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Detection of cigarettes and other tobacco products by giant African pouched rats (Cricetomys gambianus)
Journal of Veterinary Behavior 9 (2014) 262e268
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Journal of Veterinary Behavior journal homepage: www.journalvetbehavior.com
Research
Detection of cigarettes and other tobacco products by giant African pouched rats (Cricetomys gambianus) Amanda Mahoney a, *, Kate La Londe a, b, Timothy L. Edwards a, b, Christophe Cox a, Bart J. Weetjens a, Alan Poling a, b a b
Anti-Persoonsmijnen Ontmijnende Product Ontwikkeling (APOPO), Morogoro, Tanzania, Africa Western Michigan University, Kalamazoo, Michigan
a r t i c l e i n f o
a b s t r a c t
Article history: Received 13 February 2014 Received in revised form 21 May 2014 Accepted 6 June 2014 Available online 13 June 2014
If the illicit tobacco trade were eliminated, governments could gain at least $31.3 billion a year, and more than 164,000 premature deaths a year could be avoided after 2030 (Joossens, Merriman, Ross, and Raw, 2009). Dogs have been used successfully in tobacco control programs, and there is a good chance that rats could also play an important role. In the present experiment, giant African pouched rats were trained to respond to filters that had been stored together with cigarettes (i.e., soaked) and to not respond to filters that had been soaked with noncigarette items. Generalization to untrained types of tobacco was then tested. The sensitivity of 4 rats trained on filters soaked with 1 of 7 types of cigarettes ranged from 86% to 100% (mean, 95%). There was very little evidence of generalization when the rats were tested on tobacco leaves and snuff but good evidence of generalization when the rats were tested on cigarettes that had been soaked with strong-smelling additives. These findings suggest that rats may be a valuable asset in the global effort to control illicit cigarette trade. Ó 2014 Elsevier Inc. All rights reserved.
Keywords: tobacco smuggling scent detection generalization work animals pouched rats hit/positive indication rate
Introduction Smuggled tobacco, comprising 330-660 billion cigarettes annually (Action on Smoking and Health Report, 2010), undermines tobacco control policies by increasing accessibility to and consumption of harmful tobacco products (Illicit Trade Report, 2012, World Customs Organization). Moreover, smuggling prevents governments from collecting tobacco taxes and funds criminal organizations. If the illicit tobacco trade were eliminated, governments could gain at least $31.3 billion in lost revenue annually, and more than 164,000 premature deaths a year could be avoided after 2030 (Joossens, Merriman, Ross, and Raw, 2009). Customs officers and other officials use a variety of methods to search for hidden tobacco, including trained search dogs (Nagy, 2012). The United Kingdom Border Agency Detector Dog Service, for instance, used 11 dogs and handlers to detect more than 18 million cigarettes and 197 tons of other illicit tobacco during 2007/
* Address for reprint requests and correspondence: Amanda Mahoney, PhD, APOPO US, 3375 Scott Blvd, Santa Clara, CA 95054. Tel: þ1-408-398-4341. E-mail address: [email protected] (A. Mahoney). 1558-7878/$ e see front matter Ó 2014 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jveb.2014.06.002
2008 (Home Office UK Border Agency, 2008). Trained tobaccodetection dogs are available from commercial suppliers, and it is clear that Canis familiaris can detect the smell of tobacco. Like dogs, giant African pouched rats (Cricetomys gambianus) have a highly sensitive sense of smell. Previous research has shown that the rats, which are relatively large rodents (1-2 kg body weight) that live for 7-8 years in captivity, can be readily trained through operant conditioning procedures to detect land mines (e.g., Poling et al., 2010a,b; Poling et al., 2011); to detect the presence of Mycobacterium tuberculosis, the bacillus that causes tuberculosis, in human sputum (Mahoney et al., 2012; Poling et al., 2010c; Weetjens et al., 2009); and to detect salmonella bacteria in horse feces (Mahoney et al., in press). The rats are relatively inexpensive to transport, house, and maintain, are not dependent on a single human handler to work well, and maintain accurate performance during long periods of repetitive work. If the rats can reliably detect tobacco, these characteristics make them useful as alternatives or adjuncts to dogs for the detection of smuggled tobacco. The use of rats for tobacco detection could involve either a direct detection or a remote scent tracing (RST) application. Direct detection means that the rats would search in the immediate vicinity of potential objects of interest (i.e., cigarettes or
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other tobacco) and emit a trained indication response, such as scratching, when those objects are encountered. In RST, samples are taken from locations of interest (by, for instance, drawing air and particles from inside shipping containers through filters) and presenting those samples (e.g., filters) to rats in a remote location. Locations producing samples judged by the animals to be positive for the object of interest (i.e., tobacco) are then searched more thoroughly by other methods, such as direct-detection animals or humans, whereas areas producing negative samples are exempt from further inspection except for quality control. Although both systems are potentially valuable, RST has the potential to be faster and cheaper than direct detection (Jones et al., 2011). The present experiment examined whether pouched rats could accurately detect cigarettes in an RST application and whether rats trained to detect cigarettes would also detect other forms of tobacco. Often, an indication response consistently reinforced (rewarded) in the presence of one scent will, with no direct training, occur in the presence of scents that share chemical compounds with the trained scent (e.g., Mgode et al., 2012) or over scents of a different magnitude than the one trained. This process is termed stimulus generalization (Cooper et al., 2007; Cuvo, 2003) and is an important process in the formation of concepts (Herrnstein et al., 1976; Pierce and Cheney, 2013). For example, early training of tuberculosis-detection rats involves presenting sputum samples containing relatively few of the infectious bacilli and reinforcing identification responses in the presence of those samples but not in the presence of bacillus-free samples. Later, the indication responses reliably occur in the presence of samples containing fewer or many more of the bacilli (as determined by counting the bacilli under a microscope), despite the fact that such responses have never been reinforced in the presence of those concentrations. Stimulus generalization can be valuable, as when a rat trained to detect one brand of cigarettes reliably detects other brands, but it can also be a problem, as when a rat trained to detect cigarettes also emits an indication response in the presence of smells not associated with tobacco (e.g., those emanating from manila rope or a foodstuff). There are several factors that may play into whether extraneous odors come to control the response. If the target odor is established first, then blocking may prevent subsequent extraneous odors from being established, whereas if 2 or more odors are established simultaneously then overshadowing may prevent the weaker odor from exerting control (Pierce and Cheney, 2013). The nature of the substances and the concentration level of the extraneous odor also affect the results. Waggoner et al. (1998), for example, trained dogs on a target odor and then tested the dogs on the target odor with an extraneous odor present (i.e., an odor pair). They kept the target odor constant while increasing concentration levels of the extraneous odor and demonstrated across 2 odor pairs that hit/correct positive indication rates and false alarm rates remained stable until levels of the extraneous odor far exceeded levels of the target odor. For example, in the case of 1 odor pair, the level of the target odor remained around 1 ppb, and performance did not decline until the extraneous odor reached about 20 ppm. Accordingly, the primary purpose of the present study was to determine whether giant African pouched rats could be trained to emit an indication response (i.e., pausing over a sample hole) in the presence of filters that had been stored together with tobacco cigarettes in a sealed container (i.e., soaked) and not emit this indication response in the presence of filters that had been soaked with other items. A secondary purpose was to determine whether the rats would generalize to odors of new types of tobacco with no direct training and whether the rats would respond to the trained tobacco cigarettes when strong extraneous odors were present.
