Effects of different temperatures on the demography of pistachio green stink-bug, Brachynema germari Kolenati (Hemiptera:Pentatomidae), under laboratory conditions

: e pistachio green stink-bug, Brachynema germari is one of the most important pests of pistachio in Iran that annually causes signiﬁcant damages to this strategic crop. In this research, the eﬀects of diﬀerent temperatures on the demography of B. germari were investigated under laboratory conditions. e experiments were conducted at ﬁve constant temperatures [20, 25, 27.5, 30 and 35 (±1) °C], on Ohadi cultivar. e lowest and the highest mortality rates in both the eggs and nymph stages were observed at 25 °C and 35 °C, respectively. e longest and shortest immature development times were observed at 20 °C (53.75 days) and 35 °C (18.42 days), respectively. e population growth parameters were signiﬁcantly aﬀected by the experimental temperatures. e gross reproductive rate ( GRR ) was severely reduced at 35 °C and the net reproductive rate ( R 0 ) was also decreased by increasing temperature. e intrinsic rate of increase ( r m ) was 0.05, 0.06, 0.06, 0.07 and 0.09 (day -1 ) at 20, 25, 27.5, 30 and 35 °C, respectively. Moreover, the lowest ﬁnite rate of increase ( λ ) (1.05 day -1 ) was observed at 20 °C, whereas the longest and shortest mean generation time ( T ) and doubling time ( DT ) occurred at 20 °C and 35 °C, respectively. Finally, the highest and lowest reproductive parameters values, including daily and total fecundity and fertility, were obtained at 20 °C and 35 °C. e results of this research could be useful in monitoring and eﬃcient control of this pest.


INTRODUCTION
Pistachio (Pistacia vera L., Sapindales: Anacardiaceae) is a nut crop that ranks first among different exportable agricultural crops in Iran (Sheibani et al., 1996).e annual exportation of pistachio in this country is about 100,000 tons.e country's associated income ranks second aer the oil revenue (Zare-Nazari, 2018).In pistachio orchards, there are many pests that feed on trees and cause serious damages (Hashemi-Rad, 1999).Among these, pentatomid bugs must be controlled in order to get a good crop (Hashemi-Rad, 1999).e pistachio green stink-bug Brachynema germari Kolenati (Hemiptera: Pentatomidae) is one of the most important pests of pistachio in Iran that was collected for the first time in summer of 1965 on orchards from Kerman province (Nyman et al., 1967).Both adults and nymphs of this bug feed on the fruits and cause significant direct damage in the form of epicarp lesions that can result on nut drop (Daane et al., 2005).
Also, this pest is a vector of the pathogenic yeast Nematospora coryli Peglion (Saccharomycetales: Saccharomycetaceae) that concern the cork and turns the fruit somewhat bitter; thus, its quality, quantity and marketability decrease seriously (Ershad & Barkhordari, 1975).
Temperature is one of the most important abiotic factors that affects the life cycle of insects (Roy et al., 2003).Estimation of demographic parameters of an insect under different temperatures can reveal the relationship between it and growth potential of pests (Greenberg et al., 2005;Dong et al., 2007).Among the population growth parameters, the intrinsic rate of natural increase (r m ) is a useful indicator of the effects of temperature on the population growth, and can indicate the overall effects of this factor on insect's development, survival and reproductive characteristics (Southwood, 1978).e main purpose of this study was to investigate the effect of five different temperatures on the biological characteristics, life table parameters and reproductive features of B. germari, under laboratory conditions.

Insect rearing
Adults of B. germari were collected from the pistachio orchards in Rafsanjan region located in southeastern Iran.e collected individuals were immediately transferred to the Pistachio Research Center and reared in a growth chamber set at 27.5 ± 1 °C, 65 ± 5% relative humidity (RH) and a photoperiod of 16:8 (L:D) h on Ohadi pistachio cultivar.e plant was replaced every two days.Nymphs were fed until the emergence of adults, and the emerged ones were used for egg-laying.One-day eggs were used for the experiments.

