Final report for SW22-932
Project Information
For the dairy industry to be financially sustainable, management practices need to meet consumer expectations. Some traditional dairy practices, like calf housing and the use of antibiotics, are the subject of consumer criticism. This project will improve the profitability, sustainability, and quality of life for dairies by identifying ways in which animal welfare and the development of a beneficial gastrointestinal (GI) microbiome can be optimized for dairy calves and brought into line with modern consumer preferences.
This project will investigate how husbandry practices can influence neonatal calf welfare, nutrition and development of the gastrointestinal microbiota, and how these factors contribute to healthy, active and emotionally developed calves that will grow to become productive dairy cows. Recent research has demonstrated how important it is to establish a healthy microbiome early in life and the welfare benefits of paired or group-housed calves, including improved cognitive development, reduced fear responses, and increased curiosity to novel items. In this study, we will investigate the impacts that freezing colostrum has on the structure and function of the GI microbiome and will also determine the effects of paired housing on the development of a healthy microbiome and a healthy calf. We hypothesize that fresh colostrum and paired housing of calves will facilitate the establishment of a healthy GI microbiome and the development of healthy calves, which may lead to more sustainable management practices.
The impacts of feeding frozen or fresh colostrum and housing calves individually or in pairs on welfare and biological functions will be determined. By partnering with commercial dairies and Extension experts, our results will be directly applicable to working dairies and will give them multiple specific tools to develop sustainable calf housing and management systems that meet modern standards of welfare and consumer preference.
Research objectives:
- Characterize the healthy GI microbiome early in life and describe how freezing colostrum and paired housing impact the structure and function of the GI microbiome
- Define welfare benefits of paired or group-housed calves, including improved cognitive development, reduced fear responses, and increased curiosity to novel items.
- Determine if interactions exist between colostrum, social versus single housing, microbiome structure and function, and welfare in dairy calves
Education objectives:
- Educate producers about the importance of social housing and colostrum management for optimal development and long-term productivity of dairy calves, including microbiome differences between calves raised under different colostrum management practices and the effects of paired housing on calf behavior and welfare
Phase 1:
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Days |
Samples/Observations |
|
Colostrum |
0 |
Colostrum samples from each farm and colostrum type will be collected daily for microbiome determination |
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|
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Phase 2:
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|
Days |
Samples/Observations |
|
Colostrum |
1 |
Colostrum samples from each farm and colostrum type will be collected for microbiome determination and total solids/proteins Calves fed colostrum |
|
Calves |
1,3,7,14,30,60 |
Weight, average daily gain, fecal samples from calves will be collected to determine microbiome changes due to treatments Hair clips for hormone determination (cortisol, dehydroepiandrosterone (DHEA), Triiodothyronine (T3)) |
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1,3,7,14,30,60 |
Feces for microbial community profiles |
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|
3 |
Calf total serum proteins and serum IgG |
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4,9,14 |
Calf behavior video recording |
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30 |
Assessment of ruminal development by the presence of rumination |
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daily |
Feed consumption, morbidity scores, mortality, health treatments (type and duration) |
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For both phases, microbiome determination includes an assessment of the types and functions of microbes present in active microbial populations, including pathogen relative abundance, antibiotic-resistance gene types and prevalence.
Phase 3:
After the samples are processed and the findings summarized, during the third year of the study an educational program for dairy producers will be developed and implemented to share our findings about the importance of colostrum management to the development of healthy GI microbiome.
Extension efforts will include newsletter articles and regional workshops to present research results to dairy producers, calf caretakers and dairy allied industry. Website and video material will be produced to demonstrate the feasibility of research-based management practices supported by this project. Methods are detailed in the Education section, below.
