J. Dairy Sci.

104
https://doi.org/10.3168/jds.2020-18998
© 2021 American Dairy Science Association®. Published by Elsevier Inc. and Fass Inc. All rights reserved.

Late-gestation heat stress abatement in dairy heifers promotes

thermoregulation and improves productivity
B. D. Davidson,1 B. Dado-Senn,1 N. Rosa Padilla,1 T. F. Fabris,1 L. T. Casarotto,1 V. Ouellet,1 I. M. Toledo,2
G. E. Dahl,1 and J. Laporta1*†
1
Department of Animal Sciences, University of Florida, Gainesville, 32608
2
IFAS Extension, University of Florida, Gainesville, 32603

ABSTRACT 0.17°C), and unshaved SR (19.0 vs. 35.2 ± 1.9 g/m2h),

relative to HT heifers. Additionally, VT was lower in
Multiparous, nonlactating pregnant cows are nega- CL heifers during wk −4, and −2, specifically during
tively affected by heat stress, but the effect of heat early morning and early afternoon hours. When mea-
stress on more thermotolerant pregnant heifers has sured over a 36-h time interval, ST and SR were lower
received less attention. Our objective was to character- in CL heifers, when compared with HT heifers for all
ize the effect of late-gestation heat abatement on ther- weeks. Notably, ST was reduced overnight and SR was
moregulatory responses and subsequent milk produc- reduced during the daytime. Cooled heifers had higher
tion of nulliparous Holstein heifers. Pregnant heifers, milk yield (35.8 vs. 31.9 ± 1.4 kg/d), when compared
blocked by body condition score (BCS) and predicted with HT heifers. Similar to multiparous cows, our data
transmitting ability (PTA) for milk, were enrolled in indicate that actively cooling heifers in late pregnancy
either heat stress (HT, shade of freestall barn; n = is effective in promoting thermoregulation and results
16) or cooling (CL, shade of freestall barn, water soak- in elevated milk production postcalving.
ers, and fans; n = 15) environments during the last 60 Key words: cooling, heifer, milk yield, sweating rate
d of pregnancy (~8 weeks). Rectal temperature (RT;
thermometer), respiration rate (RR; breaths/min),
INTRODUCTION
sweating rate (SR; VapoMeter, Delfin Technologies,
Kuopio, Finland), and skin temperature (ST; infrared Increasing global environmental temperatures has
thermometer) were measured thrice weekly from en- detrimental effects to the dairy industry. Heat stress
rollment to calving. Vaginal temperature (VT; i-button is experienced by livestock when ambient temperature
intravaginal device) was measured every 10 min for 7 exceeds the upper critical temperature (UCT) of the
consecutive days at wk −8, −6, −4, and −2 relative to thermoneutral zone, which occurs in lactating dairy
calving and averaged hourly. Daily thermoregulatory cattle when the temperature-humidity index (THI) is
patterns assessed by SR and ST, were measured every above 68 (Zimbelman et al., 2009). Whereas young-
4 h over a 36-h time interval at wk −6, −4, and −2 stock and dry cows generate less metabolic heat (West,
relative to calving. Upon calving, milk, protein, and fat 2003) and have a higher UCT (Hahn, 1999) to maintain
yields were recorded twice daily for 15 wk. The average normal physiological functions, they remain susceptible
temperature-humidity index (Hobo Pro temperature to heat stress. Heat stress is not a short-term event.
probe, Onset Computer Corporation, Pocasset, MA) in On average in the United States, dairy cattle will ex-
the barn during the precalving period was 77 (minimum perience heat stress for 96 d out of the year (Ferreira
of 72, maximum of 82). Only heifers that gave birth to et al., 2016). In the United States, the economic loss
a female calf (CL = 12, HT = 14) were included in the from heat stress in both lactating and nonlactating
statistical analysis. In the precalving period, CL heifers dairy cows combined is estimated at more than $1.2
had lower RR (44.3 vs. 60.0 ± 1.6 breaths/min), RT billion (Key and Sneeringer, 2014) and up to $2.5 bil-
(38.7 vs. 38.8 ± 0.04°C), unshaved ST (34.7 vs. 35.3 ± lion, annually (St-Pierre et al., 2003; Ferreira et al.,
2016; Laporta et al., 2020). Whereas heat stress in both
lactating (West, 2003; Collier et al., 2006; De Rensis et
Received June 1, 2020. al., 2015) and nonlactating, dry dairy cows (do Amaral
Accepted August 24, 2020.
*Current affiliation: Department of Animal and Dairy Sciences,
et al., 2009; Tao and Dahl, 2013; Fabris et al., 2019) has
University of Wisconsin-Madison, 53706 been studied extensively, the effect on nulliparous dairy
†Corresponding author: jlaporta@​wisc​.edu heifers is unknown.
 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Physiological states, of an animal, in order from increase first lactation milk yield when compared with
highest priority for nutrient partitioning to least are herd mates exposed to heat stress.
maintenance, pregnancy, growth, lactation, and repro-
duction (Oldham and Emmans, 1989). Under stressful MATERIALS AND METHODS
conditions, behavioral, and physiological adaptations
occur to maintain basic body functions at the expense Experimental Design and Treatments
of productivity. For example, under heat stress, cows
spend more energy on heat dissipation through in- This study was approved by the Institutional Animal
creases in respiration rate (RR), body temperature, Care and Use Committee at the University of Florida
and evaporative water loss (Armstrong, 1994; Tao et and was conducted at the University of Florida’s Dairy
al., 2011). In addition to general body mass accretion Unit (Hague, FL) from June to November 2019. Thirty-
during pregnancy, heifers experience exponential mam- one nulliparous pregnant Holstein heifers blocked by
mary gland growth. It is estimated that 48 to 94% body condition and by predicted transmitting ability
mammary growth occurs during pregnancy, with most (PTA) for milk (Zoetis Services LLC, Enlight, CLARI-
of that growth occurring during the last 2 mo of gesta- FIDE, Brattleboro, VT) were enrolled in heat stress
tion (Tucker, 1969; Knight and Peaker, 1982). Simulta- (HT, shade of a freestall barn; n = 16) or cooling (CL,
neously, the developing fetus is growing at a rapid rate shade of a freestall barn, water soakers, and fans; n =
and accumulates approximately 60% of its body mass 15) treatments for 60 d before (~8 weeks) expected
during the last trimester of gestation (Bauman and calving date (average treatment duration: 55 ± 5 d).
Currie, 1980). Therefore, exposure to stressors during Prior to enrollment all heifers were managed in a single
this critical phase of development could be detrimental group raised on pasture with access to artificial shade.
in the energy partitioning toward fetal and mammary All heifers were housed in a sand-bedded freestall
growth in nulliparous heifers. barn, with HT and CL treatments separated across 4
In dry cows, the carryover of mammary epithelial pens (n = 7–8 heifers per pen). Six weeks after first en-
cells from one lactation to the next is ~50%, suggest- rollment, heifers within the HT or CL treatment were
ing development during this time is critical for milk switched to the neighboring pen to overcome potential
yield in the lactation to follow (Pitkow et al., 1972; pen effects. The HT pens were provided with shade of
Hurley, 1989). Heat stress during the dry period de- the open-sided barn, and the CL pens were provided
creases mammary gland growth and reduces milk yield with shade of the barn, water soakers over the feed line,
potential in the subsequent lactation (Tao et al., 2011, and fans. Fans (J&D Manufacturing, Eau Claire, WI)
2012; Fabris et al., 2017). Although heifers do not have ran continuously over the stalls and the water soak-
a dry period, similar processes in the mammary gland ers (Rain Bird Manufacturing, Glendale, CA) turned
tissue are taking place during late gestation in prepara- on automatically for 1.5 min at 5-min intervals when
tion for lactation; however, the effect of heat stress on ambient temperatures exceeded 21.1°C.
thermoregulation and mammary growth of nulliparous Every 15 min, ambient temperature and relative hu-
heifers is unknown. midity of the barn were recorded with Hobo Pro series
Dairy heifers are not typically considered for heat temperature probes (Onset Computer Corporation,
stress abatement and often housed on pasture occasion- Pocasset, MA). The THI was calculated for each treat-
ally with access to natural or artificial shade during ment group based on the equation recommended by
their nonproductive phase. Even though youngstock Dikmen and Hansen (2009):
have a higher UCT and generate less metabolic heat,
heat strain accumulates throughout the day in response THI = (1.8 × T + 32) – [(0.55 – 0.0055 × RH)
to extended exposure to high THI and affects thermo- × (1.8 × T – 26)],
regulatory responses of animals, reducing their overall
productivity (Beede and Collier, 1986; Renaudeau et where T = ambient temperature (°C) and RH = rela-
al., 2011). To our knowledge, there are currently no tive humidity (%). Before calving, all heifers were fed
studies examining the effects of disparate environmental a TMR (Table 1) as a group in each of the 4 pens and
conditions during late gestation on thermoregulatory pen daily feed intake was recorded by weighing TMR
responses and productivity of nulliparous dairy heifers. offered and refused. TMR fed and refused were sampled
Herein, we investigated the effects of heat stress abate- and DMI was measured once per week through the pre-
ment on late-gestation dairy heifers’ thermoregulatory calving period. Dry matter content of these samples
responses and milk production in their first lactation. was determined by oven drying at 65°C for 48 h (Amer-
We hypothesized that actively cooled nulliparous preg- ican Scientific, Columbus, OH). Water was provided
nant heifers would thermoregulate more effectively and
Journal of Dairy Science Vol. 104 No. 2, 2021
 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Table 1. Ingredient composition of TMR fed to heifers precalving (during the last 60 d of gestation) and
postcalving (15 wk in milk). Heifers were exposed to cooling (n = 12) or heat stress (n = 14) for the final 60 d
of gestation, but all animals were provided with heat stress abatement during lactation1