Demographic parameters study
e demographic parameters of B. germari were studied under 20, 25, 27.5, 30 and 35 ±1 °C in a growth chamber [65 ± 5% RH; 16:8 h (L:D)] on Ohadi pistachio cultivar.To determine the egg incubation period, a cluster of 200 one-day old eggs were selected and monitored every 12 hours and the number of hatched eggs were recorded.To determine the length of developmental period in the first and five instar nymphs, they were reared on the pistachio clusters of the cultivar.ese clusters were collected from an unsprayed orchard in the pistachio research center in Rafsanjan (Kerman province, Iran).e nymphs were reared in transparent plastic containers of 20 × 20 × 30 cm on the pistachio clusters, which were replaced every two days.e nymphs were checked daily until adults' emergence and death events in each stage were recorded separately.Aer emergence, both male and female insects were kept in pairs for oviposition within a plastic container (7 × 17 × 10 cm) with fresh pistachio clusters.e containers were revised daily and the number of laid eggs and adult's longevity were recorded.Demographic parameters were obtained under above-mentioned temperatures by constructing a life table of the insect by the method described by Carey (1993).e estimated demographic parameters are detailed on Figure 1 Fig. 1.Demographic parameters estimated in this study, sensu Carey (1993).Population growth parameters as net reproductive rate (R 0 ), intrinsic rate of natural increase (r m ), finite rate of increase (λ), generation time (T) and doubling time (DT) as well as reproductive parameters of B. germari were obtained following Birch (1948) and Carey (1993) equations under five constant temperatures.e longevity, growth, development, and reproductive period of the adults were analyzed with one-way ANOVA method.e variances and standard errors of the population growth and reproduction parameters were estimated by Jackknife technique (Meyer et al., 1986;Sokal & Rohlf, 1995).

Mortality rate
e mortality rate of eggs and nymph stages of B. germari under the studied temperatures are given in Table I.In both stages, the number of normal mortalities was separated from the abnormal mortalities (i.e.: non-inoculated eggs and dead nymphs by impact).e lowest mortality rate of the eggs (4.63%) was observed at 25 °C and with increasing temperature the mortality rate also increased, reaching 8.94% at 35 °C.Also, the mortality rates of the nymph stages under five different temperatures show that the highest value (23.68%) occurred at 35 °C, whereas the lowest (10.30%) being at 25 °C.e age-specific survival rates are shown in Figure 2. e total life cycle (egg to egg) reared at 20, 25, 27.5, 30 and 35 °C was 135, 117, 104, 80 and 57 days, respectively.II.Both the mean incubation period of eggs and the mean developmental time of nymph stages were the highest at 20 °C, and the length of these stages decreased with increasing temperature, until reaching the lowest values at 35 °C.e longest and the shortest total development times (egg to adult) also occurred at 20 °C and 35 °C, respectively.
e longevity of adult females was significantly affected by the temperatures (F=421.52,df=346, P<0.0001).In addition, there were significant differences among the studied temperatures on the longevity of adult males (F=141.41,df=346, P<0.0001).e longest and the shortest male and female adult longevity were observed at 20 °C and 35 °C, respectively (Table II).According to the results, the longevity of adult males was lower than that of the adult females under the experimental temperatures.e longest pre-oviposition period was observed at 20 °C, with significant differences with the other temperatures (F=2909.29,df=346, P<0.0001).e oviposition period also showed significant differences among different temperatures (F=524.74,df=346, P<0.0001).e post-oviposition period was significantly different at

