Review: welfare of dairy cows in continuously housed and pasture-based production systems

There is increasing interest in the use of continuous housing systems for dairy cows, with various reasons put forward to advocate such systems. However, the welfare of dairy cows is typically perceived to be better within pasture-based systems, although such judgements are often not scientifically based. The aim of this review was to interrogate the existing scientific literature to compare the welfare, including health, of dairy cows in continuously housed and pasture-based systems. Although summarising existing work, knowledge gaps and directions for future research are also identified. The scope of the review is broad, examining relevant topics under three main headings; health, behaviour and physiology. Regarding health, cows on pasture-based systems had lower levels of lameness, hoof pathologies, hock lesions, mastitis, uterine disease and mortality compared with cows on continuously housed systems. Pasture access also had benefits for dairy cow behaviour, in terms of grazing, improved lying/resting times and lower levels of aggression. Moreover, when given the choice between pasture and indoor housing, cows showed an overall preference for pasture, particularly at night. However, the review highlighted the need for a deeper understanding of cow preference and behaviour. Potential areas for concern within pasture-based systems included physiological indicators of more severe negative energy balance, and in some situations, the potential for compromised welfare with exposure to unpredictable weather conditions. In summary, the results from this review highlight that there remain considerable animal welfare benefits from incorporating pasture access into dairy production systems.     

Genetic parameters for production,health, fertility and longevity traits in dairy cows

Milk production, fertility, longevity and health records, were extracted from databases of two milk recording organisations in the United Kingdom for the first three lactations of the Holstein–Friesian breed. These included data related to health events (mastitis and lameness), voluntarily recorded on a proportion of farms. The data were analysed to calculate disease incidence levels and to estimate genetic parameters for health traits and their relationships with production and other functional traits. The resulting dataset consisted of 124 793 lactations from 75 137 animals of 1586 sires, recorded in 2434 herds. Incidence of health events increased with parity. The overall incidence of mastitis (MAS) and lameness (LAM), defined as binary traits, were 17% and 16%, respectively. Heritability estimates for MAS and LAM were 0.04 and 0.02, respectively, obtained from repeatability linear sire models. Heritability estimates of mastitis and lameness as count traits were slightly higher, 0.05 and 0.03, respectively. Genetic correlations were obtained by bivariate analyses of all pair-wise combinations between milk 305-day yield (MY), protein 305-day yield (PY), fat 305-day yield (FY), lactation average log_e transformed lactation average somatic cell count (SCS), calving interval (CI), days to first service (DFS), non-return at 56 days (NR56), number of inseminations (NINS), mastitis (MAS), number of mastitis episodes (NMAS), lameness (LAM), number of lameness episodes (NLAM) and lifespan score (LS). As expected, MAS was correlated most strongly with SCS (0.69), which supports the use of SCS as an indicator trait for mastitis. Genetic correlations between MAS and yield and fertility traits were of similar magnitude ranging from 0.27 to 0.33. Genetic correlations between MAS with LAM and LS were 0.38 and −0.59, respectively. Not all genetic correlations between LAM and other traits were significant because of fewer numbers of lameness records. LAM had significant genetic correlations with MY (0.38), PY (0.28), CI (0.35), NINS (0.38) and LS (−0.53). The heritability estimates of mastitis and lameness were low; therefore, genetic gain through direct selection alone would be slow, yet still positive and cumulative. Direct selection against mastitis and lameness as additional traits should reduce incidence of both diseases, and simultaneously improve fertility and longevity. However, both health traits had antagonistic relationships with production traits, thus genetic gain in production would be slower.     

Exposure chamber for determining the biological effects of electric and magnetic fields on dairy cows

An exposure chamber was designed to study the effects of electric and magnetic fields (EMF) on oestrous cycles, hormonal profile during gestation, pineal function, quantity and quality of milk production, feed intake, and central nervous system of dairy cattle. The chamber was 15 x 10 x 3 m; and the control system was fully computerized so that the field intensities can be varied and monitored continuously, on site or remotely. During exposure to EMF, milk production, feed consumption, and health were monitored closely and blood and cerebral spinal fluid were continuously sampled. The chamber characteristics allow use of a wide range of exposure such as electric fields (0-30 kV/m) and magnetic fields (0-100 microT) at frequencies ranging from 45 to 3000 Hz.     

