Evaluating automated infrared thermography and vulva exposure tracking as components of an estrus detection platform in a commercial dairy herd.

The primary objective of this study was to develop an automated infrared thermography platform (Estrus BenchMark) capable of measuring skin temperature and tail movements as a means of identifying cows in estrus. The secondary objective was to evaluate the accuracy of Estrus BenchMark to detect estrus compared to in-line milk progesterone (P4) analysis (Herd Navigator System) in a commercial dairy herd managed under a robotic milking system. Data were collected on forty-six cows from 45 to 120 d after calving. Cows were flagged in estrus when milk P4 fell below 5 ng/mL. The Estrus BenchMark true positive estrus alerts (Sensitivity; Se%) were compared to Herd Navigator System estrus alerts at different time-windows (±12 h, ±24 h, ±48 h, and ±72 h) relative to the Estrus BenchMark estrus alerts for all the estrus alerts (AE) and confidence-quality estrus (CQE; >80% quality) alerts identified by Herd Navigator System. The Estrus BenchMark captured skin temperature and tail movements resulting in vulva exposure (left tail movements, LTail; right tail movements, RTail; and pooled tail movements, PTail) for each milking event. Skin temperature tended to increase when the milk P4 concentration (Least-Squares Means ±â€¯SE) dropped for AE (estrus day [d 0]; P4; 3.51 ±â€¯0.05 ng/mL, Skin temperature; 33.31 ±â€¯2.38 °C) compared with d -7 (P4; 20.22 ±â€¯0.73 ng/mL; Skin temperature: 32.05 ±â€¯3.77 °C). The increase in skin temperature, however, was significant in cows with CQE > 80% at d 0 (32.75 ±â€¯0.29 °C) compared to d -7 (31.80 ±â€¯0.28 °C). The prevalence of tail movements to expose vulva was greater (P = 0.01) in AE at d 0 (LTail: 62.50%; PTail; 68.75%; and RTail: 56.25%) compared with d -7 (LTail: 18.75%; PTail: 9.37%: and RTail: 9.37%), and d +4 (LTail: 9.37%; PTail: 9.37%; and RTail: 12.5%). Moreover, the higher prevalence of tail movements at d 0 was observed in cows with CQE > 80% (LTail; 65%, PTail; 80%, and RTail; 70%) compared to those with CQE < 80%. The highest Estrus BenchMark Youden index (YJ; 0.45), diagnostic odds ratio (DOR; 9.04), and Efficiency (0.77) were achieved for AE in a ±48 h window and at ±72 h window for CQE (YJ; 0.66, DOR; 25.29, and Efficiency 0.76) relative to Herd Navigator System estrus alerts. The highest Estrus BenchMark resulted in 58% estrus detection rates for AE and 80% for cows with CQE compared to the Herd Navigator System.

Evaluation of infrared thermography combined with behavioral biometrics for estrus detection in naturally cycling dairy cows.

