Final Report for ONE11-133

Project Type: Partnership
Funds awarded in 2011: $14,445.00
Projected End Date: 12/31/2013
Region: Northeast
State: New York
Project Leader:
Dr. Michele Barrett, DVM
Keseca Veterinary Clinic
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Project Information


In 2010, several New York herds with elevated bulk tank somatic cell counts were experiencing mastitis cases that were unresponsive to treatment. Lactococcus lactis subspecies(ssp.) lactis was identified via molecular identification via PCR technique at Quality Milk Production Services(Ithaca, NY) as the pathogen causing many of these non-responding infections. At that time, it was unclear if there was a true increase in incidence of Lactococcal mastitis in these herds or if this was simply a result of more accurate testing techniques.

To further characterize this pathogen and its presence in local farms, clear and specific identification from milk samples was required, but molecular testing was perceived as cost-prohibitive by the affected herds. This study evaluated the accuracy of a less expensive test, the API 20 Strep test (bioMerieux), for identifying Lactococcus lactis (ssp.) lactis from bovine milk samples compared to molecular identification at Quality Milk Production Services. Out of the 73 isolates analyzed by both tests methods, 24 were molecularly identified as Lactococcus lactis ssp. lactis. Comparison of the API 20 Strep results at 4 hours and 24 hours post-incubation to molecular identification indicated that the API 20 Strep system is an accurate alternative to molecular testing only when read at 24 hours using the “low discrimination” cutoff. At this cut point, Sensitivity =76.2% (95%CI= 58.0-94.4), Specificity=100% (95%CI=100-100), Negative Predictive Value= 88.6% (95%CI=79.3-98), Positive Predictive Value=100% (95%CI=100-100), and test agreement was high (k=0.81). A 12-page herd survey was completed by both affected and non-affected farms in an effort to assess farm-level risk factors of Lactococcal mastitis, but statistical analysis was unrewarding since the 24 Lactococcus isolates identified represented only five different farms. An antimicrobial susceptibility library was compiled using Minimum Inhibitory Concentration testing on the 24 milk isolates and 8 total environmental isolates from the five affected farms.

Based on the above results and standard mastitis control programs, farms identified as having intramammary infections due to Lactococcus lactis ssp. lactis were instructed to identify and segregate infected animals from non-infected animals, focus on improving management of sand bedded stalls, and treat infected animals based on susceptibility patterns while the pathogen continues to be investigated. In addition to these guidelines, this study has confirmed the accuracy (under specific conditions) of a widely accessible and relatively inexpensive test to aid practitioners in identifying the disease (API 20 Strep system), provided MIC data to aid in proper treatment decisions and to spur potential field trials, and identified at least one potential environmental source of Lactococcus on dairy farms, sand bedding, with the intention of helping individually affected farms manage the disease and increase profitability.


Mastitis is one of the most costly health concerns affecting dairy farmers in the United States, accounting for approximately 1.8 billion in industry losses annually (Schroeder, 2012). Current estimates show that farmers lose approximately $200 per cow each year battling mastitis, due to reduced milk production, treatment costs, discarded milk, labor costs, and cost of replacements (Smith and Hogan, 2001). Applying these estimates to the New York dairy industry, which includes 610,000 dairy cows (USDA-NASS, 2012), the economic losses due to mastitis amount to approximately $122,000,000 annually. In addition to these direct losses, mastitis also affects milk quality of milk produced, reducing a farmer’s opportunity for milk quality premiums from cooperatives. With growing consumer concern about farming practices, as well as pressure for higher milk quality standards, farmers will need to control mastitis in order to sustain their viability in national and international markets.

In 2010, several New York herds were experiencing problems with mastitis cases that were not responding well to treatment and high bulk tank somatic cell counts. Through molecular testing at Quality Milk Production Services, several of these poorly responding mastitis cases were identified as Lactococcus lactis ssp. lactis. While Lactococcus had previously been isolated from bovine mammary glands and clinical mastitis cases, it was unclear in 2010 whether the number of cases observed indicated an increase in disease incidence and growing emergence as a pathogen, or if newer molecular testing was identifying Lactococcal infections that would have been grouped in a simpler classification system of Streptococcus species in the past. The latter was certainly possible, since Lactococci are often misidentified as other Streptococci-like bacteria based on traditional bench-top techniques (Gordoncillo et al, 2010). To understand the behavior and impact of Lactococcus lactis ssp. lactis as a mastitis pathogen, it was clear that more specific identification would be needed to identify and study the disease.