Genetic evaluation of ovulatory disorders in Austrian Fleckvieh cows: a comparison between linear models and survival analysis

The objective of this study was to compare linear models and survival analysis for genetic evaluation of ovulatory disorders, which included veterinary treatments of silent heat/anestrus and cystic ovaries. Data of 23 450 daughters of 274 Austrian Fleckvieh sires were analyzed. For linear model analyses, ovulatory disorders were defined as a binary response (presence or absence) in the time periods from calving to 150 days after calving and from calving to 300 days after calving. For survival analysis, ovulatory disorders were defined either as the number of days from calving to the day of the first treatment for an ovulatory disorder (uncensored record) or from calving to the day of culling, or the last day of the period under investigation (until 150 or 300 days after calving; censored record). Estimates of heritability were very similar (0.016 to 0.020) across methods and periods. Correlations between sire estimated breeding value from linear model and survival analysis were 0.98, whereas correlations between different time periods were somewhat lower (0.95 and 0.96). The results showed that the length of time period had a larger effect on genetic evaluation than methodology.     

Hair cortisol detection in dairy cattle by using EIA: protocol validation and correlation with faecal cortisol metabolites

Hair may be a useful matrix to detect cumulative cortisol concentrations in studies of animal welfare and chronic stress. The aim of this study was to validate a protocol for cortisol detection in hair from dairy cattle by enzyme immunoassay (EIA). Seventeen adult Holstein–Friesian dairy cows were used during the milking period. Hair cortisol concentration was assessed in 25-day-old hair samples taken from the frontal region of the head, analysing black and white coloured hair separately. Concentrations of cortisol metabolites were determined in faeces collected twice a week during the same period of time. There was a high correlation between cortisol values in faeces and cortisol in white colour hair samples but such correlation was not significant with the black colour hair samples. The intra- and inter-assay coefficients of variation were 4.9% and 10.6%, respectively. The linearity showed R²=0.98 and mean percentage error of −10.8±1.55%. The extraction efficiency was 89.0±23.52% and the parallelism test showed similar slopes. Cortisol detection in hair by using EIA seems to be a valid method to represent long-term circulating cortisol levels in dairy cattle.     

Mixture model for inferring susceptibility to mastitis in dairy cattle: a procedure for likelihood-based inference

A Gaussian mixture model with a finite number of components and correlated random effects is described. The ultimate objective is to model somatic cell count information in dairy cattle and to develop criteria for genetic selection against mastitis, an important udder disease. Parameter estimation is by maximum likelihood or by an extension of restricted maximum likelihood. A Monte Carlo expectation-maximization algorithm is used for this purpose. The expectation step is carried out using Gibbs sampling, whereas the maximization step is deterministic. Ranking rules based on the conditional probability of membership in a putative group of uninfected animals, given the somatic cell information, are discussed. Several extensions of the model are suggested.     

Quantitative underwater 3D motion analysis using submerged video cameras: accuracy analysis and trajectory reconstruction

In this study we aim at investigating the applicability of underwater 3D motion capture based on submerged video cameras in terms of 3D accuracy analysis and trajectory reconstruction. Static points with classical direct linear transform (DLT) solution, a moving wand with bundle adjustment and a moving 2D plate with Zhang's method were considered for camera calibration. As an example of the final application, we reconstructed the hand motion trajectories in different swimming styles and qualitatively compared this with Maglischo's model. Four highly trained male swimmers performed butterfly, breaststroke and freestyle tasks. The middle fingertip trajectories of both hands in the underwater phase were considered. The accuracy (mean absolute error) of the two calibration approaches (wand: 0.96 mm – 2D plate: 0.73 mm) was comparable to out of water results and highly superior to the classical DLT results (9.74 mm). Among all the swimmers, the hands' trajectories of the expert swimmer in the style were almost symmetric and in good agreement with Maglischo's model. The kinematic results highlight symmetry or asymmetry between the two hand sides, intra- and inter-subject variability in terms of the motion patterns and agreement or disagreement with the model. The two outcomes, calibration results and trajectory reconstruction, both move towards the quantitative 3D underwater motion analysis.     