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Methods Subjects Four male adult pouched rats (Crycetomys gambianus) served as subjects. The subjects (Harrison, Habreeze, Myron, and Bravo) were 1-2 years old at the start of the study and, before the start of this study, had been trained to emit an indication response to filters soaked with tobacco cigarettes and not emit that response to filters soaked with other items. The subjects were obtained from Anti-Persoonsmijnen Ontmijnende Product Ontwikkeling (APOPO)’s breeding colony and were weaned at 4 weeks of age. Socialization and training began immediately after weaning. During the study, food pellets were earned within sessions, and no food was given outside the sessions except on weekends, when sessions were not arranged. Every Friday at 2 PM, the rats were fed approximately 175 g of a mixture of apples, avocados, sardines, carrots, and peanuts. Any unconsumed food was removed on Sunday afternoon. Ethical clearance to conduct the experiments was obtained from the Tanzanian Ministry of Health. Apparatus and materials Materials for training samples included the targets (unopened cartons of cigarettes, tied plastic bags containing snuff, tied plastic bags containing fire-cured leaf tobacco, and tied plastic bags containing Virginia leaf tobacco), controls (food items wrapped in cellophane, loose tea, lemongrass, coffee, curry powder), filters (approximately 2.5 1.5 cm hand-cut pieces of polypropylene filter paper), and sample “soaking” buckets. “Soaking” is commonly used in the RST literature to refer to placing a target in contact with or in proximity to the filter used to capture the scent of that target. An effort was made to match the packaging and physical properties of the target samples with the controls as closely as possible. For example, when the 2 tobacco leaf types were introduced, 2 nontobacco leaves were introduced as novel controls. The training sample tobacco products and controls were left in their original packaging and were placed in a plastic bucket (30 27 cm) with an airtight lid for at least 5 days before being used for training or testing. On training days, the sample preparer wore latex gloves (changed between sample types) and used tweezers to grab 3 pieces of filter paper. The filter paper was put into small plastic pots (2 cm wide and 3 cm high), and lids were placed on the pots until the time of the evaluation session. Sessions were conducted in a rectangular training cage with Plexiglas walls and a stainless steel floor measuring 205 cm long, 55 cm wide, and 55 cm high. Ten holes 2 cm in diameter with sliding covers were spaced equidistant along the centerline of the floor. An infrared sensor in each hole detected when the rat’s nose was in the hole. When the rat’s nose broke the sensor for a set duration, a pellet dispenser located on the short wall of the cage (near the last hole evaluated) dispensed three 45-mg rat pellets (Test Diet Omnitreat; Test Diet, 1050 Progress Dr., Richmond, IN 47374). Pots containing soaked filters were placed beneath the holes for the rats to evaluate. The pots were held in cassettes, or bars, that fit underneath the floor of the cage and could be locked into place. After all 10 holes were evaluated, a bar was removed and replaced with the next bar. A laptop computer connected to the cage arranged sessions. When a session opened, the computer screen depicted each bar as a row of cells with each new bar starting from the left side of the screen, creating a grid. Within the grid, pots containing the positive (rewarded) sample were shaded gray. When a rat’s nose broke the sensor for the specified duration, the computer recorded that an indication had been made. Indications within shaded cells were
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considered hits. Blind samples were included in all sessions. Blind samples were positive samples but were not depicted on the computer screen or on the data sheet that was printed for each session. Indications within nonshaded hits were considered suspect because the pot could contain a blind positive sample. After each session, the status of blind positive samples was changed to positive, allowing for correct analysis of the results. Training sessions During training sessions, the rat walked on the floor of the cage, smelling each sample as it passed. The rats were trained to start at the hole on the far left and work toward the right. If an indication (i.e., the nose breaking the sensor in the hole for 1.5 seconds) was made over a known positive sample, an auditory click occurred and 3 food pellets were delivered. If an indication was made over a suspect sample, no programmed consequences were delivered. After a response to any sample (positive or negative), the trainer closed the corresponding hole’s cover. The rats evaluated each bar twice, so any samples not responded to on the first run (i.e., with the hole still open) were evaluated a second time. This was done to increase the likelihood that the rat smelled all the samples. Even if positive samples were not present in the bar, the rats evaluated the entire bar twice. The rats smelled the holes sequentially (from left to right), and there were no programmed consequences if it appeared that a hole was skipped. If the rat attempted to go backward and smell holes it had already passed, all earlier hole covers were closed until the rat turned back in the correct direction and were reopened before beginning the next run. A run was terminated when the rat passed over the 10th (last) hole. The next run began when a new bar of samples was fixed underneath the cage and the holes were opened. When training novel scents, the new targets were always presented in bars with controls but no other target types. On these training days, the rats first evaluated 5 bars (50 samples) containing 8 training targets and 42 controls and then evaluated 5 bars with 5 previously trained targets and 45 controls. When a novel target was introduced, the threshold at which the conditioned reinforcer (the click sound) was delivered initially was lowered to 500 milliseconds. This criterion change allowed even very weak responses to be reinforced. The threshold value was gradually raised across training sessions as the rats learned to pause over the novel target and not pause over the novel controls. After performance on the new target improved to 90% accuracy or above for all 4 rats, and the indication responses were at least 1.5 seconds, the new target was presented in bars with previously trained targets. During such mixed training sessions, 1-2 blind samples of each target type were included. Mixed training sessions were administered until performance remained unchanged for 5 days on all trained target types for all 4 rats, then a generalization test to a different type of tobacco or a more complex sample was conducted. Test sessions Test sessions were conducted the same as training sessions, except that untrained sample types were presented as blind samples. Test samples were always evaluated without reinforcement arranged for indication responses. In phase 1.0, there were 6 test samples in each session (37.5% of the total number of positive samples), in phase 1.1 and 2.0 four test samples (28.6% and 33.3%, respectively), and phase 3.0 and 3.1 three test samples (27.3%). The first 2 generalization tests were conducted across 2 days, and the last 3 tests were conducted across 4 days. The Table shows the sample type and number of samples presented each session during each testing stage. The ratio of blind to known positive
Table Test sessions configuration Phase
Targets
No. trained (blind)
1. 0
Cigarettes
10 (2)
1.1
Virginia leaf Fire-cured leaf Snuff Cigarettes
2.0
Virginia leaf Fire-cured leaf Snuff Cigarettes
3.0
Virginia leaf Fire-cured leaf Snuff Cigarettes þ coffee/curry Virginia leaf þ curry Fire-cured leaf þ coffee Snuff þ curry Cigarettes
2 2 2 5 (1) 5 (1) 2 2 2 (1) 2 (1) 2 (1) 2 (1)
a
Cigarettes þ coffee/curry Cigarettes wrapped in expanded polystyrene cardboard, tape
Controls
No.
Packaged food items Loose leaf tea Guava leaves Lemongrass Packaged food items Loose leaf tea Guava leaves Lemongrass Packaged food items Coffee Curry powder
78 2 2 2 84 2 2 2 80 2 2
1 1 1 1 8 (2)
Cigarettes þ coffee/curry 3.1
No. testeda
3 8 (2) 3
Packaged food items Coffee Curry powder Coffee/curry Expanded polystyrene, cardboard, tape
83 2 2 83 2
All test samples were evaluated under blind testing conditions.
samples varied between phases. In phase 1.0 and 1.1, 20% of training samples were blind, in phase 2.0, 50% of training samples were blind, and in phase 3.0 and 3.1, 25% of training samples were blind. Phase 1 Phase 1 tested generalization to 3 types of tobacco after training on 7 different brands of cigarettes. The rats were first trained on filters soaked with 1 of 7 brands of cigarettes. Subsequently, all rats received training on all 7 cigarette brands until performance remained above 90% across 3 consecutive sessions. Generalization to 3 novel types of tobaccodfilters soaked with Virginia leaf tobacco, fire-cured leaf tobacco, and snuffdwere then tested. All rats were tested on each of these new tobacco types. During each day of the test, there were 8 known samples and 2 blind samples soaked with 1 brand of cigarette, 2 blind samples soaked with Virginia leaf tobacco, 2 blind samples soaked with fire-cured leaf tobacco, and 2 blind samples soaked with snuff. There were also 78 samples soaked with previously trained controls (packaged items wrapped in cellophane similar to cigarette packaging). During the test, 1 of these trained control samples was presented on each test day. Finally, 3 untrained control samples were included in the test session; there were 2 samples of filters soaked with ground tea, 2 samples of filters soaked with guava leaves, and 2 samples of filters soaked with lemongrass. Because the rats did not generalize to the new tobacco types, 1 target, Virginia leaf tobacco, was selected for direct training. Direct training meant that reinforcement was arranged for indications. After the rats had continued training on cigarettes generalization to snuff and fire-cured leaf tobacco was then tested again. In each test session, there were 4 known samples and 1 blind sample soaked with 1 brand of cigarette and 4 known samples and 1 blind sample soaked with Virginia leaf tobacco. There were 2 samples from each of the 3 untrained controls: loose-leaf tea, guava leaves, and lemongrass. There were 84 samples soaked with packaged items, which served as trained controls.