However, from the perspective of the affected farms, use of the molecular test was still perceived as cost prohibitive. Realizing there was a need in the local farming community for more information on this pathogen, the objective of this study was evaluate a more affordable and faster method of identifying the disease (API 20 Strep), with the intent of using that information to lay the foundation for investigation of disease risk factors and potential treatment protocols. Overall, this study sought to create a foundation of knowledge concerning Lactococcal mastitis in hopes of assisting farmers both regionally and nationwide in identifying and controlling Lactococcus as a pathogen, thereby minimizing its impact on farm sustainability.

References Cited:
Gordoncillo, M. J. N., J. A. N. Bautista, M. Hikiba, I. G. Sarmago, and J. M. B. Haguingan. 2010. Comparison of conventionally identified mastitis bacterial organisms with commercially available microbial identification kit (BBL Crystal ID®). Phil. J. Vet. Med. 47(1):54-57.

Schroeder, J.W. Mastitis control programs: bovine mastitis and milking management. North Dakota State University Extension Service Bulletin. Fargo, North Dakota. July 2012.

Smith K, and J. Hogan . The world of mastitis. Proc. 2nd Int. Symp. National Mastitis Council, Madison WI: Mastitis Milk Quality, Vancouver, Canada; 2001; Pages 1–12

United States Department of Agriculture-National Agricultural Statistics Service. New York Crop and Livestock Report. Livestock_Report/2012/nycl0212.pdf. Feb 2012.

Project Objectives:

The primary goal of this study was to expand the breadth of knowledge about Lactococcous lactis subspecies lactis isolated from bovine milk samples in order to provide dairy farmers and veterinarians with practical knowledge to prevent, identify, and control the disease. This goal was to be achieved through three main objectives:

  1. Evaluation of the accuracy of the API 20 Strep system for identification of Lactococcus lactis ssp. lactis from bovine milk samples compared to molecular identification technique. Analysis of farm-level risk factors for disease based on positive Lactococus lactis ssp lactis identifications from Part 1, using on-farm records systems and a written herd survey. Compilation of an antimicrobial susceptibility database via Minimum Inhibitory Concentration (MIC) testing on Lactococus lactis ssp lactis isolates identified from milk samples and environmental sampling to guide treatment decisions and provide a basis for future treatment trials.


Click linked name(s) to expand
  • Denise Burnett
  • Mary Ellen Charter, LVT
  • Dr. Brenda Moslock Carter, DVM


Materials and methods:

Part 1: Evaluation of the API(registered trademark)20 Strep System for Identification of Lactococcus lactis ssp lactis Isolated from Bovine Milk Samples

From March 2011 through March 2012, samples submitted to the Keseca Veterinary Clinic Milk Laboratory and identified as pure cultures of non-hemolytic, esculin-positive Streptococcus species with positive reactions on Enterococcosel plates were inoculated onto API 20 Strep test strips according to manufacturer instructions (bioMerieux). API 20 Strep test results were obtained for each isolate at both 4 hours (4hr) and 24 hours (24 hr) of incubation at 36oC by entering each isolate’s specific combination of positive or negative tests on the API strip (Fig. 1) into the online manufacturer database, which produced an identification of the pathogen and a qualification (Not Valid, Unacceptable, Presumptive, Low Discrimination, Acceptable, Good, Very Good, or Excellent) for the identification. The only modification to manufacturer instructions was that all isolates were read at 24 hr, not just isolates with invalid identifications at 4h, as the manufacturer suggests. Isolates were then sent to Quality Milk Production Services (QMPS, Cornell University, Ithaca, NY) for molecular identification using PCR. Isolates with inconclusive molecular identification, or with API identifications qualified as “Unacceptable”, “Doubtful”, or “Not Valid”, were excluded from analyses. Three different cutoffs for quality of API identification were used for test evaluation. For each cutoff analyzed (Low Discrimination, Acceptable, and Good), API identifications of Lactococcus lactis ssp. lactis with quality scores equal or better to the cutoff were considered positive, and those with lower quality than the cutoff were considered negative. Two by two tables were evaluated using WinEpiscope 2.0 to calculate the Sensitivity (Se), Specificity (Sp), Positive Predictive Value (PPV), Negative Predictive Value (NPV), and kappa (k) for each cutoff versus molecular identification.