Low estrus detection rates (>50%) are associated to extended calving intervals, low economic profit and reduced longevity in Holstein dairy cows. The objective of this study was to evaluate the accuracy of infrared thermography and behavioral biometrics combined as potential estrus alerts in naturally (not induced) cycling dairy cows housed in a tie-stall barn. Eighteen first lactation cows were subjected to transrectal ultrasonography to determine spontaneous ovulation. The dominant follicle (DF) disappearance was used retrospectively as an indirect indicator of ovulation, and to establish the estrus period (48-24 h prior the DF disappearance). Raw skin temperature (Raw IR) and residual skin temperature (Res IR) were recorded using an infrared camera at the Vulva area with the tail (Vtail), Vulva area without the tail (Vnotail), and Vulva's external lips (Vlips) at AM and PM milking from Day 14 until two days after ovulation was confirmed. Behavioral biometrics were recorded on the same schedule as infrared scan. Behavioral biometrics included large hip movements (L-hip), small hip movements (S-hip), large tail movements and small tail movements to compare behavioral changes between estrus and nonestrus periods. Significant increases in Raw IR skin temperature were observed two days prior to ovulation (Vtail; 35.93 ±â€¯0.27 °C, Vnotail; 35.59 ±â€¯0.27 °C, and Vlips; 35.35 ±â€¯0.27 °C) compared to d -5 (Proestrus; Vtail; 35.29 ±â€¯0.27 °C, Vnotail; 34.93 ±â€¯0.31 °C, and Vlips; 34.68 ±â€¯0.27 °C). No significant changes were found for behavioral parameters with the exception of S-hip movements, which increased at two days before ovulation (d -2; 11.13 ±â€¯1.44 Events/5min) compared to d -5 (7.30 ±â€¯1.02 Events/5min). To evaluate the accuracy of thermal and behavioral biometrics, receiver operating characteristic curve analysis was performed using Youden index (YJ), diagnostic odds ratio, positive likelihood ratio (LR+), Sensitivity, Specificity and Positive predicted value to score the estrus alerts. The greatest accuracy achieved using thermal parameters was for Res IR Vtail PM (YJ = 0.34) and L-hip PM (YJ = 0.27) for behavioral biometrics. Combining thermal and behavioral parameters did not improve the YJ index score but reduced the false-positive occurrence observed by increasing the diagnostic odds ratio (26.62), LR+ (12.47), Specificity (0.97) and positive predicted value (0.90) in a Res IR Vtail PM, S-hip AM, S-hip PM combination. The combination of thermal and behavioral parameters increased the accuracy of estrus detection compared to either thermal or behavioral biometrics, independently in naturally cycling cows during milking.

Sexual stages of the female portion in the scallop Nodipecten nodosus (Linné, 1758) and astaxanthin quantity in each stage / Estágios sexuais da porção feminina da gônada da vieira Nodipecten nodosus (Linné, 1758) e a quantidade de astaxantina em cada estágio

This work describes the gametogenic cycle of the scallop Nodipecten nodosus kept in a culture system. To this end, during one year, samples were taken from the broodstocks every 30 days to be submitted to macroscopic and microscopic analyses and to measure the amount of astaxanthin. To perform the microscopic evaluation, 5 μ slices from the median portion of the female part of the gonad were submitted to the pattern methodology for histological analyses with paraffin and HE coloration. The remaining portion of the female gonad was lyophilised to extract and quantify the levels of astaxanthin using HPLC. The microscopic analyses revealed four well defined stages for the reproductive cycle. Analyses of data taken throughout the year indicated preferential spawning periods from December to January and from July to September. The astaxanthin analyses showed higher amounts of this carotenoid during the advanced pre-spawning and the initial spawning periods than during gametogenesis, initial pre-spawning, advanced spawning, and the spent stages. According to these results, it was possible to establish a descriptive table of the sexual stages of the female portion of the gonad and the amount of astaxanthin in the sexual stage of the scallop Nodipecten nodosus.

Este trabalho descreve o ciclo gametogênico da vieira Nodipecten nodosus mantida em ambiente de cultivo. Para isto, durante um ano, amostras de indivíduos reprodutores foram coletadas a cada 30 dias e submetidas à avaliação macroscópica e microscópica e à quantificação de astaxantina. Para a avaliação microscópica, secções de 5 μ da porção mediana feminina da gônada foram submetidas à metodologia de análise histológica padrão em parafina e coloração HE. O restante da porção feminina da gônada foi liofilizado para extração e quantificação de astaxantina em HPLC. A avaliação microscópica permitiu a descrição de quatro estágios bem definidos para o ciclo reprodutivo. Na análise ao longo do ano, foram observados períodos preferenciais de desova em dezembro e janeiro e de julho a setembro. A análise da quantidade de astaxantina, mostrou, nos estádios de pré-desova avançada e de desova inicial, uma maior quantidade desse carotenoide em comparação aos estádios de gametogênese, pré-desova inicial, desova avançada e repouso. Em função desses resultados, foi possível estabelecer um quadro descritivo dos estágios sexuais da porção feminina da gônada e quantidade de astaxantina em cada estágio sexual da vieira Nodipecten nodosus.

