Recognition of prey handedness

In every experiment the snake moved its eyes to stare at the crawling snail as soon as the latter was placed in front of the snake. While staring, the snake approached 64 of 76 (84.2%) dextrals and 26 of 38 (68.4%) sinistrals presented. In the rest of the cases, the snake averted its head and eyes from the snail and did not approach. When approaching, the snake kept staring at the snail. During this period, the snake often reoriented its head to the snail by shifting the direction and angle of head-tilting at a distance of 10 to 20 mm from the snail. These behaviors during continuous staring suggest the importance of vision for critical operation of the mandibles and upper jaws at the subsequent moment of strike. The snake struck at 52 of the 64 (81.2%) dextrals and 19 of the 26 (73.1%) sinistrals. In the rest of the approaching cases, the snake moved away from the snail without striking.

There was no difference in shell size between approached and non-approached dextral snails (Fig. 2a). In contrast, approached sinistrals were smaller than non-approached ones. This was significant in terms of the effect of interaction between shell size and handedness on this positive or negative decision for approach (F 1,110 = 5.4, p = 0.013, Fig. 2a). Similarly, dextrals that were struck did not differ in shell size from dextrals that were not struck, whereas sinistrals struck were significantly smaller than sinistrals not struck (F 1,86 = 6.0, p = 0.016). The mean shell sizes presented to the snake did not differ between these dextrals and sinistrals (see materials and methods). Therefore, the snake dinstinguished prey enatiomorphs during staring and approaching.

Figure 2 Handedness-dependent size effects on decisions for approach (a) and strike (b), and size-dependent predation on sinistrals (c). No and Yes are the negative and positive decisions, respectively. The snake strikes at a snail only after approach. Each number under the decision indicates the number of replicates. Each of plots and error bars indicates the mean and standard error. Full size image

Neither the time lengths of staring nor of approaching depended on the subsequent decisions (F 1,64 = 1.1, p = 0.29; F 1,79 = 1.3, p = 0.26, respectively) or on snail handedness (F 1,64 = 0.03, p = 0.87; F 1,79 = 0.15, p = 0.70, respectively). On average, snakes made decisions for approach in 10.3 sec ± 2.5 S.E. and for strike in 63.9 sec ± 7.9. The snake was given no choice of handedness of prey. Nevertheless, the snake frequently refrained from approaching or striking at a relatively large sinistral by staring for around 10 seconds or one minute, respectively.

Squamate reptiles use tongue-flicks for vomeronasal chemoreception30. In the present experiment, however, the snake did not flick the tongue in 85 of 111 (76.6%) cases before the decision for approach or in 54 of 88 (61.4%) cases before the decision for strike. In the rest (tongue-flicking cases), the number of tongue-flicks did not depend on the shell size (F 1,11 = 0.03, p = 0.86 before the decision for approach; F 1,20 = 0.001, p = 0.99 before the decision for strike) or on snail handedness (F 1,11 = 0.08, p = 0.78 before the decision for approach; F 1,20 = 3.5, p = 0.076 before the decision for strike).

Without flicking the tongue, the snake could not obtain odors for vomeronasal chemoreception. Nevertheless, the snake ceased staring at and did not approach relatively large sinistrals (F 1,81 = 7.1, p = 0.009, Fig. 2b). After it approached in the other cases, the snake also refrained from striking at one dextral and two sinistrals without flicking the tongue. These sinistrals not struck may have been larger than the other sinistrals struck, but more replicates are necessary for statistical validation (Fig. 2b). In this predation experiment with no choice of prey handedness, the snake avoided preying on relatively large sinistrals whereas it preyed on dextrals irrespective of the shell size (F 1,110 = 11.7, p = 0.001; Fig. 2c). This was also the case when the snake did not flick the tongue (F 1,62 = 9.1, p = 0.004). The snake therefore recognizes prey handedness without relying on vomeronasal chemoreception by tongue-flick.

The head-tilting direction for strike varied among predation events irrespective of snake individuals (Fig. 3). This direction was rightward more frequently for sinistral prey than for dextral prey (F 1,67 = 4.1, p = 0.046). This indicates that prey handedness affects the left-right direction of predatory behavior. In every strike the snake successfully captured and fed the prey, unlike P. iwasakii5,15.

