Bruising in Slaughter Cattle and Its Relationship with Creatine Kinase Levels and Beef Quality as Affected by Animal Related Factors

Description of the study site

The study was conducted in East London Abattoir which is located in Buffalo City Municipality. The abattoir is situated at longitude and latitude 32.′2° S and 27.′ 5° E in the Amathole District of the Eastern Cape Province of Republic of South Africa (RSA). The area has vegetation that ranges from grassland and thicket to forests and bush veld with Themeda trianda and Digitaria eriantha being the most dominant grass species. The area receives approximately 850 mm of rainfall per year of which most falls during the summer months (November, December and January). The day temperature during for the period of study ranged from 10°C to a high of 36°C, averaging about 22.3°C. The topography of the area is naturally flat with few steep slopes.

Animal management

Data was collected from 321 cattle over four seasons (winter, spring, summer and autumn) between the year 2011 and 2012. The seasons were categorised into the months as follows: winter (June to August), spring (September to October), summer (November to February) and autumn (March to May). All cattle were slaughtered at the East London commercial abattoir in the Eastern Cape Province. This abattoir operates from 0600 to 1700 h on Mondays till Fridays. On arrival at the abattoir the three breeds of cattle; Bonsmara, Brahman and Beefmaster were identified according records from the farmers. The numbers of animals per breed were 108 Bonsmara, 130 Beefmaster and 83 Brahman respectively. The animals used were both male and females and grouped according to age categories as follows: Group 1 (16 months), Group 2 (18 months) and Group 3 (24 months).

The cattle were transported by the owners from the farms to the abattoir. Transportation time (departure, arrival time and distance travelled), lairage duration, number of animals per pen, slaughtered animals per day and daily temperature were all recorded. On arrival at the abattoir, 27 offloading activities were monitored and cattle were given water ad-libitum prior to slaughter. Cattle were then slaughtered using commercial standard procedures of the abattoir. The captive bolt method was used to stun the animals, using a voltage of 300 V, a current of 5 A for 40 to 45 s at a pulse 12 beats/s.

Blood collection and plasma separation

The blood samples were collected from the animals using labelled disposable vacutainer tubes. The blood samples were placed in ice packs prior to creatine kinase (CK) analysis. A centrifuge machine (model: 5403 Centrifuge Geratebay Eppendorf GmbH, Engelsdorp Germany) was used to centrifuge blood samples at 3,550 rpm, 21°C for 10 min. Plasma was then separated and inserted into 1.5 mL Eppendorf tubes using pipettes and stored at −20°C.

Creatine kinase level determination

The serum samples were used to determine the levels of CK at the National Health Laboratory Services (NHLS), Port Elizabeth using Model DXC 600 machine (Beckman Coulter, Ireland). The contents included 2×61 ML CK Reagent, 1 preparation insert and the reactive ingredients included creatine phosphate, disodium Salt 461 mmol/L, nadide 30: 0 mmol/L, adenosine-5-diphosphate, monopotassium salt, dihydrate 36.0 mmol/L, glucose 24.0 mmol/L, glucose-6-phosphate dehydrogenase 46.1 kU/L and hexokinase 136 kU/L. All the ingredients were added for quantitative determination of CK activity of units per litre (U/L) in plasma.

Bruise measurements

Only carcasses that had bruises were considered. The assessment was done using a method based on the Australian Carcass Bruise Score System (Anderson and Horder, 1979). Bruise scores were calculated for each load by multiplying the number of bruises in each size class by a weighting factor: slight 1, medium 3, and heavy 5, and adding these values (Anderson and Horder, 1979). The bruise scores were then divided by the number of cattle per load to give a mean bruise score per animal for each load. Bruise age was estimated using the method of Gracey et al. (1999) as shown in Table 1.

Table 1

Colour observations used to estimate the age of bruises

Meat quality measurements

The Muscularis longisimus thoracis et Lumborum (LTL) were used for ultimate pH and colour (L*, a*, and b*) measurements. The muscle was removed by cutting a sample between the 4th and 6th ribs of the loin region while the carcass was still hanging, prior to vacuum-packing and storing in a cooler box an hour after collection and further storing in the refrigerator at −4°C.

Colour measurement

Colour of the meat (L* = lightness, a* = redness and b* = yellowness) for the Bonsmara, Beefmaster and Brahman carcasses was determined from the Muscular longissimus thoracis et. Lumborum (LTL) 24 h after slaughter using a colour-guide 45/0 BYK-Gardener GmbH machine with a 20 mm diameter measurement area and illuminant D65-day light, 100 standard observer. Three readings were taken by rotating the Colour Guide 900 machine between each measurement, in order to obtain a representative average value of the colour. This guide was calibrated each day before taking measurements using the green standard.