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Phase 2 Phase 2 tested whether, after training on filters soaked with 1 of the 4 types of tobacco, the indication response would occur over filters soaked with tobacco and an added substance. The rats were trained directly on snuff and fire-cured leaf tobacco and continued training on Virginia leaf tobacco and cigarettes. All 4 tobacco types plus controls were then mixed during training sessions and presented to the rats across 11 training sessions. While this training stage was ongoing, the experimenter soaked fresh targets of cigarettes, Virginia leaf tobacco, dark leaf tobacco, and snuff separately with filter papers along with either coffee or curry powder for 1 week. Generalization tests were then conducted with these targets. This test was conducted across 4 days. During each test day, there were 2 known samples and 1 blind sample soaked with each trained tobacco target: cigarettes, the 2 types of leaves, and snuff. There was 1 sample from each untrained tobacco target: cigarettes with coffee or curry powder, Virginia leaf tobacco with curry, firecured tobacco leaves with coffee, and snuff with curry. The same trained controls were included as well as 2 novel untrained controls, coffee and curry powder. There were 80 samples from trained controls and 2 samples from the untrained controls in each session. Phase 3 The next training phase reverted to sessions containing only cigarettes as targets for 5 days. Generalization to cigarettes soaked with either coffee or curry powder was then retested. This test was conducted across 4 days. Each test day included 8 known cigarette samples and 2 blind cigarette samples, 3 untrained samples soaked with cigarette plus coffee or curry, 83 samples from the trained controls, and 2 samples soaked with coffee or curry. A total of 12 untrained target filters were presented across this phase, a larger number than in previous phases because it was predicted that the untrained target samples might exert weak stimulus control because of the presence of the distracter odor. The larger sample size allowed for a better measure of the sensitivity of the rats on this sample type while still allowing for analysis of performance on other sample types. Training was then conducted on cigarettes plus coffee or curry powder for 6 days. The final test was conducted across 4 days. Each
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test day included 8 known and 2 blind cigarette plus coffee or curry-soaked filters, 3 untrained samples soaked with cigarettes wrapped in expanded polystyrene (commonly referred to as styrofoam) cardboard, and packaging tape, 83 samples from trained controls, and 2 samples soaked with expanded polystyrene cardboard, and packaging tape. Results The performance of individual rats across target and control samples for the 5 generalization tests are displayed in Figures 1-3. In each test, there were trained targets and trained controls, novel (test) targets, and novel (test) controls to provide a means of assuring that responses to the novel targets were because of shared characteristics with previously learned targets and not simply because of novelty. Figure 1 shows the rats’ hit (correct identification) rates on the first 2 generalization tests, and Figure 2 shows their false alarm rates on these tests. In the first test, the rats were tested on 3 other types of tobacco after training only on cigarettes. During 2 days of tests, the average percentage of hits on trained filters soaked with cigarettes was 94.7% (range, 85.7%-100%). The average percentage of false alarms (emissions of indicator responses to control samples) on trained filters soaked with cellophane-packaged items was 0.16% (range, 0%-0.63%). The average percentage of hits on untrained tobacco types (Virginia leaf tobacco, fire-cured leaf tobacco, and snuff) was 10.4% (range, 0%-25%), which may be indicative of some generalization. However, the average percentage of hits on untrained controls (loose tea, guava leaves, and lemongrass) was 16.7% (range, 0%-50%), suggesting that hits on novel tobacco were not indicative of generalization. Instead, it appeared that there was a low-to-moderate level of rat responding toward all novel stimuli during the first test session. In fact, Bravo responded to all tested targets and tested controls only once, and 5 of his hits occurred on the first presentation of that stimulus type. Myron responded to the fire-cured tobacco leaf-soaked filter once, the guava-soaked filter once, and the lemongrass-soaked filter twice. Harrison responded to the snuff-soaked filter once and the lemongrass-soaked filter
Figure 1. Phase 1 hit rate results. Percentage of hits on targets across trained targets and tested targets for 4 rats.
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Figure 2. Phase 1 false alarm results. Percentage of hits on controls across trained controls and tested controls.
once. Habreeze did not respond to any tested targets and responded to the guava-soaked filter once. The rats were then trained directly on one of the novel tobacco products, Virginia leaf. The first generalization test was then repeated with fire-cured leaf tobacco and snuff. The average hit rate
over filters soaked with cigarettes was 92.7%; Harrison and Habreeze hit all these targets, Myron hit 80% of them, and Bravo hit 90%. During the 2 test days, the average percentage of hits on filters soaked with Virginia leaf tobacco was 70%; Harrison hit all the Virginia leaf tobacco samples, Myron and Habreeze each hit 90% of
Figure 3. Phase 2 and 3 hit rate results. Percentage of hits on targets across trained targets and tested targets for 4 rats.
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them, and Bravo did not hit any of them. None of the rats responded to filters soaked with fire-cured leaf tobacco or filters soaked with snuff. One rat (Harrison) responded to 1 of the trained controls (packaged food items). Unlike the first generalization test, none of the rats responded to novel controls. No consistent indications were observed toward the novel tobacco types in the generalization tests; thus, the final 2 tobacco types were trained directly. After training was complete on all tobacco types, each training session had 1 brand of cigarette (alternated each session), Virginia leaf tobacco, fire-cured leaf tobacco, and snuff. Packaged items, loose tea, guava leaves, and lemongrass served as controls. Hit rates during phases 2 and 3, the final 3 tests, are displayed in Figure 3. False alarms are not depicted because they were very low across all 3 tests; no rat had more than 1 false alarm in a session, and no rat responded to any of the untrained controls. In the third generalization test, filters were soaked with 1 of the 4 target types along with a bag of either coffee or curry powder. Performance on the trained tobacco types was variable, except on cigarettes. All 4 rats hit all samples that had been soaked with cigarettes. Three of the 4 rats hit all samples that had been soaked with Virginia leaf tobacco. Bravo hit 2, or 50%, of the Virginia leaf targets. Harrison and Myron hit all the fire-cured leaf tobacco targets, and Habreeze hit 25% of them. Harrison and Habreeze hit all the snuff targets, whereas Myron and Bravo hit 25% of these targets. There were again very few false alarms on the trained controls; Myron and Habreeze had 1 false alarm each, and the other 2 rats had zero false alarms. Bravo responded to 2 samples soaked with cigarettes and coffee, and Harrison and Myron each responded to 1 sample soaked with cigarettes and curry powder. There was only 1 other indication response toward the targets soaked with a second item: Myron responded to 1 sample containing filters soaked with fire-cured leaf tobacco and coffee. There were no indication responses to samples soaked with coffee or curry powder. This test yielded low hit rates on filters soaked with cigarettes and either coffee or curry powder. It is unclear, however, whether the low hit rate was an outcome of withholding reinforcement for indications or whether it was a reflection of poor stimulus control. The rats were trained for 4 days with only filters soaked with cigarettes or packaged items. The test was then repeated, excluding the other 3 tobacco types. In this test, Harrison and Myron hit all the cigarette-soaked samples, whereas Habreeze hit 95% and Bravo hit 77.5% of them. Myron had 2 false alarms on the packaged items. On the filters soaked with cigarettes plus either coffee or curry powder, Harrison and Myron each hit 83.3%, whereas Habreeze and Bravo each hit 66.7%. There were no false alarms on the filters soaked with coffee or curry powder. Training sessions were conducted with reinforcement arranged for indications over filters soaked with cigarettes plus coffee or curry powder and then the final test was conducted. In this test, the test samples were filters soaked with a carton of cigarettes wrapped in a layer of expanded polystyrene then a layer of cardboard, and then a layer of masking tape. Harrison, Habreeze, and Bravo hit all the filters soaked with cigarettes plus either coffee or curry powder, whereas Myron hit 90% of these samples. Habreeze hit 11 of the 12 test samples (91.7%), whereas Harrison, Myron, and Bravo hit all the 12 test samples. Discussion In this study, giant African pouched rats were readily trained to offer an indication response to filters that had been soaked with 1 of 7 brands of tobacco cigarettes and not offer this indication response over filters soaked with other substances that shared similar
