Elisabeth Kommisrud

Behavior of lactating Holstein-Friesian cows during spontaneous cycles of estrus.

The objectives of the present study were to describe, in detail, behavior associated with standing estrus (STE) in lactating dairy cows and behavioral changes during complete estrous cycles. Estrus signs were monitored by continuous video recording of 20 Holstein-Friesian (HF) cows housed on an outdoor wood-chip pad during 1 estrous cycle (22 d). Other social behavior was recorded during STE and, for comparison, during 1 selected day when none of the cows were in estrus. Standing stationary when mounted was defined as the primary estrus sign. Anogenital sniff, chin rest, attempt to mount, and mount were defined as secondary estrus signs. Ovarian cyclicity was confirmed by progesterone measurements. This study reports short mean duration of STE (7.1±1.44h) and estrus (mount period; 12.9±1.84h) of the 13 cows expressing these signs. All mounting activities involved at least one cow in, or within 4h of, STE. The most frequent sign during STE was anogenital sniff initiated, followed by chin rest received, chin rest initiated, chase up initiated, anogenital sniff received, mount initiated, head butt, mount received, attempt to mount initiated, push away received, play rub, attempt to mount received, follow initiated, threat received, flehmen, avoid, bellow, and social lick received. Standing and mounting activity in HF cows was inconsistent during estrus, indicating that other signs could be of greater use. The frequency of secondary estrus signs initiated and received increased gradually during the last 12h before STE, revealing significant differences between periods from 4 to 6 and 1 to 3h before STE. A considerable increase in receptive behavior (secondary estrus signs received) was identified between 1 to 3h prior to STE and STE. Both frequent initiated and received behaviors were associated with STE. A significant decrease in the frequency of secondary estrus signs initiated and received occurred 3h after STE. Cows in STE simultaneously predominantly chose the other standing cow as mate and expressed secondary estrus signs more frequently. Based on the results of this study, we suggest that chase up could be regarded as a reliable indicator of estrus and that the changes in proceptive (initiated) and receptive (received) behavior could be used as predictors of different stages in estrus. Knowledge of these behavioral signs may improve heat detection rates and the ability to predict the optimum breeding time for dairy cows.

Sexually active groups in cattle-A novel estrus sign

The current study presents a novel objective measure for characterizing sexually active groups (SAG 3-5) and relates this measure to other behaviors of lactating Holstein-Friesian cows. Cows in SAG 3-5 were required to participate in a minimum of 1 estrus behavior per 5min while staying within 3m (2 cow lengths) of its partner(s) for a minimum of 5min. Twenty Holstein-Friesian cows were video-monitored continuously through 1 complete estrous cycle (22d). Standing behavior, SAG 3-5, secondary estrus signs (SEC), and other social and agonistic behaviors were recorded continuously. The period of mounting estrus (MTE) was divided into the 3 parts: prestand, standing estrus (STE), and poststand. The mean durations of MTE, prestand, STE, and poststand period were 12.9±1.84, 4.0±1.93, 7.1±1.44, and 1.8±0.57h (n=13). The fractions of time spent in SAG 3-5 during MTE, prestand, STE, and poststand period were 13, 8, 19, and 1% (n=11). During MTE, cows participated, on average, in 5.8±1.24 SAG 3-5 and initiated 9.5±2.99 mounts, with mean durations of 0.25±0.03h and 4.00±0.36s, respectively. The novel measure SAG 3-5 was a sign of long duration not confined only to groups of STE cows. On one day when no cows were in estrus and during the periods 4 to 24h before and after MTE, no SAG 3-5 behaviors were observed. Luteal-phase cows participated in SAG 3-5 only when the partner was a single cow in estrus. The time spent in SAG 3-5 increased between 1 and 3h before MTE and the prestand period (3 vs. 8%) and reached a peak level during STE. From STE to poststand, time spent in SAG 3-5 decreased considerably (19 vs. 1%). The observed decrease in nonmutual agonistic behaviors 4 to 24h before MTE is suggested as an early sign of pre-estrus. Changes in SAG 3-5, agonistic behaviors, and SEC are suggested as indicators of the specific stages of MTE. Increased SEC initiated and SAG 3-5 were indicators of late pre-estrus and early estrus (prestand). Peak levels of SAG 3-5, SEC, and social agonistic behaviors were indicators of STE. A sudden decrease in behaviors, preceded by frequent interactions, was indicative of late estrus (poststand). On the basis of the findings reported here, we propose that SAG 3-5, as well as proceptive and receptive patterns of SEC and agonistic behaviors, be included in estrus detection protocols. Updated knowledge of these behavioral interactions may assist when determining the stage of estrus and the optimal time to breed dairy cows.