Effects of local anesthetic and a nonsteroidal antiinflammatory drug on pain responses of dairy calves to hot-iron dehorning.

This study examined the effects of a nonsteroidal antiinflammatory agent (NSAID) on physiological responses of calves immediately after hot-iron dehorning (DH) and during the time that local anesthetic (LA) wears off (2 to 3 h) after this procedure. Forty-six calves (33 +/- 0.3 d of age) were randomly assigned to 6 treatments: hot-iron DH versus sham DH with either no pain mitigation, LA alone, or LA with NSAID (i.v. Meloxicam). Eye temperature (measured using infrared thermography) was recorded every 5 min for 3 h after treatments. Heart rate (HR) and heart rate variability (HRV) were recorded continuously; for analysis of HRV, short segments of 512 interbeat intervals were examined. After DH without LA or NSAID, HR increased by 35 +/- 3.0 beats/min in the first 5 min and remained elevated above baseline for 3 h. The HRV around the time of DH did not differ between treatments; however, the root mean square of successive differences decreased from 68 to 41 +/- 12.6 ms immediately following DH without pain relief, suggesting a decrease in vagal tone at this time. Between 2 and 3 h following DH with LA, there was a decrease in eye temperature (-0.6 +/- 0.1 degrees C), an increase in HR (8 +/- 3.0 beats per min) and changes in HRV. Changes in HRV at this time included a decreased high-frequency power and an increase in the low-frequency power and low-frequency/high-frequency ratio, indicating a change in sympatho-vagal balance. The changes in eye temperature, HR, and HRV between 2 and 3 h following DH with LA indicated the onset of pain coinciding with the time that the LA effects wear off. In addition, this study demonstrated that the combination of LA and NSAID mitigated the onset of pain responses when the LA wanes.

Quorum-sensing signals indicate that cystic fibrosis lungs are infected with bacterial biofilms.

The bacterium Pseudomonas aeruginosa permanently colonizes cystic fibrosis lungs despite aggressive antibiotic treatment. This suggests that P. aeruginosa might exist as biofilms--structured communities of bacteria encased in a self-produced polymeric matrix--in the cystic fibrosis lung. Consistent with this hypothesis, microscopy of cystic fibrosis sputum shows that P. aeruginosa are in biofilm-like structures. P. aeruginosa uses extracellular quorum-sensing signals (extracellular chemical signals that cue cell-density-dependent gene expression) to coordinate biofilm formation. Here we found that cystic fibrosis sputum produces the two principal P. aeruginosa quorum-sensing signals; however, the relative abundance of these signals was opposite to that of the standard P. aeruginosa strain PAO1 in laboratory broth culture. When P. aeruginosa sputum isolates were grown in broth, some showed quorum-sensing signal ratios like those of the laboratory strain. When we grew these isolates and PAO1 in a laboratory biofilm model, the signal ratios were like those in cystic fibrosis sputum. Our data support the hypothesis that P. aeruginosa are in a biofilm in cystic fibrosis sputum. Moreover, quorum-sensing signal profiling of specific P. aeruginosa strains may serve as a biomarker in screens to identify agents that interfere with biofilm development.

The effects of mutated skeletal ryanodine receptors on calreticulin and calsequestrin expression in the brain and pituitary gland of boars.