Part 2: Assessment of Farm-Level Risk Factors

Herds representing cows with Lactoccoccal intramammary infections identified in Part 1 were asked to participate in a herd survey to identify farm-specific factors contributing to incidence of Lactoccocal intramammary infections. Individual interviews with managers of Lactoccoccus “positive” farms were utilized to complete a 12-page survey, which covered key issues such as herd demographics, milking equipment, milking routine, facilities, feeds and water, and bedding (Document 2). Analysis of farm records in DairyComp305 was also conducted at this time. Unfortunately, while this created a very extensive farm information database, the number of farms identified in Part 1 containing cows with Lactoccoccal intramammary infections (n=5) was so small that statistical analysis was not possible at this time. However, surveys were also conducted on five farms from Part 1 whose cows had intramammary infections caused by pathogens other than Lactococcus lactis ssp. lactis to serve as control herds should statistical analysis be possible in the future.

Without statistical analysis to guide environmental sampling, subsequent environmental sampling was directed at the only observed commonality between all five positive farms- sand bedding. Sand bedding samples were collected through random sampling of a minimum of ten stalls per pen on each farm. Where pens exceeded 100 stalls per pen, 10% of stalls were sampled. Within each sampled stall, 3 cores of sand bedding were removed using a silage sampler to ensure 6-8 inch cores of sand were included in sample. Individual pen samples were combined to create a single farm sample for culture according to National Mastitis Council protocols. Non-hemolytic, esculin-positive, Streptococcus species isolates with positive reactions on Enterococcosel plates from these bedding samples were run on API strips prior to shipment to QMPS for molecular identification in similar fashion to milk isolates in Part 1.

Part 3: Antimicrobial Susceptibility Testing

All milk and environmental isolates verified by molecular identification at the referral lab (QMPS) as Lactococcus lactis ssp lactis were submitted for antimicrobial susceptibility testing via Minimum Inhibitory Concentration (MIC) method at QMPS according to industry standards. Results of MIC testing were entered into a database and analyzed for percent of isolates susceptible to each drug (Ampicillin, Penicillin, Erythromycin, Pirlimycin, Penicillin/Novobiocin, Tetracycline, and Cephalothin) based on cutoffs for Streptococci (CLSI, 2004, Fig. 2). MIC levels were also recorded for Oxacillin+2% NaCl, Ceftiofur, and Sulphadimethoxine, but percent sensitivities were not calculated due to lack of established cutoff values for these antimicrobials. From this information, a comparison was made between sensitivities of milk and environmental isolates from each positive farm.

Reference Cited:

Clinical Laboratory Standards Institute (CLSI): 2004, Performance standards for antimicrobial disk and dilution susceptibility tests for bacteria isolated from animals; approved standard M31-S1, vol. 17, no. 17. CLSI, Wayne, PA.

Research results and discussion:

Part 1: Evaluation of the API (registered trademark) 20 Strep System for Identification of Lactococcus lactis ssp lactis Isolated from Bovine Milk Samples

Out of the 5,261 isolates submitted to the KVC laboratory and evaluated by standard biochemical techniques during the study period, 221 (4.2%) were identified as non-hemolytic, esculin-positive, Streptococci with positive reactions on Enterococcosel plates. Of those isolates, only 79 were enrolled in the study based on purity and growth density criteria. Six isolates were excluded from analysis due to inconclusive molecular identifications. Of the remaining 73 isolates, 24 (32.9%) were identified molecularly as Lactococcus lactis ssp. lactis.

