1

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Effect of feed texture, meal frequency and pre-slaughter fasting on 2

behaviour, stomach content and carcass microbial quality in pigs 3

4

L. Saucier1, D. Bernier1, R. Bergeron1, A. Giguère2, S. Méthot2 and L. Faucitano2 5

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1Département des sciences animales, Faculté des sciences de l’agriculture et de l’alimentation, 8

Université Laval, Quebec City, QC, Canada, G1K 7P4 9

(e-mail: linda.saucier@san.ulaval.ca); 2Agricultre and Agri-Food Canada, Dairy and Swine 10

Research and Development Centre, 2000 College Street, Sherbrooke, QC, Canada, J1M 1Z3 11

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Running title: Pre-slaughter management and carcass microbial quality 13

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Abbreviations: AAFC, Agriculture and Agri-Food Canada; FT, food texture; MF, meal 15

frequency; WT, feed withdrawal time; SEM, standard error of the mean; TAM, 16

total aerobic mesophilic; MPN, Most Probable Number; Log A, Log means; 17

, mean Log values; SD, standard deviation; cfu, colony forming unit; 18

CFIA, Canadian Food Inspection Agency. 19

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Saucier, L., Bernier, D., Bergeron, R., Giguère, A., Méthot, S. and Faucitano, L. 2007. Effect of 1

feed texture, meal frequency and pre-slaughter fasting on behaviour, stomach content and 2

carcass microbial quality in pigs. Can. J. Anim. Sci. xxxxx. In this study, behaviour in lairage, 3

weight and composition of stomach contents and carcass microbial contamination were studied 4

in 96 barrows assigned to the following treatments: feed texture (FT; mash vs. pellets), meal 5

frequency (MF; 2 vs. 5 meals per day) and fasting time (WT; 4, 14 and 24 h) according to a 2 x 2 6

x 3 factorial design. Pigs fed two meals had heavier stomach weights at slaughter than those fed 7

five times per day (P = 0.01). An interaction was found between WT and FT (P = 0.002) for 8

stomach weight. With respect to the contamination of the mouth, Total Aerobic Mesophilic 9

counts were higher than 104 cfu/cm2 but not significantly different between treatments. 10

Coprophagy behaviour in lairage was not correlated with mouth contamination at slaughter. The 11

treatment resulting in the lowest Escherichia coli counts on the thoracic area was feeding the 12

pigs pellets five times per day followed by a 24 h fast. In contrast, the highest E. coli counts were 13

observed in pigs fed mash five times per day followed by a 4 h fast. Comparison a posteriori of 14

these two extreme scenarios yielded a P-value of 0.03. 15

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Key words: animal behaviour, carcass hygiene, fasting, feeding, pigs, stomach. 17 18

3

Saucier, L., Bernier, D., Bergeron, R., Giguère, A., Méthot, S. et Faucitano, L. 2007. Effet du 1

type de moulée, de la fréquence des repas et de la mise à jeun avant l’abattage sur le 2

comportement, le contenu stomacal et la contamination microbienne de la carcasse chez le 3

porc. Can. J. Anim. Sci. xxxxx. Dans cette étude, le comportement des animaux au repos, le 4

contenu de l’estomac ainsi que la contamination microbiologique des carcasses ont été évalués 5

sur 96 mâles castrés assignés aux traitements suivants : texture des aliments (FT : farine vs. 6

cubés), la fréquence des repas (MF : 2 vs. 5 fois par jour) et le temps de jeûne (WT : 4, 14 et 24 7

h) selon un plan expérimental factoriel 2 x 2 x3. Les porcs recevant deux repas par jour avaient 8

un poids d’estomac plus élevé que ceux nourris cinq fois par jour (P = 0,01). Une interaction 9

significative a été observée entre WT et FT (P = 0,002). La contamination de la gueule par les 10

aérobes mésophiles totaux était supérieure à 104 cfu/cm2, mais pas significativement différente 11

entre les traitements. Le comportement coprophage pendant la période de repos n’était pas 12

corrélé à la contamination de la gueule au moment de la saignée. Le traitement pour lequel la 13

contamination de la région thoracique par Escherichia coli était la plus faible est celui dont les 14

animaux étaient nourris avec de la moulée cubée, cinq fois par jour et suivi d’un jeûne de 24h. À 15

l’inverse, l’énumération des E. coli était plus élevée pour les porcs nourris avec de la moulée 16

non cubée, présentée cinq fois par jour et suivi d’un jeûne de 4 h. La comparaison a posteriori de 17

ces deux scénarios renvoie une probabilité de 0.03. 18

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Mots clés : comportement animal, hygiène des carcasses, mise à jeun, diète, porc, estomac. 20