Genome‐wide association analysis and gene set enrichment analysis with SNP data identify genes associated with 305‐day milk yield in Holstein dairy cows

Milk production traits, such as 305‐day milk yield (305MY), have been under direct selection to improve production in dairy cows. Over the past 50 years, the average milk yield has nearly doubled, and over 56% of the increase is attributable to genetic improvement. As such, additional improvements in milk yield are still possible as new loci are identified. The objectives of this study were to detect SNPs and gene sets associated with 305MY in order to identify new candidate genes contributing to variation in milk production. A population of 781 primiparous Holstein cows from six central Washington dairies with records of 305MY and energy corrected milk were used to perform a genome‐wide association analysis (GWAA) using the Illumina BovineHD BeadChip (777 962 SNPs) to identify QTL associated with 305MY (P < 1.0 × 10^?5). A gene set enrichment analysis with SNP data (GSEA‐SNP) was performed to identify gene sets (normalized enrichment score > 3.0) and leading edge genes (LEGs) influencing 305MY. The GWAA identified three QTL comprising 34 SNPs and 30 positional candidate genes. In the GSEA‐SNP, five gene sets with 58 unique and 24 shared LEGs contributed to 305MY. Identification of QTL and LEGs associated with 305MY can provide additional targets for genomic selection to continue to improve 305MY in dairy cattle.     

Combined analysis of data from two granddaughter designs: A simple strategy for QTL confirmation and increasing experimental power in dairy cattle

A joint analysis of five paternal half-sib Holstein families that were part of two different granddaughter designs (ADR- or Inra-design) was carried out for five milk production traits and somatic cell score in order to conduct a QTL confirmation study and to increase the experimental power. Data were exchanged in a coded and standardised form. The combined data set (JOINT-design) consisted of on average 231 sires per grandsire. Genetic maps were calculated for 133 markers distributed over nine chromosomes. QTL analyses were performed separately for each design and each trait. The results revealed QTL for milk production on chromosome 14, for milk yield on chromosome 5, and for fat content on chromosome 19 in both the ADR- and the Inra-design (confirmed within this study). Some QTL could only be mapped in either the ADR- or in the Inra-design (not confirmed within this study). Additional QTL previously undetected in the single designs were mapped in the JOINT-design for fat yield (chromosome 19 and 26), protein yield (chromosome 26), protein content (chromosome 5), and somatic cell score (chromosome 2 and 19) with genomewide significance. This study demonstrated the potential benefits of a combined analysis of data from different granddaughter designs.     

Step behaviour and autonomic nervous system activity in multiparous dairy cows during milking in a herringbone milking system

Behavioural and cardiac responses of multiparous dairy cows (n=24) during milking in a 2×4 stall herringbone milking system were evaluated in this study. Heart rate (HR), parasympathetic tone index (high frequency component, HF) of heart rate variability and sympathovagal balance indicator LF/HF ratio (the ratio of the low frequency (LF) and the HF component) were analysed. Measurement periods were established as follows: (1) standing calm (baseline), (2) udder preparation, (3) milking, (4) waiting after milking in the milking stall and (5) in the night (2 h after milking). Step behaviour was recorded and calculated per minute for the three phases of the milking process (udder preparation, milking and waiting after milking). HR was higher during udder preparation and milking compared with baseline (P=0.03, 0.027, respectively). HF was significantly lower than baseline levels during waiting in the milking stall after milking (P=0.009), however, during udder preparation, milking and 2 h after milking did not differ from baseline (P>0.05, in either case). LF/HF during the three phases of the milking process differed neither from baseline levels nor from each other. Steps occurred more often during waiting after milking than during udder preparation (P=0.042) or during milking (23; P=0.017). Our results suggest that the milking procedure itself was not stressful for these animals. After milking (following the removal of the last teat cup and before leaving the milking stall), both decreased parasympathetic tone (lower HF) and increased stepping rate indicated a sensitive period for animals during this phase.     