Determination of meat pH

Meat pH 24 from Bonsmara, Beefmaster and Brahman of carcasses was measured from the Muscularis longissimus thoracis et lumborum muscle at the level of the 10th rib 24 h after slaughter on carcasses that were stored at 0 to 3°C. The measurement was carried out using a portable pH meter (CRISON pH 25, CRISON Instruments, SA Spain). The pH meter was calibrated using pH 4, pH 7 and pH 9 standard solutions (CRISON Instrument, SA Spain) before each day’s measurement. The measurements were then performed with a sharpened metal sheath to prevent probe breakage and meat contamination.

Statistical analysis

The PROC GLM of Statistical Analysis System (SAS) Institute (2003) was used to analyse the effect of breed, age, gender and season on the activity of creatine kinase and bruising score. The model used was as follows:

where,

Significant differences between least square group means for CK and animal factors were compared using the Duncan’s procedure. Pair-wise comparison of LS means for creatine kinase concentration, sex, breed and animal age at slaughter were done using PDIFF.

The effect of breed, age group and season on CK, pH_u, L,* a*, and b*, were analysed using Proc GLM of SAS (2003). The stastical model used as follows:

where,

The significant difference between least square group means for CK levels and beef quality parameters were compared using Duncan multiple range procedure. Correlations were computed using PROC GLM of Statistical Analysis System (SAS) (2003) to determine the relationship between pre-slaughter condition on CK and meat quality attributes.

The effect of breed on the levels of plasma creatine kinase

The Bonsmara (Table 2) breed had significantly higher (705.3±80.57 U/L) creatine kinase (CK) levels than Brahman (461.8±80.63 U/L), and Beefmaster breed (657.4±80.63 U/L) respectively. The higher CK activity in Bonsmara indicate that the breed was more susceptible to stress before slaughter and also could have been due to rough driving and mishandling before and after transportation (Broom et al., 1996; Averos et al., 2008). Warries et al. (1995) and Early et al. (2010) highlighted that animals transported for periods of between 5 to 15 h are susceptible to stress which increases the activity of creatine kinase. The reaction of this breed to longer transportation is perplexing because it has been highlighted that Bonsmara are able to withstand harsh and unfamiliar conditions (Ndlovu et al., 2008). However, if animals are transported for longer hours they have reduced plasma CK levels if they are given feed and water at the lairages.

Table 2

Least square means (±SE) for the effect of breed on activity of creatine kinase cattle at slaughter

The lower the CK levels in Brahman and Beefmaster showed that these two breeds were not susceptible to stress during transportation as compared to Bonsmara breed. This is because these are synthetic breeds which can adapt well to handling and adverse environmental conditions (Beatty et al., 2006). The activity of CK increases in the blood after tissue damage, poor muscular tissue reperfusion, decreased heat and fatigue; apparently as a result of an increase in the permeability of the muscle membranes induced by capture, loading and transportation (Lopez et al., 2006; Guardia et al., 2009). Furthermore, other animals also are susceptible to transport stress leading to highest levels of CK in the blood (Yu et al., 2009).

Effect of sex and age on plasma creatine kinase levels

Male animals had the highest CK values (p<001) compared to female animals as shown in Table 3. Males were more stressed than females and this agrees with the study by Mota-Rojas et al. (2006) which indicated that the sex of the animal has an effect on the response of the animal to stress. This is because male animals are more excitable, aggressive and stress sensitive than females. Therefore, that could be the reason of increased plasma activity of CK found in steers transported for 12 to 16 h. Similar results were reported by Neal et al. (2009) who reported that male humans had higher levels of CK.

Table 3

Least square means (±SE) for the effect of sex and age of animal on the activity of creatine kinase levels of cattle at slaughter

Creatine kinase levels were higher (p<0.05) in 18 months old animals than in 16 and 24 months old animals as shown in Table 3. According to Smith et al. (1998) the different levels of CK in the older animals is not well understood although it might be due to muscle fatigue as a result of strenuous exercise. Similarly, the CK levels in 16 month old animals was higher (p<0.05) than in 24 month old. Young animals are poorly adapted to cope with transportation related stress thus it results in high rates of injuries. Young animals respond to transportation stress by increasing their body temperature, heart rate and plasma cortisol concentration (Steinhardt and Thielscher, 1999). The findings of the current study are similar to those reported by Parrot et al. (1994), who observed that young animals are at risk of more bruising than older animals resulting in high levels of CK. This is also linked with CK in muscles as it is expressed in higher levels in males and younger animals (Neal et al., 2009).