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properties (e.g., guava leaves and loose leaf tea were used as distractors when training leaf tobacco). After initial training, the rats did not generalize to any of 3 other types of tobacco. This was perhaps unsurprising considering that the probability of an indication response occurring is inversely related to the degree of disparity between the trained and tested stimuli (Ghirlanda and Enquist, 2003; Honig and Urcuioli, 1981), and the odor of tobacco cigarettes is likely to differ substantially from that of tobacco leaves or snuff. Generalization to fire-cured tobacco leaves and snuff also failed to occur after the rats were trained to detect Virginia leaf tobacco. They were then trained to detect all tobacco types. The rats learned to identify the 3 new tobacco types when there was only 1 target presented in a session; however, performance dropped for some rats when the different tobacco types were presented together in a session. For example, when cigarettes, the 2 leaf types, and snuff were trained separately, Bravo’s hit rat was more than 90% on all types. When the targets were put together in a session, Bravo’s hit rat on the 2 leaves and snuff fell lower than 50%. Myron’s performance on snuff and Habreeze’s performance on firecured leaves also dropped lower than 50%. In behavioral terminology, stimuli that all evoke a common response are said to comprise a functional stimulus class (e.g., Dougher and Markham, 1994). In this study, although all 4 tobacco types evoked an indication response, the strength of the response across stimuli varied when they were presented in a mixed training format and training on 1 or more targets did not result in generalization to other prospective members of the class. The failure of the 4 types of tobacco to form a functional class could be because of the training procedures. If, for example, all tobacco types had been trained as positive samples for an equal period or if a match-tosample format had been used, emergent responding over other forms of tobacco may have been observed (e.g., Hayes and Barnes, 1997; Wirth and Chase, 2002). It is also possible that the odor bouquet of the different types of tobacco is not more alike than the bouquet of the control substances, in which case generalization would not be expected. The final phase tested whether the rats would continue to respond to target filters when an extraneous odor was present. Three of the 4 rats responded to the filters soaked with cigarettes plus coffee or curry but did not respond to the control filters soaked with coffee or curry alone. None of the rats responded to the other 3 tobacco types when soaked with coffee or curry, again suggesting that stimulus control of these target types over indication responses was not very robust. The test was then repeated excluding the 3 noncigarette tobacco types, essentially creating a less complex discrimination task. In this test, all 4 rats responded to the target filters despite the presence of an extraneous odor and did not respond to the control filters. In the final test, all 4 rats responded to the novel target type (i.e., cigarettes wrapped in expanded polystyrene cardboard, and packing tape) and again did not respond to the novel controls. This study suggests that rats may be useful for detecting illicit cigarettes; there is a lot of work to be done, however, before a firm conclusion can be drawn. This study used passive sampling, in which paper filters were soaked in a sealed bucket with the target samples for at least 5 days and up to several weeks, whereas an operational situation would use active sampling, in which device a pump would suck air from the container through a filter. The filter would then be removed from the device and presented to the rats. It is unclear whether the density of molecules trapped from the cigarette products would be sufficient to make them detectable by the rats. Extraneous molecules would also be trapped, and the quantity and variety of these molecules might have to be mimicked in control samples to adequately train the rats. If so, the preparation of control samples would be complex. Another consideration is the
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number of reward samples needed to maintain rat performance in an operational setting. In this study, 8%-13% of targets evaluated were positive, whereas in reality there may be thousands of containers that do not contain cigarettes for every container that does. To address this issue, APOPO will need to devise a way of implanting known positive targets among samples being tested, while making sure these planted positive samples cannot be discriminated from unidentified positive samples. In short, this study demonstrated that pouched rats were readily trained to detect various tobacco types and, after early training stages, the rats responded to targets containing novel additives. Cigarettes are of paramount importance in the tobacco control effort. An estimated 10.7% of global sales of tobacco are illicit, representing 600 billion cigarettes. This has historically represented revenue losses of between $US40 and $US50 billion to governments around the world (WHO FCTC Protocol, 2003). For this reason, APOPO’s training protocol for tobacco-detection rats will focus on detection of cigarettes, and the data described herein provide evidence that the rats may be a useful supplement to the global effort to control illicit cigarette trade. Acknowledgments The APOPO has Animal Welfare Assurance (no. A5720-01) from the Office of Laboratory Animal Welfare, National Institutes of Health, USA. Ethical considerations The subjects met best practice standards as set by the Animal Welfare Assurance Committee Office of Laboratory Animal Welfare, National Institutes of Health, USA. The research protocol was reviewed and approved by APOPO’s Institutional Animal Care and Use Committee formed at the Sokoine University of Agriculture in Tanzania. Conflict of interest The authors declare no conflict of interest. The idea for the article was conceived by Christophe Cox and Bart Weetjens. The experiments were designed by Amanda Mahoney. The experiments were performed and the data were analyzed by Amanda Mahoney, Kate LaLonde, and Timothy Edwards. The article was written by Amanda Mahoney and Alan Poling. There was no external funding source for this article. References Action on Smoking and Health. 2010. Essential information on tobacco smuggling. ASH Report.
Cooper, J.O., Heron, T.E., Heward, W.L., 2007. Applied Behavior Analysis. Merrill, Columbus, OH. Cuvo, J.A., 2003. On stimulus generalization and stimulus classes. J. Behav. Educ. 12, 77e83. Dougher, M.J., Markham, M.R., 1994. Stimulus equivalence, functional equivalence, and the transfer of function. In: Hayes, S.C., Hayes, L.J., Sato, M., Oko, K. (Eds.), Behavior Analysis of Language and Cognition. Context Press, Reno, NV, pp. 71e90. Ghirlanda, S., Enquist, M., 2003. A century of generalization. Anim. Behav. 66, 15e36. Hayes, S.C., Barnes, D., 1997. Analyzing derived stimulus relations requires more than the concept of a stimulus class. J. Exp. Anal. Behav. 68, 235e270. Herrnstein, R.J., Loveland, D.H., Cable, C., 1976. Natural concepts in pigeons. J. Exp. Psychol.: Anim. Behav. Process. 2, 285. Home Office UK Border Agency, 2008. Tackling smuggled tobacco together. Available at: http://www.hmrc.gov.uk/pbr2008/tobacco-2800.pdf. Accessed August 19, 2011. Honig, W.K., Urcuioli, P.J., 1981. The legacy of Guttman and Kalish (1956): 25 years of research on stimulus generalization. J. Exp. Anal. Behav. 36, 405e445. Jones, B., Fjellanger, R., Cox, C., Poling, A., 2011. Remote explosive scent tracing of explosive remnants of war: a perspective from the 2010 Morogoro Workshop. J. Explos. Remn. War Min. Action 15.1, 55e73. Joossens, L., Merriman, D., Ross, M., Raw, M., 2009. How eliminating the global illicit cigarette trade would increase tax revenue and save lives. The International Union Against Tuberculosis and Lung Disease, Paris, France. Available at: http:// global.tobaccofreekids.org/files/pdfs/en/ILL_global_cig_trade_full_en.pdf. Mahoney, A., Edwards, T., LaLonde, K., Beyene, N., Cox, C., Weetjens, B., Poling, A., 2014. Pouched rats’ (Cricetomys gambianus’s) detection of Salmonella in horse feces. J. Vet. Behav.: Clin. Appl. Res. 9, 124e126. Mahoney, A., Weetjens, B.J., Cox, C., Beyene, N., Reither, K., Makingi, G., Jubitana, M., Kazwala, R., Mfinanga, G.S., Kahwa, A., Durgin, A., Poling, A., 2012. Detection of tuberculosis in human sputum: comparison to culturing and polymerase chain reaction. Tuberc. Res. Treat. http://dx.doi.org/10.1155/2012/716989. Mgode, G.F., Weetjens, B.J., Cox, C., Jubitana, M., Machang’u, R.S., Lazar, D., Weiner, J., Van Geertruyden, J.P., Kaufmann, S.H., 2012. Ability of Cricetomys rats to detect Mycobacterium tuberculosis and discriminate it from other microorganisms. Tuberculosis 92, 182e186. Nagy, J., 2012. Tackling cigarette smuggling with enforcement: case studies reviewing the experience in Hungary, Romania, and the United Kingdom. World Cust. J. 6, 29e40. Pierce, W.D., Cheney, C.D., 2013. Behavior Analysis and Learning, 5th ed. Psychology Press, New York City, NY, pp. 223e254. Poling, A., Weetjens, B.J., Cox, C., Beyene, N., Bach, H., Sully, A., 2010a. Teaching giant African pouched rats to find landmines: operant conditioning with real consequences. Behav. Anal. Pract. 3, 19e25. Poling, A., Weetjens, B., Cox, C., Beyene, N., Sully, A., 2010b. Using giant African pouched rats (Cricetomys gambianus) to detect landmines. Psychol. Rec. 60, 715e727. Poling, A., Weetjens, B.J., Cox, C., Mgode, G., Jubitana, M., Kazwala, R., Mfinanga, G., Huis in ’t Veld, D., 2010c. Using giant African rats to detect tuberculosis in human sputum samples: 2009 findings. Am. J. Trop. Med. Hyg. 83, 1308e1310. Poling, A., Weetjens, B.J., Cox, C., Beyene, N.W., Sully, A., 2011. Using trained pouched rats (Cricetomys gambianus) to detect land mines: another victory for operant conditioning. J. Appl. Behav. Anal. 44, 351e355. Waggoner, L.P., Jones, M., Williams, M., Johnston, J.M., Edge, C., Petrousky, J.A., 1998. Effects of extraneous odors on canine detection. Paper presented at Enforcement and Security Technologies, Boston, MA http://dx.doi.org/10.1117/ 12.335008. Weetjens, B.J., Mgode, G.F., Machang’u, R.S., Kazwala, R., Mfinanga, G., Lwilla, F., Cox, C., Jubitana, M., Kanyagha, H., Mtandu, R., Kahwas, A., Mwessongo, J., Makingi, G., Mfaume, S., Steenberge, V., Beyene, N.W., Billet, M., Verhagen, R., 2009. African pouched rats for the detection of pulmonary tuberculosis in sputum samples. Int. J. Tuberc. Lung Dis. 13, 737e743. FCTC Protocol, 2003 Framework Convention on Tobacco Control. WHO report. Illicit trade report, 2012, World Customs Organization. Wirth, O., Chase, P., 2002. Stability of functional equivalence and stimulus equivalence: effects of baseline reversals. J. Exp. Anal. Behav. 77, 29e47.