Effects of stage of pregnancy on variance components,daily milk yields and 305-day milk yield in Holstein cows,as estimated by using a test-day model

Pregnancy and calving are elements indispensable for dairy production, but the daily milk yield of cows decline as pregnancy progresses, especially during the late stages. Therefore, the effect of stage of pregnancy on daily milk yield must be clarified to accurately estimate the breeding values and lifetime productivity of cows. To improve the genetic evaluation model for daily milk yield and determine the effect of the timing of pregnancy on productivity, we used a test-day model to assess the effects of stage of pregnancy on variance component estimates, daily milk yields and 305-day milk yield during the first three lactations of Holstein cows. Data were 10 646 333 test-day records for the first lactation; 8 222 661 records for the second; and 5 513 039 records for the third. The data were analyzed within each lactation by using three single-trait random regression animal models: one model that did not account for the stage of pregnancy effect and two models that did. The effect of stage of pregnancy on test-day milk yield was included in the model by applying a regression on days pregnant or fitting a separate lactation curve for each days open (days from calving to pregnancy) class (eight levels). Stage of pregnancy did not affect the heritability estimates of daily milk yield, although the additive genetic and permanent environmental variances in late lactation were decreased by accounting for the stage of pregnancy effect. The effects of days pregnant on daily milk yield during late lactation were larger in the second and third lactations than in the first lactation. The rates of reduction of the 305-day milk yield of cows that conceived fewer than 90 days after the second or third calving were significantly (P<0.05) greater than that after the first calving. Therefore, we conclude that differences between the negative effects of early pregnancy in the first, compared with later, lactations should be included when determining the optimal number of days open to maximize lifetime productivity in dairy cows.     

山东省某地区奶牛源大肠埃希菌的血清型、耐药特性及分子特性

【目的】旨在对从山东省某地区4个健康奶牛养殖场分离到的大肠埃希菌进行优势血清型、耐药特性、Ⅰ类整合子基因盒携带情况以及系统进化群分析。【方法】采集194份来自山东省某地区4个规模化奶牛场奶牛新鲜粪便样品,进行大肠埃希菌分离和鉴定,利用常用大肠埃希菌诊断血清进行血清型鉴定;利用10%的绵羊血平板检测溶血性;利用K-B法检测对14种常规抗菌药物的敏感性;利用聚合酶链式反应(PCR)检测革兰阴性菌常见的6大类24种耐药基因、Ⅰ类整合子基因盒结构并对目的条带测序分析;利用细菌多位点序列分型(Multilocus sequence typing,MLST)技术分析大肠埃希菌的ST型并使用eBURST v3软件分析菌株之间的克隆关系。【结果】从194份新鲜粪便样品中分离到171株大肠埃希菌,其中主要为致病性(19.9%)和侵袭性大肠埃希菌(17.0%),优势血清型分别为O128:K67(12/171)和O143:K7(12/171)。另外,具有溶血性的大肠埃希菌阳性率为9.4%(16/171);药敏试验结果显示多重耐药菌株的比率为22.2%,其中对氨苄西林耐药率最高为33.9%,四环素次之,为24.0%;PCR检测耐药基因和整合子结果显示,59.1%的菌株携带β-内酰胺类耐药基因blaTEM,59.1%的菌株携带氨基糖苷类耐药基因ant(2′),未检测到四环素耐药基因tetA和tetB;Ⅰ类整合子的阳性率为4.1%(7/171),dfrA12-aadA2-sul1为优势基因盒结构(4/171);MLST将大肠埃希菌分为8种ST型,其中,ST155(10/171)和ST58(45/171)形成一个克隆复合物且没有发现新的ST型。【结论】本研究证实,从该地区规模化健康奶牛场新鲜粪便中分离到的大肠埃希菌优势血清型为O128:K67和O143:K7;少部分大肠埃希菌具有溶血性;仅对氨苄西林、四环素等具有较高的耐药率;优势基因盒结构为dfrA12-aadA2-sul1;MLST分型显示不同奶牛场分离出亲缘关系较近的菌株,其分布具有多态性,血清型与ST型之间无相关性。本研究表明源自表观健康的奶牛的大肠埃希菌存在多重耐药现象,具有食品公共卫生安全隐患,该研究对于提升规模化奶牛场奶制品的安全生产与质量评估具有一定的理论指导意义。     

Evaluation of the impact of a Herd Health and Production Management programme in organic dairy cattle farms: a process evaluation approach