Effects of season on bruising score and the levels of plasma creatine kinase

Results on the effect of season on the bruising score and CK levels are shown in Table 4. Creatine kinase (CK) was higher during the winter season (704.1±43.85 U/L) and lower in the summer season (181.3±124.09 U/L), while spring (698.3±59.19 U/L) and autumn (658.0±116.25 U/L) had intermediate CK levels. Knowles et al. (1998) observed similar results and found that CK levels were higher in winter season due to poor pre-slaughter conditions. However, other authors reported an increased CK values in lambs as a result of physical stress during summer (Lowe et al., 2002; Miranda-de la Lama et al., 2011). Wilson et al. (1990) and Liu et al. (2008) reported that vigorous physical activity and high temperatures had an effect on CK level.

Table 4

Least square means (±SE) for effects of season on bruising score and CK levels of cattle

Autumn had the highest (p<0.05) bruising score (1.86±1.07), while winter (1.44±0.13) and spring (1.32±0.54) had intermediate scores and the summer season had the least bruising score (1.12±0.11) as shown in Table 3 and 4. Seasonal factors mainly temperature and humidity also seem to play a role on the prevalence of bruises. Results from this study concur with Gosalvez et al. (2006) and dalla Costa et al. (2007), who found that cold weather combined with precipitation, increased the mean number of bruises per animal, although the reasons are still unclear.

Effect of breed on meat quality

Ultimate pH was significantly affected by the breed of cattle as shown in Table 5. Beef from Beefmaster and Brahman breeds had significantly higher (p<0.05) pH_u values of 6.3±0.05 and 6.1±0.06, respectively compared to that of the Bonsmara breed which had an average value of 5.6±0.05. The pH_u levels of Bonsmara were within the optimal range and similar to those reported by Muchenje et al. (2008). Different temperament found within an individual breed can influence meat quality due to inherent genetic parameters. The findings of this study are similar to the findings by Grandin (1997), Mormede et al. (2002), Fazio and Ferlazo (2003) who reported that breed temperament influence the degree to which animals respond to stress.

Table 5

Effects of breed on ultimate pH, colour of meat and the activity of creatine kinase from beef cattle (±SEM)

The higher pH values obtained from Beefmaster and Brahman could be due to stress prior to slaughter, rough handling, inclement temperatures or anything that causes the animal to draw on its glycogen reserves before slaughter. These two breeds are known to have aggressive behaviour and are more prone to injuries resulting in carcass damage and poorer meat. The current results are similar to observations by Voisinet et al. (1997) and King et al. (2006) who found that cattle which are highly temperamental had a high incidence of borderline dark cutters than calm cattle. Brahman breed had a significant effect (p<0.05) on L* values, with darker meat than Bonsmara and Beefmaster breeds. The darker meat produced by the Brahman breed in comparison to the Bonsmara show similar trends to the findings by O’Neill et al. (2006). O’Neill et al. (2006) observed that indigenous breeds such as Bonsmara released more catecholamines than exotic breeds during the pre-slaughter period, causing the depletion of glycogen.

Bonsmara and Beefmaster had the highest a* values with Brahman having the least a* values. The a* values ranged from 17.5±0.53 in meat from Bonsmara to 13.2±0.66 in meat from Brahman. The a* values from these breeds were within the normal range. This is because pH_u values were fairly normal and colour is not greatly affected by pre-slaughter conditions such as transportation. Bonsmara and Beefmaster breeds had the highest b* values (12.8±0.53) while the lowest b* values (9.6±0.61) were found from the Brahman breed. This could be due to the extent of stress during handling and transportation depending on the temperament, handling during the journey, condition of the animal, and duration of the journey resulting in pale colour of animals.

Effect of age on meat quality

The effect of age on meat quality attributes is shown in Table 6. Meat from 16, 18, and 24 months old animals had no effect on pH_u. The pH_u values were 6.1 in meat from 16, 18, and 24 months old. The pH_u values from 16, 18, and 24 months old cattle were above the normal pH range. Meat with an ultimate pH that is above 6.0 is categorised as a dark firm dry (DFD) meat (Bartos et al., 1993; Mounier et al., 2006) with increased tenderness (Silva et al., 1999) and poor palatability (Viljoen et al., 2002; Wulf et al., 2002). This can also be attributed to the fact that an animal’s behaviour differs with its age (Terlouw, 2005). The age of the group had an effect (p<0.05) on L* values (Table 3 and 6). The L* values ranged from 25.2±0.71, 26.6±0.83, and 20.9±0.73 for 16, 18, and 24 months old cattle respectively.