Contents lists available at ScienceDirect
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Research
Detection of cigarettes and other tobacco products by giant African pouched rats (Cricetomys gambianus) Amanda Mahoney a, *, Kate La Londe a, b, Timothy L. Edwards a, b, Christophe Cox a, Bart J. Weetjens a, Alan Poling a, b a b
Anti-Persoonsmijnen Ontmijnende Product Ontwikkeling (APOPO), Morogoro, Tanzania, Africa Western Michigan University, Kalamazoo, Michigan
a r t i c l e i n f o
a b s t r a c t
Article history: Received 13 February 2014 Received in revised form 21 May 2014 Accepted 6 June 2014 Available online 13 June 2014
If the illicit tobacco trade were eliminated, governments could gain at least $31.3 billion a year, and more than 164,000 premature deaths a year could be avoided after 2030 (Joossens, Merriman, Ross, and Raw, 2009). Dogs have been used successfully in tobacco control programs, and there is a good chance that rats could also play an important role. In the present experiment, giant African pouched rats were trained to respond to filters that had been stored together with cigarettes (i.e., soaked) and to not respond to filters that had been soaked with noncigarette items. Generalization to untrained types of tobacco was then tested. The sensitivity of 4 rats trained on filters soaked with 1 of 7 types of cigarettes ranged from 86% to 100% (mean, 95%). There was very little evidence of generalization when the rats were tested on tobacco leaves and snuff but good evidence of generalization when the rats were tested on cigarettes that had been soaked with strong-smelling additives. These findings suggest that rats may be a valuable asset in the global effort to control illicit cigarette trade. Ó 2014 Elsevier Inc. All rights reserved.
Keywords: tobacco smuggling scent detection generalization work animals pouched rats hit/positive indication rate
Introduction Smuggled tobacco, comprising 330-660 billion cigarettes annually (Action on Smoking and Health Report, 2010), undermines tobacco control policies by increasing accessibility to and consumption of harmful tobacco products (Illicit Trade Report, 2012, World Customs Organization). Moreover, smuggling prevents governments from collecting tobacco taxes and funds criminal organizations. If the illicit tobacco trade were eliminated, governments could gain at least $31.3 billion in lost revenue annually, and more than 164,000 premature deaths a year could be avoided after 2030 (Joossens, Merriman, Ross, and Raw, 2009). Customs officers and other officials use a variety of methods to search for hidden tobacco, including trained search dogs (Nagy, 2012). The United Kingdom Border Agency Detector Dog Service, for instance, used 11 dogs and handlers to detect more than 18 million cigarettes and 197 tons of other illicit tobacco during 2007/
* Address for reprint requests and correspondence: Amanda Mahoney, PhD, APOPO US, 3375 Scott Blvd, Santa Clara, CA 95054. Tel: þ1-408-398-4341. E-mail address: [email protected] (A. Mahoney). 1558-7878/$ e see front matter Ó 2014 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jveb.2014.06.002
2008 (Home Office UK Border Agency, 2008). Trained tobaccodetection dogs are available from commercial suppliers, and it is clear that Canis familiaris can detect the smell of tobacco. Like dogs, giant African pouched rats (Cricetomys gambianus) have a highly sensitive sense of smell. Previous research has shown that the rats, which are relatively large rodents (1-2 kg body weight) that live for 7-8 years in captivity, can be readily trained through operant conditioning procedures to detect land mines (e.g., Poling et al., 2010a,b; Poling et al., 2011); to detect the presence of Mycobacterium tuberculosis, the bacillus that causes tuberculosis, in human sputum (Mahoney et al., 2012; Poling et al., 2010c; Weetjens et al., 2009); and to detect salmonella bacteria in horse feces (Mahoney et al., in press). The rats are relatively inexpensive to transport, house, and maintain, are not dependent on a single human handler to work well, and maintain accurate performance during long periods of repetitive work. If the rats can reliably detect tobacco, these characteristics make them useful as alternatives or adjuncts to dogs for the detection of smuggled tobacco. The use of rats for tobacco detection could involve either a direct detection or a remote scent tracing (RST) application. Direct detection means that the rats would search in the immediate vicinity of potential objects of interest (i.e., cigarettes or
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other tobacco) and emit a trained indication response, such as scratching, when those objects are encountered. In RST, samples are taken from locations of interest (by, for instance, drawing air and particles from inside shipping containers through filters) and presenting those samples (e.g., filters) to rats in a remote location. Locations producing samples judged by the animals to be positive for the object of interest (i.e., tobacco) are then searched more thoroughly by other methods, such as direct-detection animals or humans, whereas areas producing negative samples are exempt from further inspection except for quality control. Although both systems are potentially valuable, RST has the potential to be faster and cheaper than direct detection (Jones et al., 2011). The present experiment examined whether pouched rats could accurately detect cigarettes in an RST application and whether rats trained to detect cigarettes would also detect other forms of tobacco. Often, an indication response consistently reinforced (rewarded) in the presence of one scent will, with no direct training, occur in the presence of scents that share chemical compounds with the trained scent (e.g., Mgode et al., 2012) or over scents of a different magnitude than the one trained. This process is termed stimulus generalization (Cooper et al., 2007; Cuvo, 2003) and is an important process in the formation of concepts (Herrnstein et al., 1976; Pierce and Cheney, 2013). For example, early training of tuberculosis-detection rats involves presenting sputum samples containing relatively few of the infectious bacilli and reinforcing identification responses in the presence of those samples but not in the presence of bacillus-free samples. Later, the indication responses reliably occur in the presence of samples containing fewer or many more of the bacilli (as determined by counting the bacilli under a microscope), despite the fact that such responses have never been reinforced in the presence of those concentrations. Stimulus generalization can be valuable, as when a rat trained to detect one brand of cigarettes reliably detects other brands, but it can also be a problem, as when a rat trained to detect cigarettes also emits an indication response in the presence of smells not associated with tobacco (e.g., those emanating from manila rope or a foodstuff). There are several factors that may play into whether extraneous odors come to control the response. If the target odor is established first, then blocking may prevent subsequent extraneous odors from being established, whereas if 2 or more odors are established simultaneously then overshadowing may prevent the weaker odor from exerting control (Pierce and Cheney, 2013). The nature of the substances and the concentration level of the extraneous odor also affect the results. Waggoner et al. (1998), for example, trained dogs on a target odor and then tested the dogs on the target odor with an extraneous odor present (i.e., an odor pair). They kept the target odor constant while increasing concentration levels of the extraneous odor and demonstrated across 2 odor pairs that hit/correct positive indication rates and false alarm rates remained stable until levels of the extraneous odor far exceeded levels of the target odor. For example, in the case of 1 odor pair, the level of the target odor remained around 1 ppb, and performance did not decline until the extraneous odor reached about 20 ppm. Accordingly, the primary purpose of the present study was to determine whether giant African pouched rats could be trained to emit an indication response (i.e., pausing over a sample hole) in the presence of filters that had been stored together with tobacco cigarettes in a sealed container (i.e., soaked) and not emit this indication response in the presence of filters that had been soaked with other items. A secondary purpose was to determine whether the rats would generalize to odors of new types of tobacco with no direct training and whether the rats would respond to the trained tobacco cigarettes when strong extraneous odors were present.