Mutations in skeletal ryanodine receptors (sRyR) result in malignant hyperthermia in humans and Porcine Stress Syndrome (PSS) in pigs. Whether the sRyR is expressed in neuronal tissue and what impact it has on neuronal function is relatively unexplored. We have hypothesized that the presence of mutated sRyR may be accompanied by compensatory changes in Ca(2+)-binding protein expression. We were interested in whether pigs heterozygous for mutated sRyR would show changes in the expression of Ca(2+)-binding proteins, in specific regions of the brain, and whether changes in this expression would be accompanied by the presence of sRyR within that region. The objectives of the current experiments were to determine (1) whether calreticulin (CR) and calsequestrin (CS) are expressed in the pituitary gland and brain of the pig, (2) if boars heterozygous for mutated sRyR differed from wild-type boars in the expression of CR or CS, and (3) if altered Ca(2+)-binding protein expression would be accompanied by the presence of sRyR mRNA. Boars either heterozygous or wild-type (n=6) for the mutation in sRyR known to cause PSS, were euthanized and the pituitary gland and brains were collected for western blotting for CR and CS. An additional four wild-type boars were sacrificed and brains were collected for in situ hybridization for sRyR mRNA. Immunoreactive CR was expressed in porcine tissues with highest (p<0.0001) expression in the pituitary gland and lower but equivalent expression in the hypothalamus, frontal cortex, and hippocampus. Immunoreactive CS was not detectable in the pituitary gland while low levels were observed in the hypothalamus and frontal cortex. Dramatically higher (p<0.0001) levels of CS were found in the hippocampus. Genotype did not affect CR expression in the pituitary gland or any brain region examined. Immunoreactive CS levels were lower (p<0.002) in the hippocampus of heterozygous compared to wild-type boars. In situ hybridization experiments revealed the presence of sRyR mRNA in the hippocampus equally distributed across all cell subfields. In conclusion, both CR and CS were expressed in the porcine brain with specific patterns of expression across the brain regions examined. Boars heterozygous for mutated sRyR had lower CS in the hippocampus, which was accompanied by the expression of mRNA for sRyR.

Distributions of conjugated linoleic acid (CLA) isomers in tissue lipid classes of pigs fed a commercial CLA mixture determined by gas chromatography and silver ion-high-performance liquid chromatography.

Pigs were fed a commercial conjugated linoleic acid (CLA) mixture, prepared by alkali isomerization of sunflower oil, at 2% of the basal diet, from 61.5 to 106 kg live weight, and were compared to pigs fed the same basal diet with 2% added sunflower oil. The total lipids from liver, heart, inner back fat, and omental fat of pigs fed the CLA diet were analyzed for the incorporation of CLA isomers into all the tissue lipid classes. A total of 10 lipid classes were isolated by three-directional thin-layer chromatography and analyzed by gas chromatography (GC) on long capillary columns and by silver-ion high-performance liquid chromatography (Ag+-HPLC); cholesterol was determined spectrophotometrically. Only trace amounts (<0.1%; by GC) of the 9,11-18:2 cis/trans and trans,trans isomers were observed in pigs fed the control diet. Ten and twelve CLA isomers in the diet and in pig tissue lipids were separated by GC and Ag+- HPLC, respectively. The relative concentration of all the CLA isomers in the different lipid classes ranged from 1 to 6% of the total fatty acids. The four major cis/trans isomers (18.9% 11 cis,13 trans-18:2; 26.3% 10 trans,12 cis-18:2; 20.4% 9 cis,11 trans-18:2; and 16.1% 8 trans, 10 cis-18:2) constituted 82% of the total CLA isomers in the dietary CLA mixture, and smaller amounts of the corresponding cis,cis (7.4%) and trans,trans (10.1%) isomers were present. The distribution of CLA isomers in inner back fat and in omental fat of the pigs was similar to that found in the diet. The liver triacylglycerols (TAG), free fatty acids (FFA), and cholesteryl esters showed a similar pattern to that found in the diet. The major liver phospholipids showed a marked increase of 9 cis,11 trans-18:2, ranging from 36 to 54%, compared to that present in the diet. However, liver diphosphatidylglycerol (DPG) showed a high incorporation of the 11 cis,13 trans-18:2 isomer (43%). All heart lipid classes, except TAG, showed a high content of 11 cis,13 trans-18:2, which was in marked contrast to results in the liver. The relative proportion of 11 cis,13 trans-18:2 ranged from 30% in the FFA to 77% in DPG. The second major isomer in all heart lipids was 9 cis,11 trans-18:2. In both liver and heart lipids the relative proportions of both 10 trans,12 cis-18:2 and 8 trans, 10 cis-18:2 were significantly lower compared to that found in the diet. The FFA in liver and heart showed the highest content of trans,trans isomers (31 to 36%) among all the lipid classes. The preferential accumulation of the 11 cis,13 trans-18:2 into cardiac lipids, and in particular the major phospholipid in the inner mitochondrial membrane, DPG, in both heart and liver, appears unique and may be of concern. The levels of 11 cis,13 trans-18:2 naturally found in foods have not been established.