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It is well known that pre-slaughter management influences the final quality of meat (Gispert et 1

al. 2000; Faucitano 2001; Guàrdia et al. 2004; 2005). To obtain better quality and safer pork, 2

feed withdrawal of 16-24 h before slaughter is recommended (Eikelenboom et al. 1991). Indeed, 3

fasting pigs before slaughter reduces the volume of stomach contents at evisceration, preventing 4

the release and spread of microbial contamination (e.g., Salmonella) through the spillage of 5

viscera contents due to inadvertent puncture during the carcass dressing process (Berends et al. 6

1996; Miller et al. 1997). However, industry reports and some studies have revealed a high 7

variability in stomach weights at slaughter, even among pigs that were subjected to fasting 8

before slaughter (Guise et al. 1995; Turgeon 2003). According to Guise et al. (1995), smaller 9

feed particle size and pelleting may increase gastric emptying rates in pigs. Gregory et al. (1990) 10

also showed that feeding finely-ground feed favoured gastric emptying to the same extent as 11

liquid feeding. The lower feed intake associated with pellet feeding, which is generally more 12

finely ground (Fekete et al. 1983), could also influence gut content and its weight. Another study 13

reported lower whole stomach and content weights in pigs fed ad libitum compared to those 14

subjected to restricted feeding (Magras et al. 2000). This difference can be explained by the 15

intake of smaller feed portions at each meal in pigs fed ad libitum, which favours food digestion 16

and consequently accelerates stomach emptying (Laplace et al. 1986). Similarly, lower stomach 17

weights have been reported for pigs fed ad libitum, compared to pigs restricted to three meals per 18

day (Turgeon, 2003). 19

The results on the effects of fasting time on carcass microbial contamination remain 20

scarce. Nattress and Murray (2000) observed a reduction (lower than one Log unit) in stomach 21

cell counts, for coliforms and Escherichia coli biotype 1 when fasting time was extended from 0-22

1 to 4-5 h during lairage. Microbial counts in the caecum were unaffected by fasting while feed 23

5

withdrawal only decreased stomach cell counts when performed on the farm rather than at the 1

abattoir. However, Isaacson et al. (1999) reported a decrease in the proportion of pigs positive 2

for the presence of Salmonella in the ileocaecal contents at slaughter when a 24-h feed 3

withdrawal was applied, but the effect was only observed in combination with a 4-h transport. 4

The pig’s mouth has been reported to be another important source of cross-contamination 5

from coliforms and E. coli on the dressing line (Gill and Jones 1997). The environmental factors 6

contributing to the contamination of the mouth are not fully understood. However, it is expected 7

that mouth contamination can result, at least in part, from the pigs’ contact with manure present 8

on the pen floor or walls while performing their natural exploratory behaviour (Hurd et al. 2005). 9

The presence of manure on the truck or lairage pen floor is a likely source of fecal bacteria (e.g., 10

E. coli, Salmonella) and shedding rate of animals is known to be influenced by feed withdrawal 11

time and stress (Gregory 1998; Reid et al. 2002). 12

This experiment was conducted to determine the combined effects of different feeding 13

practices (5 vs. 2 meals per day, mash vs. pellets) prior to transportation to the abattoir and 14

various feed withdrawal times (4, 14 or 24 h) on stomach weight and carcass contamination level 15

at slaughter. Animal behaviour in lairage was observed in order to identify its contribution to 16

stomach weight variation and carcass microbial quality. 17

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MATERIAL AND METHODS 1

Animals and treatments 2

A total of ninety six (96) Duroc x (Landrace x Large White) crossbred barrows weighing 3

approximately 63 kg (approximately 16 weeks of age) were randomly divided into four groups 4

(or replicates) of 24 pigs each, and kept in pens of six animals on a commercial swine facility. 5

Animals were cared for according to the recommended code of practice of Agriculture and Agri-6

Food Canada (AAFC;1993) and following the guidelines of the Canadian Council on Animal 7

Care (1993). Over the last six weeks of the finishing period, pens of pigs were either fed pellets 8

or mash (feed texture factor: FT), and were fed either twice or five times per day (meal 9

frequency factor: MF). Animals fed twice a day received their meals at 0800h and 1600h, 10

whereas the others received 5 meals every 3 h, starting at 0800h. Pigs were fed a corn-soybean 11

based finisher diet containing 16 % CP and 3300 kcal ME/kg. At 110.0 (± 2.8) kg BW, pigs were 12

withdrawn from feed and shipped (1.5 h transport time or 120 Km) in groups of 24 pigs (six pigs 13

per combination of factors FT and MF) to the experimental swine unit of AAFC in Sherbrooke 14