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Methods Subjects Four male adult pouched rats (Crycetomys gambianus) served as subjects. The subjects (Harrison, Habreeze, Myron, and Bravo) were 1-2 years old at the start of the study and, before the start of this study, had been trained to emit an indication response to filters soaked with tobacco cigarettes and not emit that response to filters soaked with other items. The subjects were obtained from Anti-Persoonsmijnen Ontmijnende Product Ontwikkeling (APOPO)’s breeding colony and were weaned at 4 weeks of age. Socialization and training began immediately after weaning. During the study, food pellets were earned within sessions, and no food was given outside the sessions except on weekends, when sessions were not arranged. Every Friday at 2 PM, the rats were fed approximately 175 g of a mixture of apples, avocados, sardines, carrots, and peanuts. Any unconsumed food was removed on Sunday afternoon. Ethical clearance to conduct the experiments was obtained from the Tanzanian Ministry of Health. Apparatus and materials Materials for training samples included the targets (unopened cartons of cigarettes, tied plastic bags containing snuff, tied plastic bags containing fire-cured leaf tobacco, and tied plastic bags containing Virginia leaf tobacco), controls (food items wrapped in cellophane, loose tea, lemongrass, coffee, curry powder), filters (approximately 2.5 1.5 cm hand-cut pieces of polypropylene filter paper), and sample “soaking” buckets. “Soaking” is commonly used in the RST literature to refer to placing a target in contact with or in proximity to the filter used to capture the scent of that target. An effort was made to match the packaging and physical properties of the target samples with the controls as closely as possible. For example, when the 2 tobacco leaf types were introduced, 2 nontobacco leaves were introduced as novel controls. The training sample tobacco products and controls were left in their original packaging and were placed in a plastic bucket (30 27 cm) with an airtight lid for at least 5 days before being used for training or testing. On training days, the sample preparer wore latex gloves (changed between sample types) and used tweezers to grab 3 pieces of filter paper. The filter paper was put into small plastic pots (2 cm wide and 3 cm high), and lids were placed on the pots until the time of the evaluation session. Sessions were conducted in a rectangular training cage with Plexiglas walls and a stainless steel floor measuring 205 cm long, 55 cm wide, and 55 cm high. Ten holes 2 cm in diameter with sliding covers were spaced equidistant along the centerline of the floor. An infrared sensor in each hole detected when the rat’s nose was in the hole. When the rat’s nose broke the sensor for a set duration, a pellet dispenser located on the short wall of the cage (near the last hole evaluated) dispensed three 45-mg rat pellets (Test Diet Omnitreat; Test Diet, 1050 Progress Dr., Richmond, IN 47374). Pots containing soaked filters were placed beneath the holes for the rats to evaluate. The pots were held in cassettes, or bars, that fit underneath the floor of the cage and could be locked into place. After all 10 holes were evaluated, a bar was removed and replaced with the next bar. A laptop computer connected to the cage arranged sessions. When a session opened, the computer screen depicted each bar as a row of cells with each new bar starting from the left side of the screen, creating a grid. Within the grid, pots containing the positive (rewarded) sample were shaded gray. When a rat’s nose broke the sensor for the specified duration, the computer recorded that an indication had been made. Indications within shaded cells were
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considered hits. Blind samples were included in all sessions. Blind samples were positive samples but were not depicted on the computer screen or on the data sheet that was printed for each session. Indications within nonshaded hits were considered suspect because the pot could contain a blind positive sample. After each session, the status of blind positive samples was changed to positive, allowing for correct analysis of the results. Training sessions During training sessions, the rat walked on the floor of the cage, smelling each sample as it passed. The rats were trained to start at the hole on the far left and work toward the right. If an indication (i.e., the nose breaking the sensor in the hole for 1.5 seconds) was made over a known positive sample, an auditory click occurred and 3 food pellets were delivered. If an indication was made over a suspect sample, no programmed consequences were delivered. After a response to any sample (positive or negative), the trainer closed the corresponding hole’s cover. The rats evaluated each bar twice, so any samples not responded to on the first run (i.e., with the hole still open) were evaluated a second time. This was done to increase the likelihood that the rat smelled all the samples. Even if positive samples were not present in the bar, the rats evaluated the entire bar twice. The rats smelled the holes sequentially (from left to right), and there were no programmed consequences if it appeared that a hole was skipped. If the rat attempted to go backward and smell holes it had already passed, all earlier hole covers were closed until the rat turned back in the correct direction and were reopened before beginning the next run. A run was terminated when the rat passed over the 10th (last) hole. The next run began when a new bar of samples was fixed underneath the cage and the holes were opened. When training novel scents, the new targets were always presented in bars with controls but no other target types. On these training days, the rats first evaluated 5 bars (50 samples) containing 8 training targets and 42 controls and then evaluated 5 bars with 5 previously trained targets and 45 controls. When a novel target was introduced, the threshold at which the conditioned reinforcer (the click sound) was delivered initially was lowered to 500 milliseconds. This criterion change allowed even very weak responses to be reinforced. The threshold value was gradually raised across training sessions as the rats learned to pause over the novel target and not pause over the novel controls. After performance on the new target improved to 90% accuracy or above for all 4 rats, and the indication responses were at least 1.5 seconds, the new target was presented in bars with previously trained targets. During such mixed training sessions, 1-2 blind samples of each target type were included. Mixed training sessions were administered until performance remained unchanged for 5 days on all trained target types for all 4 rats, then a generalization test to a different type of tobacco or a more complex sample was conducted. Test sessions Test sessions were conducted the same as training sessions, except that untrained sample types were presented as blind samples. Test samples were always evaluated without reinforcement arranged for indication responses. In phase 1.0, there were 6 test samples in each session (37.5% of the total number of positive samples), in phase 1.1 and 2.0 four test samples (28.6% and 33.3%, respectively), and phase 3.0 and 3.1 three test samples (27.3%). The first 2 generalization tests were conducted across 2 days, and the last 3 tests were conducted across 4 days. The Table shows the sample type and number of samples presented each session during each testing stage. The ratio of blind to known positive
Table Test sessions configuration Phase
Targets
No. trained (blind)
1. 0
Cigarettes
10 (2)
1.1
Virginia leaf Fire-cured leaf Snuff Cigarettes
2.0
Virginia leaf Fire-cured leaf Snuff Cigarettes
3.0
Virginia leaf Fire-cured leaf Snuff Cigarettes þ coffee/curry Virginia leaf þ curry Fire-cured leaf þ coffee Snuff þ curry Cigarettes
2 2 2 5 (1) 5 (1) 2 2 2 (1) 2 (1) 2 (1) 2 (1)
a
Cigarettes þ coffee/curry Cigarettes wrapped in expanded polystyrene cardboard, tape
Controls
No.
