East African Journal of Agriculture and Biotechnology Effects of Temperature on the Development and Survival of Cricket Species; Acheta domesticus and Gryllus bimaculatus (Orthoptera: Gryllidae)

Temperature plays an important role in the development and survival of insects. The effects of temperature on the development of two cricket species, Acheta domesticus and Gryllus bimaculatus were determined at six constant temperatures (18, 22, 26, 30, 34 and 38 0 C). Parameters for stage-specific development such as fecundity, weight and fat content, structural body size, sex ratio, development time, and longevity were investigated. Relative humidity, light intensity, and photoperiod were set at 60-90 %, 500 ± 25 Lux, and 12:12 L:D, respectively. The results indicated that the duration of


INTRODUCTION
Climate variability has imposed large fitness costs on insects, showing diapause and other life cycle responses, threatening population persistence (Lopatina et al., 2014;Gillooly et al., 2002). Phenotypic plasticity, the capacity of a single genotype to exhibit variable phenotypes in different environments, is common in insects and is often highly adaptive (Tomberlin & Sheppard, 2002). Most studies have concluded that insects would become more abundant as temperatures increase through a number of interrelated processes, including range extensions and phenological changes (Reznik & Virghina, 2011). Insects are poikilotherms, significantly affected by climatic factors, with the temperature being the most prominent environmental factor with a marked influence on insect biology and behaviour (Aksit et al., 2007;Infante, 2000). However, individual species responses vary when exposed to stressful conditions (Mori et al., 2005;Miller & Paustial, 1992). Insects respond either through a change in behaviour to avoid stress by migration or through changed activity patterns (Hagstrum & Miliken, 1988). These insects can continuously adapt to stress conditions through selection or by plastic responses, by changes in morphology, life history, or physiology (Manrique et al., 2012;Lopatina et al., 2007).
Insect development parameters such as sex ratio, longevity, and fecundity are significantly affected by temperature (Infante, 2000). Since temperature affects the population size and variation of edible insects, under various situations, knowledge of the thermal requirements of crickets is crucial for conservation initiatives.
The insect order, orthoptera, forms part of the hemi metabolic insect groups, characterized by developing nymphal instars resembling the mature adult (Otte, 2007). Egg, nymph, and adult make up the lifecycle of crickets, which undergo incomplete metamorphosis (Chapman et al., 2013;Resh et al., 2009). Depending on their environment they have a lifecycle that lasts between two to three months and a life span of more than six weeks (Otte, 2007). Temperatures between 27 and 32 0 C are ideal for these crickets growth. To mate, the male crickets chirp their wings together. The female cricket has a long needle-like protrusion (ovipositor) used for laying eggs in addition to two cerci and can lay up to 200 eggs at a time in any available damp substrate (Huber, 1989).
Crickets play an important role in maintaining the balance of ecosystems (Umpold & Schulter, 2013). They break down plant material, renew soil minerals, and are an important source of protein for many households, reducing the pressure on fish resources that have been used to formulate poultry feeds (FAO, 2010). The adult cricket is composed of 47% crude protein, 10% carbon, and 25% fat; food nutrients on a dry weight basis (Ayieko et al., 2016). In addition, the insect contains a variety of minerals and vitamins. When the diet is enriched with fish offal, the adults are rich in omega-3 and essential unsaturated fatty acids (Umpold & Schluter, 2013). The estimated value of adults as feed when dried is similar to that of soybean or meat and bone meal. If they are used live as a special form of feed, their worth as a product can be higher (FAO, 2010).
A cricket begins its existence as an egg, then cracks the egg capsule and burrows its way out of the substrate. It will have developed into a nymph after roughly 14 days. Nymphs resemble adult crickets with a few exceptions: they lack wings at first and females lack ovipositors (Chapman et al., 2013). These juvenile crickets are frequently eaten by larger crickets and other insects (Chapman et al., 2013;Resh et al., 2009). To grow, a nymph loose its hard exoskeleton and replace it with a new one that is soft and milky white at first but hardens within hours. Moulting occurs every eight to ten times. This process is called moulting and happens eight to ten times (Resh et al., 2009). After roughly a month, a nymph will begin to grow wings. When a cricket achieves maturity, its wings are fully formed, and it has just two goals: eating and mating (Hardy et al., 1983). A male will endeavour to attract fertile females. After mating has occurred, a female will spend her time looking for good spots to lay her eggs (Resh et al., 2009).
In spite of the economic importance of edible crickets little is known about the effects of temperature on their development and survival. This study was conducted to determine the effects of temperature on the development and survival of two cricket species, Acheta domesticus and Gryllus bimaculatus under laboratory conditions in order to create heat accumulation-based forecasting models for the edible insect. In addition, knowing the best temperature for the primary phonological traits of crickets (nymphal weight, growth rate, development, fecundity, and longevity) would aid in the effective conservation of this edible insect and further used to predict the potential range of the crickets.