(QC). In total, 4 separate shipments of 24 pigs were made, followed each time by a 5-d 15

acclimatization period before slaughter. This transfer was done to ensure better control of the 16

fasting intervals to be applied before slaughter. At the AAFC swine unit, animals within each 17

feeding treatment combination were kept in small pens of two pigs, for a period of five days, in 18

order to allow them to recover from transportation stress (Brown et al. 1999). On their arrival at 19

the AAFC experimental farm, pigs immediately had access to feed and water, and as feed and 20

water intake did not decline during this 5-day period (data not shown), it was assumed that they 21

had recovered from transportation and adapted to their new environment. The four feeding 22

treatments continued to be applied at the AAFC finishing site until the day the animals were 23

7

transported to be slaughtered. Pig’s body weight was recorded two days before shipment to the 1

abattoir. Prior to shipping, the three pens within each feeding treatment were randomly assigned 2

to one of three fasting treatments (feed withdrawal time: WT). Feed was either not withdrawn (0 3

h) or withdrawn at 10 or 20 h prior to transportation. The resulting 12 treatment combinations of 4

the 2 x 2 x 3 factorial design are presented in Table 1. 5

Four animals, two per treatment combination, were transported in two different 6

compartments (one in the front and one in the rear of the truck) for 30 min in a single-decked 7

trailer in six separate shipments and were given 3.5 h rest at the AAFC experimental abattoir 8

before slaughter. The position of treatment groups during transport was randomized to limit the 9

effects of compartment position in the truck on the response of pigs to transport stress (Barton-10

Gade et al. 1996). Animals were housed in pairs (within the same treatment combination) at the 11

abattoir. Total fasting time (from last feed to slaughter) was 4, 14 or 24 h, but water was 12

available all the time. At slaughter, pigs were immobilised in a typical “V” type restrainer and 13

were electrically stunned (head-only; 300 V for 4 sec) and exsanguinated in a prone position. 14

15

Assessment of behaviour 16

Behavioural observations were conducted by visual scan sampling during the first hour of 17

lairage. One observer slowly walked beside each pen and noted the posture (standing, sitting, 18

lying down) and activity of each animal, at 60-s intervals (60 scans per pair of pigs per hour). 19

The activities measured were: licking the floor (< 15 cm or > 15 cm, approximately, from fecal 20

matter), licking or eating feces (coproghagy), drinking, and immobile/inactive (while lying). Any 21

activity that did not fall into these categories was termed “other”. To determine the effects of the 22

treatment combinations on the rate of defecation during transport, feces were collected from each 23

8

compartment of two pigs on the truck floor and weighed after each journey. A flow-meter (Le 1

Compte ltée, St-Hyacinthe, Quebec, Canada) was installed on the water line in each pen to 2

evaluate the total water used that is the water actually drunk or otherwise wasted, by pigs in 3

lairage. 4

5

Carcass dressing 6

After slaughter, carcasses were individually processed for scalding and dehairing (Scaldomatic 7

B20-GT Standard, Oskar Baumann Fleischereimaschinenbau Gmbh & Co., Stuttgart, Germany) 8

for 6 min at 60oC and then manually singed to remove any residual hair. Carcasses were then 9

eviscerated, split and chilled (1oC, air velocity of 1m/s) according to standard commercial 10

practices. Despite the fact that scalding destroys most of the bacteria on the carcass surface (Gill 11

2000), and that water is not changed but simply replenished during industrial scalding, the tank 12

was emptied and rinsed between each shipment as an extra precaution. 13

14

Stomach weight 15

Stomachs were removed directly on the dressing line during evisceration and placed in a plastic 16

bag to avoid evaporation and spillage of content. They were identified and stored at 4oC until 17

they were weighed the next day, full and emptied of their contents. Quantity of water in the 18

stomach contents was evaluated by weight difference between the wet and dry contents. The 19

percentage of dry matter was calculated after lyophilization at 50°C for 18 h. 20

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Microbial analysis 1

Microbial sampling of the mouth and thoracic area was performed using a sterile sponge kept in 2

a sterile Whirl-pakTM sampling bag (#B01245, Nasco, Fort Atkinson, WI) soaked with 10 ml of 3

0.1% peptone water supplemented with 1% Tween 80 (Gill and Jones 1997; 2000). The mouth 4

was swabbed immediately after bleeding. Using sterile gloves, one side of the sponge was used 5

to sample the exterior of the upper jaw, on each side, and the other side of the sponge was used 6

to sample the palate for a total average surface of 236 cm2. Immediately after evisceration, the 7

thoracic area was swabbed. Inside the carcass, ribs were swabbed on both sides with one side of 8

the sponge. The other side of the sponge was used to swab the flanks for an average surface of 9

926 cm2. All samples were kept on ice for transportation and were maintained at 4oC pending 10

analysis. Total Aerobic Mesophilic bacteria (TAM), coliform bacteria and E. coli counts were 11

performed using hydrophobic grid membrane filtration (HGMF) techniques as described by Gill 12

and Jones (2000) using the Spreadfilter from Filtaflex Ltd (Almonte, ON, Canada). Samples 13

were homogenized for 2 min using a Stomacher (Lab blender 400, Seward Medical, London, 14

