Cannibalism and intraguild predation involved in the intra- and inter-specic interactions of the invasive fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae) and lepidopteran maize stemborers

Intra- and interspecic competitions play important role in determing spread and survival/death (cannibalism or intraguild predation) of organisms which share the same resource. However, the trends of relation between cannibalism and intraguild predation, and the costs and benets of such behaviours are dicult to establish within insect communities and a little is known about how such behaviours are affected by invasive species. The present study was aimed at assessing the interactions between larvae of fall armyworm (Spodoptera frugiperda) and maize stemborers (Busseola fusca, Sesamia calamistis and Chilo partellus) in relation to cannibalism and intraguild predation when they are subjected to utilize the same resource. Experiments involving treatments with either single species of S. frugiperda or any of the stemborers or pairwise species combinations were conducted under laboratory conditions. The experimental insect larvae were reared on maize leaves and monitored until the last developmental stage where larval survival, mortality, and cannibalism and/or intraguild predation were recorded. Results of the intra-specic interaction indicated that S. frugiperda exhibited cannibalism to a larger degree than the stemborers species, especially at the late instars stages. The higher cannibalism trait in S. frugiperda turned however to competitive advantage as it led to a higher degree of intraguild predation when they cohabit with other stemborer species. Overall, interaction with S. frugiperda is thus detrimental for the stemborer species and may be an important factor to explain the invasive success of the pest. Such knowledge is essential in designing successful integrated pest management strategies.


Introduction
Resource availability varies spatially and temporally either within or across developmental stages and competitive exclusion should result when resources are limited. Such exclusion depends on the biological characteristics of the interacting organisms, where some species may have better competitive abilities.
Intra-and interspeci c competitions play an important role in determing survival/death and spread of organisms which share the same niche (Cameron et al. 2007; Gurevitch et al. 1992). The interaction between organisms can be expressed as either exploitative (as use of the resource by one species reduces the availability for the other) or interference competition (when one competing species behaviourally restricts the other species from access limited resources supplies). In the most extreme form of interference competition, the interaction leads to the death of one of the involved organisms and could be de ned as a form of either cannibalism or intraguild predation.
Cannibalism, also called intraspeci c predation, is known as killing and consumption of a conspeci cs (Richardson et al. 2010) and is a widespread behaviour in phytophagous insects, mostly in lepidopteran species (Richerdson et al. 2010). It is often favored under high population densities (Sokame et al. 2022) and when resources are limited (Elgar & Crespi 1992) but can occur even when food is not limited (Polis 1981). Intraguild predation (IGP) occurs between different species within the same trophic level (Bentivenha et al. 2016a). Polis & Holt (1992) reported that although combining elements of predation and competition, intraguild predation is distinct from competition because one participant (the predator) accrues immediate energetic gains. It is also distinct from classical predation, because the act reduces potential competition besides the actual energetic gains. Intraguild predation can act as mechanisms of population regulation or as determinants of community structure in phytophagous insect communities (Holt & Polis 1997;Bentivenha et al. 2016a;. Irrespective of whether it is cannibalism or intraguild predation, these behaviours are costly in terms of energy, time, risk of injury and death (Briffa & Elwood 2004;Kelly & Godin 2001) for competing individuals that may have repercussions for an individual's tness. The balance between cannibalism and intraguild predation depends on involved species. For instance, it was reported that intraguild predation is less frenquent than cannibalism for coccinellid species Ciccinella spetempunctata L. and Hippodamia convergens Guerin-Meneville as compared to the predatory bug Geocoris bullatus Say when cannibalism versus intraguild predation was compared among the three species (Takizawa & Snyder 2011). Furthermore, Rasekh & Osawa (2020) reported that two coccinellid species Harmonia axyridis and H. yedoensis were exposed to either cannibalism or IGP, and H. axyridis had longer development time while H. yedoensis presented shorter development and lighter adult weight. Therefore, the trends of relation between cannibalism and intraguild predation, and the costs and bene ts of such behaviours is di cult to establish. Moreover, there is still little known about how such behaviours are affected by invasive species and how such that could affect crop pests that share the same resources.  Cruz and Turpin 1982) leading them to share the same ecological niche with possible intraguild competition. A previous study reported competition among these four maize pests when they utilize the same resource in a restricted space (Sokame et al. 2020) and the decrease of stemborer density at maize reproductive stage in maize elds following the invasion of FAW in the system (Sokame et al. 2021a) indicates competitive exclusion by FAW since its introduction in Africa. However, the potential of cannibalism and intraguild predation within the community is not well known.