Colony Establishment
The insect colony was established at the Insect Farm of Jaramogi Oginga Odinga University of Science and Technology (JOOUST). The collected insects were fed on poultry growers mash and reared in the insect-rearing unit at JOOUST. Wild adults (2000 individuals, assessed by weight) were gathered in Western Kenya from the designated habitats. The crickets were raised in 60 L plastic buckets, each containing approximately 100 crickets. The buckets were covered with mosquito nettting to keep predators out and crickets in (Mellisa, 2014;Clifford & Woodring, 1990). Drinking water was offered ad libitum in a 16 cm diameter saucer using a moistened cotton wool. To serve as hideouts, egg trays measuring 29 cm x 29.5 cm were placed vertically in the buckets (Mellisa, 2014;Wineriter & Walker, 1988).

Experimental Design
Newly laid cricket eggs were collected randomly from the laboratory colonies to create six sets of crickets colonies for each species; Acheta domesticus and Gryllus bimaculatus and reared in an incubator under constant conditions. There were six treatments of temperature regimes on two cricket species. For this experiment, relative humidity, light intensity, and photoperiod were set at 60-90 %, 500 ± 25 Lux, and 12:12, respectively (Das et al., 2012).
The treatments were as follows:

Fecundity
Adults of Acheta domesticus and Gryllus bimaculatus were selected and coupled in well ventilated transparent containers (20 cm x 20 cm x 15 cm) and treated to varying temperature regimes (Calvo & Molina, 2005). A total of ten pairings were chosen and an oviposition substrate consisting of a moistened cotton ball was placed within a sterile petri dish in each container for egg laying. The number of eggs in each vial was counted under a stereomicroscope using a fine camel's hair brush (Otieno et al., 2019).

Weight and Fat Content
To remove any contaminants, the crickets were cleaned. Fresh body weight was measured at birth and once a week until death with an electronic scale (OHAUS Pioneer TM) set to the nearest 0.1 mg (Zebino et al., 2016). Adult males, virgin females and mated females were freeze killed for dry weights, and the specimens were maintained thermo-resistant glass vials and microwaved at 45 0 C for 72 hours (Mellisa, 2014). The dry weights were calculated by using the dried specimens. Five grams of each specimen were put into Eppednorf ® tube (1.5 mL) in the soxhlet extractor for fat extraction. This was immersed in a 1:1 (vol:vol) petroleum ether solution and heated for 6 hours. The condensing unit was removed from the extraction unit and the sample was allowed to cool down. The samples were then dried once more for 48 hours and weighed after that (lean dry weight) (Wigglesworth, 1972;Mellisa, 2014). To a precision of 10 -5 g, every weight was measured using analytical Sartorius ® balance (Infante, 2000). Fat content was calculated using the equation: Crude fat = weight of tube with sample − weight of tube weight of specimen

Structural Body Size and Sex Ratio
The length of the elytron, which is the distance between the apex and base, the width of the protonum at its widest point, and the hind leg's femur were used to estimate the structural body size. A digital calliper was used to measure each proportion with an accuracy of 0.01mm (Honnek, 1993). The sex ratio of adults was recorded.

Development Time and Longevity
The time to complete each life stage (egg, nymph, and adult) of the crickets was determined. The newly enclosed nymphs were marked with a finetipped brush with a small dot of ink in the protonum using non-toxic permanent ink pens (EDDING 751 band) due to its excellent adhesion, quick drying, and good visibility to estimate cricket longevity (Das et al., 2012). Different colours were used for each day of emergence (Infante, 2000;Bowling, 1955). Marked nymphs were returned to their respective buckets, daily observations made, and the waste removed and observed under a stereomicroscope to quantify the number of marked nymphs that had died (Bowling, 1955).