Figure 3: Frequencies of leftward and rightward strikes at dextral and sinistral prey. Each value indicates the number of replicates. Predation succeeded in every striking occasion. Full size image

While handling the prey after strike, the snake held the ventral outer surface of the last whorl (umbilicus side) with the upper jaws and the soft body with the mandibles inserted into the aperture (Fig. 4). When the prey was dextral, the left mandible was at the peripheral side of the whorl and the right mandible at the side of the shell columellar (umbilicus) (Fig. 4a). On the other hand, whenever preying on the sinistral, the snake oppositely positioned the left and right mandibles with no change of the upper jaws’ location on the ventral shell surface (Fig. 4b). This means that the snake laterally reverses the manner of prey handling according to the direction of shell asymmetry. Otherwise, in the case of sinistral prey, the snake would have directed the upper jaws toward the dorsal outer surface of the shell aperture and often fail in prey capture, as known for P. iwasakii5,15.

Figure 4: Left-right reversal of handling behavior in Pareas carinatus according to prey handedness. (a) Handling of a dextral snail (Cryptozona siamensis). (b) Handling of a sinistral snail (Dyakia salangana). Left and right mandibles are inserted into the aperture in reverse relative to the structure of shell whorl while the upper jaws hold the ventral outer surface (umbilicus side) of the shell. Full size image

Different snail species have different odors31. However, our results rule out olfactory recognition of sinistral species by the vomeronasal system, which is typically important for squamates’ chemical recognition30. In the distribution range of Pareas, prey reversal frequently evolved in response to specialized dextral-predation by the snakes15. These phylogenetically independent sinistral lineages reversed by a single gene15,24 would be unlikely to evolve to release sinistral-specific odors if any. Of 29 sinistrals of Dyakia salangana given to the snake, which must have shared species-specific odors, the snake only approached 17 that were smaller in the mean shell size (14.6 mm ± 1.6 S.E.) than the rest (23.7 ± 0.87; F 1,27 = 8.8, p = 0.006). After approach, the snake struck at the smaller 10 (11.3 ± 1.6) but not at the other larger sinistrals (22.1 ± 1.1; F 1,27 = 8.7, p = 0.006). These size-dependent decisions for predation on conspecific prey are not ascribable to prey odor differences.

For predation success in extracting the soft body, which is otherwise withdrawn into the shell, snail-eating snakes must locate the mouthparts properly onto the asymmetric shell at the moment of strike5. Pareas carinatus reverses such definite orientation of the apparatus by staring at sinistral prey (Fig. 4). Pareas snakes have a developed optical system with the large eyes for night vision as well as other nocturnal reptiles do32,33,34,35. These suggest that visual structure perception is necessary for their chirally specialized predation and overrides chemical odor distinction36 where prey is visible. Dextral and sinistral shells are physically discrete in coiling direction and lateral location of the aperture, through which the soft body is extracted (Fig. 1). These major differences in shell structure may serve as a visual cue for the snake to distinguish between prey enantiomorphs.

Feeding efficiency

The snake took a longer time to finish feeding on a larger snail regardless of prey handedness (F 1,64 = 4.6, p = 0.036) and retracted the mandibles for a larger number of times while taking longer (F 1,64 = 057, p < 0.001). However, the number of retractions increased only with the dextral prey size (F 1,47 = 31, p < 0.001), but did not with the sinistral prey size (F 1,17 = 0.65, p = 0.43) (Fig. 5a). Thus, difference in the number of retractions between prey enantiomorphs depended on the prey size (F 1,64 = 9.7, p = 0.003).

Figure 5: Size-dependent efficiencies and benefits of preying on a dextral. (a) Size dependent increase of the number of mandibular retractions only in dextral-predation. (b) Higher retraction frequency in dextral-predation than in sinistral-predation. (c) Size-dependent increase of soft-body mass gained per retraction only in dextral-predation. (d) Size-dependent increase of relative benefit only in dextral-predation. Time is the feeding time in seconds. Mass is the prey weight reduction in grams after predation. Solid and open circles indicate predations on dextrals and sinistrals, respectively. The regression (interrupted) line for sinistrals is illustrated in each case to indicate the intersection with that (solid line) for dextrals, though the slope for the former was not significant (see Table S1 for regression statistics). Full size image

The snake rectracted the mandibles more frequently while preying on the dextral than the sinistral (F 1,64 = 5.3, p = 0.024) (Fig. 5b). The prey soft-body mass gained per retraction increased with the dextral prey size (F 1,47 = 66, p = 0.024) but did not with the sinistral prey size (F 1,17 = 0.008, p = 0.93) (Fig. 5c). The interaction between the size and handedness was accordingly significant (F 1,30 = 8.1, p = 0.008). Thus, when preying on the dextral, mandibular retractions are not only more frequent but also increasingly more efficient with the prey size in terms of soft-body gain per retraction.