From these 73 isolates, 11 were excluded from 4 hr analysis and 13 were excluded from 24 hr analysis for poor quality of API identification. At 4 hours, each of the Good, Acceptable, and Low Discrimination cutoffs had unacceptable Se (< 4.3%), high Sp (100%) and little agreement with molecular identification (< 0.05). At 24 hours, the Good and Acceptable cutoffs had similar results (Se= 9.5% (95%CI=0-22.1), Sp=100% (95%CI= 100-100), PPV= 100% (95%CI= 100-100), NPV=67.2% (95%CI= 55.2-79.3), k=0.12]. However, the 24hr Low Discrimination cutoff had markedly increased Se, while Sp remained high [Se=76.2% (95%CI= 58.0-94.4), Sp= 100% (95%CI=100-100)]. This increased NPV, and yielded the highest observed agreement between API test and molecular identification for all cutoffs and time periods when evaluating to the subspecies [PPV= 100% (95%CI=100-100), NPV=88.6% (95%CI=79.3-98), k=0.81]. At the species level, test agreement was even higher at the 24h Low Discrimination cutoff (k=0.85).

According to bioMerieux technical services and test instructions, 4hr API identifications qualified as “acceptable”, “good”, or “very good” do not need to be re-read at 24 hours. However, 25% of isolates identified molecularly as Lactococcus lactis ssp. lactis in this study would have been misidentified if those instructions were followed, as 6 isolates were identified as other bacteria at 4hr with the qualification of “Good Identification” (Leuconostoc sp. (n=5), Lactococcus lactis ssp. cremoris(n=1)).

Yet, these same 6 isolates were identified by API 20 Strep test at 24 hours as Lactococcus lactis ssp. lactis with a “low discrimination” qualification. It is for this reason that API identification guidelines were slightly modified to read all isolates at both 4 hr and 24 hr. As described above, analysis shows that using results obtained at 24 hr with a “low discrimination” cut off (e.g. results qualified as low discrimination, acceptable, good, very good, or excellent =API positive) is the optimal interpretation protocol for this pathogen. This would indicate that practitioners intending to utilize this tool for identifying Lactococcus lactis ssp lactis specifically should rely on 24 hour results over 4 hour results for better test accuracy. However, practitioners or laboratories considering using this testing method must also consider the impact of moderate test sensitivity, as it results in lower negative predictive values in populations with higher disease prevalence.

There are several possible explanations as to why 24hr API test results are more accurate for Lactoccoccus lactis ssp lactis than 4 hr results. The first is that the bacterial population in each well may not be large enough at 4 hr to produce the biochemical reaction needed to trigger the color indicator for each of the individual tests within the API 20 Strep system. This is less likely since McFarland standards were utilized to ensure appropriate levels of bacterial growth in media prior to test inoculation. The second explanation is that, even with sufficient bacterial load in each well, more than a four hour time period is needed for Lactococcus lactis ssp. lactis to complete the biochemical processes required to turn a test positive within the API 20 Strep system. Thus, something about incubation conditions or Lactococcus lactisi ssp. lactis’ metabolism itself may be contributing to the delay in color change within wells. To examine this further, the percent of study isolates positive on each individual test within the API 20 Strep system at 4h and 24 h were compared to similar information in the API manufacturer’s literature for Lactococcus lactis ssp. lactis (Fig. 3). There are four specific tests for which the API reference data falls within the confidence intervals of this study’s data at 24 hours, but not at 4 hours (ADH, MAN, TRE and AMD). Fifteen of the 21 isolates molecularly identified as Lactococcus lactis ssp. lactis were negative at 4 hours and positive at 24 hours on at least one of these four tests. Therefore, it is likely that a delayed positive result of these four tests contributes to the higher sensitivity of the API 20 Strep system at 24 hours, although the exact reason for this delay is not clear at this time.