UK) prior to cell enumeration. Serial dilutions were filtered and membranes (ISO-GRIDTM, 15

Neogen, Lansing, MI) were incubated on appropriate agar medium. TAM counts were performed 16

on Tryptic Soy Agar (TSA; Becton Dickinson, Missisauga, ON) incubated at 35oC for 48 h and 17

colonies were stained with a 0.1% solution of Triphenyltetrazolium chloride (Sigma Aldrich, 18

Oakville, ON) for 15 min. Coliforms were enumerated on Lactose Monesin Glucuronate agar 19

(LMG; QA Life Sciences, San Diego CA) after an incubation of 24 h at 35°C. The filters were 20

then transferred to buffered 4-methylumbeliferyl-β-D glucuronide agar (BMA; QA Life 21

Sciences) and incubated for 2 h at 35°C for E. coli enumeration after illumination of the grid 22

with a long-wave UV light. Grids were photographed (Coolpix995, Nikon Corp., Tokyo, Japan) 23

10

and the Most Probable Number (MPN) was determined using the software HGMF Interpreter 1

(Filtaflex Ltd). 2

3

Data Analysis 4

Cell counts, expressed as MPN, were transformed to Log values and results were expressed as 5

the Log means (Log A) according to the formula Log A = + Logn10 . (SD2/2) where and SD 6

are respectively the mean and the standard deviation of the Log-transformed values (Gill and 7

Jones 2000). Mean Log values ( ) were compared in the analysis of variance using the MIXED 8

procedure of SAS, version 8.02 (SAS Institute, Inc. 2000). E. coli counts for thoracic 9

contamination were analysed using the Tobit regression model (Tobin 1958) with the LIFEREG 10

procedure of SAS. Data on animal behaviour for each pair of pigs were averaged, for a total of 11

48 experimental units analysed according to a Three-Way Factorial analysis of variance with 12

week of measurement as a blocking factor. Results are reported as percentages of observations. 13

Spearman correlation coefficients were calculated to establish the relationship between water 14

consumption, time spent drinking, weight of water in stomach and microbial contamination. 15

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RESULTS AND DISCUSSION 17

Behavioural observations in lairage and total water used 18

The amount of feces collected on the truck following transportation was on average 0.94 ± 0.46 19

kg and was not affected by treatment combinations (P = 0.17). This does not support our 20

hypothesis that fasting reduces feces excretion during transport and hence potential 21

contamination, as was suggested by Chevillon (1994). The short transportation period (30 min) 22

might explain the lack of feces excretion variation among treatment combinations. Longer 23

11

transport period would have to be tested to confirm Chevillon’s hypothesis (Chevillon 1994). In 1

lairage, no treatment combination significantly affected time spent standing up (61.8± 17.3 % of 2

scans, P = 0.29), lying down (38.2 ± 17.3 % of scans, P = 0.29), or lying down inactive (32.4 ± 3

15.8 % of scans, P = 0.11; Table 2). 4

These results were unexpected since time spent lying down has been observed to be 5

reduced in pigs fasted for 24 h and exposed to an unfamiliar animal for 30-min in a novel 6

environment (Fernandez et al. 1995). The discrepancy between the present experiment and the 7

latter may be partly explained by the fact that pigs were kept with familiar animals in lairage and 8

did not fight, as is often the case at commercial abattoirs. Time spent licking the floor was also 9

not affected by treatments (< 15 cm of feces: 1.0 ± 1.0, P = 0.80; > 15 cm of feces: 4.4 ± 3.6 % 10

of scans, P = 0.77). We expected fasted pigs to perform more exploratory behaviour (Kelley et 11

al. 1980) such as licking. Licking or eating fecal matter (coprophagy) was observed in 12

approximately 0.5% of scans, but this frequency was too low for meaningful comparison 13

between treatments. A significant interaction (P = 0.03) between MF and FT was found for 14

drinking behaviour. Pigs fed twice per day were observed at the drinker more often when fed 15

pellets instead of mash (8.6 % vs. 4.5% of scans; SEM: 1.67). However, pigs fed 5 meals per day 16

were observed at the drinker more often when fed pellets instead of mash (5.3 % vs. 8.7% of 17

scans; SEM: 1.67). This interaction is difficult to explain and was not observed for the variable 18

total water used. Indeed, a significant interaction was also obtained for total water used, but 19

between treatments FT and WT (P = 0.01; Fig. 1). Total water used decreased with increasing 20

fasting time when pigs were fed mash. In contrast, it was lowest at 14 h and highest at 24 h 21

fasting time in pigs fed pellets. A higher drinking rate in long-term fasted pigs (24 h vs. 16 h and 22

8 h) was also reported by Brown et al. (1999) and may be related to hunger stimulating drinking 23

