding for development of obesity need further exploration.

Insulin signalling mechanisms for triacylglycerol storage.

Insulin signalling is uniquely required for storing energy as fat in humans. While de novo synthesis of fatty acids and triacylglycerol occurs mostly in liver, adipose tissue is the primary site for triacylglycerol storage. Insulin signalling mechanisms in adipose tissue that stimulate hydrolysis of circulating triacylglycerol, uptake of the released fatty acids and their conversion to triacylglycerol are poorly understood. New findings include (1) activation of DNA-dependent protein kinase to stimulate upstream stimulatory factor (USF)1/USF2 heterodimers, enhancing the lipogenic transcription factor sterol regulatory element binding protein 1c (SREBP1c); (2) stimulation of fatty acid synthase through AMP kinase modulation; (3) mobilisation of lipid droplet proteins to promote retention of triacylglycerol; and (4) upregulation of a novel carbohydrate response element binding protein ß isoform that potently stimulates transcription of lipogenic enzymes. Additionally, insulin signalling through mammalian target of rapamycin to activate transcription and processing of SREBP1c described in liver may apply to adipose tissue. Paradoxically, insulin resistance in obesity and type 2 diabetes is associated with increased triacylglycerol synthesis in liver, while it is decreased in adipose tissue. This and other mysteries about insulin signalling and insulin resistance in adipose tissue make this topic especially fertile for future research.

Effect of exogenous phytase on degradation of inositol phosphate in dairy cows.

The effect of exogenous phytase on inositol phosphate degradation in the rumen of dairy cows was investigated in a 4 × 4 Latin square design. Four lactating Danish Holstein cows fitted with ruminal, duodenal, and ileal cannulas were offered a total mixed ration (TMR) with a high content of inositol phosphate and supplemented with 1 of 4 concentrations of phytase [none, low, medium, or high, corresponding to 23, 2,023, 3,982, and 6,015 phytase units/kg of dry matter (DM)]. Exogenous phytase lead to a higher rumen pool of phytase. Inositol phosphate content in digesta samples from rumen, duodenum, ileum, and feces was almost entirely composed of myo-inositol hexakisphosphate (InsP(6)), indicating that degradation of this compound is the rate-limiting step in inositol phosphate degradation in the digestive tract. Ruminal and total-tract degradations of InsP(6) were higher when exogenous phytase was added to the TMR. Degradation of InsP(6) occurred mainly before the duodenum. The ruminal degradability of InsP(6) was increased with increasing dietary concentrations of phytase: 86.4, 93.7, 94.5, and 96.3% for none, low, medium, or high, respectively. A comparison of the InsP(6) content in individual feedstuffs and in samples of the TMR revealed that the exogenous phytase started degrading the inositol phosphate when feeds and phytase were mixed, and thus the InsP(6) phosphorus (InsP(6)-P) content in the TMR was found to decrease with higher doses of phytase (1.69, 1.51, 1.39, and 1.25 g/kg of DM for the none, low, medium, and high phytase doses, respectively). It was not possible to distinguish between the degradation of inositol phosphate occurring in the TMR and in the rumen. Exogenous phytase had no effect on total P intake or flow of total P to the duodenum and ileum, whereas exogenous phytase increased flow of microbial P to the duodenum and total fecal P excretion. None of the investigated rumen variables (pH, degradability of neutral detergent fiber, and rumen kinetics for neutral detergent fiber) were affected by treatment. Rumen and total-tract degradations of inositol phosphate were increased when exogenous phytase was added to the TMR, which offers the potential for reducing P excretion through reduced dietary P.

Milk fatty acid composition and production performance of Danish Holstein and Danish Jersey cows fed different amounts of linseed and rapeseed.

Fat supplements are used in diets for dairy cows to increase energy intake and milk production and the fatty acid composition of the feed affects milk fatty acid composition. A total of 74 Danish Holstein and 41 Danish Jersey cows were divided into 4 groups and the cows within each group were fed a mixed ration supplemented with 0, 3.5, 6.8, or 10.2% of dry matter of a linseed:rapeseed (1:3) mixture during lactation wk 6 to 30. Milk yield, fat, and lactose contents were not affected by treatments for Danish Holsteins, whereas these parameters increased when increased amounts of oilseeds were fed to Danish Jerseys. For both breeds, milk protein content decreased when increased amounts of oilseeds were fed. The milk fatty acid composition showed higher concentrations of saturated fatty acids and lower concentrations of unsaturated fatty acids in milk fat from Danish Jerseys compared with Danish Holsteins. Increased amounts of oilseeds in feed increased milk fat concentration of all C18 fatty acids except C18:2 n-6, whereas the content of C6 to C14, C11 to C17, and in particular, C16, decreased. This effect was more pronounced for Danish Holsteins than for Danish Jerseys. The apparent recovery of C18:2 n-6 and C18:3 n-3 decreased when increased amounts of oilseeds were fed; however, this was most likely due to increased amounts of fatty acid from feed used for other energy demands than milk production. It was concluded that up to 6.8% of oilseed supplementation can be fed without production problems and, in many cases, with positive production responses, including an improved milk fatty acid profile.