Use of immobilized cryopreserved bovine semen in a blind artificial insemination trial

To make timing of artificial insemination (AI) relative to ovulation less critical, methods for prolonging shelf life of spermatozoa in vivo after AI have been attempted to be developed. Encapsulation of sperm cells is a documented technology, and recently, a technology in which sperm cells are embedded in alginate gel has been introduced and commercialized. In this study, standard processed semen with the Biladyl extender (control) was compared with semen processed by sperm immobilization technology developed by SpermVital AS in a blind field trial. Moreover, in vitro acrosome and plasma membrane integrity was assessed and compared with AI fertility data for possible correlation. Semen from 16 Norwegian Red young bulls with unknown fertility was collected and processed after splitting the semen in two aliquots. These aliquots were processed with the standard Biladyl extender or the SpermVital extender to a final number of 12 × 10(6) and 25 × 10(6) spermatozoa/dose, respectively. In total, 2000 semen doses were produced from each bull, divided equally by treatment. Artificial insemination doses were set up to design a blinded AI regime; 5 + 5 straws from each extender within ejaculates in ten-straw goblets were distributed to AI technicians and veterinarians all over Norway. Outcomes of the inseminations were measured as 56-day nonreturn rate (NRR). Postthaw sperm quality was assessed by flow cytometry using propidium iodide and Alexa 488-conjugated peanut agglutinin to assess the proportion of plasma membrane and acrosome-intact sperm cells, respectively. In total, data from 14,125 first inseminations performed over a 12-month period, 7081 with Biladyl and 7044 with SpermVital semen, were used in the statistical analyses. There was no significant difference in 56-day NRR for the two semen categories, overall NRR being 72.5% and 72.7% for Biladyl and SpermVital, respectively. The flow cytometric results revealed a significant higher level of acrosome-intact live spermatozoa in Biladyl-processed semen compared to SpermVital semen. The results indicate that the level of acrosome-intact live spermatozoa in the AI dose did not affect the 56-day NRR for the two semen processing methods. In conclusion, this study has showed that immobilized spermatozoa provide equal fertility results as standard processed semen when AI is performed in a blinded field trial, although the immobilization procedure caused increased sperm damage evaluated in vitro compared to standard semen processing procedure.

Comparison of electronic volume and forward scatter principles of cell selection using flow cytometry for the evaluation of acrosome and plasma membrane integrity of bull spermatozoa

The objective of the study was to compare two different flow cytometers to reveal if there are differences between them and to find the most suitable protocol for analysis of spermatozoa. These two flow cytometers; Cell Lab Quanta™ and Coulter Epics XL, have different principles to calculate cell size, electric volume, and forward scatter (FS), respectively. Flow cytometry is a valuable tool to assess various spermatozoa quality traits simultaneously, such as plasma membrane and acrosome integrity. A double‐ and triple‐stain combination was performed to compare evaluation of these two parameters by both flow cytometers and to assess the need of a fluorescent probe to identify the spermatozoa. Propidium iodide was used to assess the proportion of dead spermatozoa, whereas Alexa Fluor® 488 conjugated peanut agglutinin (PNA‐ Alexa 488) was used to evaluate the percentage of acrosome intact and acrosome–reacted cells or degenerated cells. In the triple‐stain protocol, MitoTracker® Orange (MO) was included to test the capacity of this probe to discriminate spermatozoa from egg yolk and debris particles present in the semen sample. Cryopreserved semen from 13 Norwegian Red bulls was included in the study and the semen was evaluated immediately after thawing and after 3 hr incubation at 37°C. The results show that there is good agreement between the instruments. Nevertheless, a significant difference was found in percentages of acrosome intact live spermatozoa (% AIL) when including MO as a spermatozoa identification probe, compared to assessment without MO, with the Coulter Epics XL, while no significant difference was found when including the probe with the Cell Lab Quanta. In conclusion, the results show that cell size measurement based on electronic volume used by the Cell Lab Quanta flow cytometer is more accurate than FS used by the Coulter Epics XL flow cytometer in identification of spermatozoa.

Comparison of Holstein-Friesian and Norwegian Red dairy cattle for estrus length and estrous signs