Cooperators
- - Producer
- - Producer
- - Producer
Research
Objectives
Phase 1: Characterize and compare viable microbiome in fresh and frozen colostrum from conventional and organic dairies
- determination of types and functions of microbes present in living microbial populations,
- relative abundance of pathogens
- antimicrobial-resistance gene types and prevalence
- statistical comparisons by treatment type
Phase 2: Compare the effects of feeding the two most contrasting colostrum types to calves in paired- versus single-housing
- Colostrum quality
- Calf health
- Weight, feed consumption and average daily weight gain
- Total Serum Proteins
- Morbidity and mortality scores
- Type and duration of treatments
- Calf welfare
- Non-invasive endocrinological analysis of welfare hormones
- Behavioral observations
- Characterization of the GI microbiome
- Microbial community profiles
- Census of bacterial taxa (phyla, family, genera) in each group
- Statistical comparisons of specific taxa & diversity metrics
- Definition of normal/healthy flora based on correlations with health outcomes
Phase 3: Develop and implement educational program regarding importance of colostrum management, development of healthy GI microbiome, and benefits of proper social development
- The project team, including producers and led by extension representative Betsy Karle, will conduct webinars, field days, workshops on-farm and on-campus, and distribute materials such as fact sheets to dairy stakeholders.
- Evaluation of outreach program effectiveness will be conducted with standard program evaluation criteria guided by logic model and tools such as pre/post surveys of producer attitudes and willingness to implement or suggest changes to housing or colostrum management. The primary goal is to increase knowledge of educational program participants and increase the dairy producer intent to adopt research-based management practices. Actual adoption of practices in this study will be measured on the collaborating farms.
Materials and Methods
The study was conducted on 20 partnering commercial dairies. Five of these producer-partner farms follow organic practices and the remainder are conventional dairies. The study included two research phases followed by a third phase dedicated to an educational plan designed to increase knowledge regarding the importance of responsible antibiotic use, calf social development, and the development and management of a healthy GI microbiome. Research results have been shared via multiple technical and non-technical presentations to various audiences as detailed below. Results and recommendations from this project will continue to be publicized to the dairy farming and food animal medicine communities via outreach and technical publications.
For Phase 1, we used a 2 x 2 study design: colostrum was collected from two farm types (organic or conventional) and analyzed as fresh or frozen. A total of 228 colostrum samples were collected from conventional farms and 116 from organic farms. Fresh samples were immediately transported back to the lab and split into two aliquots - one immediately processed and one frozen. To measure the microbial load in all samples, two methods were used - aerobic incubations with 3M Petri Film in a most-probable number (MPN) approach, and a previously validated quantitative-PCR assay targeting the 16S rRNA genes of viable cells. To characterize the microbial communities in each sample, a standard 16S rRNA amplicon sequencing approach was used with barcoded PCRs performed in our lab and raw sequence data generated by the UC Riverside Genomics Core using the MiSeq PE300 chemistry.
Bacterial isolates (i.e. axenic cultures of individual strains) were also obtained from a random subset of 20 organic and 60 conventional colostrum samples. Cultures were streaked to purity, and strains were archived as glycerol stocks stored at -80 deg C. identified taxonomically via the DNA sequence of the 16S rRNA gene. Students subsequently performed a literature review for each strain to determine what evidence existed for it to be classified as a pathogen, commensal, or potential probiotic strain.
The second phase of the project was originally planned to include two treatments in a 2 x 2 study: Colostrum (fresh or frozen) and Housing (paired or individual) on four farms. Due to the outbreak of Highly Pathogenic Avian Influenza (HPAI) that affected dairy cows, our access to dairies was severely limited but we were able to collect samples from two farms. With the help of five DVM students employed by the project, we collected samples from 44 calves at a conventional dairy in California and 40 calves at an organic dairy in Colorado over the course of 60 days. The following sample types were collected: colostrum (day 1), fecal samples (days 1, 3, 7, 14, 30, 60), hair (days 1, 14), blood (day 3), body weight (days 1, 14). Respiratory scores, fecal scores, and milk consumption were also monitored daily for days 1-14. Colostrum and fecal samples were shipped on ice to the lab immediately after collection. Video recordings of calf behavior in the hutches were made at days 4, 9, and 14.
