The main goal of this study was to develop, apply, and validate a new method to predict an indicator for CH4 eructed by dairy cows using milk mid-infrared (MIR) spectra. A novel feature of this model was the consideration of lactation stage to reflect changes in the metabolic status of the cow. A total of 446 daily CH4 measurements were obtained using the SF6 method on 142 Jersey, Holstein, and Holstein-Jersey cows. The corresponding milk samples were collected during these CH4 measurements and were analyzed using MIR spectroscopy. A first derivative was applied to the milk MIR spectra. To validate the novel calibration equation incorporating days in milk (DIM), 2 calibration processes were developed: the first was based only on CH4 measurements and milk MIR spectra (independent of lactation stage; ILS); the second included milk MIR spectra and DIM information (dependent on lactation stage; DLS) by using linear and quadratic modified Legendre polynomials. The coefficients of determination of ILS and DLS equations were 0.77 and 0.75, respectively, with standard error of calibration of 63g/d of CH4 for both calibration equations. These equations were applied to 1,674,763 milk MIR spectra from Holstein cows in the first 3 parities and between 5 and 365 DIM. The average CH4 indicators were 428, 444, and 448g/d by ILS and 444, 467, and 471g/d by DLS for cows in first, second, and third lactation, respectively. Behavior of the DLS indicator throughout the lactations was in agreement with the literature with values increasing between 0 and 100 DIM and decreasing thereafter. Conversely, the ILS indicator of CH4 emission decreased at the beginning of the lactation and increased until the end of the lactation, which differs from the literature. Therefore, the DLS indicator seems to better reflect biological processes that drive CH4 emissions than the ILS indicator. The ILS and DLS equations were applied to an independent data set, which included 59 respiration chamber measurements of CH4 obtained from animals of a different breed across a different production system. Results indicated that the DLS equation was much more robust than the ILS equation allowing development of indicators of CH4 emissions by dairy cows. Integration of DIM information into the prediction equation was found to be a good strategy to obtain biologically meaningful CH4 values from lactating cows by accounting for biological changes that occur throughout the lactation.

This study investigates the feasibility to predict individual methane (CH 4 ) emissions from dairy cows using milk mid-infrared (MIR) spectra. To have a large variability of milk composition, two experiments were conducted on 11 lactating Holstein cows (two primiparous and nine multiparous). The first experiment aimed to induce a large variation in CH 4 emission by feeding two different diets: the first one was mainly composed of fresh grass and sugar beet pulp and the second one of maize silage and hay. The second experiment consisted of grass and corn silage with cracked corn, soybean meal and dried pulp. For each milking period, the milk yields were recorded twice daily and a milk sample of 50 ml was collected from each cow and analyzed by MIR spectrometry. Individual CH 4 emissions were measured daily using the sulfur hexafluoride method during a 7-day period. CH 4 daily emissions ranged from 10.2 to 47.1 g CH 4 /kg of milk. The spectral data were transformed to represent an average daily milk spectrum (AMS), which was related to the recorded daily CH 4 data. By assuming a delay before the production of fermentation products in the rumen and their use to produce milk components, five different calculations were used: AMS at days 0, 0.5, 1, 1.5 and 2 compared with the CH 4 measurement. The equations were built using Partial Least Squares regression. From the calculated R 2 cv , it appears that the accuracy of CH 4 prediction by MIR changed in function of the milking days. In our experimental conditions, the AMS at day 1.5 compared with the measure of CH 4 emissions gave the best results. The R 2 and s.e. of the cross-validation were equal to 0.79 and 5.14 g of CH 4 /kg of milk. The multiple correlation analysis performed in this study showed the existence of a close relationship between milk fatty acid (FA) profile and CH 4 emission at day 1.5. The lower R 2 (R 2 5 0.76) obtained between FA profile and CH 4 emission compared with the one corresponding to the obtained calibration (R 2 c 5 0.87) shows the interest to apply directly the developed CH 4 equation instead of the use of correlations between FA and CH 4 . In conclusion, our preliminary results suggest the feasibility of direct CH 4 prediction from milk MIR spectra. Additional research has the potential to improve the calibrations even further. This alternative method could be useful to predict the individual CH 4 emissions at farm level or at the regional scale and it also could be used to identify low-CH 4 -emitting cows.