Data Analysis
Differences in fecundity, weight and fat content, structural body size and sex ratio, development time, and longevity among temperature treatments were tested using analysis of variance (ANOVA). The significance of pair-wise correlations amongst the measured parameters was tested using Pearson's correlation coefficient (r). The significance of the correlation of the phenotypic factors in relation to changes in temperature was assessed using Spearman's (ρ) correlation coefficient (Pinheiro et al., 2018). All the analyses were performed in the R environment (R -Core Team 2017).

Fecundity
The effects of six constant temperatures on the fecundity of two cricket species (Acheta domesticus and Gryllus bimaculatus) differed significantly (F5, 205 = 272; p < 0.001) (  (Table 1). At all temperatures, males lived longer than females, but no discernible differences were recorded between the sexes.

Sex Ratio
There was a significant difference (p-value ≤ 0.05) in the analysis of the sex ratio between the two cricket species (

Adult Weight
The effects of temperature on adult weight of the crickets differed significantly (F5, 54 = 2.1; p < 0.001). The highest body weight (22.04 g and 20.47 g) for Acheta domesticus was recorded at 26˚C for both females and males, respectively ( Table 2). In Gryllus bimaculatus, the highest adult weight of 23.42 g and 21.61 g was recorded at 30 o C for females and males, respectively.

Fat Content
The temperature treatment resulted in significantly different fat contents at the significance level of p<0.05) between the two cricket species (Table 2). Acheta domesticus reared at 22 o C recorded the highest fat content (19.43 and 17.71 g /100g dry weight) for females and males, respectively. Gryllus bimaculatus recorded the highest fat content (11.78 and 9.51 g/100g dry weight) at 30 0 C for females and males, respectively. The lowest fat content was recorded in crickets reared at 38 o C for both species. Significant (p < 0.05) temperature x species interaction effects on fat content were identified, albeit the magnitude of the interaction was relatively small, indicating that temperature treatment influenced fat content more than species.

Effect of Temperature On Structural Body Length
Both temperature and cricket species had significant effects on structural body length (pvalue = 0.0008 and p-value <.0001, respectively). Acheta domesticus reared at 26 o C recorded higher lengths: body lengths of 18.3 mm and 18.4 mm for males and females, respectively. Length of tegmina; 9.2 mm and 13.4 mm; Length of the femur of hind leg: 11.0 mm and 12.0 mm for males and females, respectively. Gryllus bimaculatus had a higher (24.1 mm and 24.8 mm) body length at 30 o C for males and females, respectively. The lowest lengths were recorded in crickets reared at 18 o C and 38 o C (Table 3). The interaction effect of temperature and species on structural body length was significant. Crickets with the highest body length (24.8 mm) were obtained from Gryllus bimaculatus reared at 30 o C with the shortest (16.0mm) lengths recorded in Acheta domesticus reared at 18 o C ( Table 3).

Effect of Temperature on Development of the Different Growth Stages of Crickets
The developmental times for each stage of the two cricket species, Acheta domesticus and Gryllus bimaculatus at six constant temperatures are presented in Table 4. There were significant differences (p-value ≤ 0.05) in the analysis of development amongst the temperature treatments. The average developmental time for each stage was significantly shortened as the temperature increased. The highest number of moults (10  moults

Correlations Among the Variables
The correlations between different characters of cricket development and survival are shown in Table 5. Both positive and negative association amongst the variables were identified. Development time was positively correlated with longevity at 30 o C (r=0.4546). Adult weight and fat content are positively and closely correlated (r = 0.8692, p-value = 0.004) while fecundity and adult longevity were negatively correlated (r = -0.0953). Increased female longevity has been recorded as a result of reduced egg production in Drosophila species (Colin & Spurgeon, 2019). The decrease in female longevity may be due to higher energy diverted towards the reproductive machinery. Significant positive correlations between fecundity and adult weight (r = 0.8424, p-value < 0.0001) were recorded (Table 5). Body size and fecundity are a function of genetics and the environment. Large females have higher fecundity; therefore, selection should favour increased body size (Honek, 1993). A significant negative correlation was observed between development times (r = 0.7316, p-value = 0.003) and adult sizes, suggesting that an increase in development rate resulted in reduced body size and weight. Correlations between development times and adult weights shows that there is a lot of potential for using them to assess the calibre of insects reared (Lande & Arnold, 1983). This is because insects with lower development rates have high adult weights and sizes and accumulate more fat content. A decline in size was followed by an increase in inhibition of reproductive maturation, as reflected by the decline in fecundity. The increased inhibition of the reproductive cells has been found to be followed by a decrease in size and weight in several insects. Reduced reproductive development associated with a decline in fecundity under low temperatures is caused by poorly developed ovaries.
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Regression Model
Employing a regression model where the dependent variables are development and survival and all the other factors are treated as independent, relationships between cricket development and survival and the other variables were further investigated. When using backward selection, nonsignificant variables were gradually eliminated. At a 0.05 percent significance level, any parameter still included in the model is significant. The model had a high level of significance and was able to explain 91.3 percent of the data's variability, according to the results shown in Table 6, which summarizes the model. According to the model's output, fecundity, adult lifespan, and body weight were the important factors that accounted for the variation in cricket development and survival. This suggests that there is a lot of promise for utilizing them to monitor the growth of crickets. All parameters left in the model are significant at a 0.05 significance level.