Ingredient2 (% of DM) Precalving #1 Postcalving #2 Postcalving

Corn silage 67.6 42.1 43.9
Soybean meal 44 solvent 14.6 13.9 13.2
Grass hay 8.30 — —
Novasil 0.60 — —
Close up mineral 4.70 — —
Biochlor 4.20 — —
Corn grain ground fine — 20.1 16.5
Cottonseed — 4.10 4.10
Palmit 80 — 1.10 0.90
Fresh cow mineral — 5.20 5.20
Citrus pulp dry — 13.5 13.5
Lactating mineral — — 2.70
1
#1 Postcalving diet was fed to the lactating cows until August 27, 2019; the diet then changed to #2
Postcalving for the remaining duration of the experiment and included the same feed ingredients as the previ-
ous diet with lactating mineral added.
2
Grass hay contained 10% CP, 67% NDF, and 8% lignin NDF; Novasil Plus (BASF, Florham Park, NJ); close
up mineral fed during the precalving period contained (DM basis) 35.2% calcium carbonate, 16.2% magnesium
sulfate 7H2O, 11.8% calcium sulfate dihydrate, 11.4% ReaShure Choline (Balchem, New Hampton, NY), 9.9%
wheat middlings, 6.1% PDS SE 58 Zinpro Premix (Zinpro, Eden Prairie, MN), 4.9% white salt, 2.7% magne-
sium oxide, 1.3% prescriptive yeast, and 0.4% Rumensin 90 (Elanco Animal Health, Greenfield, IN); Bio-Chlor
(Arm and Hammer Animal Nutrition Inc., Trenton, NJ); Palmit 80 (GlobalAgriTrade Corporation, Rancho
Dominguez, CA); fresh cow mineral fed during the postcalving period contained (DM basis) 23.8% sodium
bicarbonate, 19.8% corn grain ground fine, 19.5% PDS SE AA Pack, 11.7% calcium carbonate, 7.8% dicalcium
phosphate, 6.4% PDS SE L65 Mintrex Premix, 5.2% white salt, 4.1% magnesium oxide, 0.9% prescriptive en-
zymes, 0.8% prescriptive yeast, and 0.2% Rumensin 90 (Elanco Animal Health); lactating mineral fed during
the postcalving period contained (DM basis) 77.0% corn grain ground fine, 7.1% sodium bicarbonate, 4.8%
dicalcium phosphate, 2.7% molasses cane, 2.6% PDS SE l65 Mintrex Premix, 2.4% white salt, 2.3% magnesium
oxide, 0.9% calcium carbonate, 0.3% prescriptive yeast, 0.1% Rumensin 90 (Elanco Animal Health).