Cannibalism and intraguild predation could contribute to FAW rapid establishment and for interactions in these maize lepidopteran pest communities. In the native range of FAW, cannibalism has been reported (Chapman et al. 1999b;Andow et al. 2015) as well as intraguild predation between FAW and Helicoverpa zea (Bentivenha et al. 2016a). Cannibalism has also been reported in C. partellus, but only at high larval densities (Bonhof & Overholt 2001). Nevertheless, there is still much to be clari ed about the behaviour of the other stemborer species, and interspeci c interactions between FAW and these stemborers when they co-occur and the fast and successful establishment of FAW in Africa. A better understanding of the intraand interspeci c interactions will be useful to understand how competitive displacement occurs between those pest species sharing a given ecological niche, and consequently might help to enhance designing successful integrated pest management strategies (Benelli 2015). In this context, the present study aimed at assessing the potential of cannibalism and intraguild predation of the four lepidopteran maize pests species when they are subjected to utilize the same resource and if this can lead to competitive exclusion.

Insects
The stemborer and FAW larvae were provided by the Animal Rearing and Containment Unit (ARCU) at icipe, Nairobi, Kenya. Larvae were reared on arti cial diet in cylindrical plastic jars (16.5 cm high and 9 cm in diameter) with about 200 ml of diet per jar. After inoculation, the plastic jars were tightly sealed using perforated lids with galvanized mesh. They were covered with tissue paper and kept in a rearing room at 26 ± 1°C, 60 ± 5% RH., and L12 and D12 photoperiod. Twice a year, each colony was rejuvenated with eld-collected stemborer or FAW larvae. All insect laboratory rearing was carried out at 25 ± 2 o C; 50-70% RH and a photoperiod of 12:12 (L:D) h.

Experimental Set Up
Experiment 1: Cannibalism and intraguild predation involved in fall armyworm and stemborer species larval age-grouped pairing communities To understand the potential of cannibalism and intraguild predation, an experiment was settled involving treatments with either single species of FAW or any of the stemborers (B. fusca: Bf, S. calamistis: Sc and C. partellus: Cp) or pairwise species combinations between FAW and each of the stemborers. The experiments were conducted in plastic containers (16 cm × 10 cm × 7cm) lined with a paper towel to absorb excess moisture containing 6 pieces of freshly cut maize leaves (10 cm × 6 cm). The larval instars were structured in three age groups: neonate to second as small size, third to fourth as medium size and fth and sixth instar as large size. The single-species infestation treatments consisted of 40, 20 and 10 larvae for small, medium, and large size groups, respectively. Those densities were chosen as these densities are usually encountered in the eld (Sokame BM. Pers. Obs.). Similar to the method of Bonhof & Overholt (2001) to assess cannibalism between stemborer species, the pairing-species combination involved 20, 10 and 5 larvae per container of each species for FAW+Bf, FAW+Sc and FAW+Cp pairings for each small, medium and large size group, respectively ( Table 1). The survival, relative growth rate, mortality (dead larvae found in full corpse), and cannibalism coupled with intra-guild predation (dead larvae found in partial corpse or disappeared corpse) were recorded four (4) days after the settlement (corresponding to the mean time from one instar to another until L5 for FAW) and after ve (5) days from L5 to L6, the last instar for pupation. To check if cannibalism or intraguild predation continues in all instars and whether individuals that have fed on conspeci cs gain some advantages, a second experiment was conducted. It involved single species infestation of either fall armyworm (FAW) or any of the stemborers (B. fusca: Bf, S. calamistis: Sc and C. partellus: Cp) larvae and pairwise multi-species infestation of FAW and stemborers was settled with larval neonate instar. The single-species infestation treatments consisted of 20 neonates while the pairing-species combination involved 10 neonates of each species per container. The same parameters as described in experiment 1 above were evaluated from neonate to the last instar larvae for pupation with four (4) days interval.