Packaged food items Loose leaf tea Guava leaves Lemongrass Packaged food items Loose leaf tea Guava leaves Lemongrass Packaged food items Coffee Curry powder
78 2 2 2 84 2 2 2 80 2 2
1 1 1 1 8 (2)
Cigarettes þ coffee/curry 3.1
No. testeda
3 8 (2) 3
Packaged food items Coffee Curry powder Coffee/curry Expanded polystyrene, cardboard, tape
83 2 2 83 2
All test samples were evaluated under blind testing conditions.
samples varied between phases. In phase 1.0 and 1.1, 20% of training samples were blind, in phase 2.0, 50% of training samples were blind, and in phase 3.0 and 3.1, 25% of training samples were blind. Phase 1 Phase 1 tested generalization to 3 types of tobacco after training on 7 different brands of cigarettes. The rats were first trained on filters soaked with 1 of 7 brands of cigarettes. Subsequently, all rats received training on all 7 cigarette brands until performance remained above 90% across 3 consecutive sessions. Generalization to 3 novel types of tobaccodfilters soaked with Virginia leaf tobacco, fire-cured leaf tobacco, and snuffdwere then tested. All rats were tested on each of these new tobacco types. During each day of the test, there were 8 known samples and 2 blind samples soaked with 1 brand of cigarette, 2 blind samples soaked with Virginia leaf tobacco, 2 blind samples soaked with fire-cured leaf tobacco, and 2 blind samples soaked with snuff. There were also 78 samples soaked with previously trained controls (packaged items wrapped in cellophane similar to cigarette packaging). During the test, 1 of these trained control samples was presented on each test day. Finally, 3 untrained control samples were included in the test session; there were 2 samples of filters soaked with ground tea, 2 samples of filters soaked with guava leaves, and 2 samples of filters soaked with lemongrass. Because the rats did not generalize to the new tobacco types, 1 target, Virginia leaf tobacco, was selected for direct training. Direct training meant that reinforcement was arranged for indications. After the rats had continued training on cigarettes generalization to snuff and fire-cured leaf tobacco was then tested again. In each test session, there were 4 known samples and 1 blind sample soaked with 1 brand of cigarette and 4 known samples and 1 blind sample soaked with Virginia leaf tobacco. There were 2 samples from each of the 3 untrained controls: loose-leaf tea, guava leaves, and lemongrass. There were 84 samples soaked with packaged items, which served as trained controls.
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Phase 2 Phase 2 tested whether, after training on filters soaked with 1 of the 4 types of tobacco, the indication response would occur over filters soaked with tobacco and an added substance. The rats were trained directly on snuff and fire-cured leaf tobacco and continued training on Virginia leaf tobacco and cigarettes. All 4 tobacco types plus controls were then mixed during training sessions and presented to the rats across 11 training sessions. While this training stage was ongoing, the experimenter soaked fresh targets of cigarettes, Virginia leaf tobacco, dark leaf tobacco, and snuff separately with filter papers along with either coffee or curry powder for 1 week. Generalization tests were then conducted with these targets. This test was conducted across 4 days. During each test day, there were 2 known samples and 1 blind sample soaked with each trained tobacco target: cigarettes, the 2 types of leaves, and snuff. There was 1 sample from each untrained tobacco target: cigarettes with coffee or curry powder, Virginia leaf tobacco with curry, firecured tobacco leaves with coffee, and snuff with curry. The same trained controls were included as well as 2 novel untrained controls, coffee and curry powder. There were 80 samples from trained controls and 2 samples from the untrained controls in each session. Phase 3 The next training phase reverted to sessions containing only cigarettes as targets for 5 days. Generalization to cigarettes soaked with either coffee or curry powder was then retested. This test was conducted across 4 days. Each test day included 8 known cigarette samples and 2 blind cigarette samples, 3 untrained samples soaked with cigarette plus coffee or curry, 83 samples from the trained controls, and 2 samples soaked with coffee or curry. A total of 12 untrained target filters were presented across this phase, a larger number than in previous phases because it was predicted that the untrained target samples might exert weak stimulus control because of the presence of the distracter odor. The larger sample size allowed for a better measure of the sensitivity of the rats on this sample type while still allowing for analysis of performance on other sample types. Training was then conducted on cigarettes plus coffee or curry powder for 6 days. The final test was conducted across 4 days. Each
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test day included 8 known and 2 blind cigarette plus coffee or curry-soaked filters, 3 untrained samples soaked with cigarettes wrapped in expanded polystyrene (commonly referred to as styrofoam) cardboard, and packaging tape, 83 samples from trained controls, and 2 samples soaked with expanded polystyrene cardboard, and packaging tape. Results The performance of individual rats across target and control samples for the 5 generalization tests are displayed in Figures 1-3. In each test, there were trained targets and trained controls, novel (test) targets, and novel (test) controls to provide a means of assuring that responses to the novel targets were because of shared characteristics with previously learned targets and not simply because of novelty. Figure 1 shows the rats’ hit (correct identification) rates on the first 2 generalization tests, and Figure 2 shows their false alarm rates on these tests. In the first test, the rats were tested on 3 other types of tobacco after training only on cigarettes. During 2 days of tests, the average percentage of hits on trained filters soaked with cigarettes was 94.7% (range, 85.7%-100%). The average percentage of false alarms (emissions of indicator responses to control samples) on trained filters soaked with cellophane-packaged items was 0.16% (range, 0%-0.63%). The average percentage of hits on untrained tobacco types (Virginia leaf tobacco, fire-cured leaf tobacco, and snuff) was 10.4% (range, 0%-25%), which may be indicative of some generalization. However, the average percentage of hits on untrained controls (loose tea, guava leaves, and lemongrass) was 16.7% (range, 0%-50%), suggesting that hits on novel tobacco were not indicative of generalization. Instead, it appeared that there was a low-to-moderate level of rat responding toward all novel stimuli during the first test session. In fact, Bravo responded to all tested targets and tested controls only once, and 5 of his hits occurred on the first presentation of that stimulus type. Myron responded to the fire-cured tobacco leaf-soaked filter once, the guava-soaked filter once, and the lemongrass-soaked filter twice. Harrison responded to the snuff-soaked filter once and the lemongrass-soaked filter
Figure 1. Phase 1 hit rate results. Percentage of hits on targets across trained targets and tested targets for 4 rats.
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Figure 2. Phase 1 false alarm results. Percentage of hits on controls across trained controls and tested controls.
once. Habreeze did not respond to any tested targets and responded to the guava-soaked filter once. The rats were then trained directly on one of the novel tobacco products, Virginia leaf. The first generalization test was then repeated with fire-cured leaf tobacco and snuff. The average hit rate
over filters soaked with cigarettes was 92.7%; Harrison and Habreeze hit all these targets, Myron hit 80% of them, and Bravo hit 90%. During the 2 test days, the average percentage of hits on filters soaked with Virginia leaf tobacco was 70%; Harrison hit all the Virginia leaf tobacco samples, Myron and Habreeze each hit 90% of
Figure 3. Phase 2 and 3 hit rate results. Percentage of hits on targets across trained targets and tested targets for 4 rats.