Analysis of random and site-directed mutations in rhII, a Pseudomonas aeruginosa gene encoding an acylhomoserine lactone synthase.

The opportunistic human pathogen Pseudomonas aeruginosa possesses two cell density-dependent genetic regulatory systems that control expression of a number of secreted virulence factors. These two systems, the lasI-lasR and rhII-rhIR gene pairs, are members of the luxI-luxR family of quorum-sensing signal generators and signal receptors. The rhII gene in P. aeruginosa encodes a 201-amino-acid protein that catalyses the synthesis of an autoinducer, butyrylhomoserine lactone. Through a programme of random and site-specific mutagenesis of rhII we have gained a better understanding of how its protein product functions. Eight residues critical to butyrylhomoserine lactone synthesis by RhiI were identified by random mutagenesis, and all mapped to a conserved region that spans residues 24-104. Seven of the eight residues were charged amino acids and the other was a glycine. By using site-specific mutagenesis we showed that an active-site cysteine or serine was not required for butyrylhomoserine lactone synthesis, and that two conserved aromatic amino acids in the postulated active site region could be altered without complete loss of RhII activity. Furthermore, two residues towards the C-terminus that align with critical residues in LuxI can be altered in RhII without loss of activity. These studies suggest that as opposed to the current models for acyl substrate binding to quorum-sensing signal generators, charged amino acid residues participate directly in the catalysis of butyrylhomoserine lactone synthesis rather than cysteines, serines or hydrophobic amino acids.

Generation of cell-to-cell signals in quorum sensing: acyl homoserine lactone synthase activity of a purified Vibrio fischeri LuxI protein.

Many bacteria use acyl homoserine lactone signals to monitor cell density in a type of gene regulation termed quorum sensing and response. Synthesis of these signals is directed by homologs of the luxi gene of Vibrio fischeri. This communication resolves two critical issues concerning the synthesis of the V. fischeri signal. (i) The luxI product is directly involved in signal synthesis-the protein is an acyl homoserine lactone synthase; and (ii) the substrates for acyl homoserine lactone synthesis are not amino acids from biosynthetic pathways or fatty acid degradation products, but rather they are S-adenosylmethionine (SAM) and an acylated acyl carrier protein (ACP) from the fatty acid biosynthesis pathway. We purified a maltose binding protein-LuxI fusion polypeptide and showed that, when provided with the appropriate substrates, it catalyzes the synthesis of an acyl homoserine lactone. In V. fischeri, luxi directs the synthesis of N-(3-oxohexanoyl) homoserine lactone and hexanoyl homoserine lactone. The purified maltose binding protein-LuxI fusion protein catalyzes the synthesis of hexanoyl homoserine lactone from hexanoyl-ACP and SAM. There is a high level of specificity for hexanoyl-ACP over ACPs with differing acyl group lengths, and hexanoyl homoserine lactone was not synthesized when SAM was replaced with other amino acids, such as methionine, S-adenosylhomocysteine, homoserine, or homoserine lactone, or when hexanoyl-SAM was provided as the substrate. This provides direct evidence that the LuxI protein is an auto-inducer synthase that catalyzes the formation of an amide bond between SAM and a fatty acyl-ACP and then catalyzes the formation of the acyl homoserine lactone from the acyl-SAM intermediate.

Are they ready? Discharge planning for older surgical patients.

1. With ever-increasing cost containment, patients are being discharged from the hospital earlier. 2. The majority (96%) of elderly patients (average age 70 years) in this study, after abdominal and cardiac surgery, thought they were ready to be discharged. 3. Surgical patients' level of functional ability was significantly improved from predischarge to postdischarge. There were no significant differences in their pain, strength, knowledge, and mood levels. 4. Predominantly non-nursing functions (eg, transportation, housekeeping) were identified as areas of needed help at home for elderly patients after surgery.