For both the experiments after the infestation, plastic containers were closed with perforated plastic lids until the experiment terminated. The insects in the container were kept in a rearing room at a temperature of 25 ± 0.05°C, RH of 58.5 ± 0.4%, and a photoperiod of L12:D12. Each treatment was replicated twenty times. Like the method of Bonhof & Overholt (2001), after 4 days, containers were opened to record the number of surviving larvae (Ns) and the number of dead larvae (Nd) of each species. The cannibalism (C) in single-species communities and both cannibalism and intraguild predation (C&IGP) in multi-species communities were calculated as follow: Where N denoted the initial number of larvae inoculated in each container.
The relative growth rate (RGR) for each species was calculated using the following equation (

RGR = Masspersurvivinglarva − Initialmassperlarva Numberofdaysafterpairingspecies
Data Analyses The number of surviving larvae, the number of dead larvae and the number of larvae that have been consumed (cannibalism and intraguild predation) from either experiment 1 or experiment 2, were analysed using a generalized linear model (GLM) with negative binomial procedure. Signi cant differences were separated using Tukey's multiple comparison test performed using the R package "lsmeans" (Lenth 2016). The relative growth rate (RGR) of the larvae from the two experiments were analysed using Kruskal-Wallis nonparametric procedure with dunn.test R package after being tested for normality and homogeneity of variance using Shapiro-Wilk and Bartlett tests, respectively. All analyses were performed with R software version 3.5.1 (R Core Team 2018).

Results
Survival, mortality, and cannibalism of fall armyworm and stemborers in single-species combination At small larval size, FAW had the lowest survival rate while B. fusca had the highest (LR = 26.38, df = 3; P ≤ 0.0001). At medium larval size group, no signi cant difference was recorded between the four species (LR = 3.01, df = 3; P = 0.39) while at large larval size group, the survival rates of the three stemborer species were signi cantly higher than those of FAW (LR = 31.66, df = 3; P ≤ 0.0001) (Fig. 1A). Between larval size groups for a given species (Fig. 1A), the survival rate of FAW was signi cantly lower at large size group as compared to the smaller size groups (χ 2 = 22.30; df = 2; P ≤ 0.0001) in opposition to C.
In terms of mortality (Fig. 1B), there was no signi cant difference between the four species at small larval size group (χ 2 = 7.57; df = 3; P = 0.06). However, at medium size group, B. fusca presented a signi cantly higher mortality rate than FAW (χ 2 = 22.23; df = 3; P ≤ 0.0001), while at large size group, C. partellus mortality was signi cantly lower as compared to FAW and B. fusca (χ 2 = 25.37; df = 3; P ≤ 0.0001). Between larval size groups for a given species, large size of FAW presented a signi cant higher mortality as compared to small and medium size groups (χ 2 = 16.62; df = 2; P ≤ 0.0001). The mortality rates of B.
Survival, mortality, and cannibalism coupled with intraguild predation in pairing-species combination between fall armyworm and stemborers For all combinations, FAW survival rate was signi cantly higher than that of its corresponding competing species at all larval size groups: FAW+Bf ( Fig. 2A, Table 2), FAW+Sc (Fig. 2B, Table 2) and FAW+Cp (Fig. 2C, Table 2). There was no signi cant difference for mortality within species in all combinations at small larval size group: FAW+Bf (Fig. 2D, Table 2), FAW+Sc (Fig. 2E, Table 2) and FAW+Cp (Fig. 2E, Table   2). However, the mortality of FAW was signi cantly higher in the pairing with B. fusca at medium and large larval size groups (Fig. 2D, Table 2), with S. calamistis at medium and large larval size groups (Fig. 2E, Table 2) and with C. partellus at medium and large larval size groups (Fig. 2F, Table 2). The cannibalism coupled with intraguild predation of stemborer species were signi cantly higher than that of FAW in all combinations at all larval size groups: FAW+Bf (Fig. 2G, Table 2), FAW+Sc (Fig. 2H, Table 2) and FAW+Cp (Fig. 2I, Table 2). For this comparison purpose, the survival, mortality, and cannibalism and/or intraguild predation rates were calculated for the overall larvae regardless of the species present in each test for pairing-species combinations. All stemborer single-species combinations exhibited a signi cant higher survival rate than their respective pairing-species combinations with FAW at all larval size groups (Fig. 3, small: LR = 202.62; df = 6; P ≤ 0.0001, medium: LR = 146.56; df = 6; P ≤ 0.0001 and large: LR = 127.78; df = 6; P ≤ 0.0001).