to all heifers ad libitum. After calving, all cows were vaginally to measure core body temperatures every 10
managed as a single group and housed in a shaded, min and averaged hourly for 7 consecutive days. Probes
sand-bedded freestall barn equipped with water soakers were placed at enrollment, removed for 1 wk and rein-
and fans. Lactating cows were fed a TMR ad libitum serted on wk −6, −4, and −2 relative to calving.
to meet NRC requirements for lactating cows (NRC, Thrice weekly during the 8-wk precalving period,
2001; Table 1) and were milked twice daily according thermal indices of heifers (i.e., RT, ST, SR, RR), wind
to the standard operating procedures of the University speed, and air quality of the pens were recorded at
of Florida Dairy Unit. 1400 h. Respiration rates were measured by counting
flank movements for 1 min (breaths per min; bpm).
Physiological, Environmental, and Milk Wind speed (m/s) and air flow (m3/s) were recorded
Yield Measurements using an anemometer (MS6252A Digital Anemometer
System, Proster, Miller Place, NY) and air quality was
At enrollment (wk −8 relative to calving), baseline measured using an air quality meter (LKC-1000S+ Air
BW, BCS, sweating rates (SR), skin (ST), and rectal Quality Monitor; Temtop, Milpitas, CA). Air qual-
temperatures (RT) were measured. Rectal tempera- ity measurements were obtained at the pen level and
ture was measured with a digital thermometer (GLA included particle matter (fine particle: PM2.5, and
M900, accuracy ± 0.1°C, GLA Agricultural Electronics, regular particle: PM10), number of particles (per L),
San Luis Obispo, CA), SR with a VapoMeter (Delfin air quality index (AQI), formaldehyde (HCHO), and
Technologies, Kuopio, Finland), and ST with an in- total volatile organic carbons (TVOC). To assess pat-
frared thermometer (Raytek MiniTemo MT6 Infrared terns of thermoregulatory responses, SR, and ST were
Thermometer; Instrumart, South Burlington, VT). The recorded every 4 h during a 36-h period (d 1 = 0600
SR and ST were recorded from a 5-cm2 shaved and un- to 2200 h, d 2 = 0200 to 1800 h) at wk −6, −4, and
shaved area over the right front shoulder blade. Blank −2 relative to calving. After calving, colostrum yield,
controlled internal drug release devices were fitted with daily milk yield, and components (i.e., fat and protein
a temperature probe (i-button cat. #DS1922-F5, ac- percentage and yield), health events, and body weights
curacy ± 0.0625°C; Maxim, Irving, TX) and inserted were retrieved from AfiFarm Dairy Herd Management

