Investigation Into the Humaneness of Slaughter Methods for Guinea Pigs (Cavia porcelus) in the Andean Region (2024)

Current methods

The methods practiced involved cervical neck dislocation either before or after a unilateral cut to the ventral aspect of the neck without stunning. Each animal was individually restrained and slaughtered by the same operator. Method 1 involved a unilateral cut to the ventral aspect of the neck severing the left carotid artery and jugular vein followed by cervical neck dislocation (neck cut-cervical neck dislocation [NC-CND]) by extending the head 90° to the spine and pulling of the hind legs (n=30). Method 2 involved cervical neck dislocation, as described, followed by the neck cut (cervical neck dislocation-neck cut [CND-NC]; n=30).

Immediately following neck dislocation with both methods, the animals were assessed for behavioral and brainstem signs associated with sensibility/insensibility. The following parameters were assessed/recorded: adequate restraint of animals, the time from restraint to neck cut or dislocation (depending on the method used), number of attempts to dislocate the neck, body flaccidity, convulsions, corneal reflex, eyeball rotation, nystagmus, jaw relaxation, vocalization, rhythmic breathing, response to handling, cleat response by pressing between the digits of a hind leg (spinal reflex), and absence of righting reflex.

Immediately after slaughter, the neck cut was examined to assess the damage to the carotid arteries, jugular veins, spinal cord, trachea, and esophagus, as well as the position of neck dislocation.

Electrical stunning

Eighty-three animals were electrically stunned with a head-only 50Hz alternating current (AC) (sinusoidal waveform) constant voltage electrical stunner with pin electrodes (Whitehead Engineering Ltd., Bath, UK) mounted on a table and activated by a foot switch.

The operator restrained each animal by holding the animal with one hand against their body and supporting the head with the free hand. Stunning electrodes were placed between the eyes and the ears, spanning the brain. During the first 5s after stunning, the presence of rhythmic breathing, corneal reflex, and cleat response were assessed. Next the animals were immediately slaughtered with a bilateral cut to the ventral surfaces of the neck.

A subset of animals (n=10 males; n=10 females) were electrically stunned as described and were allowed to recover. This allowed for assessment of the duration of induced insensibility. Once the stun was assessed, the following behaviors, signs, and reflexes were recorded until the return of sensibility: onset of rhythmic breathing, corneal reflex, neck tension, and balance. Immediately after the return of three out of the four signs, the animals were electrically stunned and slaughtered by bilateral severance of the carotid arteries and jugular veins.

Peak current, current profile, peak voltage, stun duration, and time to reach 140mA (minimum recommended stunning current in the European Union for rabbits; Anil et al., 1998) were recorded for all animals using a 199C Fluke Scope meter and 179 Fluke multimeter (Fluke Corporation, Everett, WA). All current and voltage recordings are root mean square values.

CO₂ stunning

Two groups of guinea pigs were stunned with a rising concentration of CO₂ (cylinder concentration of 99%) into a chamber (custom-made; Solutions for Research, Silsoe, UK) at a flow rate of 20% CO₂ (n=9) or 30% CO₂ (n=12) of the chamber volume per minute. The total volume of the chamber was 27L (size 30cm×30cm×30cm), with two CO₂ sensors (COZIR Sensor, Gas Sensing Solutions Ltd., Cumbernauld, UK) positioned 50mm and 20mm from the bottom and perpendicular from the CO₂ inlet. The inlet tube was positioned 35mm from the top of the chamber and delivered CO₂ at 5.4L/min or 8.1L/min (air flow meter, Platon, Domont, France) to obtain a fill rate of 20% or 30% of the chamber volume per minute, respectively.

A 14-mm diameter outlet for the release of displaced gas was on the opposing side to the inlet of the chamber. The CO₂ concentration was measured every second, and the results were stored on a laptop computer (Dell Inc., Austin, TX) using custom-developed software (Solutions for Research, Silsoe, UK). Data were exported to Excel (Microsoft, Redmond, WA) for data analysis.

The animals were individually placed in the chamber, with the lid firmly secured. The concentration of CO₂ was gradually increased, with a fill rate of either 20% or 30% CO₂ per minute. Prior to CO₂ filling, the animals underwent a 30-s acclimatization period, during which baseline behaviors were recorded. Video was continuously recorded for each animal from the acclimatization period, during CO₂ stunning, until removal from the chamber.

Onset and duration of spontaneous and evoked behaviors were assessed at a later date from the videos using the ethogram presented in Table ​Table1.1. These data were analyzed by the same trained operator, who was blinded to the treatments and animal type. The ethogram was developed from the assessment of behavior of guinea pigs and other species during CO₂ stunning. Immediately following cardiac arrest or after negative responses to evoked stimuli, all animals were slaughtered by bilateral severance of the carotid arteries and jugular veins. Carcasses were examined for bruises, damage to internal organs, and muscle hemorrhages.