Abstract. Mitigating the proportion of energy intake lost as methane could improve the sustainability and profitability of dairy production. As widespread measurement of methane emissions is precluded by current in vivo methods, the development of an easily measured proxy is desirable. An equation has been developed to predict methane from the mid-infrared (MIR) spectra of milk within routine milk-recording programs. The main goals of this study were to improve the prediction equation for methane emissions from milk MIR spectra and to illustrate its already available usefulness as a high throughput phenotypic screening tool. A total of 532 methane measurements considered as reference data (430 AE 129 g of methane/day) linked with milk MIR spectra were obtained from 165 cows using the SF 6 technique. A first derivative was applied to the MIR spectra. Constant (P0), linear (P1) and quadratic (P2) modified Legendre polynomials were computed from each cows stage of lactation (days in milk), at the day of SF 6 methane measurement. The calibration model was developed using a modified partial least-squares regression on first derivative MIR data points · P0, first derivative MIR data points · P1, and first derivative MIR data points · P2 as variables. The MIR-predicted methane emissions (g/day) showed a calibration coefficient of determination of 0.74, a cross-validation coefficient of determination of 0.70 and a standard error of calibration of 66 g/day. When applied to milk MIR spectra recorded in the Walloon Region of Belgium (%2 000 000 records), this equation was useful to study lactational, annual, seasonal, and regional methane emissions. We conclude that milk MIR spectra has potential to be used to conduct high throughput screening of lactating dairy cattle for methane emissions. The data generated enable monitoring of methane emissions and production characteristics across and within herds. Milk MIR spectra could now be used for widespread screening of dairy herds in order to develop management and genetic selection tools to reduce methane emissions.

The challenge of managing and breeding dairy cows is permanently adapting to changing production circumstances under socio-economic constraints. If managing and breeding address different timeframes of action, both need relevant phenotypes that allow for precise monitoring of the status of the cows, and their health, behavior, and well-being as well as their environmental impact and the quality of their products (i.e., milk and subsequently dairy products). Milk composition has been identified as an important source of information because it could reflect, at least partially, all these elements. Major conventional milk components such as fat, protein, urea, and lactose contents are routinely predicted by mid-infrared (MIR) spectrometry and have been widely used for these purposes. But, milk composition is much more complex and other nonconventional milk components, potentially predicted by MIR, might be informative. Such new milk-based phenotypes should be considered given that they are cheap, rapidly obtained, usable on a large scale, robust, and reliable. In a first approach, new phenotypes can be predicted from MIR spectra using techniques based on classical prediction equations. This method was used successfully for many novel traits (e.g., fatty acids, lactoferrin, minerals, milk technological properties, citrate) that can be then useful for management and breeding purposes. An innovation was to consider the longitudinal nature of the relationship between the trait of interest and the MIR spectra (e.g., to predict methane from MIR). By avoiding intermediate steps, prediction errors can be minimized when traits of interest (e.g., methane, energy balance, ketosis) are predicted directly from MIR spectra. In a second approach, research is ongoing to detect and exploit patterns in an innovative manner, by comparing observed with expected MIR spectra directly (e.g., pregnancy). All of these traits can then be used to define best practices, adjust feeding and health management, improve animal welfare, improve milk quality, and mitigate environmental impact. Under the condition that MIR data are available on a large scale, phenotypes for these traits will allow genetic and genomic evaluations. Introduction of novel traits into the breeding objectives will need additional research to clarify socio-economic weights and genetic correlations with other traits of interest.

Unbalanced metabolic status in the weeks after calving predisposes dairy cows to metabolic and infectious diseases. Blood glucose, IGF-I, non-esterified fatty acids (NEFA) and β-hydroxybutyrate (BHB) are used as indicators of the metabolic status of cows. This work aims to (1) evaluate the potential of milk mid-IR spectra to predict these blood components individually and (2) to evaluate the possibility of predicting the metabolic status of cows based on the clustering of these blood components. Blood samples were collected from 241 Holstein cows on six experimental farms, at days 14 and 35 after calving. Blood samples were analyzed by reference analysis and metabolic status was defined by k-means clustering (k=3) based on the four blood components. Milk mid-IR analyses were undertaken on different instruments and the spectra were harmonized into a common standardized format. Quantitative models predicting blood components were developed using partial least squares regression and discriminant models aiming to differentiate the metabolic status were developed with partial least squares discriminant analysis. Cross-validations were performed for both quantitative and discriminant models using four subsets randomly constituted. Blood glucose, IGF-I, NEFA and BHB were predicted with respective R 2 of calibration of 0.55, 0.69, 0.49 and 0.77, and R 2 of cross-validation of 0.44, 0.61, 0.39 and 0.70. Although these models were not able to provide precise quantitative values, they allow for screening of individual milk samples for high or low values. The clustering methodology led to the sharing out of the data set into three groups of cows representing healthy, moderately impacted and imbalanced metabolic status. The discriminant models allow to fairly classify the three groups, with a global percentage of correct classification up to 74%. When discriminating the cows with imbalanced metabolic status from cows with healthy and moderately impacted metabolic status, the models were able to distinguish imbalanced group with a global percentage of correct classification up to 92%. The performances were satisfactory considering the variables are not present in milk, and consequently predicted indirectly. This work showed the potential of milk mid-IR analysis to provide new metabolic status indicators based on individual blood components or a combination of these variables into a global status. Models have been developed within a standardized spectral format, and although robustness should preferably be improved with additional data integrating different geographic regions, diets and breeds, they constitute rapid, cost-effective and large-scale tools for management and breeding of dairy cows.