12

of water (Yang et al. 1981; Robert et al. 1993). However, pigs fed mash and fasted for 24 h did 1

not show such an increase in the total water used. Furthermore, mash fed pigs did not show an 2

increase, but rather a reduction in their total water used as fasting time increased, which does not 3

support the hunger hypothesis in these animals. Even though a significant correlation was 4

obtained between drinking behaviour and total water used (r = 0.64; P < 0.0001), it was lower 5

than previously reported (r = 0.81) by Ange et al. (2000), suggesting a lack of precision in either 6

behavioural recording or measurement of total water used in the present experiment. 7

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Stomach weight 9

The average weight of the stomachs harvested in this study was 1205 g (range from 451 g to 10

3308 g; Table 3). This average weight meets the abattoir specification for maximum stomach 11

weight (1400 g; Chevillon 1994). However, 25 % of the stomachs evaluated in this study 12

exceeded this weight and, as reported by Faucitano et al. (2006), 51 % had contents weighing 13

more than 500 g (671.3 g on average), which is the recommended threshold to prevent carcass 14

contamination at slaughter (Chevillon 1994). 15

A significant interaction between FT and WT was observed for full stomach weights 16

(P = 0.002; SEM = 98.62), as well as stomach water and dry matter content weights (P = 0.003; 17

SEM = 69.52 and P = 0.0004; SEM = 22.73, respectively; Fig. 2 a, b, c). All three variables 18

decreased with increasing fasting time in pigs fed mash. In pigs fed pellets, however, full 19

stomach weights and water content were lower at 14 h compared to 4 h fasting, but were higher 20

at 24 h fasting. Dry matter content was slightly reduced at 24 h compared to 14 h. These 21

interactions indicate that the increase in stomach weight observed in pigs fed pellets at 24 h 22

compared to 14 h is mainly due to a higher water content, which could be the result of the higher 23

13

quantity of water used by these animals. Indeed, a significant correlation was found between the 1

total water used and the weight of water content in the stomach (r = 0.40; P < 0.004). A possible 2

explanation is that pigs fed pellets were more hungry during lairage because their stomachs were 3

on average less full than those of pigs fed mash (P < 0.001). The enhanced hunger would have 4

led to an increased water intake after 24h of fasting. This is supported by the fact that pellet 5

feeding has been reported to reduce feed intake (Fekete et al. 1983), and to cause a faster gastric 6

emptying (Gregory et al. 1990; Guise et al. 1995). 7

Full stomach weights tended to be higher (P = 0.09) and dry matter content weights of the 8

stomach were significantly higher (P = 0.01) for pigs fed two meals per day compared to animals 9

fed 5 times a day (Table 3). Similar results have been found by Turgeon (2003), with pigs fed 10

three meals a day having heavier stomach weights at slaughter than pigs fed ad libitum. Hence, 11

feeding pigs more frequently would be a better feeding practice than feeding them twice a day 12

for the reduction of stomach weight at slaughter. 13

14

Carcass microbial contamination 15

Cell counts for TAM, coliforms and E. coli on the mouth followed a Log-normal distribution. 16

TAM counts (Log A) ranged from 5.43 to 4.73, coliforms from 2.44 to 1.38 and E. coli from 1.83 17

to 0.98 Log cfu/cm2 (Table 4). Enumeration variations between treatments were all below one 18

Log unit except for the coliforms where a variation of 1.06 Log units was observed between 19

treatment 3 and treatment 7. For coliform counts, the lowest observed value occurred in the 20

mouth of pigs fed pellets twice per day and fasted for 24 h while the highest counts were 21

observed in the mouth of pigs fed mash twice per day and fasted for 4 h. The analysis of variance 22

revealed a significant overall difference between treatments for the coliforms (P = 0.001) and the 23

14

E. coli counts (P = 0.02) but not for the TAM (P = 0.1). The FT x MF interaction influenced 1

TAM and coliforms counts (P = 0.01 and P = 0.02, respectively). For pigs fed mash, TAM 2

counts were higher when they were fed 5 times a day while for pellet fed pigs, TAM counts were 3

higher when they were fed two times a day. As for coliforms counts, they were higher in pigs fed 4

mash compared to those receiving pellets two times per day but were similar when pigs were fed 5

5 times per day. Furthermore, there was a MF x WT interaction for coliform and E. coli counts 6

(P < 0.001 and P = 0.003, respectively), with coliform and E. coli counts decreasing with 7

increasing fasting time in pigs fed two times per day. For the animals fed 5 times a day, the 8

lowest coliform and E. coli counts on the mouth were recorded for animals fasted for 4 h. 9

However, these interactions are of little practical value as all mean values used to evaluate them 10

are within one Log unit difference (Gill et al. 2002; Gill and Baker 1998; Jarvis, 1989). 11

Coliforms and E. coli counts for the mouth are below the threshold level recommended in 12

the guidelines of the Meat Hygiene Manual (CFIA 2006) for raw meat. All Log mean and mean 13