Part 2: Assessment of Farm-Level Risk Factors

The 24 Lactococcus lactis ssp. lactis isolates from Part 1 represent five farms in Western New York. Unfortunately, this number of farms was too small to statistically assess risk factors with confidence. Still, the survey information collected from these five farms as well as five other “control” herds from Part 1 was entered into a database for future analysis should more positive farms be identified. Relevant demographic data for the five positive farms is described in Figure 4. Positive herds varied in size, milk production per cow, and milk quality profile. Farms A and C had the highest rates of chronic mastitis issues, while new infection rates were fairly similar between all farms. Non-hemolytic, esculin-positive Streptococcus species represented a higher percent of total farm milk cultures for Farms A and E, compared to Farms B, C, and D. Furthermore, Farms A and B represented the highest number of isolates identified as Lactococcus lactis ssp. lactis during Part 1 of the study.

Without strong statistical analysis to guide environmental sampling, the focus of this portion of the study was directed at the one easily identified commonality between Lactococcus lactis ssp. lactis positive farms- sand bedding. Cultures of sand bedding samples on Farms A, B, C, D, and E all produced Lactococcus lactis ssp lactis. While no causative relationship can be confirmed by our study design, the conclusion that Lactococcus lactis ssp. lactis can, at the very least, survive in sand bedding may be the most important conclusion provided by Part 2 of this study. Further investigation is needed to determine if Lactococcus lactis ssp. lactis is found in other types of bedding (e.g. sawdust, straw, etc), as well as in sand bedding on farms without identified Lactococcal intramammary infections. Despite the need for further study, isolation of Lactoccoccus in the sand bedding was enough to encourage affected farms to revisit their bedding management, and more specifically, their stall grooming protocols, to reduce general pathogen load within bedding and to minimize movement of bacterial populations from deep bedding to the stall surface.

Part 3: Antimicrobial Susceptibility Testing

Antimicrobial susceptibility testing was completed on the 24 Lactococcus lactis ssp. lactismilk isolates identified in Part 1. Eight environmental samples were also evaluated for susceptibility, with 3 samples from Farm A, 2 samples from Farm D, and 1 sample each from Farms B,C, and E. Overall, MIC results indicate that the 24 milk isolates of Lactococcus lactis ssp. lactis were highly susceptible to Ampicillin, Erythromycin, Pirlimycin, Penicillin/Novobiocin combination, and Cephalothin, but only had moderate sensitivity to Tetracycline and poor sensitivity to Penicillin (Fig. 5).

MICs of environmental isolates showed similar trends (Fig. 6). However, it is important to note that in vitro results do not always equate to in vivo activity, and as previously mentioned, farms reported poor treatment response in cases identified prior to this study. Still, considering current available intramammary treatments for dairy cattle, this in-vitro sensitivity provides practitioners with multiple potential treatment choices for cows with clinical disease. While treatment evaluations were not a goal or objective of this study, this information is one of the largest sets of susceptibility data for Lactococcus lactis ssp. lactis isolated from bovine milk and may serve as a starting point for future drug treatment trials.

A comparison of MIC profiles of milk isolates and environmental isolates for each farm was also conducted (Fig 7). The percent of milk isolates matching the individual test susceptibilities of the environmental isolates varied widely between individual drug and by farm. This highlights the need for further investigation concerning the association between milk and environmental strains, possibly through DNA sequencing.

Research conclusions:

The impacts of study results are best understood when reexamining the objectives of the study- more economical identification of the disease, identification of potential risk factors, and MIC data to guide treatment decisions and support future treatment efficacy studies. Identification of the API 20 Strep system as an easily accessible, economically feasible, and accurate testing strategy for Lactococcus lactis ssp lactis from bovine milk provides practitioners with a new tool for identifying Lactococcal mastitis within their client herds. The quick turn-around time (24 hours) continues to provide a faster service than use of a referral laboratory, and its easy accessibility means practitioners can begin to use it as a means of identifying the true extent of Lactococcal mastitis within the U.S. Dairy population, as long as 24 hour results are utilized. Identification of the disease will be a necessary first step to address the many areas identified in this project as needing further investigation. Additionally, results from Parts 2 and 3 provide a foundation for further research concerning on farm management of the disease once more cases and more affected farms are identified. While causality cannot be proven by our study design, the identification of Lactococcus in sand bedding on Lactococcus lactis ssp lactis positive farms sets the stage for further studies regarding the environmental sources of Lactococcus on farms.The MIC data from Part 3 similarly lays a foundation for potential randomized controlled field trials for treatment efficacy investigations.