Captive bolt

Ten animals were shot with a Dick KTBG spring-powered, captive-bolt gun (Friedr Dick GmbH & Co., Deizisau, Germany). One operator restrained the animal on a metal table, with the head held firmly against the table, while a second operator placed the captive-bolt gun on the head. The shot position was on the midline between the eyes and ears, with the objective that the bolt should penetrate the thalamus/midbrain regions. Immediately after shooting, the animals were assessed for rhythmic breathing, convulsions, corneal reflex, nystagmus, jaw relaxation, body flaccidity, response to handling, cleat response, and righting reflex.

Next the animals were immediately slaughtered by bilateral severance of the carotid arteries and jugular veins. Any animals who showed either rhythmic breathing or a corneal reflex were immediately dispatched by cervical neck dislocation, followed by a ventral neck cut. The heads of three animals were removed for postmortem examination of captive-bolt injury.

Statistical analysis

All statistical analyses were carried out using the statistical package Stata, Version 11 (Stata Corporation, College Station, TX). Descriptive statistics were performed on all variables evaluating brain and spinal function for each of the different methods assessed.

Current methods

Sixty animals were evaluated using the current methods: 30 animals with NC-CND (mean live weight, 1,072±194g) and 30 with CND-NC (mean live weight, 1,046±184g; Table ​Table2).2). Thirteen (43%) and 4 (13%) guinea pigs in the CND-NC and NC-CND groups, respectively, responded to handling (p=.010). Body flaccidity was absent in 6 (20%) and 14 (47%) animals in the NC-CND and CND-NC groups, respectively (p=.028). Almost half (47%) of the animals who received the NC-CND and more than half (60%) who received CND-NC presented with jaw tension after their respective treatments. Twelve (40%) and 9 (30%) animals in the NC-CND and CND-NC groups, respectively, had positive corneal reflexes, and more than half of the animals had positive cleat responses during both methods. In addition, 27 (90%) animals in the NC-CND group and 29 (97%) animals in the CND-NC group had at least one behavioral or cranial/spinal response.

All animals in both treatment groups had at least the spinal cord partially severed. However, it was not possible to assess in detail the extent of the damage in each animal due to the importance of retaining the connection between the head and the carcass during processing.

Electrical stunning

Eighty-five guinea pigs (40 females and 46 males) were evaluated after head-only electric stunning (mean live weight, 1,098±219g), with 2 (2%) animals being classified as mis-stunned. Both animals had rhythmic breathing. In addition, 3 out of 80 had a positive cleat response. None of them presented with a corneal reflex. Mean peak current and voltage were 583±116mA and 117±2V, respectively. The time to reach the peak current was 1.2s to 9.9s (M=4.9±1.4s; Table ​Table33).

Four current profiles were identified (Figure ​(Figure1):1): climbing satisfactory (n=59; 68%), poor initial contact (n=22; 26%), interrupted current (n=7; 8%), and failure to maintain current flow (n=9; 11%). Climbing satisfactory and poor initial contact were recorded as being mutually exclusive; however, each could occur in the same animal along with interrupted current or failure to maintain contact. Time to reach peak current was significantly quicker when a climbing-satisfactory profile was achieved (p<.001). There was no significant difference in peak current reached between animals who were mis-stunned and those who were properly stunned (p=.65).

The four behavioral parameters measured in a subset of animals to assess the duration of insensibility were highly correlated (data not shown). The mean time to onset of rhythmic breathing and balance was shorter in males than females (p=.02 and p=.04, respectively; Table ​Table4).4). However, there was no significant difference in the mean duration of insensibility between sexes (females, 47±3.5s; males, 39±2.9s; p=.09). Peak current reached in the subset was 340mA to 820mA (M=591±121mA), and the mean peak voltage was 119±2V (range=115–122V).

The duration of induced insensibility was 30s to 69s (M=43±11s). There was a significant difference in the peak current reached between males and females (p=.04), with males receiving, on average, higher current levels than females (Table ​(Table4).4). Meanwhile, females were, on average, heavier than males (p<.001). After adjusting peak current for the animal live weight, the effect of current becomes nonsignificant (Table ​(Table5),5), suggesting that live weight was the main determining factor for duration of insensibility (Table ​(Table55).

CO₂ stunning

Twenty-one animals were evaluated during CO₂ stunning. Nine animals (7 males and 2 females; mean live weight=993±218g) were stunned with a CO₂ flow rate of 20% per minute and 12 animals (8 males and 4 females; mean live weight=1,109±236g) were stunned with a flow rate of 30% CO₂ per minute. Time to onset of the spontaneous and evoked behaviors is presented in Table ​Table66.