Logs for TAM are between the marginal (m = 104) and maximum values accepted (M = 106) by 14

these guidelines. These results confirm previous observations by Gill and Jones (1997) who 15

indicated that the mouth is an important source of cross-contamination between carcasses. No 16

correlation between licking/eating feces behaviour and the E. coli counts was found, suggesting 17

that under the experimental conditions of this study, contact of the pigs with manure in the 18

lairage pen alone could not explain the variation among treatments. According to Nattress and 19

Murray (2000), feed withdrawal increased the E. coli counts in the caecum but not in the 20

stomach. Variation in the gastrointestinal microflora is expected to influence carcass 21

contamination but this remains to be elucidated further especially if the evisceration is performed 22

adequately and that the viscera are not punctured during the dressing process. 23

15

For cell counts obtained on the thoracic area, TAM counts (Log A) ranged from 2.44 to 1

2.00 Log cfu/cm2, coliforms from 2.60 to 1.95 Log cfu/cm2, and a maximum of 3.55 Log 2

cfu/926cm2 for the E. coli counts was obtained on the carcass of pigs fed mash 5 times a day and 3

fasted for 24 h (Table 5). TAM counts were higher (P = 0.004) for pigs fed twice a day 4

compared to those receiving 5 meals per day (2.19 vs. 2.01 cfu/cm2; SEM: 0.04). TAM counts 5

were also different with respect to fasting (P = 0.004). The counts were significantly higher 6

(P = 0.003) for a feed withdrawal of 14 h compared to 4 h (2.23 vs. 1.99 cfu/cm2; SEM: 0.05) but 7

to a lesser extent (P = 0.07) when compared to 24 h (2.08 cfu/cm2). These results suggest that 8

there is a particular fasting time interval that is most detrimental with respect to carcass hygiene 9

expressed as TAM counts. To our knowledge, this is the first report to specifically address the 10

effect of fasting prior to slaughter on carcass hygiene. Although variation in the gastrointestinal 11

microflora is expected to influence carcass hygiene, it is not the only source of contamination 12

(i.e. mouth; Gill, 2000). Therefore, it would be misleading to compare previous results on 13

shedding with cell counts on the carcass. Again, because differences were below one Log unit, 14

these effects are of little practical value. 15

Forty samples out of 96 were below detection level (1.70 Log cfu/926cm2) for the E. coli 16

counts on the thoracic area. The global analysis of the original model returned no significant 17

results; data were therefore analysed using the Tobit regression model which is an appropriate 18

method for dealing with left-censored data (SAS Institute, Inc. 2000). We noticed that the lowest 19

E. coli counts were recorded for pigs fed pellets 5 times per day and submitted to a 24 h fasting 20

prior to slaughter, while the highest values were found in pigs fed mash 5 times per day and 21

fasted for 4 h before slaughter. An a posteriori Tobit analysis on these two treatment 22

combinations revealed a significant difference (P = 0.03). 23

16

No correlation was found between stomach weight and carcass microbial contamination 1

as well as between E. coli contamination of the thoracic area and feces weight collected on the 2

truck floor after transportation. The low stomach weight and the careful evisceration procedure 3

we were able to carry out under the controlled conditions of an experimental abattoir limited the 4

risk for stomach puncture and subsequent contamination due to stomach content spillage. The 5

lack of relationship between the E. coli counts on the thoracic area and defecation rate (weight of 6

feces) on the truck can be explained by the behaviour of pigs during the transportation between 7

the farm and the abattoir (data not shown). During the 30 minute journey, pigs always stood and 8

very seldom touched the floor with their nose (data not shown). The pigs’ standing behaviour has 9

been reported during short journeys in a number of previous studies (Lambooy 1988; Bradshaw 10

et al. 1996; Riches et al. 1996) and can be explained either by the lack of sufficient time to lie 11

down or by the uncomfortable travel conditions (i.e., high amount of vibrations) which prevented 12

pigs from lying down. The standing position and the scarce exploratory behaviour seemed to 13

have prevented the contamination of pigs’ body or mouth with the feces present on the vehicle 14

floor. Furthermore, despite this behaviour, the short exposure of pigs to manure might have 15

limited their contamination anyway as it is known that the risk for carcass fecal contamination 16

increases with the animals’ exposure time to manure in the surrounding environment (Hurd et al. 17

2005). 18

All TAM and E. coli counts taken from the thoracic area meet the CFIA guidelines 19

(2006). However, for no obvious reasons, 67% of all the coliforms counts on the third replicate 20

were between the marginal (m = 500) and maximum values accepted (M = 103) by the CFIA 21

guidelines (2006), independently of the treatments. Variation in the incidence of contaminated 22

carcasses between different sampling days both at the same abattoir and between abattoirs has 23