DISCUSSION
According to Colin and Spurgeon (2019), extreme heat could result in either temporary or permanent infertility or the inactivation of sperm stored in the spermatheca, reducing fertility. High temperatures are known to frequently hasten pre-imaginal development in insects that overwinter as adults, ensuring the timely development of the diapausing stage before the start of winter (Manrique et al., 2012). Nonetheless, the maturation of different species is restricted naturally and consequently, miniature and diaphanous adults are usually produced with accelerated development (Lamb et al., 2009). On the other hand, successful overwintering depends on sufficient fat and glycogen reserves, which are often positively correlated with body weight (Garcia- Barros, 2000). Several researchers have recognized the importance of larger weight in enhancing fat content in insects.
High-temperature acceleration of pre-imaginal development combined with its inhibition of reproductive maturation is recorded for several species. Our investigation showed that cricket females stopped laying eggs at 18°C, indicating that low temperatures also caused sterility in these species (Calvo & Molina, 2005). Both extremes of temperatures resulted in moribund ovaries leading to very low or no egg production. In addition, large females have a greater potential for fecundity and some other selective advantages (Padmavathi et al., 2008;Aksit et al., 2007). Thus, an insect faces two seemingly opposite challenges: increase the adult weight or speed up pre-adult development. Fast development results in small adults, as in the cotton bollworm, Helicoverpa armigera (Hubner) and some other species of insects.
An essential aspect that has a significant impact on how insects develop is the temperature (Neven, 2000). Crickets are not an exception. The development of insects due to fluctuating temperatures differs among species (Padmavathi et al., 2008). A decrease in the speed of development with reduced temperatures is common, with a marked increase in the period taken in every stage (Ikemoto & Takai, 2000). The findings of this study show that when the temperature rose, the length of time that various stages of crickets took to develop 187 | This work is licensed under a Creative Commons Attribution 4.0 International License.
decreased. However, most eggs did not hatch when the temperature was 18 0 C, and neither the eggs nor the nymphs matured when the temperature was 38 0 C. The outcomes from this study were consistent with Colin & Spurgeon's (2019) findings that insects could not finish their normal development at 18 or 38 degrees Celsius. Thus, both low and high temperatures were harmful to the growth of crickets. Under laboratory conditions, a temperature range of 26 to 34 0 C proved acceptable for the development of crickets. The growth rate of an insect and temperature show a positive correlation when the temperature range is appropriate, becoming sigmoid over the complete temperature ranges through which insects are capable of developing.

CONCLUSION AND RECOMMENDATIONS
The results provide important information regarding the thermal requirements of Acheta domesticus and Gryllus bimaculatus. The development of Acheta domesticus from egg to adult would occur between the thermal range 22 o C and 30 o C, while that of Gryllus bimaculatus would occur between the thermal range 26 o C, and 34 o C. The optimal growth rate was observed at 26 o C for Acheta domesticus and 30 o C for Gryllus bimaculatus. This study therefore shows that cricket development, survival, and distribution could be affected by future temperature increases.
This data can be used to model development in the wild and estimate potential distribution limits. In addition, parameters such as adult weight, and fecundity, under different temperatures could be used to optimize production under mass rearing.
However, nature does not have a steady temperature; it can change by roughly 10 o C in a day. Therefore, further investigation is needed to determine whether the severe temperatures in this study location prevent some cricket species from spreading in the wild.

ACKNOWLEDGEMENT
The staff at Egerton University and Jaramogi Oginga Odinga University of Science and Technology patiently took their time to assist the researchers throughout the study. The researchers also grateful to Insects for Food and Feeds (INSEFOODS), financed by the World Bank for sponsoring this research.