Repeated measurement of mechanical properties in viable osteochondral explants following a single blunt impact injury.

The objective of this work was to develop a method for repeated same-site measurement of mechanical properties suitable for the detection of degenerative changes in a biologically active explant model after a single blunt impact injury. Focal blunt impact injuries to articular surfaces lead to local cartilage degeneration and loss of mechanical properties. We employed a repeated measurement methodology to determine variations in mechanical same-site properties before and after injury in living cartilage, with the hypothesis that normalization with initial mechanical properties may provide a clearer evaluation of impact effects and improve our understanding of the biologic responses to impact injury. Bovine osteochondral explants were cultured for up to 14 days after impact injury. Indentation tests were performed before and after impact injury to assess relative changes in mechanical properties. Creep strain increased significantly in impacted explants after 7 days and in both impacted and control explants after 14 days. Further analysis at 14 days revealed decreases in stretch factor beta, creep time constant and local compressive modulus. A repeated measures methodology reliably detected changes in the mechanical behaviour of viable osteochondral explants after a single impact injury.

Time-dependent effects of Prkce deletion on glucose homeostasis and hepatic lipid metabolism on dietary lipid oversupply in mice.

AIMS/HYPOTHESIS: We examined the time-dependent effects of deletion of the gene encoding protein kinase C epsilon (Prkce) on glucose homeostasis, insulin secretion and hepatic lipid metabolism in fat-fed mice. METHODS: Prkce(-/-) and wild-type (WT) mice were fed a high-fat diet for 1 to 16 weeks and subjected to i.p. glucose tolerance tests (ipGTT) and indirect calorimetry. We also investigated gene expression and protein levels by RT-PCR, quantitative protein profiling (isobaric tag for relative and absolute quantification; iTRAQ) and immunoblotting. Lipid levels, mitochondrial oxidative capacity and lipid metabolism were assessed in liver and primary hepatocytes. RESULTS: While fat-fed WT mice became glucose intolerant after 1 week, Prkce(-/-) mice exhibited normal glucose and insulin levels. iTRAQ suggested differences in lipid metabolism and oxidative phosphorylation between fat-fed WT and Prkce(-/-) animals. Liver triacylglycerols were increased in fat-fed Prkce(-/-) mice, resulting from altered lipid partitioning which promoted esterification of fatty acids in hepatocytes. In WT mice, fat feeding elevated oxygen consumption in vivo and in isolated liver mitochondria, but these increases were not seen in Prkce(-/-) mice. Prkce(-/-) hepatocytes also exhibited reduced production of reactive oxygen species (ROS) in the presence of palmitate. After 16 weeks of fat feeding, however, the improved glucose tolerance in fat-fed Prkce(-/-) mice was instead associated with increased insulin secretion during ipGTT, as we have previously reported. CONCLUSIONS/INTERPRETATION: Prkce deletion ameliorates diet-induced glucose intolerance via two temporally distinct phenotypes. Protection against insulin resistance is associated with changes in hepatic lipid partitioning, which may reduce the acute inhibitory effects of fatty acid catabolism, such as ROS generation. In the longer term, enhancement of glucose-stimulated insulin secretion prevails.

Effect of exogenous phytase on feed inositol phosphate hydrolysis in an in vitro rumen fluid buffer system.

Three in vitro experiments using a rumen fluid buffer system were performed to investigate the effect of addition of 4 experimental phytases (Phy1, Phy2, Phy3, and Phy4) compared with no addition of phytase on feed inositol phosphate hydrolysis in wheat and rapeseed cake to determine which of the 4 phytases was most suitable under rumen-like conditions. The feedstuffs were incubated with a mixture of physiological buffer, ruminal fluid, and exogenous phytase at pH 6.2, after which the samples were incubated for different periods. Incubations were stopped using HCl, and the samples were analyzed for inositol phosphates via high performance ion chromatography. Addition of phytase (Phy1) resulted in enhanced degradation of myo-inositol hexakisphosphate (InsP(6)) in rapeseed cake, whereas addition of exogenous phytase did not improve the degradation of InsP(6) in wheat. Only rapeseed cake was therefore used subsequently. All 4 phytases increased degradation of InsP(6) in rapeseed cake in the in vitro system, and degradability of InsP(6) increased with higher incubation time and higher phytase dosages, independent of phytase. Addition of 2 units of phytase per gram of substrate of the phytases Phy1, Phy2, Phy3, and Phy4 led to an undegraded InsP(6) content of 56, 49, 70, and 18%, respectively, when incubated with rapeseed cake for 6h, indicating that Phy2 and Phy4 were the most effective phytases. However, Phy2 had a higher specific activity than Phy4, as 60% of the original InsP(6) content was remaining after 3h when 5mg of enzyme protein per gram of substrate of Phy2 was added to rapeseed cake, whereas 150 mg of enzyme protein per gram of substrate of Phy4 was necessary to achieve a similar result. Therefore, Phy2 appeared to be most applicable under rumen-like conditions.