Animal health planning activities are not always providing a satisfactory positive impact on herd health and welfare. Moreover, evaluating the impact of advisory programmes is complex due to multiple interacting elements that influence its outcome. Therefore, measuring solely health outcomes is not sufficient: the whole process of the implementation and use of such programmes should be evaluated. In order to evaluate the impact of an intervention with a Herd Health and Production Management (HHPM) programme a process evaluation framework was designed and used. The intervention involved 20 organic dairy cattle farmers and their advisors, in both France and Sweden. In both countries 20 organic dairy farms were selected as control herds. The evaluation of the HHPM programme was based on: (a) the compliance to the programme; (b) the programme’s functions influencing herd health management practices and stimulating dialogue between farmers and advisors; (c) its effectiveness in terms of improving herd health compared with control farms. Complete compliance to the programme was fulfilled by 21 out of 40 farmers–advisors. Results from a questionnaire showed that the programme functioned as intended (e.g. by allowing early identification of herd health problems), stimulated change in farmers’ herd health management practices and farmer–advisor dialogue. Even though the majority of the users perceived that the programme contributed to herd health improvements, no significant differences in health outcomes were found when compared with control farms 12 months after the start of the intervention. The programme allowed creating an environment promoting the exchange of information between farmers and advisors, necessary to define pertinent advice in a farm-specific situation. Future research should aim at improving methods for the evaluation of the effect of advisory programmes, by identifying early indicators for effective advice and developing methods to evaluate the quality of advisory situations without interfering with them.     

Welfare implication of measuring heart rate and heart rate variability in dairy cattle: literature review and conclusions for future research

Heart rate (HR) measurements have been used to determine stress in livestock species since the beginning of the 1970s. However, according to the latest studies in veterinary and behaviour–physiological sciences, heart rate variability (HRV) proved to be more precise for studying the activity of the autonomic nervous system. In dairy cattle, HR and HRV indices have been used to detect stress caused by routine management practices, pain or milking. This review provides the significance of HR and HRV measurements in dairy cattle by summarising current knowledge and research results in this area. First, the biological background and the interrelation of the autonomic regulation of cardiovascular function, stress, HR and HRV are discussed. Equipment and methodological approaches developed to measure interbeat intervals and estimate HRV in dairy cattle are described. The methods of HRV analysis in time, frequency and non-linear domains are also explained in detail emphasising their physiological background. Finally, the most important scientific results and potential possibilities for future research are presented.     

Graph-based impact analysis as a framework for incorporating practitioner knowledge in dairy herd health management

Production diseases in dairy cows are multifactorial, which means they emerge from complex interactions between many different farm variables. Variables with a large impact on production diseases can be identified for groups of farms using statistical models, but these methods cannot be used to identify highly influential variables in individual farms. This, however, is necessary for herd health planning, because farm conditions and associated health problems vary largely between farms. The aim of this study was to rank variables according to their anticipated effect on production diseases on the farm level by applying a graph-based impact analysis on 192 European organic dairy farms. Direct impacts between 13 pre-defined variables were estimated for each farm during a round-table discussion attended by practitioners, that is farmer, veterinarian and herd advisor. Indirect impacts were elaborated through graph analysis taking into account impact strengths. Across farms, factors supposedly exerting the most influence on production diseases were ‘feeding’, ‘hygiene’ and ‘treatment’ (direct impacts), as well as ‘knowledge and skills’ and ‘herd health monitoring’ (indirect impacts). Factors strongly influenced by production diseases were ‘milk performance’, ‘financial resources’ and ‘labour capacity’ (directly and indirectly). Ranking of variables on the farm level revealed considerable differences between farms in terms of their most influential and most influenced farm factors. Consequently, very different strategies may be required to reduce production diseases in these farms. The method is based on perceptions and estimations and thus prone to errors. From our point of view, however, this weakness is clearly outweighed by the ability to assess and to analyse farm-specific relationships and thus to complement general knowledge with contextual knowledge. Therefore, we conclude that graph-based impact analysis represents a promising decision support tool for herd health planning. The next steps include testing the method using more specific and problem-oriented variables as well as evaluating its effectiveness.     