Journal of Dairy Science Vol. 104 No. 2, 2021

 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Software (Afimilk Ltd., Kibbutz, Afikim, Israel) from main effects of treatment, day as the repeated measure,
calving to 105 DIM (15 wk). Gestation length (GL) and their interactions. All other variables were analyzed
was calculated by subtracting calving date from the using PROC MIXED. The model included fixed effects
artificial insemination date before confirmation of preg- of treatment, time (hours, week, or day), all possible
nancy. interactions, and animal ID nested within treatment
was used as the random effect. Pre- and postcalving
Blood Collection and Processing data were analyzed separately. Measurements obtained
on enrollment day were used as covariates in the mod-
Blood samples were collected at enrollment and els for precalving BW, blood electrolytes and acid-base
weekly until calving. Samples were collected from the indicators, BCS, and precalving TP. The model to
coccygeal vessels using 2 vacutainer tubes (10 mL, analyze milk performance (i.e., yield, fat, and protein)
Becton Dickinson, Franklin Lakes, NJ), 1 containing included PTA for milk as a covariate. These heifers
clot activator gel (BD cat. #366430, Becton Dickin- were bred to sexed semen, so heifers that gave birth
son; placed at room temperature for ~30 min), and to bull calves (2 CL and 2 HT) or twins (1 CL) were
1 containing sodium-heparin anticoagulant (BD cat. excluded from all statistical analyses. Significance was
#366480, Becton Dickinson; placed on ice). To deter- declared at P ≤ 0.05 and tendency was declared at 0.05
mine the metabolic effects of heat stress in dairy heif- < P ≤ 0.10. Data are presented as least squares means
ers, we evaluated the acid-base and electrolyte balance ± standard error, unless otherwise stated.
and the circulating metabolites. To do so, whole blood
from the sodium-heparin tube was immediately ana- RESULTS
lyzed with the hand-held i-STAT 1 machine using the
CHM8+ cartridge (Abbott cat. #09P31–25, Abbott, Environmental Measures
Princeton, NJ). Variables of interest included sodium,
potassium, chloride, total carbon dioxide (TCO2), Temperature-humidity index of the pens did not differ
anion gap, ionized calcium, glucose, BUN, creatinine, for the duration of the treatment period and averaged
hematocrit (HCT), and hemoglobin (HGB). Serum 77.33 and 77.34 ± 0.20 for CL and HT, respectively
and plasma were separated through centrifugation at (P = 0.99). Air quality measurements, including PM2.5,
3,000 × g for 20 min at 4°C. Serum was aliquoted and PM10, number of particles, AQI, HCHO, and TVOC,
stored at −20°C. Total protein (TP) was assessed from were not different among treatment pens (all P ≥ 0.34)
plasma using a digital Brix refractometer (MA871; and all were within acceptable ranges. The AQI aver-
Milwaukee Instruments, Rocky Mount, NC) before it aged 29.6 for HT and 25.1 for CL pens; and fell within
was aliquoted and stored at −20°C. Postcalving blood the good category according to the standards of the
samples were collected at 14, 28, and 42 DIM. Hema- US Environmental Protection Agency Clean Air Act
tocrit was assessed from whole blood from the sodium- (AirNow, 2019; EPA, 2019). Wind speed and air flow
heparinized tube. Capillary tubes (Fisher Scientific, were higher in the CL pens (4.23 vs. 0.71 ± 0.28 m/s
Pittsburgh, PA) filled with whole blood were centri- and 8,740 vs. 1,557 ± 584 m3/s, both P < 0.0001),
fuged at 2,240 × g for 3 min at 20°C (room tempera- when compared with HT pens for the duration of the
ture) in a microhematocrit centrifuge and read with a precalving period.
circular microcapillary hematocrit reader. Remaining
serum and plasma were processed, TP was measured Growth Measurements and Feed Intake
from plasma, and they were stored at −20°C.
Providing cooling during the last 8 weeks of gestation
did not affect precalving BCS or DMI (P ≥ 0.73; Table
Statistical Analysis
2). Gestation length was longer for CL heifers compared
All statistical analyses were performed in SAS (ver- with HT heifers (276.4 vs. 272.7 ± 1.4 d, respectively; P
sion 9.4, SAS Institute Inc., Cary, NC). Data were = 0.01; Table 2). Precalving BW was similar between
tested for covariance (Levene’s test) and normality was CL and HT heifers, but an effect of week was detected
tested by evaluating the Shapiro-Wilk statistic using (P < 0.0001; Table 2), in which both groups increased
the UNIVARIATE procedure. To meet the homoge- BW as they approached calving. There was a treatment
neity of variance criteria, raw data were transformed by week interaction for postcalving BW (P = 0.0002;
[log10(x) or 1/x] when deemed appropriate. Air quality, Table 2), in which the loss of BW for the CL heifers was
THI, and DMI were analyzed by pen using generalized more pronounced in wk 2 through 5, with a tendency
linear mixed models using PROC MIXED including for lower BW in the CL heifers during wk 6 and 15.

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 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Table 2. Body weight, BCS, DMI, gestation length (GL), milk fat percentage, milk protein percentage, milk
fat yield, and milk protein yield from nulliparous heifers exposed to either cooling (n = 12) or heat stress (n =
14) environments during the last 60 d of gestation

Treatment2 P-value*

Heat Treatment
Variable1 Cooled stressed SEM Treatment Week × week
Precalving
Pre-BW, kg 599.9 610.0 7.10 0.16 <0.0001 0.47
BCS 3.57 3.59 0.04 0.73 0.32 0.38
DMI, kg/d 7.99 8.34 1.03 0.76 0.14 0.98
Postcalving
GL, d 276.4 272.7 1.41 0.01 — —
Post-BW 573.0 589.8 16.5 0.32 <0.0001 0.0002
Fat, % 3.64 3.69 0.10 0.65 <0.0001 0.05
Protein, % 2.88 2.90 0.05 0.72 <0.0001 0.01
Fat yield, kg/d 1.30 1.18 0.06 0.08 <0.0001 0.50
Protein yield, kg/d 1.04 0.94 0.04 0.01 <0.0001 0.003
1
Pre-BW was collected weekly from enrollment through calving; BCS was recorded weekly during the last 60 d
of gestation; before calving, DMI was calculated per cow by dividing the daily sub pen intakes by the number
of cows in the pen on the day of measurement; Post-BW was collected weekly from calving to 15 wk in milk.
2
Heifers were cooled (shade of freestall barn, fans, soakers) or heat stressed (shade of freestall barn).
*Significance was declared at P ≤ 0.05; tendency was declared at 0.10 ≥ P > 0.05.

Thermoregulatory Responses by week by hour interactions. Cooled heifers had lower