Impacts on local farmers have been observed, and will only continue to grow with continued investigation. Not only does validation of the API 20 Strep system for identifying Lactococcus from milk samples mean that farmers now have a less expensive test for identifying this disease, but the quick turn-around time means also means faster return to production, a reduction in discarded milk, and better milk quality for cows that are identified, treated, and cleared of disease. Together, this means more profitability for the farm. Additionally, basic knowledge gained from the three sections of the study, combined with standard mastitis control interventions, allowed the authors to hypothesize some preliminary management guidelines to help manage Lactococcal intramammary infections on farms while additional knowledge on this pathogen continues to be investigated:

Management Guideline #1- Although considered an environmental pathogen, DNA sequencing on isolates from the 2010 (pre-study) Lactoccoccus mastitis isolates indicated instances of potential contagious transmission. As a result, it was suggested that cows diagnosed with Lactococcal intramammary infection should be identified with leg bands and moved to a segregated pen at the end of milking line in order to limit environmental contamination from known infected cows, as well as to prevent any potential transmission of disease from infected to non-infected cows during milking. This serves to minimize transmission while disease dynamics of Lactococcal intramammary continue to be investigated.

Management Guideline #2- Focus on Best Management Practices concerning sand bedding management in order to reduce bacterial load, including Lactococcus, within sand bedding. These Best Management Practices included:

  • Use of new sand instead of recycled sand for bedding when possible, and especially in high risk groups (i.e. fresh cows and pre-fresh cows) More frequent cleaning of stalls to remove organic debris For farms that rake sand stalls to counteract compaction, reduce the depth of raking to minimize moving bacterial populations from deep sand to shallow sand near cow udders (Video 1).

Management Guideline #3: Based on MIC data and available intramammary treatments for cattle, the authors suggested treatment of new infections with hetacillin. For resistant/nonresponsive infections, pirlimycin was suggested as a second-line antibiotic. Finally, treatment at dry off with a penicillin/novobiocin combination product was recommended for cows experiencing Lactococcal intrammammary infections during lactation. The goal of this recommendation was not only to increase cure rates, but also to minimize unnecessary or ineffective treatments through targeted therapy.

The goal of this study was to lay the foundation of knowledge to support future investigations into assessing the success of management suggestion as well as treatment success rates, and not to assess these specific issues in the current study. However, these basic recommendations were suggested to these farms as part of a comprehensive milk quality control program while more information on this disease continues to be investigated. Four out of five Lactococcus farms adopted new infection protocols, but only two farms have adopted the dry treatment recommendations. Adoption of sand bedding management measures were more accepted on farms where milk quality was a major current economic concern (i.e. Farm A).

Farm A was the only farm to adopt all 3 management guidelines. These guidelines were incorporated into their overall milk quality improvement plan in 2011. Through a well rounded and aggressive approach, Farm A has increased the milk quality premiums from an annual average of $0.07 per 100 lbs milk (cwt) in 2010 to an average of $0.28/cwt in 2012 through reduction in bulk tank somatic cell levels, equaling an approximate increase of $75,600 annually from milk quality premiums alone. While this increase in milk quality cannot be contributed solely to the recommended guidelines alone, it highlights the potential economic benefit for farms that are able to improve udder health and milk quality.

Additional impacts of this project include various improvements in veterinarian-client communication. The herd survey used for Part 2 of the project has been slightly modified for use in annual and semi-annual herd reviews to aid veterinarians in identifying key health and management concerns on farms where a veterinarian may only visit once or twice a year. This helps in maintaining the Veterinarian-Client-Patient relationship (VCPR) required by law for veterinarians to prescribe and ensure proper use of antibiotics on farms. Furthermore, presentation of study results at client meetings has prompted milk quality discussions with 5 different clients at subsequent herd health visits, enabling veterinarians to discuss herd-specific issues even when unrelated to Lactococcus.