Understanding the reproductive performance of a dairy cattle herd by using both analytical and systemic approaches: a case study based on a system experiment

Reproductive performance has recently been a growing concern in cattle dairy systems, but few research methodologies are available to address it as a complex problem in a livestock farming system. The aim of this paper is to propose a methodology that combines both systemic and analytical approaches in order to better understand and improve reproductive performance in a cattle dairy system. The first phase of our methodology consists in a systemic approach to build the terms of the problem. It results in formalising a set of potential risk factors relevant for the particular system under consideration. The second phase is based on an analytical approach that involves both analysing the shapes of the individual lactation curves and carrying out logistic regression procedures to study the links between reproductive performance and the previously identified potential risk factors. It makes it possible to formulate hypotheses about the biotechnical phenomena underpinning reproductive performance. The last phase is another systemic approach that aims at suggesting new practices to improve the situation. It pays particular attention to the consistency of those suggestions with the farmer's general objectives. This methodology was applied to a French system experiment based on an organic low-input grazing system. It finally suggested to slightly modify the dates of the breeding period so as to improve reproductive performance. The formulated hypotheses leading to this suggestion involved both the breed (Holstein or Montbéliarde cows), the parity, the year and the calving date with regard to the turnout date as the identified risk factors of impaired performance. Possible use of such a methodology in any commercial farm encountering a biotechnical problem is discussed.     

Efficacy of decreasing the dose of GnRH used in a protocol for synchronization of ovulation and timed AI in lactating dairy cows

To determine the efficacy of reducing the dosage of GnRH used in a protocol for synchronization of ovulation and timed AI, primiparous and multiparous lactating Holstein cows (n=237) were randomly assigned to 1 of 2 treatment groups. Ovulation was synchronized for cows in the first group using intramuscular injections of GnRH and PGF₂ as follows: Day 0, 100 μg GnRH; Day 7, 25 mg PGF₂; Day 9, 100 μg GnRH. Ovulation was synchronized in the second group of cows using the same injection schedule and dosage of PGF₂ but only 50 μg GnRH per injection. All cows underwent a timed AI at 12 to 18 h after the second GnRH injection. The proportion of cows ovulating in response to the second GnRH injection (synchronization rate) and pregnancy status at 28 and 56 d post AI were determined using transrectal ultrasonography. The synchronization rate, double-ovulation rate, conception rate at 28 and 56 d post AI, and pregnancy loss from 28 to 56 d post AI did not differ statistically between treatment groups. For all cows, synchronization rate was 84.0%, and double-ovulation rate was 14.1%. Conception rates calculated using all cows receiving synchronization of ovulation were 41.1% at 28 d and 34.4% at 56 d post AI. Conception rates calculated for only synchronized cows were 47.6% at 28 d and 40.1% at 56 d post AI. For all cows, pregnancy loss from 28 to 56 d post AI was 13.5%, with an attrition rate of 0.5% per day. Estimated savings in hormone costs using 50 rather than 100 μg GnRH per injection for synchronizing ovulation were $6.40 per cow and $20.27 per pregnancy. Thus, decreasing the dosage of GnRH used for synchronization of ovulation and timed AI in lactating dairy cows reduces synchronization costs per cow and per pregnancy without compromising the efficacy of the synchronization protocol.     

Variation in the faecal shedding of Salmonella and E. coli O157:H7 in lactating dairy cattle and examination of Salmonella genotypes using pulsed-field gel electrophoresis

AIMS: To examine the variability in faecal shedding of Salmonella and Escherichia coli O157:H7 in healthy lactating dairy cattle and to evaluate the genetic relatedness of Salmonella isolates. METHODS: Faecal samples were obtained from lactating Holstein dairy cattle on four commercial farms in the southwestern US. All farms were within an 8-km radius and were sampled in August 2001, January 2002 and August 2002 (60 cows per farm per sampling; n = 720 total samples). Samples were cultured for E. coli O157:H7 and Salmonella and a portion of the recovered Salmonella isolates were examined for genetic relatedness using pulsed-field gel electrophoresis (PFGE). RESULTS: Faecal shedding of E. coli O157:H7 and Salmonella varied considerably between farms and at the different sampling times. Large fluctuations in the percentage of positive animals were observed from summer to summer for both of these pathogens. Similarly, Salmonella serotype and serotype prevalence varied from farm to farm and within farm from one sampling time to another. Multiple Salmonella genotypes were detected for a number of serotypes and identical genotypes were found on different farms with one genotype of Salmonella Senftenberg identified on three of the four farms. Significance and Impact of the STUDY: This study demonstrated the wide variability in pathogen shedding within and among dairy farms all located in a small geographical region and highlights the complexity of pathogen control at the farm level.     