Mean time to onset of increased respiratory effort occurred earlier in the 30% CO₂ flow-rate group compared with the 20% CO₂ flow-rate group (p=.02). In both groups, the onset of increased respiratory effort occurred before head recumbency (M=62±11s in the 20% flow-rate group; M=60±8s in the 30% CO₂ flow-rate group). There was a significant difference in evoked behaviors between the two groups (negative righting reflex, p=.01; negative corneal reflex, p=.004).

During the adaptation period, most animals displayed exploratory behavior; this behavior was absent during CO₂ stunning. A third of the animals in the 30% CO₂ flow-rate group displayed nostril flaring. Only one animal showed escape-like behavior, and this behavior only occurred once. None of the animals in either treatment group presented with excessive salivation, ataxia, convulsions, or freezing behavior (Table ​(Table7).7). The hazard ratio of ceasing breathing was 1.2 higher in guinea pigs stunned with a flow rate of 20% CO₂ (95% CI [1.01, 1.3]; p=.03; Figure ​Figure22).

The data were fitted with a cubic model to assess the relationship between respiratory rate and CO₂ concentration (R²=.89; Figure ​Figure3).3). The respiratory rate was predicted by the following equation:

Fill-rate group and sex were not significant (p=.7 and p=.4, respectively) and therefore were not considered in the final model.

Captive bolt

Ten animals were shot with a spring-powered captive-bolt gun. Only one animal presented nystagmus, rhythmic breathing, cleat response, and righting reflex (Table ​(Table8).8). The shot in this animal was misplaced, with the animal being shot at the back of the head with the bolt angling left laterally and only superficially damaging the left occipital lobe. No other animals showed any signs of sensibility or return of sensibility after captive-bolt shooting. The depth of penetration was 10mm to 13mm.

Neck dislocation

The Code of Practice for Humane Killing recommends dislocation of the neck as an effective method of dispatching rodents up to 500g (Code of practice, 1997), while the AVMA guidelines consider cervical dislocation as an acceptable method of dispatching animals weighing<200g (AVMA Members of the Panel on Euthanasia, 2013). However, for heavier animals, the large muscle mass in the cervical region makes neck dislocation difficult to perform proficiently, thereby increasing the risk for failing to completely sever the spinal cord.

In this study, the minimum live weight was 796g. Forty percent of the animals in the NC-CND group and 30% in the CND-ND groups had positive corneal reflex; 47% and 60%, respectively, presented jaw tension; and more than half of the animals had a positive cleat response after both treatments. Although all animals in both treatment groups had the spinal cord at least partially severed, the presence of cranial and spinal responses suggests that ascending and descending spinal pathways were at least partially intact and functional in most animals.

These findings confirm that in practice, it is difficult to induce complete insensibility and eliminate brainstem reflexes when using cervical neck dislocation. Therefore, cervical neck dislocation should not be recommended as a dispatch method for guinea pigs for meat production, which raises the question of whether it should be considered an effective and humane method for dispatching guinea pigs in the laboratory.

Electrical stunning

Two of the 85 animals in the study had rhythmic breathing immediately after the stun. In both cases, the peak current was below the overall mean peak current, and in 1 animal, the current profile was categorized as being interrupted and failing to maintain contact. Similarly, in 2 of the 3 animals who had a positive cleat response, the current profile identified was poor initial contact and a peak current below the mean.

The application of an adequate current across the brain is essential for ensuring animals are rendered immediately insensible when using head-only electrical stunning. The reduced area of contact at the surface of the head can result in heating effects enhancing the buildup of carbon on the electrodes and therefore increasing the resistance (Sparrey & Wotton, 1997). In addition, the amount of fur on the guinea pigs' heads might have hindered appropriate contact in some animals. As a result, in some animals, the delivery of electrical current to the head and brain was insufficient to stun.

The minimum recommended stunning current for rabbits is 140mA (Anil et al., 1998). A previous study in guinea pigs showed that the higher voltages produced a better stun (Mariño Frias, 2010). However, that study did not report the current delivered to the head. Wotton and O'Callaghan (2002) found that the magnitude of the applied voltage was positively correlated with the current delivered, and high initial voltages are important for breaking down the initial high impedance of biological tissues. However, it is the current delivered to the head and not voltage alone that is responsible for inducing grand mal epileptic activity within the brain and the resulting period of insensibility (Gregory, 2007).

Head-only electrical stunning usually produces a stun that is reversible; therefore, the animals need to undergo a subsequent dispatching procedure soon after stunning to ensure that they do not recover. Previous studies with broilers have shown an increase in the duration of insensibility when the current delivered to the head increased (Gregory & Wotton, 1990b; Raj & O'Callaghan, 2004); however, this effect was not observed in hens (Gregory & Wotton, 1994).