17

been reported by Botteldoorn et al. (2003). Carcass contamination at that time may have been 1

caused by an unexpected event independent of the treatments under study. 2

3

CONCLUSION 4

TAM counts on the mouth were not influenced significantly by treatments, but results confirm 5

previous reports on the risk of contamination resulting from the manipulation of the head and 6

touching the mouth of the animal. Hence, precautionary measures must be implemented to avoid 7

spreading this source of contamination throughout the carcass. Feeding pellets 5 times per day 8

combined to a 24 h feed withdrawal minimized E. coli contamination on the thoracic area. A 9

higher frequency of feeding appears to be a better practice to reduce stomach weight and TAM 10

counts on the thoracic area. These data were collected using short transport times (30 min) and 11

animals were slaughtered in an experimental abattoir. More work is needed to fully evaluate the 12

impact of feed texture, meal frequency and pre-slaughter fasting on pig behaviour and carcass 13

microbial quality, notably on longer times for transport, as greater variations may be observed in 14

commercial settings. 15

16

17

18

ACKNOWLEDGEMENTS 1

The authors are grateful to the staff of the AAFC Swine Complex for care of the animals at the 2

farm, at slaughter and for their assistance in the carcass evisceration work. The authors thank M.-3

J. Gagné and T. Tulkki for their technical assistance, and Dr C.O. Gill, for his valuable input 4

during the preparation of the manuscript. Sincere thanks are addressed to the swine company F. 5

Ménard for providing the animals, feed and manpower and to Agriculture and Agri-Food Canada 6

for the financial support. 7

8

9

19

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8

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25

LEGEND OF FIGURES 1

Fig.1. Effects of feed texture (FT) and fasting time (WT) on total water used in lairage (SEM: 2

2.37). 3

4

Fig. 2. Effects of feed texture (FT) and fasting time (WT) on full stomach weights (a; SEM: 5

98.62), water content (b; SEM: 69.52) and dry matter content (c; SEM: 22.73). 6

7

8

26

Table 1. Description of experimental treatments 1

Treatments Feed type Meal frequency ( # feedings/day)

Fasting time before slaughter

(h) 1 Pelleted 2 4 2 Pelleted 2 14 3 Pelleted 2 24 4 Pelleted 5 4 5 Pelleted 5 14 6 Pelleted 5 24 7 Mash 2 4 8 Mash 2 14 9 Mash 2 24

10 Mash 5 4 11 Mash 5 14 12 Mash 5 24

2

27

Table 2. Effects of feed texture (FT), meal frequency (MF) and fasting time (WT) on pig behaviour (% of scans) 1

during the first hour of lairage 2

3 Feed texture Meal frequency

(meals / d) Fasting time

(h) P-values

P M SEM 2 5 SEM 0 14 24 SEM

FT

MF

F

FT*MF

FT*

F

MF*

F

FT*MF*

F

Standing 61.2 62.5 3.8 58.1 65.6 3.8 61.1 58.8 65.6 4.5 0.78 0.12 0.51 0.30 0.41 0.31 0.12

Lying down

38.8 37.5 3.8 41.9 34.4 3.8 38.9 41.1 34.4 4.5 0.79 0.13 0.50 0.30 0.41 0.32 0.12

Inactivez 33.7 31.2 3.2 36.0 28.9 3.2 32.9 35.6 28.8 3.8 0.55 0.10 0.41 0.40 0.27 0.13 0.06

Licking < 15 cm

0.8 1.1 0.2 0.8 1.1 0.2 1.2 0.8 1.0 0.3 0.39 0.39 0.57 0.20 0.72 0.52 0.82

Licking > 15 cm

4.1 4.8 0.9 4.1 4.8 0.9 3.4 4.5 5.4 1.1 0.54 0.54 0.32 0.98 0.36 0.80 0.47

zInactive=observed lying. 4 P=pellets; M=mash. 5

6

7

28

Table 3. Weights (g) of full stomach and stomach content (dry matter and water) at 1

slaughter for pigs fed in 2 or 5 meals per day over the last six weeks of the finishing 2

period 3

4 Meal frequency

2 5 SEM P Full stomach (g) 1237.2 1122.0 46.8 0.09

Stomach content (g) 696.7 598.0 43.1 0.11

Water content (g) 527.0 466.3 35.1 0.22

Dry matter content (g)

169.7 131.7 10.1 0.01

5 6

29

Table 4. Descriptive statisticsz for sets of TAM, coliforms and E. coli counts (cfu/cm

2) 1

taken from the mouths of all the animals (n = 96) immediately after exsanguination 2

TAM (cfu/cm2) Coliforms (cfu/cm2) E. coli (cfu/cm2)