Baseline Sensitivity of Cercospora zeae-maydis to Quinone Outside Inhibitor Fungicides.

Cercospora zeae-maydis, the causal agent of gray leaf spot on corn (Zea mays), can cause severe yield loss in the United States. Quinone outside inhibitor (QoI) fungicides are effective tools that can be used to manage gray leaf spot, and their use has increased in corn production in the United States. In total, 61 C. zeae-maydis isolates collected from fields in which QoI fungicides had never been applied were tested in vitro using azoxystrobin-, pyraclostrobin-, or trifloxystrobin-amended medium to determine the effective fungicide concentration at which 50% of the conidial germination was inhibited (EC50). The effect of salicylhydroxamic acid (SHAM) also was evaluated for seven isolates to determine whether C. zeae-maydis is capable of using alternative respiration in azoxystrobin-amended medium. All seven C. zeae-maydis isolates tested had significantly greater (P < 0.02) EC50 values when SHAM was not included in medium amended with azoxystrobin, indicating that C. zeae-maydis has the potential to utilize alternative respiration to overcome QoI fungicide inhibition in vitro. Baseline EC50 values of azoxystrobin ranged from 0.003 to 0.031 µg/ml, with mean and median values of 0.018 and 0.019 µg/ml, respectively. Baseline EC50 values of pyraclostrobin ranged from 0.0003 to 0.0025 µg/ml, with mean and median values of 0.0010 and 0.0010 µg/ml, respectively. Baseline EC50 values of trifloxystrobin ranged from 0.0004 to 0.0034 µg/ml, with mean and median values of 0.0023 and 0.0024 µg/ml, respectively. These baseline sensitivity values will be used in a fungicide resistance monitoring program to determine whether shifts in sensitivity to QoI fungicides are occurring in C. zeae-maydis populations.

First Report of Charcoal Rot Caused by Macrophomina phaseolina on Sunflower in Illinois.

In September 2009, sunflower (Helianthus annuus L.) plants (cv. Mycogen 8C451) from a University of Illinois field research trial in Fayette County, Illinois exhibited silvery gray girdling lesions on the lower stems and premature death. When lower stems and roots were split open, the pith tissue was compressed into layers. Black microsclerotia (90 to 180 µm) were present on the outside of the lower stem tissue and in the stem vascular tissue. Five pieces (approximately 1 cm long) of symptomatic stem tissue from five different affected plants (25 pieces total) were soaked in a 0.5% solution of NaOCl for 30 s, rinsed with sterile distilled water, and placed on potato dextrose agar (PDA; Becton, Dickinson, and Company, Franklin Lakes, NJ). Gray hyphae grew from all of the stem pieces, which subsequently turned black and formed black microsclerotia (75 to 175 µm). On the basis of plant symptoms and size and color of the microsclerotia, the disease was diagnosed as charcoal rot caused by Macrophomina phaseolina (Tassi) Goid (2). To confirm that the isolated fungus was M. phaseolina, DNA was extracted from the pure culture, and PCR amplification of a subunit rDNA and internal transcribed spacer (ITS) region with primers EF3RCNL and ITS4 was performed (3). The Keck Biotechnology Center at the University of Illinois, Urbana sequenced the PCR product. The resulting nucleotide sequence shared the highest homology (99%) with sequences of M. phaseolina when compared with the subunit rDNA and ITS sequences in the nucleotide database ( http://www.ncbi.nlm.nih.gov ). A greenhouse experiment was conducted to confirm pathogenicity; the greenhouse temperature was approximately 27°C and sunflower plants (cv. Cargill 270) were grown in pots and watered daily to maintain adequate soil moisture for growth. Sterile toothpicks were infested with M. phaseolina and placed through the stems (10 cm above the soil surface) of five 40-day-old sunflower plants that were approximately at growth stage R4 (1,4). Five sterile, noninfested toothpicks were similarly placed through sunflower plants to act as controls. Parafilm was used to hold the toothpick in the stem and seal the stem injury. Thirty-five days after inoculation, the mean lesion length on stems inoculated with M. phaseolina was 595 mm and no lesions developed on the control plants. M. phaseolina-inoculated plants also began to wilt and die. Cultures identical to the original M. phaseolina isolate were reisolated from stem lesions of the M. phaseolina-inoculated plants. This is the first report of charcoal rot on sunflower in Illinois to our knowledge. Sunflower is currently not a major crop grown in Illinois, but on-going research is focused on evaluating sunflower as a potential late-planted crop to follow winter wheat. If sunflower production increases in Illinois, growers may need to take precautions to manage charcoal rot. References: (1) L. K. Edmunds. Phytopathology 54:514, 1964. (2) T. Gulya et al. Page 263 in: Sunflower Technology and Production. American Society of Agronomy, Madison, WI, 1997. (3) N. S. Lord et al. FEMS Microbiol. Ecol. 42:327, 2002. (4) A. A. Schneiter and J. F. Miller. Crop Sci. 21:901, 1981.