Three-Dimensional (3D) cell culture monitoring: Opportunities and challenges for impedance spectroscopy

Three-dimensional (3D) cell culture has developed rapidly over the past 5–10 years with the goal of better replicating human physiology and tissue complexity in the laboratory. Quantifying cellular responses is fundamental in understanding how cells and tissues respond during their growth cycle and in response to external stimuli. There is a need to develop and validate tools that can give insight into cell number, viability, and distribution in real-time, nondestructively and without the use of stains or other labelling processes. Impedance spectroscopy can address all of these challenges and is currently used both commercially and in academic laboratories to measure cellular processes in 2D cell culture systems. However, its use in 3D cultures is not straight forward due to the complexity of the electrical circuit model of 3D tissues. In addition, there are challenges in the design and integration of electrodes within 3D cell culture systems. Researchers have used a range of strategies to implement impedance spectroscopy in 3D systems. This review examines electrode design, integration, and outcomes of a range of impedance spectroscopy studies and multiparametric systems relevant to 3D cell cultures. While these systems provide whole culture data, impedance tomography approaches have shown how this technique can be used to achieve spatial resolution. This review demonstrates how impedance spectroscopy and tomography can be used to provide real-time sensing in 3D cell cultures, but challenges remain in integrating electrodes without affecting cell culture functionality. If these challenges can be addressed and more realistic electrical models for 3D tissues developed, the implementation of impedance-based systems will be able to provide real-time, quantitative tracking of 3D cell culture systems.     

N-alkanes v. ytterbium/faecal index as two methods for estimating herbage intake of dairy cows fed on diets differing in the herbage: maize silage ratio and feeding level

The aim of this study was to compare the n-alkanes and the ytterbium (Yb)/faecal index techniques as two methods for estimating the herbage intake of dairy cows fed indoors on different herbage : supplement ratios and feeding levels. The supplement was a mixture of maize silage and soyabean meal (ratio of 87 : 13 on a dry matter (DM) basis). In all, four treatments were studied. The herbage : supplement ratio in the diet was 25 : 75, 50 : 50, 75 : 25 and 50 : 50 for treatments 1, 2, 3 and 4, respectively. Animals were offered for treatments 1, 2 and 3, 100% of ad libitum intake measured before the experiment and 70% of ad libitum intake for treatment 4. Cows were fed herbage in the morning and supplement in the evening. A total of six lactating Holstein dairy cows were used in a 4 × 4 Latin square with four 14-day periods. Herbage and supplement intakes, faecal output (FO), in vivo organic matter (OM) digestibility and faecal recovery of markers were measured on the last 5 days of each period. Intake was estimated with the two methods and from two faecal sampling techniques, that is, total faecal collection v. grab sampling during milking. Mean herbage intake as fed, or estimated from n-alkanes or from the Yb/faecal index was 7.7, 8.1 and 10.2 kg DM, respectively. The mean prediction error, expressed as a fraction of actual herbage intake, was 0.10 and 0.50 for the n-alkanes and Yb/faecal index methods, respectively. The n-alkanes method clearly showed much better accuracy than the Yb/faecal index method for estimating intake, irrespective of the faecal sampling method, herbage : silage proportion or feeding level. For the n-alkanes method, herbage intake was slightly overestimated (7%) when herbage proportion in the diet was high, due to a ratio of faecal C33 : C32 recovery >1. The high bias for the Yb/faecal index was due to the cumulative effect of overestimation of FO (mean recovery of Yb = 0.92) and underestimation of the diet indigestible fraction (-8%). Between-treatment variations of FO were on average well estimated by Yb. Between-treatment variations of OM digestibility estimated using the faecal index technique were lower than those observed in vivo. It is concluded that intake of grazing dairy cows receiving high levels of maize silage supplement should be estimated using the n-alkanes method.