1. Ketone and NEFA testing as diagnostic tools in assessing transition dairy cows Stephen LeBlanc OABP/OABA meeting April 14, 2005

2. Monitoring Programs for Transition Cows 1.Monitor Current Transition Cow Program –HERD LEVEL –Track success and compliance with existing program –Early detection of problems 2.Monitor for Subclinical Disease - INDIVIDUAL LEVEL –Early treatment to prevent clinical disease Helps to quantify problems and direct investigation

3. Options for Monitoring or Investigating Clinical disease incidence Milk production DMI –Why doesn’t it get done?? –Group average; distribution within group –Target > 12 kg DMI average in close-up (heifers & cows; 3 weeks before due) –Fresh group Metabolic tests

4. Daily Dry Matter Intake Around Calving * CALVING

5. Re-esterified  triglyceride Stored in liver Exported in VLDL Gluconeo- genesis Propionate AA Glycerol NEFA Glucose Fetus Mammary gland Incompletely oxidized  ketones Completely oxidized  energy BHB Acetoacetate Acetone

6. Re-esterified  triglyceride Stored in liver  Exported in VLDL Gluconeo- genesis Propionate AA NEFA Glucose Fetus Mammary gland Incompletely oxidized  ketones Completely oxidized  energy BHB Acetoacetate Acetone Unsuccessful response to NEB – Ketosis and Fatty liver

7. Typical patterns of DMI and NEFA Overton/Burhans, 2001

8. Associations with health and performance Pre-partum NEFA associated with: –~ 4X increased risk of LDA (Cameron et al, 1998; LeBlanc et al, 2005) –~ 1.5X increased risk of RP (Dyk, 1995; LeBlanc et al, 2004) –2 – 3 X increased risk of subclinical ketosis (Osborne, 2003; Gooijer et al, 2004)

9. Median time to diagnosis of clinical ketosis = 11 DIM Duffield, 2000 Incidence of Subclinical Ketosis

10. Prevalence of Subclinical Ketosis Duffield et al 1998 Oetzel, 2003

11. Clinical ketosis treatment rate is a poor estimate of ketosis (Duffield et al 1998)

12. Associations with health and performance BHB (subclinical ketosis) in early lactation is associated with: –4-8X increased risk of LDA (Geishauser, 2000; LeBlanc et al, 2005) –Decreased milk production (Duffield, 2000) –Increased severity of mastitis (Suriyasathaporn et al, 2000) –50% decrease in pregnancy at first AI (Walsh et al, 2004)

13. Effect of subclinical ketosis in week 2 on CR at 1 st AI (Walsh et al, 2004)

14. Cow-side tests for ketosis (relative to serum BHB ≥1400 µmol/L) Milk Keto-Test 100 µmol/L –Sensitivity = 83% –Specificity = 82% 200 µmol/L –Sensitivity = 54% –Specificity = 94% Oetzel, 2004 Powder lacks sensitivity Urine Ketostix (read at 5 seconds) “small” (15µmol/L) –Sensitivity = 79% –Specificity = 96% Carrier et al, 2004 Acetest tablet lacks specificity

15. Subclinical Ketosis Monitoring Programs (True Prevalence = 20%) TestPV +PV-Apparent Prevalence Keto-Test (100 umol/L) 62%93%23% Keto-Test (200 umol/L) 80%87%11% Ketocheck (Milk)90%86%8% Acetest(Urine)38%100%53%

16. Sampling logistics In a herd with 50 to 1000 cows, if a prevalence of “positive” tests –e.g. NEFA ≥ 0.5 or BHB ≥ 1400 And ≥ 10% is the threshold of interest And you wish to be 75% confident of detecting this level of problem, then… 13 samples are required Oetzel proposes using 12 samples for NEFA and BHB blood testing for investigations

18. Metabolic Predictors of LDA 1184 animals in 20 herds Weekly visit by technician Same day, same time (AM) Cows enrolled 4 - 10 d prior to expected calving Sampled weekly until the week after calving (Total of 2 - 4 samples) LeBlanc et al, 2005

21. Prepartum DA model Among all variables measured in last week before calving: OR 95% CI P NEFA  0.5 mEq/L3.51.9 – 7.1.0001 Sensitivity = 64%Specificity = 66% LeBlanc et al, 2005

22. Simple Association of NEFA 4-10 d before DUE with LDA NEFAORSeSpLR  0.32.363561.4  0.42.650721.8  0.54.146822.6  0.63.030892.6  0.82.617932.5  1.04.115963.8 LeBlanc et al, 2005

23. Simple Association of NEFA 4-10 d before DUE with LDA NEFAOROR*SeSe*SpSp*LRLR*  0.32.32.6636156611.41.6  0.42.62.9504772771.82.0  0.54.15.1464382872.63.3  0.63.03.7302689922.63.2  0.82.63.0171293962.53.1  1.04.15.1151296983.85.2 * Excluding cows within 2 days of actual calving LeBlanc et al, 2005

24. Postpartum DA model VariableOR95% CIP RP1.71.1 – 2.7.01 Metritis4.82.0 – 11.2.0003 BHB per 100  mol/L  *1.081.06 – 1.1.0001 NEFA per 1.0 mEq/L  *2.41.4 – 4.3.002 Season, parity, MF, twins all NS Minimum significant cut-points in the model: BHB  1000  mol/L ; NEFA  0.6 mEq/L LeBlanc et al, 2005

25. Postpartum Simple Associations with DA TestCutpointORSeSpLR NEFA0.64.886431.5 0.83.968641.9 1.04.856792.6 BHB10006.369742.6 12008.063823.5 14008.053884.3 LeBlanc et al, 2005

26. Postpartum Simple Associations with DA TestCutpointORSeSpLR Milk BHB 1002.864621.7 2003.448802.4 LeBlanc et al, 2005

27. Sample handling Serum (red top) or plasma (purple top) Avoid hemolysis Ideal: keep chilled, separate within a few hours, ship chilled to arrive at lab in 1-2 days Serum can be frozen for at least 1 month What you could get away with: delay of < 24 h to separate; serum at room temp for < 24 h or in fridge for < 3 days (Stokol & Nydam, 2004)

28. Monitoring Energy Metabolism in Transition Cows Pre-Calving - NEFA Post-Calving -Ketones –Routine monitoring (milk or urine)

29. Monitoring Energy Metabolism in Transition Cows Helps to direct investigation –What is the problem? –Where/when is the problem? Rarely answers “WHY?” –Need to look further and test hypotheses

30. Evaluation of a Rapid, On- Site Serum NEFA Test 10 Guelph-area farms Prepartum blood sample (-7 to –4 days) Harvested serum and aliquoted Measure NEFA concentrations: –Animal Health Laboratory ( Hitachi 911 analyzer ) –DVM NEFA Gooijer et al, ICPD 2004

31. Correlation between tests Pearson’s r = 0.89 Gooijer et al, ICPD 2004

32. DVM NEFA AHL NEFA  0.5<0.5 >0.41337140  0.425 185210 158192350 Test Characteristics of DVM NEFA (Gold Standard = AHL > 0.4 mEq/L) Sensitivity = 84% Specificity = 96% Gooijer et al, ICPD 2004

33. Maintaining Peripartum DMI Fresh feed daily Adequate bunk space (>60 cm) > 100 ft 2 /cow of pack < 100% stocking Separate heifer groups Moderate BCS (3.5) Adaptation to new rations (3-4 weeks) Adequate eNDF Minimize group/pen changes Heat abatement –THI > 72 –T > 27 C Free choice water 0.5 – 0.75% BW in concentrates 60:40 Forage:concentrate Rumensin CRC