1,2,3-triazolyl amino acids as AMPA receptor ligands.

The central nervous system glutamate receptors are an important target for drug discovery. Herein we report initial investigations into the synthesis and glutamate receptor activity of 1,2,3-triazolyl amino acids. Two compounds were found to be selective AMPA receptor ligands, which warrant further investigation.

Occurrences of Diplodia Leaf Streak Caused by Stenocarpella macrospora on Corn (Zea mays) in Illinois.

In August 2008, long and narrow lesions were observed on leaves of corn (Zea mays L.) growing in a field in Pope County, Illinois. Lesions were 10 to 35 × 50 to 250 mm and were cream to tan. Dark pycnidia inside the lesions were immersed and approximately 350 µm in diameter. Affected leaves were collected and placed into a moist chamber to encourage the development of conidia. Conidia developed in cirri and were dark, one septate, and 7 to 11 × 59 to 87 µm. Cirri were streaked onto potato dextrose agar (PDA; Becton, Dickinson, and Company, Franklin Lakes, NJ) and cultures arising from single conidia were transferred and maintained. On the basis of the corn leaf symptoms and the morphological characteristics of the pycnidia and conidia, the fungus was tentatively identified as Stenocarpella macrospora (Earle) Sutton (1). To complete Koch's postulates, 'Garst 84H80-3000GT' corn was inoculated in the greenhouse. Conidia were produced by placing a S. macrospora isolate from Pope County, IL onto water agar containing autoclaved corn leaves and incubating at room temperature until pycnidia and conidia were produced (approximately 3 weeks). A conidial suspension was used to inoculate the leaf whorls of corn plants (approximately at the V4 growth stage). Control plants were mock inoculated with sterile water. The experiment was repeated once over time. Twenty days after inoculation, all plants inoculated with S. macrospora conidia developed lesions similar to those observed in the field, and mock-inoculated plants remained symptomless. The fungus was reisolated on PDA from the symptomatic leaves. In August 2009, symptomatic leaves similar to those observed in Pope County, IL in 2008 were observed and collected from corn fields in Gallatin and Vermillion counties. Pycnidia and conidia from these lesions were similar to those described above, and isolates from single conidia were obtained from these samples. To confirm the identity of all isolates collected, PCR amplification of the small subunit rDNA and internal transcribed spacer (ITS) region with primers EF3RCNL and ITS4 was conducted (3). The PCR product was sequenced with these primers at the Keck Biotechnology Center at the University of Illinois, Urbana. The resulting nucleotide sequence was compared with small subunit rDNA and ITS sequences deposited in the GenBank nucleotide database, which revealed 99% homology to sequences of S. macrospora. In total, six of our S. macrospora isolates from Gallatin, Pope, and Vermillion counties were submitted to the United States Department of Agriculture-Agriculture Research Service Culture Collection in Peoria, IL, where they have received NRRL Accession Nos. 54190-54195. To our knowledge, this is the first report of S. macrospora affecting corn in Illinois. Although not observed in the Illinois corn fields described above, S. macrospora has been reported to infect stalks and ears (2). Because of the large leaf lesions caused by S. macrospora and its reported aggressiveness in causing disease on leaves, ears, and stalks, this pathogen has the potential to cause severe yield and quality losses to corn in the United States (2). References: (1) M. L. Carson. Diseases of minor importance or limited occurrence. Page 23 in: Compendium of Corn Diseases. 3rd ed. The American Phytopathological Society, St. Paul, MN, 1999. (2) F. M. Latterell and A. E. Rossi. Plant Dis. 67:725, 1983. (3) N. S. Lord et al. FEMS Microbiol. Ecol. 42:327, 2002.