(P < 0.0001; Figure 3A) SR during wk −4 and −2
Heifers with access to active cooling during the last and tended to have lower SR during wk −6, relative to
8 weeks of gestation had significantly lower RT (38.7 HT heifers. Specifically, during wk −6, CL heifers, in
vs. 38.8 ± 0.04°C, P = 0.007; Figure 1B), unshaved ST comparison to HT, tended to have lower SR at 1000 h,
(34.7 vs. 35.3 ± 0.17°C, P = 0.002; Figure 1C), and and SR was significantly lower at 1400 h on d 1 (Figure
shaved ST (35.5 vs. 36.0 ± 0.16°C, P = 0.006; Figure 3A). During wk −4 and −2, CL heifers had lower SR
1D) relative to their heat-stressed counterparts. Ad- between 1000 to 1800 h on d 1 and between 1000 to
ditionally, unshaved SR (19.0 vs. 35.2 ± 1.90 g/m2h, P 1800 h on d 2, relative to HT heifers (Figure 3A). Dur-
< 0.0001; Figure 1E) and shaved SR (19.2 vs. 32.8 ± ing wk −4, CL heifer SR tended to be lower at 2200
2.23 g/m2h, P < 0.0001; Figure 1F) were lower in the h of d 1 and at 0600 h of d 2, in comparison with HT
CL heifers, relative to the HT heifers. For these ther- heifers (Figure 3A).
moregulatory indices there was also a significant effect Skin temperatures were lower in the CL heifers (P <
of week (all P ≤ 0.05; Figure 1) but no treatment by 0.0001; Figure 3B) during all weeks, relative to HT heif-
week interactions (all P ≥ 0.12). There was a tendency ers. Specifically, during wk −6, CL heifers had lower ST
for a treatment by week interaction for RR (P = 0.08; at 0600 h, between 1800 and 2200 h of d 1, and between
Figure 1A), whereby CL heifers had significantly lower 0200 and 0600 h of d 2 (Figure 3B). During wk −4, CL
RR from wk −8 through −1, relative to calving, when heifers had lower ST at 0600 h of d 1 and between 0200
compared with HT heifers. Vaginal temperatures had and 0600 h of d 2 (Figure 3B). Finally, during wk −2,
a significant treatment by week by time interaction (P CL heifers had significantly lower ST at 0600 and 2200
= 0.02; Figure 2), wherein during wk −4 and −2, CL h of d 1, and between 0200 and 1000 h on d 2, relative
heifers had lower VT. Specifically, relative to HT, VT to HT heifers (Figure 3B).
was consistently lower in the CL heifers on wk −4 at
0000 h, between 0400 to 0600 h, 1100 to 1200 h, and at
Milk Yield and Components
2300 h (Figure 2). During wk −2, VT was lower in the
CL heifers relative to HT, from 0000 to 0600 h, 1000 to There was a treatment by week interaction for milk
1400 h, and at 1600 and 2200 h (Figure 2). yield (P = 0.02; Figure 4), whereby CL heifers produce
To assess patterns of thermoregulatory responses, SR more milk than HT heifers during wk 3 to 4 and wk 6
and ST were recorded every 4 h during a 36-h period to 15, with a tendency for higher production at wk 5 (P
(d 1 = 0600 to 2200 h, d 2 = 0200 to 1800 h) at wk ≤ 0.08; Figure 4). Fat percentage and protein percent-
−6, −4 and −2 relative to calving. For unshaved SR age were similar (P ≥ 0.65; Table 2) between treat-
and unshaved ST during wk −6, −4, and −2 (Figure ments but displayed treatment by week interactions (P
3A, 3B, respectively) there were significant treatment
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 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Figure 1. Respiration rate (A; bpm = breaths per minute), rectal temperature (B), skin temperature (C, D), and sweating rate (E, F) mea-
surements from cooled (shade of a freestall barn, fans, and soakers; n = 12) or heat stressed (only shade; n = 14) nulliparous heifers during the
last 8 wk of gestation. Unshaved skin temperature (C), shaved skin temperature (D), unshaved sweating rate (E), and shaved sweating rate (F)
were obtained from the same unshaved and shaved areas over the front right shoulder blade. Relative to calving, measurements were obtained
3 times weekly for 8 wk and averaged by week. ** indicates significance (P ≤ 0.05). Data are graphed using the LSM ± SE of the interaction
[treatment (trt) by time].

≤ 0.05; Table 2). Fat percentage tended to be higher from −4 to −2, then decrease from −2 to calving. In
in the HT heifers during wk 12, and protein percentage addition, there was a week effect for sodium, glucose,
tended to be higher in HT heifers during wk 5, rela- creatinine, and chlorine concentrations (all P < 0.0001;
tive to CL. Fat yield tended to be higher in CL heifers Table 3). Sodium increased from wk −2 to calving,
compared with HT (1.30 vs. 1.18 kg/d, P = 0.08; Table relative to wk −6, glucose levels decreased from wk
2). Protein yield had a treatment by week interaction −4 to −2 and increased from wk −2 to calving, and
(P = 0.003; Table 2) whereby the CL heifers, relative to creatinine concentrations increased from wk −6 to −4,
HT heifers, had higher protein yields during wk 3 and decreased from wk −4 to −2, and increased from wk
4 and wk 7 through 15, with a tendency during wk 6. −2 to calving. Chloride continually increased from wk
−4 to calving, relative to wk −6 concentrations (Table
Blood Electrolytes and Acid-Base Balance 3). Total carbon dioxide tended to be higher in CL
heifers (P = 0.06; Table 3) and BUN was significantly
Anion gap was not different between treatments (18.6 higher (13.1 vs. 11.1 ± 0.73 mg/dL, P = 0.01; Table
vs. 18.7 ± 0.41 mmol/L, CL vs. HT, P = 0.80; Table 3) in the CL heifers, relative to HT heifers. An effect
3). Potassium and ionized calcium had a tendency for of week was observed for TCO2 (P = 0.009; Table 3),
a week effect (P ≤ 0.10; Table 3), whereby concentra- wherein levels peaked at wk −4, decreased from wk −4
tions tended to decrease from wk −6 to −4, increase to −2, and further decreased at calving. There was a

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 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

treatment by time interaction for precalving HCT and wk −8 through −1. Postcalving HCT was not differ-
HGB (P ≤ 0.02; Table 3). Both precalving HCT and ent between treatments (30.0 vs. 30.5 ± 0.68% for CL
HGB were higher in CL heifers, relative to HT, during vs. HT respectively, P = 0.45; data not shown). There
wk −6 and tended to be higher in HT heifers at calving. was a day effect (P = 0.02) for postcalving HCT by
Precalving TP was not different between treatments which levels decreased from calving to postcalving d 42.
(8.87 vs. 8.95 ± 0.15 mg/dL, CL vs. HT respectively, P Postcalving TP was not different between treatments
= 0.59; data not shown) but there was an effect of week (9.34 vs. 9.46 ± 0.17 mg/dL, CL vs. HT respectively,
(P < 0.0001), whereby TP continually decreased from P = 0.48), but there was a day effect (P < 0.0001)
whereby TP increased from calving to postcalving d 28
and decreased from postcalving d 28 to 42.