In this study, live weight was the main factor associated with the duration of insensibility, with heavier animals having a longer period of insensibility. Potentially, the increased soft tissue over the skull in heavier animals could have resulted in improved electrical contact with the pin electrodes as they depressed or pierced the skin, thereby reducing the impedance of the head to current flow. The period of induced insensibility varies between species. In this study, the first signs of potential recovery appeared at 20s (one animal showing rhythmic breathing).

The time to hypoxemia-induced brain failure following bleeding was not assessed in this study, so the required duration of unconsciousness cannot be estimated. However, the results do suggest that the period of induced insensibility in guinea pigs is relatively short. Therefore, exsanguination should occur as soon as possible after head-only electrical stunning to minimize the potential for recovery during the bleeding period.

If head-only electrical stunning is to be adopted, the initial cost of the stunner, the cost of electricity, and the fact that blackouts are common in countries where guinea pigs are kept for human consumption should be considered before adopting this method. The blackouts could be addressed by using battery-powered electrical stunning systems.

CO₂ stunning

Initial CO₂ concentration and fill rate of the chamber have been reported as important factors for respiratory distress and time to loss of consciousness (Gerritzen, Lambooij, Hillebrand, Lankhaar, & Pieterse, 2000; Velarde et al., 2007). In this study, increased respiratory effort and heaving were observed before head recumbency in both groups, suggesting that there was a period of respiratory distress before loss of consciousness. CO₂ is a potent activator of hyperventilation and respiratory distress (Raj & Gregory, 1995). The mean time to the end of respiratory distress and cessation of breathing was significantly shorter in the 30% CO₂ flow-rate group (p=.03). Lee and Weary (2007) found aversion to CO₂ in rats at a concentration of 17% to 20%; the authors suggested the avoidance was in response to severe dyspnea.

Adverse reactions such as seizures, convulsive chewing, nasal hemorrhage, sero-sanguineous nasal discharge, and excessive salivation have been reported in mice and rats prior to death from exposure to CO₂ (Conlee et al., 2004) and are commonly used as indicators of mucosal irritation and discomfort. In mink, convulsions during CO₂ euthanasia have been reported after they become recumbent and when all movements (including respiration) have ceased (Enggaard-Hansen, Creutzberg, & Simonsen, 1991).

In this study, none of the animals convulsed; however, a third of the animals had nostril flaring in the 30% CO₂ flow-rate group. A number of animals in the study urinated and defecated during CO₂ stunning. Previous studies have shown urination and defecation as signs of distress. However, caution must be taken with this interpretation, as alternatively, the animals could have urinated and defecated due to relaxation of sympathetic and parasympathetic tone (Conlee et al., 2004).

Although there were some signs of respiratory distress during CO₂ stunning, the overall lack of negative behavioral responses and signs of suffering suggests that the distress associated with CO₂ stunning prior to insensibility was less than the distress reported in other rodent species. Based on these results, CO₂ stunning would likely represent an improvement in terms of welfare over the currently used methods of neck dislocation and unilateral neck cut. However, due to the prolonged time to complete cessation of respiratory activity at the tested flow rates and the lack of recycling of the gas, it would be impractical and costly for slaughterhouses to use a similar single-animal system in Peru today. A more practical approach would be batch stunning of animals either with recycling CO₂ or lowering batches into a prefilled chamber.

Spring-powered captive bolt

Of the 10 animals shot with the spring-powered captive bolt, only one failed to be rendered immediately insensible. Postmortem examination revealed that the bolt only superficially damaged the left occipital lobe, suggesting that the cause of incomplete concussion was due to misplacement of the shot. Gibson et al. (2012, 2015) reported that for sheep and alpacas, a shot with a penetrative captive bolt that damages the thalamus and brainstem is essential in ensuring irrecoverable insensibility (Gibson et al., 2012, 2015).

The mean peak velocities of spring-powered, captive-bolt guns are significantly lower than the cartridge-powered guns, and the maximum kinetic energy delivered is only 5.01J (Sharp et al., 2014). With low-velocity/kinetic energy captive-bolt guns, it is thought that the direct focal damage to specific brain regions rather than concussive trauma produces brain dysfunction and loss of sensibility. Contrary to results found in kangaroo in-pouch young (Sharp et al., 2014), the spring-powered captive bolt was sufficient to cause immediate insensibility in adult guinea pigs.

Stunning using the nonpenetrative captive bolt has been demonstrated to be a humane stunning method for rabbits, turkeys, and piglets (Casey-Trott et al., 2013; Dennis et al., 1988; Erasmus et al., 2010). In the current study, although only 10 animals were assessed, the results suggest that the spring-powered captive bolt is an effective method for rendering guinea pigs insensible prior to slaughter. If this method is adopted, it would be essential that the guns are routinely inspected, tested for performance, and calibrated. Over time, the performance of the springs and rubber absorbers in spring-powered captive-bolt guns can degrade due to normal wear and tear.

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