Data collected from each sample type was as follows: Colostrum and Fecal samples were used to assess total microbial load and microbiome analysis via aerobic culturing, qPCR, and DNA sequencing as described above for phase 1. Hair samples were analyzed for cortisol concentration. Hair cortisol concentrations, as well as from other steroid hormones linked to welfare, reproduction and resilience or metabolism, allow monitoring long-term retrospective levels of the circulating hormone. Steroid concentrations in hair are a potential tool for dairy cattle welfare and production research by providing a useful and practical tool for long-term steroid monitoring. The number of sample collected for analysis was: hair cortisol (n=165), colostrum (n=96) and fecal (n=457).
To analyze the video data, a standardized ethogram was developed to define behaviors, ensuring consistency across observers. Behavioral data were extracted and organized into individual tracking sheets, and videos were subsequently analyzed using an open source AI software package called LabGym for automated behavior identification and quantification. We also were able to obtain expert manual annotation of calf behaviors for all of our video footage.
For the 3rd phase of our project, research results were used to develop and implement educational materials regarding the importance of colostrum management, development of healthy GI microbiome, and benefits of proper social development.
One full-time employee and three DVM students were hired in 2023 to do field work and lab work. Samples were collected from 20 dairies (five organic and 15 conventional). In 2024, one employee continued at full-time and five DVM students were hired for the summer to do field work. In 2025, one employee continued at full-time, two DVM students worked part-time during the academic year, and one DVM student worked full-time during the summer. One undergraduate and one Masters student from Cal Poly Pomona and an undergraduate student from UC Davis were also partially supported to do lab work and data analysis during 2024-2025. One part-time employee, supported by the WesternU CVM Research Office, also contributed to the project.
The results obtained for each project objective are shown below.
Determination of types and functions of microbes present in living microbial populations
Prior to collecting samples for Phase 1, we collected colostrum samples from several local dairies, and first optimized and validated DNA extraction methods and the viability quantitative-PCR protocols. Following these methodological validation steps, we collected fresh colostrum samples from 20 dairies (15 conventional and 5 organic) and compared fresh to frozen paired aliquots of each sample as described above. We spent several months optimizing our barcoded PCR protocols and submitted all of these samples (346 total including fresh vs frozen) as proposed for high-throughput DNA sequencing (HTS) in April 2024 for Phase 1. For Phase 2, we finished all of our lab work in October of 2025 and submitted all of these samples (553 colostrum and fecal samples) as proposed for high-throughput DNA sequencing (HTS) in October 2025. This 2nd set of sequence data is still being analyzed.
Colostrum Quality Differences Between Organic and Conventional Production Systems
We found that colostrum from organic dairies had higher Brix values than conventional dairy colostrum (mean of 27 versus 22) and that these differences were significant according to a pairwise t-test (p<0.001).
Colostrum Microbiome Differences Between Organic and Conventional Production Systems
We found no significant differences in alpha-diversity between the two production systems as measured by the observed number of microbial sequence types and Shannon Diversity Index. However, beta-diversity analyses showed that there were statistically significant (PERMANOVA, p<0.0001) and clearly distinct microbial "fingerprints" for each farm and that the bacterial communities from organic farms clustered together and were distinct from those of conventional farms.
From our work to build and characterize a strain library, the most common sequence type in colostrum from conventional dairies that was identified from 16S rRNA gene sequence data was most closely-related to Staphylococcus gallinarum. In organic colostrum, strains most closely-related to S. aureus and Lactococcus lactis were most common.
Our preliminary results indicate that potentially beneficial strains (identified from 16S rRNA gene sequence data as Bacillus aerius, Bacillus valezensis, and Lactococcus lactis) were only found in samples from organic dairies.
We also adopted and used a Mycoplasma bovis-specific PCR assay to screen a subset of colostrum samples for this important pathogen. All samples screened were negative.
Effects of Freezing on Colostrum Microbiome
Our high-throughput DNA sequencing showed that freezing of colostrum at -20 deg C for periods up to 144 hours (6 days) did not have a significant effect on the taxonomic composition of the colostrum microbiome. Beta-diversity analyses showed that there were no statistically significant effects of freezing on the composition and relative abundance of colostrum microbes. We also measured the effects of freezing on the number of viable cells using two independent methods - aerobic cultivation and viability quantitative-PCR, and found no significant declines in the number of viable microbes with freezing. Viable cell counts remained between 10^5 - 10^7 CFU per mL of colostrum across all timepoints as measured by both methods.