Overall, the results of this study have both local and far-reaching impacts. While the results have provided hypothesized management guidelines to help individually affected farms manage the disease and increase profitability, the study has also confirmed the accuracy of a widely accessible and relatively inexpensive test to aid practitioners in identifying the disease, provided MIC data to aid in proper treatment decisions and spur potential field trials, and identified at least one potential environmental source of Lactococcus on dairy farms, thereby addressing the key objectives outlined for the study.

Participation Summary

Education & Outreach Activities and Participation Summary

Participation Summary

Education/outreach description:
  • Communication was maintained both during and beyond the study period with the five farms identified as having Lactococcal intramammary infections in Part 1 of the study regarding individual cow treatment decisions and farm management protocols.

    A research summary written in collaboration with faculty at Cornell University’s College of Veterinary Medicine regarding the evaluation of the API 20 Strep 20 System was selected for presentation at the National Convention for the American Association of Bovine Practitioners(AABP) in Milwaukee, WI, in September 2013. In addition to an oral presentation at this conference, the research summary will be published in the conference proceedings and distributed to the nearly 6,000 members AABP. (

    Preliminary results were presented to approximately 75 farmers at the Keseca Veterinary Clinic Annual Client Meeting in March 2012.

    Final results were presented to approximately 60 farmers at the Keseca Veterinary Clinic Annual Client Meeting in March 2013 and prompted discussions at subsequent herd visits as described above.
    Information regarding this project was shared on the Keseca Veterinary Clinic website (

    Two out-of-state herds dealing with intramammary infections caused by Lactococcus lactis ssp. lactis contacted Keseca Veterinary Clinic in 2013 after finding this project in the SARE database. Both herds were looking for guidance on control of the disease and insight from the authors’ clinical experience. Phone and email consultations were conducted to assist these herds and their veterinarians in addressing Lactococcal intramammary infections

Project Outcomes

Project outcomes:

Economic analysis was not a main objective for this study, but the investigation concerning the validity of the API tests was based on the premise that the API system provided practitioners a less expensive alternative to traditional molecular testing. Based on expenditures for this study, the cost of running an individual API 20 Strep test strip was approximately $15.93, and included cost of test strips, required reagents, bacterial strains for quality control, sterile glycerol, and shipping charges for all supplies listed. This estimate reflects the cost to run an API 20 Strep test in a clinic already equipped for basic culturing, and does not include labor.

Therefore, a practitioner or lab must consider this when estimating the cost of running API 20 Strep tests in his or her particular practice or lab. In contrast, at the time of sample processing (2011-2012) the gold standard of molecular identification cost $35/isolate at the referral lab used in this project. This cost does not include the shipping cost involved in using a referral lab, which will vary by region. Thus, as long as the labor required to run the test is less than $19.07+ estimated shipping costs for use of referral lab, the API 20 Strep is a less expensive test based on direct costs.

Additional economic advantage of the API 20 Strep concerns the indirect costs of waiting for results from a referral laboratory. With a quick turnaround time of 24 hours, clients are given a result and can begin interventions with 48 hours of sampling the cow (24 hours to initial colony growth, additional 24 hours for API test) rather than a week or more with molecular testing. This equates to quicker return to production, less milk loss, and therefore reduced cost per case of mastitis to the client. Potential economic gains for clients who are able to manage Lactococcus mastitis issues on their farm through identification of infected cows and management interventions has been outline in “Impacts and Outcomes”.

Assessment of Project Approach and Areas of Further Study:

Areas needing additional study

This study highlighted several areas needing further investigation, including:

  • Statistical farm-level risk assessment following identification of additional Lactococcus-positive farms. Evaluation of on-farm treatment success rates Use of DNA identification techniques to compare bovine and environmental strains within a farm to discern environmental or contagious disease dynamics Continued outreach to the professional community, including application to present at the National Mastitis Council meeting in January 2014 and submission of a scientific paper with study results to the Journal of the American Veterinary Medical Association.
Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.