DISCUSSION

Whereas heat stress in mature dairy cows has been

studied extensively, both during lactation and the dry
period, the effect of heat stress on nulliparous, preg-
nant dairy heifers has received no attention. Herein, we
investigated the effects of providing heat stress abate-
ment in the form of shade, fans, and water soakers to
late pregnant heifers on thermoregulatory responses,
blood electrolytes and acid-base balance, growth, and
milk yield in their first lactation. Our data indicate that
actively cooling dairy heifers before calving (i.e., final
60 d of gestation) is effective in promoting thermoregu-
latory responses and results in greater milk production
after calving.
Homeotherms experience heat stress when the
metabolic and environmental heat load accumulate
beyond their capacity to dissipate heat in the environ-
ment (Bernabucci et al., 2010). During heat stress,
physiological processes (i.e., respiration rates, sweating
rates, and core body temperature) must be altered to
maintain homeostasis (Page et al., 1959). Early studies
have shown that heat stress is experienced by Holstein
cows when THI exceeds 72 (Armstrong, 1994). How-
ever, more recent studies indicate that performance of
high-producing cows begins to decline when THI levels
exceed 68 (Zimbelman et al., 2009). Although THI
thresholds that are detrimental to productivity have
been established for high-producing dairy cows, the
UCT and THI thresholds for nonlactating heifers is un-
known. A greater tolerance for increased environmental
thermoneutral zone (Hahn, 1999) and less metabolic
heat production (West, 2003) prolongs the onset of
physiological responses to heat stress in dry cows, preg-
nant heifers, and calves, but does not prevent them
from experiencing heat stress (Moran, 2002; Piccione et
al., 2003). A growing body of literature has evaluated
the effectiveness of cooling dry, nonlactating cows (Av-
Figure 2. Vaginal temperature measurements from cooled (shade endaño-Reyes et al., 2006; Tao and Dahl, 2013; Ferreira
of a freestall barn, fans, and soakers; n = 12) or heat stressed (only et al., 2016), but little research on cooling youngstock,
shade; n = 14) nulliparous heifers during the last 8 wk of gestation. such as pre-and postweaned calves, and pregnant heif-
Measurements were obtained every 10 min and averaged per hour per
day on wk −8, −6, −4, and −2, relative to calving. Data are graphed ers is available (Neuwirth et al., 1979; Spain and Spiers,
using the LSM ± SE of the interaction [treatment (trt) by time]. 1996; Dado-Senn et al., 2020).
Journal of Dairy Science Vol. 104 No. 2, 2021
 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Figure 3. Sweating rate (A) and skin temperature (B) measurements from cooled (shade of a freestall barn, fans, and soakers; n = 12) or
heat stressed (only shade; n = 14) nulliparous heifers during the last 8 wk of gestation. Relative to calving, measurements were obtained from an
unshaved portion over the right front shoulder blade every 4 h over a 36-h time interval on weeks −6, −4, and −2. **, # indicates significance
(P ≤ 0.05) and tendencies (0.10 ≥ P > 0.05), respectively. Data are graphed using the LSM ± SE of the interaction [treatment (trt) by time].

Not typically considered for heat stress abatement, cooled heifers. Dry (do Amaral et al., 2009; Tao et al.,
dairy heifers are often housed on pasture, exposed to 2011; Fabris et al., 2019) and lactating (Gao et al.,
solar radiation, and provided limited access to natural 2017) cooled cows have lower RT and RR when com-
or artificial shade. In the current experiment, all heifers pared with their heat-stressed counterparts. For dry
were housed under the shade of a freestall barn and ex- and lactating cows, RT differs approximately 0.36°C
perienced similar levels of THI, which was consistently (do Amaral et al., 2009; Tao et al., 2011; Fabris et al.,
above 68 for the duration of the trial. We demonstrated 2019) and 1.5°C (Gao et al., 2017), respectively. Herein,
the effectiveness of actively cooling pregnant heifers CL heifers had 0.12°C lower RT relative to HT heif-
through significant reductions in thermal indicators, ers, which is less than the observed in dry cows under
such as RR, RT, ST, VT, and SR, compared with non- comparable experimental conditions. In addition, RR is
reported to be higher in HT cows, compared with CL,
by approximately 33 bpm in dry cows (Tao et al., 2011)
and 45 bpm in lactating cows (Gao et al., 2017). In
the current study, HT heifers had a 16 bpm higher RR
relative to their CL counterparts. Although actively
cooled multiparous mature cows, both dry and lactat-
ing, had an average RR range between 30 and 52 bpm,
those under HT conditions had a higher RR range of
64 to 78 bpm (Tao et al., 2011; Gao et al., 2017; Fabris
et al., 2019). Herein, heifers under HT conditions had
RR of approximately 60 bpm, which aligns with the
threshold for heat stress in dry cows (Toledo et al.,
2020). Although the direction of our findings are in
accordance with results from dry and lactating cows
under heat stress, the magnitude of the physiological
differences in heifers is less. One possible explanation
Figure 4. Milk yield from cooled (shade of a freestall barn, fans,
and soakers; n = 12) or heat stressed (only shade; n = 14) nulliparous is that heifers generate less metabolic heat per unit
heifers during the last 8 wk of gestation. Production was recorded of surface area and have less heat to dissipate at this
twice daily (Afimilk Ltd., Kibbutz, Afikim, Israel) and averaged by stage of life (West, 2003). Heifers could also potentially
week. **, # indicates significance (P ≤ 0.05) and tendencies (0.10 ≥
P > 0.05), respectively. Data are graphed using the LSM ± SE of the be better at dissipating heat, as a direct benefit from a
interaction [treatment (trt) by time]. lower amount of total surface area.