Treatments SD Log A Log N No SD Log A Log N No SD Log A Log N No

1 5.00 0.61 5.43 6.61 8 1.77 0.37 1.93 2.84 8 1.31 0.43 1.52 2.41 8

2 4.68 0.44 4.91 5.75 8 1.41 0.27 1.50 2.37 8 1.30 0.27 1.38 2.28 8

3 4.76 0.45 5.00 5.88 8 1.10 0.49 1.38 2.23 8 0.93 0.51 1.22 2.11 8

4 4.66 0.40 4.84 5.72 8 1.41 0.65 1.89 2.77 8 0.96 0.52 1.28 2.03 7

5 4.69 0.32 4.81 5.68 8 1.62 0.69 2.17 2.84 8 1.31 0.50 1.60 2.40 8

6 4.69 0.19 4.73 5.65 8 1.61 0.78 2.31 3.04 8 1.00 0.60 1.41 2.28 8

7 4.62 0.39 4.79 5.63 8 1.97 0.64 2.44 3.23 8 1.55 0.49 1.83 2.61 8

8 4.52 0.46 4.77 5.59 8 1.67 0.58 2.06 2.86 8 1.28 0.37 1.44 2.34 8

9 4.82 0.28 4.91 5.81 8 1.41 0.81 2.17 3.03 8 1.05 0.41 1.25 2.11 8

10 4.81 0.35 4.96 5.83 8 1.32 0.58 1.70 2.55 8 0.84 0.36 0.98 1.84 8

11 4.76 0.29 4.86 5.76 8 1.60 0.33 1.73 2.66 8 1.35 0.47 1.60 2.54 8

12 4.87 0.28 4.96 5.88 8 1.47 0.61 1.91 2.65 8 1.25 0.68 1.78 2.52 8 Z See section on Data analysis in the Material and Methods for further details with respect to , 3

SD, log A, log N and No. 4

5

6

_ X

_ X

_ X

_ X

30

Table 5. Descriptive statisticsz for sets of TAM (cfu/cm

2), coliforms (cfu/cm

2) and E. coli 1

counts (cfu/926cm2) obtained by swabbing the inside rib cage and the flank regions of all 2

the animals (n = 96) immediately after evisceration 3

TAM (cfu/cm2) Coliforms (cfu/cm2) E. coli (cfu/926cm2)

treatments SD log_A Log_N No SD log_A Log_N No SD log_A Log_N Noy

1 2.08 0.29 2.18 3.06 8 1.87 0.52 2.18 2.70 8 1.77 0.14 1.80 2.70 3

2 2.31 0.31 2.42 3.28 8 1.85 0.69 2.40 3.30 8 1.89 0.54 2.22 3.30 2

3 2.18 0.33 2.31 3.20 8 1.88 0.67 2.39 3.31 8 1.99 0.55 2.34 3.31 2

4 2.01 0.32 2.13 3.02 8 1.81 0.56 2.17 2.78 8 1.79 0.25 1.86 2.78 3

5 2.04 0.31 2.15 3.01 8 1.82 0.74 2.46 2.85 8 1.80 0.30 1.91 2.85 2

6 1.97 0.44 2.19 3.04 8 1.74 0.73 2.36 3.26 8 2.07 0.49 2.35 3.26 7

7 2.02 0.28 2.12 3.01 8 1.66 0.50 1.95 2.78 8 1.81 0.22 1.87 2.78 3

8 2.39 0.20 2.44 3.33 8 1.91 0.77 2.60 3.45 8 2.18 0.60 2.60 3.45 4

9 2.14 0.25 2.21 3.10 8 1.85 0.63 2.30 2.81 8 1.83 0.25 1.91 2.81 2

10 1.85 0.36 2.00 2.88 8 1.70 0.63 2.16 2.81 8 1.80 0.28 1.88 2.81 1

11 2.20 0.35 2.34 3.23 8 1.72 0.78 2.43 2.95 8 1.92 0.34 2.05 2.95 5

12 2.01 0.40 2.19 3.06 8 1.77 0.55 2.12 4.88 8 2.10 1.12 3.55 4.88 2 Z See section on Data analysis in the Material and Methods for further details with respect to , 4

SD, log A, log N and No. 5

Y No is not equal to 8 because several counts were below detection level (1.70 Log cfu/926cm2). 6

7

_ X

_ X

_ X

_ X

31

Fig. 1 1

0

2

4

6

8

10

12

14

pellets mash

To

tal

wate

r u

sed

(L

)

4h fasting

14h fasting

24h fasting

2

3

32

Fig. 2 1

a

0

500

1000

1500

2000

2500

pellets mash

Fu

ll s

tom

ach

weig

ht

(g)

4h fasting

14h fasting

24h fasting

2

b

0

500

1000

pellets mash

Wate

r co

nte

nt

(g)

4h fasting

14h fasting

24h fasting

3

c

0

500

1000

pellets mash

Dry

matt

er

co

nte

nt

(g)

4h fasting

14h fasting

24h fasting

4

5