Effects of Housing Systems on Stress Physiology, Microbiome, and Behavior
We found that hair cortisol was not correlated with fecal microbial load at day 1 or day 14 (Spearman's ρ, p<0.05), hair cortisol was not correlated with colostrum quality at day 1 or day 14 (Spearman's ρ, p<0.05) or serum total protein at day 1 or day 14 (Spearman's ρ, p<0.05).
Both individually-housed and group-housed calves showed a significant cortisol decrease from day 1 to day 14 (Kruskal-Wallis, p<0.05; Dunn's test, α=0.05), with group-housed calves having a larger magnitude of cortisol reduction compared to individually-housed calves.
We also found a rapid establishment and stabilization of the fecal microbiome by day 3 of life. This was determined both by a molecular assessment (qPCR) which showed that microbial load increased from day 1 to day 3, then plateaued through day 60 and by a cultivation-based assessment (3M Petrifilm) which gave concordant results showing stabilization of microbial load by day 3. Analyses of both data sets with Kruskal-Wallis and Dunn's post-hoc tests (α=0.05) confirmed significant differences between day 1-3, but not thereafter.
For the analysis of calf behaviors, the automated AI software LabGym was able to successfully identify postural behaviors (i.e. laying, standing), while more active behaviors (i.e. locomotion, nursing) were also detected, but at lower accuracy. These analyses are still ongoing and represent one of the first applications of LabGym for the automated analysis of calf behavior. We intend to use the expert-curated manual annotation of calf behaviors from our footage as a 'gold-standard' to compare to the AI-based automatic annotation for behavior analysis.
Research outcomes
This research provides evidence-based recommendations for sustainable dairy production in the Western United States by demonstrating that: (1) organic production systems yield superior colostrum quality without requiring frozen storage compromises, (2) group housing reduces chronic stress while maintaining herd health, and (3) early-life microbiome establishment follows predictable trajectories that can be managed through targeted interventions. These findings directly support economically viable, welfare-forward practices that reduce reliance on therapeutic antimicrobials, improve calf survival and performance, and align with consumer demand for ethical animal agriculture.
Discuss the application of your results to sustainable agricultural production in the Western U.S. Describe and assess how your project has affected agricultural sustainability or will contribute to future sustainability.
Our results provide support for group housing as a welfare-promoting and economically viable practice. Individual hutch housing dominates Western U.S. calf-rearing (60-70% of operations per USDA NAHMS) justified by disease control, ease of monitoring, and perceived cost-effectiveness. However, consumer welfare concerns, retailer requirements (e.g., Costco), and regulatory trends (EU ban on individual housing <8 weeks) are pressuring U.S. producers to adapt. Our result that group-housed calves had a larger magnitude of cortisol reduction (from day 1 to day 14) compared to individually-housed calves, may support a social buffering hypothesis and if this result is borne out by future research, could lead to significant improvements in welfare. Lower cortisol predicts improved immune function, reduced disease susceptibility, and thereby enhanced welfare. Group-housed calves have also been shown to exhibit improved social skills, reduced fear responses, and better adaptation to group housing post-weaning (De Paula Vieira et al. 2010). Finally, group housing is aligned with the "freedom to express normal behavior" via social interaction.
Importantly, our results showed correlation between housing and microbial load (i.e. the fecal microbiome was established equally in both systems), supporting the conclusion that group housing does not increase disease risk when properly managed
Recommendations for sustainable agricultural production and future research
Based on the results of this project, future research could focus on:
- Forage quality and diversity: Organic dairies often utilize pasture grazing and diverse forage programs; polyphenol-rich diets may enhance mammary immune function and colostrum IgG concentration. Such studies could help understand mechanisms contributing to the significantly higher Brix values observed on organic farms by our study.
- Dry period management: Organic systems may employ longer dry periods, optimizing colostrum synthesis.