At small size group, stemborer single-species communities presented a higher mortality than their respective pairing-species combination with FAW (LR = 24.86; df = 6; P = 0.0003). At medium size group, while FAW had lower mortality, B. fusca single-species had signi cantly higher mortality as compared to its pairing-species combination counterpart mortality (LR = 28.69; df = 6; P ≤ 0.0001). At large size group, S. calamistis and C. partellus single-species combinations exhibited signi cantly lower mortalities as compared to the mortalities of their pairing-species combination counterparts (LR = 35.14; df = 6; P ≤ 0.0001).
Life-time survival, relative growth rate, mortality, and cannibalism of fall armyworm and stemborers across larval instars in single-species combination Across larval instars, the three stemborers species presented similar survival evolution while signi cantly higher than those of FAW species from third to sixth instar (Fig. 4A, χ 2 = 308.17; df = 5; P = 0.0001). The FAW presented the greater relative growth rate as compared to those of stemborers at third and fourth instars with an optimum of 15mg/day (Fig. 4B).
The In pairing-species combination between fall armyworm and stemborers, none of stemborer species survived at the last instar to pupate (Fig. 5A1-A3). The survival rate of either FAW or stemborers signi cantly decreased along larval development instars (χ 2 = 42.52; df = 5; P =0.001) and the survival of FAW was signi cantly higher than those of stemborers in each combination (χ 2 = 57.13; df = 3; P =0.004). B. fusca and S. calamistis reached 0% at L4 instar and C. partellus at L5 instar while about 10% of FAW survived at last instar to pupate. The growth rate of stemborer species did not reach the optimum before they disappeared in the combinations (Fig. 5B1-B3). The optimum of FAW growth rate ranged from 40 to 50mg/day across the different combinations and it has mostly decreased after stemborer species were no longer present in the combinations (Fig. 5B1-B3). In each pairing-species combination, the mortality rate of stemborers was signi cantly (χ 2 = 32.34; df = 3; P =0.001) higher than those of FAW along larval development instars before the optimum and thereafter, the mortality rate of FAW come over those of stemborer species (Fig. 5C1-C3). on the other hand, the cannibalism coupled with intra-guild predation of stemborers in each combination was signi cantly higher than those of FAW along larval development instars (χ 2 = 54.21; df = 3; P ≤0.0001). However, when they nished in the system, the cannibalism of FAW presented exponential trend (Fig. 5D1-D3).

Discussion
The Invasive species are generally less frequently infected in introduced environment as compared to native conspeci c populations (Torchin & Mitchell 2004). Successful introduced species often invade without their native parasites and accumulate relatively few new ones in the invaded areas. This suggest that costs-and bene ts of cannibalism differ in native and invaded area depending on whether they experience less or more parasites and pathogens in these areas. In the other hand, it has been reported that FAW cannibalism rates were around 18%, when con ned to maize seedlings and can increase up to 34% in case of limited food and even reach 100% when the larvae are con ned in Petri dishes (Bentivenha et al. 2016;Raffa 1987). The suitability of food in terms of quality on which the insect larvae were reared is also a source of cannibalism increase. Therefore, it would be interesting to study how costbene ts of cannibalism/intraguild predation could differ depending on host plant species.