Journal of Dairy Science Vol. 104 No. 2, 2021

 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Furthermore, in the present study we found that the

Treatment = heifers were cooled (shade of freestall barn, fans, soakers) or heat stressed (shade of freestall barn); weeks = relative to calving, measurements were obtained on weeks
Table 3. Sodium (Na), potassium (K), chloride (Cl), total carbon dioxide (TCO2), anion gap, ionized calcium (iCa), glucose (Glu), BUN/urea, creatinine (Crea), hematocrit

Treatment
magnitude of differences in SR and ST throughout a

× time

0.53
0.89
0.37
0.84
0.75
0.51
0.02

0.02
0.27
0.83
0.33
(HCT), hemoglobin (HGB) concentrations from nulliparous heifers exposed to either cooling (n = 12) or heat stress (n = 14) environments during the last 60 d of gestation1

36-h period was large, but also the temporal changes

across the weeks of treatment were vastly different.
Sweating and movement of blood to the skin surface are
forms of evaporative cooling used to dissipate excess

<0.0001
<0.0001
<0.0001
<0.0001

<0.0001
<0.0001

<0.0001
P-value*

0.009
heat. In our study, heifers under HT had elevated SR
Time

0.50
0.10
0.07
and ST relative to CL heifers and changes in ST and SR
were expected through the day, as demonstrated with
body temperatures patterns for the bovine (Wrenn et
Treatment

al., 1961). However, these treatment differences were

0.06
0.80
0.77
0.98
0.01
0.48
0.53

0.57
0.69
0.47
0.27

smaller and the patterns were similar in the first week

after enrollment, suggesting an acclimation period to
the new environments as the heifers transitioned into
the freestall barn from pasture. As parturition ap-
SEM

0.61
0.41
0.01
8.22
0.74
0.04
0.35

0.12
0.49
0.05
0.59

proached, CL heifers still exhibited lower ST and SR

compared with HT but again the differences between
Calving

treatments were smaller and the heifers appear to be

1.20

8.91
1.21
4.16

23.9
19.1

118.0

29.3

10.0
143.2

105.3

thermoregulating less effectively. It is possible that the

exponential growth of the fetus, coupled with increased
energy demands and simultaneous mammary gland
1.22

9.65
0.99

8.38
4.38
wk −2

colostrogenesis could add to the overall heat load and

24.0
18.5

71.1

24.7
141.8

104.5
Heat stressed

decrease the ability of the heifers to effectively dissipate

that heat in late gestation.
With increased sweating rates, panting, and drool-
1.21

1.04

8.53
4.14
wk −4

25.2
19.0

71.8
14.6

25.1
140.8

101.6

ing as means of evaporative heat loss, changes in

the acid-base balance of the body lead to electrolyte
losses. Potassium is the primary cation lost in sweat
1.23

0.99

8.52
4.24
wk −6
Treatment and week

(McEwan Jenkinson and Mabon, 1973) and loss of the

24.6
18.0

70.8
11.4

25.1
140.9

103.2

*Significance was declared at P ≤ 0.05; tendency was declared at 0.10 ≥ P > 0.05.

other electrolytes is possible through drooling (Shalit et

al., 1991). Bicarbonate (through panting) and sodium
(through urine) excretion increases in hot climatic con-
ditions (Collier et al., 1982a), and consequently, cows
Calving

1.22

1.24

9.52
4.14

24.2
18.6

111.6
11.6

28.0
144.4

106.6

are at higher risk of respiratory alkalosis. Excessive

moisture loss and total carbon dioxide reductions are
observed as a result of increased respiration rates in
1.22

1.03

8.46
4.39
wk −2

heat-stressed cows (Schneider et al., 1988) and goats

25.0
18.5

71.8
12.2

24.9
141.3

103.2

(Hamzaoui et al., 2013). To determine the metabolic

Cooled

effects of heat stress or heat abatement in dairy heif-

ers, we evaluated the acid-base and electrolyte balance
1.20

1.11

8.68
4.24
wk −4

27.5
18.7

73.5
16.3

25.5
141.5

100.4

and circulating metabolites. Although no significant

differences in blood electrolytes were detected between
groups, HT heifers had reduced levels of sodium, potas-
1.21

0.99

9.05
4.36
wk −6

sium, and chloride, relative to CL. These reductions,

25.6
18.3

73.9
12.3

26.7
140.4

101.8

coupled with increased sweating rates in the HT heif-

ers, suggest that HT heifers are negatively affected by
−6, −4, −2, and at calving.

heat stress as indicated by a loss of minerals in sweat.

Increased water consumption is a hallmark of heat
Anion gap,* mmol/L

HCT, % packed cell

stress. Indeed, dry and lactating cows under heat stress

BUN/urea, mg/dL

consume up to approximately 50% more water rela-

TCO2, mmol/L

Crea, mg/dL
iCa, mmol/L

tive to those under active cooling (Fabris et al., 2017;