- Stress reduction: Lower stocking densities and outdoor access in organic systems may reduce prepartum cortisol, enhancing colostrum quality
- Microbiome diversity: Our data show farm-specific colostrum microbiome signatures, suggesting that organic farms may harbor more diverse microbial consortia due to environmental exposures or other factors.
Education and Outreach
Participation summary:
Research results have been shared with industry partners (the collaborating dairies and the broader community) by UCANR extension sharing of our results via newsletter articles for Dairymen, and posting to Extension websites and social media accounts.
https://ucanr.edu/sites/default/files/2022-05/Dairy_Newsletter93311.pdf
Specific educational materials (Fact Sheets) were prepared by our project team to translate our research results to the broader community. The Fact Sheets included in this report have been shared as described with one example shown here:
https://www.facebook.com/uccedairy/
These Fact Sheets are also attached to this Report.
One full-time employee and three DVM students were hired in 2023 to do field work and lab work. In 2024, one employee continued at full-time and five DVM students were hired for the summer to do field work. In 2025, one employee continued at full-time, two DVM students worked part-time during the academic year, and one DVM student worked full-time during the summer. One undergraduate and one Masters student from Cal Poly Pomona and an undergraduate student from UC Davis were also partially supported to do lab work and data analysis during 2024-2025. One part-time employee, supported by the WesternU CVM Research Office, also contributed to the project.
A total of 14 students were hired by this project with three poster presentations and one oral presentation. The places of presentation, titles, and authorship of each poster is listed below. Student names are shown in bold with student presenting author starred.
- National Veterinary Scholars Symposium. 2024. Puerto Rico. Comparison of Microbial Differences in Fresh and Frozen Colostrum from California Dairy Farms. Victoria Ngo*, Allison Warnick, Samantha Lund, Sarah Warda, Brian B. Oakley, James Reynolds, Jose M. Peralta.
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National Veterinary Scholars Symposium. 2025. Spokane, WA. Effects of colostrum and housing on health and welfare of calves in conventional and organic dairy farms. Lyssabeth Lorico*, Emma Felde-Medina, Christopher Haggard, Kevin Lin, Brooke St. George, Sarah Warda, Hoangvi Le, Madison Miranda, Jadeanna Martinez, Lauren Cheng, Isabella Fajardo, Jose Peralta, James Reynolds, Brian B. Oakley
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Cal Poly Pomona Undergraduate Research Day. 2025. Evaluating the Effects of Calf Husbandry and Colostrum Freezing on the Gastrointestinal Microbiome and Calf Health. Jadeanna Martinez*, Lyssabeth Lorico, Emma Felde-Medina, Christopher Haggard, Kevin Lin, Brooke St. George, Sarah Warda, Hoangvi Le, Madison Miranda, Jose Peralta, Jim Reynolds, Brian B. Oakley
- Western University of Health Sciences Research Day. 2025. Identification and Quantification of Calf Behavior Using LabGym Artificial Intelligence. Lauren Cheng*, Lyssabeth Lorico, Emma Felde-Medina, Christopher Haggard, Kevin Lin, Brooke St. George, Sarah Warda, Hoangvi Le, Jose Peralta, Jim Reynolds, Brian B. Oakley. Podium presentation, winner of best presentation by a non-DVM student.
Education and Outreach Outcomes
We recommend and plan to continue sharing these results. The main recommendations as stated in the attached Fact Sheet are:
- Freezing colostrum for up to 6 days does not significantly reduce the number or variety of bacteria in it — it remains a safe storage practice
- Your farm environment is the main factor shaping the microbial community in your colostrum — not whether you freeze it
- Always check colostrum quality with a Brix refractometer (≥22% = good) before the first calf feeding
- Higher-quality colostrum directly translates to better immune protection for calves in the first days of life
- Consider allowing calves to have social contact — group or pair housing measurably reduces chronic stress
This portion of the project was somewhat impacted by changing job roles and retirements, but in addition to the materials already produced, Betsy Karle and Dr. Oakley still plan to produce at least one more newsletter article and/or a webinar or workshop to be shared as education and outreach activities in the near future.
- Microbiological understanding, and evidence-based calf-rearing strategies grounded in current research