In interspeci c interactions, the higher cannibalism in FAW turns however to competitive advantage when they cohabit with stemborer species. Cannibalism/intraguild predation is higher on the stemborers, and survival is always higher for FAW. The FAW larvae might possess defence mechanism in detriment of other species. Bentivenha et al. (2017a) studied intraspeci c interactions of FAW larvae and reported that they exhibited either high attack (head touching) or defense (recoiling) as the prevalent movement and that the frequency of these movements increased with larval development because of their increased interaction with other larvae when they become bigger and more mobile. These results suggest that there is a great chance that FAW can prevail over these other species and colonize the niche (Bentivenha et al. 2016b; Bentivenha et al. 2017b), when these species occur in maize elds (on plants). Intraguild studies of FAW and H. zea showed further advantage of FAW in interaction against H. zea in morphological characteristic such as integument or mandible architecture (Bentivenha et al. 2016a), which also could explain the success of FAW during intraguild competition. The intrinsic cannibal character of FAW might prepare the species to a high cost of ght within themselves (actions of defence and attack) in the situation of conspeci c cannibalism which explain the lower relative growth rate. In interspeci c competition, the actions of the co-existing stemborers which probably go into defense behaviour rather attack, in ict lower ght cost to FAW by the lack of attack actions. Therefore, FAW take the advantage by reducing the cannibalism on their conspeci c and concentrate on intraguild predation on their heterospeci c species where it is more easy to feed on stemborer larvae with less energy invested, explaining the great relative growth rate gained when reared together with stemborer species than when reared lonely as single species. Arnott & Elwood (2009) have studied those variations of the weight by assessing ghting ability in animal contests and reported that the gain of the weight of the superior from the competition is positively correlated to the contest duration and the cost or intensity of the ght. The competition between FAW and stemborers may be short duration with low-cost than between FAW conspeci c species.
The optimum of FAW growth rate ranged from 40 to 50mg/day in cannibalism coupled with intraguild predation with stemborer species while when reared lonely as single species, the growth rate reached an optimal of 30mg/day only. and not in competition with other species was signi cantly greater than the weight of those that surviving the competition with other heterospeci c species. However, in the present study, the competition was asymmetric that favour FAW because, while FAW exhibited simultaneously defense and attack behaviour, stemborer species might exhibit only defense behaviour.
The higher intraguild predation of FAW on stemborer species when they are forced to co-habitat indicates a potential competitive advantage of FAW and could be a factor explaining its success as an invasive pest species. However, this analysis needs to be done at large scale as a complex system because other parameters such as agricultural landscape (Bentivenha et al. 2016a), the food quality of the host plant and larval behaviour (Dial & Adler 1990) may in uence also intraguild interactions. Intraguild predation may be an important factor during invasion to decrease competition from other species, but in the present study, it does not seem to lead to a higher absolute survival rate of FAW, only a relatively better survival in comparison to the other species. This can be due to that FAW reverses cannibalism behaviour in detriment of intraguild predation when the heterospeci c species disappears/is eradicated to the recent evaluations in maize and sorghum cropping systems in Uganda showed a signi cant decline of the infestations in maize elds in detriment to sorghum elds from 2016, suggestion an early arrival of fall armyworm in Uganda, indicating a possible displacement of stemborer from maize onto sorghum (Hailu et al. 2021). Furthermore, through intense intraguild predation, fall armyworm can reduce the population density and impact of stemborer guild members in maize elds, as the larvae preferred to feed on stemborers rather than their conspeci c in interspeci c communities. However, if the guild of fall armyworm is to contribute to the suppression of stemborer species population in the context of integrated pest management (IPM) programme, then it necessary that its impact on those stemborers is reliably assessed at an appropriate ecological scale so that the impact on the population dynamics of the pest can be quanti ed and integrated into decision-making processes for management.
Overall, interaction among FAW and stemborer species in the case of this study affect their survival and

Declarations
Authors contributions BMS, P-AC, PA, and GKKdesigned the studies. BMS, and BM collected the data. BMS analysed the data and wrote the manuscript. All authors reviewed and approved the manuscript before submission.    Survival (A), mortality (B) and cannibalism coupled with intraguild predation rates (C) of all larvae regardless of the species between single-species and pairing-species combinations at different larval stages: small size (neonate to second instar), medium size (third to fourth instar) and large size ( fth to sixth instar). Statistical comparisons were only considered between single-and corresponding multispecies pairings. Means (±SE) were compared using Tukey's comparisons tests performed using lsmeans R package, following generalized linear model (GLM) with negative binomial procedure. Spodoptera frugiperda (FAW), Busseola fusca (Bf), Sesamia calamistis (Sc), and Chilo partellus (Cp).