Table 6

Effect of age on ultimate pH, colour of meat and the activity of creatine kinase from cattle slaughtered at the abattoir (±SEM)

Darker L* values were observed in 24 months old animals indicating meat becomes darker with increasing animal age. These results are similar to findings by Funghi et al. (1994) who found that L* values decreased with an increase in slaughtering age. Zhang et al. (2005) found that high pH meat had lower L* (lightness), a* (redness) and b* (yellowness) values than normal pH meat, indicating that high pH meat is darker and less brown than in normal pH meat. Meat from 16 month old animals had the highest a* and b* (17.9±0.53) and (12.9±0.49) and the meat from 24 months old animals had the least values of (12.9±0.54) and (9.3±0.51) respectively. Differences in meat colour has been associated with variations in intramuscular fat, moisture content and is age dependent on the muscle myoglobin content (Lawry, 1974).

Effect of breed on bruising scores of beef animals

Effect of breed on the bruising scores of slaughtered beef animals are shown in Figure 1. The Bonsmara breed had the highest percentage of non-bruised meat (88%) and the Brahman had the lowest none bruised meat (49%). Bonsmara also had highest percentage 20% of slightly bright red bruised meat (0 to 10 h old) while Brahman had lowest percentage of bright red bruised meat 4%. Susceptibility to bruising and temperament differs between individual animals. Fordyce et al. (1985) concluded that bruising score was beyond the breed differences. It appears that breed differences can be attributed to differences in behaviour and either being horned or hornless (Minka and Ayo, 2007).

Figure 1

Effect of breed on bruising scores of slaughtered beef animals. None = 0 h; slightly = 0 to 10 h old, red and hemorrhagic (bright-red); medium = 24 h old, dark- red colour; severe = 24 to 38 h old, watery consistency.

Bonsmara had the highest percentage 10% medium-bruised dark red (24 h old) with Brahman having the lowest bruised dark red meat 5%. This could have been due several injuries associated with road transportation. The level of bruising might increase with the length of journey travelled by the animals (Weeks et al., 2002). Brahman and Beefmaster breeds had the lowest percentage of severe deep bleeding bruising or injuries (24 to 38 h old).

Effect of age on bruising scores of beef animals

The highest number of slightly bright red bruises was observed in 16 month cattle while the 24 month old cattle had the lowest slightly bright bruised meat (Figure 2). The highest medium dark red bruises was observed in 16 month and lowest medium dark red bruise was observed in the 24 month old cattle. Young animals are more prone to bruising than old animals and this might have been due to increased poor handling (Strappini et al., 2009). Similar results were reported by von Holleben et al. (2009) indicating that the number heifers that had severe bruises were higher than either bulls or cows. However, contradicting results reported by Wythes and Shorthose (1991) highlighted that heavy animals had more bruising than young animals and mature ones and between old cows and old steers of the group. This therefore could mean that cattle bruising does not depend on the age of the animal but is mainly determined by the type of vehicle, duration of transport and handling methods used prior and after transport.

Figure 2

Effect of age on the bruising scores of cattle. None = 0 h; slightly = 0 to 10 h old (red and hemorrhagic bright-red); medium = 24 h old (dark-red colour); severe = 24 to 38 h old (watery consistency).

Effect of sex on bruising scores of slaughtered beef animals

Highest levels of slightly bruised meat and dark bruised meat were observed in female animals, while male animals had the lowest slightly bright bruised meat and medium dark bruised meat (Figure 3). In the literature there is evidence that female animals had higher chances for bruising than male animals (Yeh et al., 1978; Javis et al., 1995). These results are similar with Yeh et al. (1978) who found that heifers had significantly more bruises than steers and also when animals kept as separate groups, cows bruise significantly more than steers and bulls. Cows were the only class of animal where the amount of bruising increased with duration of the journey. Both male and female animals had a lower percentage of severe deep bruises.

Figure 3

Effect of sex on bruising scores of cattle. None = 0 h; slightly = 0 to 10 h old (red and hemorrhagic bright-red); medium = 24 h old (dark- red colour); severe = 24 to 38 h old (watery consistency).