Na, mmol/L

Glu, mg/dL
Cl, mmol/L

HGB, g/dL
K, mmol/L

McDonald et al., 2020). Moreover, heat stress expo-

Variable

volume

sure can alter HCT and HGB levels in cattle (Sosa

et al., 2010; Tao et al., 2012; Dado-Senn et al., 2020).
1

Journal of Dairy Science Vol. 104 No. 2, 2021

 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

Herein, CL heifers had higher HCT percentages and pregnancy for success in the subsequent lactation.
HGB levels during the precalving period, higher levels Multiparous cows exposed to heat stress during the
of TCO2, and lower RR. Although water consumption dry period has been linked to altered redevelopment
was not recorded in the present study, the decreased and compromised involution of the mammary gland,
hydration status and oxidative status coupled with the specifically by reduced autophagy and epithelial cell
reduced blood TCO2, from higher RR, and reduced proliferation during early and late dry period, respec-
HCO3− experienced by HT heifers might limit the abil- tively (Dado-Senn et al., 2018; Wohlgemuth et al., 2016;
ity of these animals to thermoregulate and carry out Tao et al., 2011). In addition, Dado-Senn et al. (2019)
important metabolic processes efficiently. In the current reported carryover effects of dry period heat stress ex-
experiment, BUN levels were within the normal range posure on mammary gland tissue microstructure (i.e.,
of 10 to 20 mg/dL (Ferguson et al.,1993); however, fewer alveoli and mammary epithelial cells in the HT
BUN tended to be higher in the CL heifers relative to cows compared with CL). Thus, impaired mammary
HT. These results are not consistent with those from growth and alterations in tissue microstructure of the
lactating cows, where BUN is typically higher in HT growing pregnant heifer might explain the reductions
cows relative to CL (Cowley et al., 2015; Gao et al., in milk yield in the subsequent lactation. Although
2017). It is suggested that lower intakes of energy and late-gestation heifers are not undergoing mammary
protein for HT lactating cows is related to the increased involution, the mammary gland is growing exponen-
whole-body amino acid utilization, a decrease in rumen tially. After first conception and continuing through
microbial crude protein synthesis, and decreased deliv- gestation, ductal growth increases and secretory cells
ery of amino acids to the mammary gland, which can proliferate to form lobulo-alveolar tissue (Knight and
result in higher BUN levels (Radkowska and Herbut, Peaker, 1982). Cell differentiation allows for epithelial
2014; Cowley et al., 2015; Gao et al., 2017). However, in cell number and surface area to increase, beginning
the present study, CL heifers experienced higher BUN mid-pregnancy as the mammary gland is initiating
levels even though DMI was not different. This sug- the secretory phase (Richert et al., 2000). It appears
gests that DMI, regardless of treatment or lactating that exposure to heat stress during this critical phase
status, plays a role in BUN levels, which in turn affects of mammary gland development has direct detrimental
metabolic processes of the animals. consequences to subsequent production. Another factor
Heat abatement through the entire dry period results affecting mammary development under heat stress is
in increases of 5 to 7.5 kg/d of milk in the subsequent the reduced blood flow to the mammary gland. Up to
lactation relative to HT in dry cows (do Amaral et al., a 14% decline in blood flow has been observed in ther-
2009; Tao et al., 2011; Thompson et al., 2014; Fabris mally stressed animals (McGuire et al., 1989; Lough
et al., 2017). Moreover, if heat stress abatement is only et al., 1990), as blood flow is redirected to peripheral
provided to cows during the final portion of the dry tissues for cooling (McGuire et al., 1989). Thus, we
period (i.e., last 21 d), milk production improvement speculate that providing heat abatement during late
is 1.4 kg/d (Urdaz et al., 2006; Gomes et al., 2013). gestation might positively affect mammary growth and
Meanwhile, lactating cows provided heat stress abate- blood flow, improving lactational output after calving.
ment increased milk production by 2 kg/cow per day, Future studies are needed to test this hypothesis.
relative to HT lactating cows whose milk production Overall, our study demonstrates a strong and
can further decrease by 0.23 up to 0.59 kg/cow per day positive thermoregulatory response of dairy heifers
with each unit increase of THI above 68 and based on to active cooling with fans and water soakers in late
geographic location (Igono et al., 1987; Bouraoui et al., gestation. Although the magnitude of those responses
2002; Bohmanova et al., 2007). In the present study, are less pronounced than that of mature multiparous
heifers cooled during the last 60 d of gestation average dry cows, these alterations translate into production
a 3.9 kg/d increase in milk in the first 15 wk of lacta- improvements postcalving. More studies are needed to
tion relative to HT heifers, indicating that CL heifers confirm the production responses, perhaps with larger
benefited from their active cooling system. numbers of animals, and to unravel the molecular and
Initial mammary gland involution and later redevel- metabolic implications of the observed responses result-
opment during the dry period is critical for maximal ing in higher yields.
subsequent production (Wolfenson et al., 1988; Capuco
et al., 2001). Indeed, it is estimated that 50% of mam- ACKNOWLEDGMENTS
mary epithelial cells are carried over from one lactation
to the next (Pitkow et al., 1972; Hurley, 1989), sug- We thank the University of Florida Dairy Unit staff
gesting mammary cell turnover, through the processes and veterinarians for their assistance in animal care
of involution and redevelopment, is crucial during late and data collection. Further acknowledgment goes to
Journal of Dairy Science Vol. 104 No. 2, 2021
 Davidson et al.: DAIRY HEIFER HEAT STRESS DURING LATE GESTATION

the research interns and volunteers who assisted on this J. Dairy Sci. 103:4822–4837. https:​/​/​doi​.org/​10​.3168/​jds​.2019​
-17926.
experiment. This study was funded by the Harriet B. De Rensis, F., I. Garcia-Ispierto, and F. López-Gatius. 2015. Seasonal
Weeks endowment to G.E. Dahl and the USDA-NIFA heat stress: Clinical implications and hormone treatments for the
AFRI Foundational Program Award 2019-67015-29445 fertility of dairy cows. Theriogenology 84:659–666. https:​/​/​doi​
.org/​10​.1016/​j​.theriogenology​.2015​.04​.021.
to J. Laporta and USDA-NIFA Hatch 1021876 project Dikmen, S., and P. J. Hansen. 2009. Is the temperature-humidity in-
to G.E. Dahl. The authors have not stated any conflicts dex the best indicator of heat stress in lactating dairy cows in
of interest. a subtropical environment? J. Dairy Sci. 92:109–116. https:​/​/​doi​
.org/​10​.3168/​jds​.2008​-1370.
do Amaral, B. C., E. E. Connor, S. Tao, M. J. Hayen, J. W. Bubolz,
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