This study addressed the effect of breed on estrus length and estrous behavior by observing 20 Holstein-Friesian (HF) and 20 Norwegian Red (NRF) cows on an outdoor wood-chip pad through 1 estrous cycle (22d). Detailed behavioral data were collected by continuous (24 h) video monitoring of all cows. Accurate estimation of duration of estrous periods, behavioral signs (sum per period and counts per hour), and duration and number of sexually active groups were reported through all stages of mount estrus (prestand, standing estrus, and poststand). These dependent variables were analyzed with a basic statistical model that included fixed effects for breed and lactation group. Other independent variables (milk yield, body condition score, and number of cows in standing estrus) were added to the basic model one by one and included in an expanded model if they had an effect on the respective dependent variables. Estrus duration was considerably shorter in HF compared with NRF cows for all the major periods: mount estrus (11.2 ± 3.0 vs. 21.3 ± 2.7 h), standing estrus (7.1 ± 1.4 vs. 11.7 ± 1.3 h), mounting period (6.9 ± 2.7 vs. 18.2 ± 2.4 h), and mounted period (9.2 ± 2.8 vs. 17.5 ± 2.6 h). Additionally, the NRF cows spent more time in sexually active groups (36.1 ± 4.0 vs. 17.6 ± 4.8%) during standing estrus compared with HF cows. The NRF cows participated in a greater number of sexually active groups (9.6 ± 1.3 vs. 5.5 ± 1.3) with longer average duration (0.42 ± 0.04 vs. 0.20 ± 0.04 h) and continued to be more active in these groups through late stages of estrus (poststand) compared with the HF breed. Mounting activity differed between breeds as NRF mounted more times in total (46.3 ± 6.2 vs. 18.1 ± 6.3) and per hour (2.6 ± 0.4 vs. 1.5 ± 0.5) during mount estrus. In addition, NRF tended to express the primary estrous sign, standing when mounted, more often during standing estrus (32.4 ± 5.0 vs. 18.5 ± 5.2). The HF initiated more unsuccessful mounts (1.6 ± 0.3 vs. 0.6 ± 0.3) per hour than did NRF during mount estrus. A significant effect of milk yield was demonstrated only on this behavior. For other estrous signs, HF cows initiated chase-up (2.0 ± 0.5 vs. 0.5 ± 0.4) and anogenital sniff (3.7 ± 0.6 vs. 2.0 ± 0.5) more frequently (counts per hour), whereas NRF expressed more total head butt behavior (32.3 ± 4.7 vs. 14.2 ± 4.8) during mount estrus. Body condition score had a significant effect on receptive behavior. Measures of estrus duration, sexually active group activity, and behavior related to estrus should be subjected to larger studies for improved heat detection and possible implementation in breeding programs.

Impact of brominated flame retardants on embryo development of Atlantic Cod (Gadus Morhua) during early life stages

Brominated flame retardants (BFRs) are ubiquitous industrial chemicals likely to persistently exist in the environment, bioaccumulate in food chains, and even may cause adverse health effects in human. Borgundfjorden fjord system, an important spawning ground for the Norwegian coastal cod (Gadus morhua) stock, was contaminated by significant levels of pollutants such as BFRs, due to the local previous industrial activities. In this study, we demonstrated high level of the BFRs polybrominated biphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) in cod liver and gonad samples from Borgundfjorden using mass spectroscopy (MS) detectors by Norwegian National Institute of Nutrition and Seafood Research (NIFES). Acute embryo toxicity test was further conducted using fertilized cod eggs. The eggs were short-term exposed to serial dilutions of five BFRs mixtures, BDE-47, or PCB mixture Aroclor 1254. At a concentration 10 times that detected in cod liver, the mixture of the five BFRs significantly reduced the embryo survival rate (p <; 0.01). Correspondingly, at 224 μg/L, which was around 10 times of that detected in cod liver, BDE-47 exhibited obvious cod embryo toxicity (p <; 0.01). As a positive control, Aroclor 1254 significantly reduced the embryo survival rate at 400 and 1600 μg/L (p <;0.001). This experiment has laid the foundation for further research on environmentally hazardous impact on the reproductive capacity of aquatic organisms, which will directly influence the fish stocks growing potential and thereby the Norwegian fishery activity.

Stress Resilience of Spermatozoa and Blood Mononuclear Cells without Prion Protein

The cellular prion protein PrPC is highly expressed in neurons, but also present in non-neuronal tissues, including the testicles and spermatozoa. Most immune cells and their bone marrow precursors also express PrPC. Clearly, this protein operates in highly diverse cellular contexts. Investigations into putative stress-protective roles for PrPC have resulted in an array of functions, such as inhibition of apoptosis, stimulation of anti-oxidant enzymes, scavenging roles, and a role in nuclear DNA repair. We have studied stress resilience of spermatozoa and peripheral blood mononuclear cells (PBMCs) derived from non-transgenic goats that lack PrPC (PRNPTer/Ter) compared with cells from normal (PRNP+/+) goats. Spermatozoa were analyzed for freeze tolerance, DNA integrity, viability, motility, ATP levels, and acrosome intactness at rest and after acute stress, induced by Cu2+ ions, as well as levels of reactive oxygen species (ROS) after exposure to FeSO4 and H2O2. Surprisingly, PrPC-negative spermatozoa reacted similarly to normal spermatozoa in all read-outs. Moreover, in vitro exposure of PBMCs to Doxorubicin, H2O2 and methyl methanesulfonate (MMS), revealed no effect of PrPC on cellular survival or global accumulation of DNA damage. Similar results were obtained with human neuroblastoma (SH-SY5Y) cell lines stably expressing varying levels of PrPC. RNA sequencing of PBMCs (n = 8 of PRNP+/+ and PRNPTer/Ter) showed that basal level expression of genes encoding DNA repair enzymes, ROS scavenging, and antioxidant enzymes were unaffected by the absence of PrPC. Data presented here questions the in vitro cytoprotective roles previously attributed to PrPC, although not excluding such functions in other cell types or tissues during inflammatory stress.