Superior performances in feeding on dextral prey in terms of retraction frequency and efficiency synergistically resulted in a significantly larger gain of soft body per time than that achieved by feeding on sinistral prey (F 1,30 = 8.5, p = 0.007) (Fig. 5d). This benefit of preying on the dextral instead of the sinistral increased with the shell size (F 1,30 = 8.0, p = 0.008), as the gain per time positively depended on the dextral’s size (F 1,47 = 57, p = 0.001) but not on the sinistral’s size (F 1,17 = 0.15, p = 0.70).

In Fig. 5a,c,d, regression lines for the dextral and sinistral cases cross at the shell sizes of 11.4, 12.1 and 12.7 mm, respectively (see Table S1 for the regression statistics). This predicts that the relative value of sinistral prey declines with the increase of the size. In practice, the snake preyed on all of the sinistrals smaller than 12.4 mm. However, the snake did not strike at 18 of the 26 (69.2%) sinistrals larger than this size. These cases of avoidance do not appear in Fig. 5, but nevertheless correspond to the range beyond the predicted threshold size of around 12 mm. These results support the hypothesis that the size-dependent increase of cost for preying on a sinistral instead of a dextral has driven the evolution of prey-handedness recognition and size-dependent avoidance of sinistral-predation.

In the distribution range of P. carinatus, sinistral species reach 17.0% in the total of 900 pulmonate species, exclusive of those that are too small for the snake to prey (see methods; Table S2). Our field records demonstrate that this arboreal snake is frequently active on trees where tree snails of pulmonates co-occur (Table S3). These tree snail species are almost invariably sinistral (subgenus Syndromus) or chirally dimorphic within populations (subgenus Amphidromus). Their high abundances are well established26,27,37,38. On the other hand, no sinistral tree snail co-occurs with a congeneric snake P. iwasakii, which lives on the islands with only one ground-dwelling sinistral and 22 dextral species and thus would rarely encounter sinistral prey. Thus, it would be of little advantage to evolve an ability to distinguish between prey enantiomorphs. This explains the frequent failure of P. iwasakii to capture a given sinistral after striking. In contrast, P. carinatus lives with abundant sinistral snails, where avoiding predatory attempts on costly sinistrals should be advantageous.

The predator in this case does not evolve to exploit sinistrals by arms race. Instead the snake has shifted to avoid a cost of attempting unsuccessful or inefficient sinistral-predation because the easier prey type (dextrals) still remains abundant. This behavioral response by visual recognition reduces both a risk for the snake to expend foraging time and energy to handle unsuitable prey and a risk for sinistral snails to undergo physical attacks by the snake. Sinistrality therefore functions as a warning sign to the predator, instead of sheltering the prey. Predator’s recognition of prey handedness, which benefits both the snake predator and sinistral prey, could further accelarate ecological prey speciation39 by a reversal gene.

Many studies have shown the association of ecological performance with the direction of asymmetry in morphology and/or behavior4,5,11,20,40,41,42. We do not know, however, how important it is for a predator to be so asymmetric for chirally specialized predation. The previous studies ascribed the reduced efficiencies of sinistral-predation by P. iwasakii to its leftward-fixed strike with no prey-handedness recognition and to its most pronounced dental asymmetry in Pareas5. In contrast, the present snake P. carinaus does not fail in dextral or sinistral-predation either by leftward or rightward striking. Moreover, the mean dental asymmetry among four of the six snakes used in this study was almost minimal (4.5%) in the genus29. Nevertheless, efficiencies of dextral-predation were obviously superior to those of sinistral-predation. Accordingly, specialized handling of asymmetric prey does not necessarily require so manifest directional asymmetry in striking behavior or mandibular morphology as expected from the previous studies.

However, P. iwasakii fed on the dextral wild-type of Bradybaena similaris more efficiently than its sinistral mutants5, which were as small as the present threshold size (Fig. 5). Pareas carinatus in contrast feeds on dextral and sinistral prey of this size range with equivalent efficiency. Thus, the efficiency of feeding on dextral prey relative to that on sinistral prey may depend on the strength of dental asymmetry. If this is the case, weaker dental asymmetry may represent weaker specialization in dextral-predation. This would be advantangeous for Pareas snakes where frequently coming across sinistral snails, because snails of even large species can be small enough to prey when they are young. Strongly right-handed dentition on the other hand would benefit them for strong specialization in habitats with few sinistrals. Testing this hypothesis requires explicit comparison of predation performance among snail-eating snakes that differ in dental asymmetry. Our results therefore have important implications for further investigation of functional significance of predator’s asymmetry for chirally specialized predation.

A single gene is responsbile for the reversal of primary and secondary asymmetries in pulmonate snails8,18. The snail-eating snake P. carinatus notices this reversal by staring at a snail, though people often do not unless told so. Our study demonstrates that a chirally specialized predator can evolve an ability to recognize the left-right reversal of prey asymmetry where advantageous.