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them, and Bravo did not hit any of them. None of the rats responded to filters soaked with fire-cured leaf tobacco or filters soaked with snuff. One rat (Harrison) responded to 1 of the trained controls (packaged food items). Unlike the first generalization test, none of the rats responded to novel controls. No consistent indications were observed toward the novel tobacco types in the generalization tests; thus, the final 2 tobacco types were trained directly. After training was complete on all tobacco types, each training session had 1 brand of cigarette (alternated each session), Virginia leaf tobacco, fire-cured leaf tobacco, and snuff. Packaged items, loose tea, guava leaves, and lemongrass served as controls. Hit rates during phases 2 and 3, the final 3 tests, are displayed in Figure 3. False alarms are not depicted because they were very low across all 3 tests; no rat had more than 1 false alarm in a session, and no rat responded to any of the untrained controls. In the third generalization test, filters were soaked with 1 of the 4 target types along with a bag of either coffee or curry powder. Performance on the trained tobacco types was variable, except on cigarettes. All 4 rats hit all samples that had been soaked with cigarettes. Three of the 4 rats hit all samples that had been soaked with Virginia leaf tobacco. Bravo hit 2, or 50%, of the Virginia leaf targets. Harrison and Myron hit all the fire-cured leaf tobacco targets, and Habreeze hit 25% of them. Harrison and Habreeze hit all the snuff targets, whereas Myron and Bravo hit 25% of these targets. There were again very few false alarms on the trained controls; Myron and Habreeze had 1 false alarm each, and the other 2 rats had zero false alarms. Bravo responded to 2 samples soaked with cigarettes and coffee, and Harrison and Myron each responded to 1 sample soaked with cigarettes and curry powder. There was only 1 other indication response toward the targets soaked with a second item: Myron responded to 1 sample containing filters soaked with fire-cured leaf tobacco and coffee. There were no indication responses to samples soaked with coffee or curry powder. This test yielded low hit rates on filters soaked with cigarettes and either coffee or curry powder. It is unclear, however, whether the low hit rate was an outcome of withholding reinforcement for indications or whether it was a reflection of poor stimulus control. The rats were trained for 4 days with only filters soaked with cigarettes or packaged items. The test was then repeated, excluding the other 3 tobacco types. In this test, Harrison and Myron hit all the cigarette-soaked samples, whereas Habreeze hit 95% and Bravo hit 77.5% of them. Myron had 2 false alarms on the packaged items. On the filters soaked with cigarettes plus either coffee or curry powder, Harrison and Myron each hit 83.3%, whereas Habreeze and Bravo each hit 66.7%. There were no false alarms on the filters soaked with coffee or curry powder. Training sessions were conducted with reinforcement arranged for indications over filters soaked with cigarettes plus coffee or curry powder and then the final test was conducted. In this test, the test samples were filters soaked with a carton of cigarettes wrapped in a layer of expanded polystyrene then a layer of cardboard, and then a layer of masking tape. Harrison, Habreeze, and Bravo hit all the filters soaked with cigarettes plus either coffee or curry powder, whereas Myron hit 90% of these samples. Habreeze hit 11 of the 12 test samples (91.7%), whereas Harrison, Myron, and Bravo hit all the 12 test samples. Discussion In this study, giant African pouched rats were readily trained to offer an indication response to filters that had been soaked with 1 of 7 brands of tobacco cigarettes and not offer this indication response over filters soaked with other substances that shared similar
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properties (e.g., guava leaves and loose leaf tea were used as distractors when training leaf tobacco). After initial training, the rats did not generalize to any of 3 other types of tobacco. This was perhaps unsurprising considering that the probability of an indication response occurring is inversely related to the degree of disparity between the trained and tested stimuli (Ghirlanda and Enquist, 2003; Honig and Urcuioli, 1981), and the odor of tobacco cigarettes is likely to differ substantially from that of tobacco leaves or snuff. Generalization to fire-cured tobacco leaves and snuff also failed to occur after the rats were trained to detect Virginia leaf tobacco. They were then trained to detect all tobacco types. The rats learned to identify the 3 new tobacco types when there was only 1 target presented in a session; however, performance dropped for some rats when the different tobacco types were presented together in a session. For example, when cigarettes, the 2 leaf types, and snuff were trained separately, Bravo’s hit rat was more than 90% on all types. When the targets were put together in a session, Bravo’s hit rat on the 2 leaves and snuff fell lower than 50%. Myron’s performance on snuff and Habreeze’s performance on firecured leaves also dropped lower than 50%. In behavioral terminology, stimuli that all evoke a common response are said to comprise a functional stimulus class (e.g., Dougher and Markham, 1994). In this study, although all 4 tobacco types evoked an indication response, the strength of the response across stimuli varied when they were presented in a mixed training format and training on 1 or more targets did not result in generalization to other prospective members of the class. The failure of the 4 types of tobacco to form a functional class could be because of the training procedures. If, for example, all tobacco types had been trained as positive samples for an equal period or if a match-tosample format had been used, emergent responding over other forms of tobacco may have been observed (e.g., Hayes and Barnes, 1997; Wirth and Chase, 2002). It is also possible that the odor bouquet of the different types of tobacco is not more alike than the bouquet of the control substances, in which case generalization would not be expected. The final phase tested whether the rats would continue to respond to target filters when an extraneous odor was present. Three of the 4 rats responded to the filters soaked with cigarettes plus coffee or curry but did not respond to the control filters soaked with coffee or curry alone. None of the rats responded to the other 3 tobacco types when soaked with coffee or curry, again suggesting that stimulus control of these target types over indication responses was not very robust. The test was then repeated excluding the 3 noncigarette tobacco types, essentially creating a less complex discrimination task. In this test, all 4 rats responded to the target filters despite the presence of an extraneous odor and did not respond to the control filters. In the final test, all 4 rats responded to the novel target type (i.e., cigarettes wrapped in expanded polystyrene cardboard, and packing tape) and again did not respond to the novel controls. This study suggests that rats may be useful for detecting illicit cigarettes; there is a lot of work to be done, however, before a firm conclusion can be drawn. This study used passive sampling, in which paper filters were soaked in a sealed bucket with the target samples for at least 5 days and up to several weeks, whereas an operational situation would use active sampling, in which device a pump would suck air from the container through a filter. The filter would then be removed from the device and presented to the rats. It is unclear whether the density of molecules trapped from the cigarette products would be sufficient to make them detectable by the rats. Extraneous molecules would also be trapped, and the quantity and variety of these molecules might have to be mimicked in control samples to adequately train the rats. If so, the preparation of control samples would be complex. Another consideration is the
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number of reward samples needed to maintain rat performance in an operational setting. In this study, 8%-13% of targets evaluated were positive, whereas in reality there may be thousands of containers that do not contain cigarettes for every container that does. To address this issue, APOPO will need to devise a way of implanting known positive targets among samples being tested, while making sure these planted positive samples cannot be discriminated from unidentified positive samples. In short, this study demonstrated that pouched rats were readily trained to detect various tobacco types and, after early training stages, the rats responded to targets containing novel additives. Cigarettes are of paramount importance in the tobacco control effort. An estimated 10.7% of global sales of tobacco are illicit, representing 600 billion cigarettes. This has historically represented revenue losses of between $US40 and $US50 billion to governments around the world (WHO FCTC Protocol, 2003). For this reason, APOPO’s training protocol for tobacco-detection rats will focus on detection of cigarettes, and the data described herein provide evidence that the rats may be a useful supplement to the global effort to control illicit cigarette trade. Acknowledgments The APOPO has Animal Welfare Assurance (no. A5720-01) from the Office of Laboratory Animal Welfare, National Institutes of Health, USA. Ethical considerations The subjects met best practice standards as set by the Animal Welfare Assurance Committee Office of Laboratory Animal Welfare, National Institutes of Health, USA. The research protocol was reviewed and approved by APOPO’s Institutional Animal Care and Use Committee formed at the Sokoine University of Agriculture in Tanzania. Conflict of interest The authors declare no conflict of interest. The idea for the article was conceived by Christophe Cox and Bart Weetjens. The experiments were designed by Amanda Mahoney. The experiments were performed and the data were analyzed by Amanda Mahoney, Kate LaLonde, and Timothy Edwards. The article was written by Amanda Mahoney and Alan Poling. There was no external funding source for this article. References Action on Smoking and Health. 2010. Essential information on tobacco smuggling. ASH Report.
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