Population growth parameters
e population growth parameters of B. germari under the studied temperatures are given in Table III.e highest gross reproductive rate was observed at 20 °C, and showed significant differences with the other temperatures (F=2737, df=346, P<0.0001).e highest net reproductive rate (R 0 ) was 66.53 at 25 °C, and showed significant differences with the others (F=1659, df=346, P<0.0001).e intrinsic rates of natural increase (r m ) were 0.05, 0.06, 0.06, 0.07 and 0.09 (day -1 ) at 20, 25, 27.5, 30 and 35 °C, respectively.Also, there were significant differences in the finite rate of increase (λ) among five temperatures (F=1076, df=346, P<0.0001), being highest at 35 ºC and lowest at 20 ºC.e highest values of mean generation time (T) and doubling time (DT) were also observed at 20 °C, and these parameters decreased by increasing the temperature from 20 °C to 35 °C (F=2.68,df=346, P<0.0001; F=1155, df=346, P<0.0001).

Reproductive parameters
e reproductive parameters are presented in Table IV.e gross fecundity rate showed significant differences under different temperatures (F=2636, df=346, P<0.0001).e highest and the lowest gross fecundity rates were observed at 20 °C and 35 °C (273.33 and 124.96 eggs per female), respectively.Also, the net fecundity and net fertility rates were significantly different among the five temperatures (F=1495, df=346, P<0.0001; F=1371, df=346, P<0.0001).e highest and the lowest gross fertility rates (251.64 and 119.49eggs per female, respectively) were observed at 20 °C and 35 °C.e daily fecundity (number of eggs laid per female per day) was the highest at 35 °C, and it was significantly different from the other temperatures (F=3087, df =346, P<0.0001).e highest daily fertility (number of fertile eggs laid per female per day) was observed at 35 °C, and was different from the other temperatures (F=4030, df=346, P<0.0001).
Table III.Population growth parameters of Brachynema germari under five constant temperatures.Mean of values that followed by the same letter in each row are not significantly different (Tukey test, P<0.05).
Table IV.Production parameters of Brachynema germari under five constant temperatures.
Mean of values that followed by the same letter in each row are not significantly different (Tukey test, P<0.05).

DISCUSSION
e suitable temperature for pistachio growth is 25 °C to 35 °C, and under this environmental condition some of its main pests are active and can cause important damage (Hokmabadi, 2011).Our results provided useful information about the growth, survival and reproduction of B. germari feeding on pistachio at five constant temperatures under laboratory conditions.ese results can help us to understand the ecology and biology of this pest.In addition, estimation of life table parameters can forecast the relationship between temperature and growth potential (Greenberg et al., 2005;Dong et al., 2007).We found that temperature affects the development time of immature stages of B. germari, agreeing with other researchers that have reported similar results (Bommireddy et al., 2004;Zamani et al., 2006;Liu & Meng, 2007;Da Silva & Daane, 2014;Basirat et al., 2016).In this research, the development time of immature stages of B. germari decreased with increasing temperature from 20 °C to 35 °C, and the shortest growth period was observed at 35 °C.In contrast, the highest mortality rate and the lowest survival rate in the immature stages were observed at this temperature, indicating that the survival rate decrease at higher temperatures.ese results are also in agreement with findings previously reported (Bommireddy et al., 2004;Da Silva & Daane, 2014;Basirat et al., 2016).Temperature had a significant effect on the longevity of males and females of this pest, and this feature decreased with increasing temperature from 20 °C to 35 °C, similar to that reported by Bommireddy et al. (2004).e reproduction parameters were also significantly affected by the different temperatures.e highest reproduction parameters were observed at 20 °C.A similar result has been reported by Mehrnejad (1998) on Agonoscena pistaciae Burkhhardt & Lauterer (Homoptera: Sternorrhyncha).e intrinsic rate of natural increase (r m ) is the most informative demographic parameter that denotes pest's increasing capacity (Andrewartha & Birch, 1954;Wang et al., 2000).is rate has been considered as the most complete parameter for comparing population growth under various environmental conditions (Aldyhim & Khalil, 1993;Tsai & Liu, 1998;Wang & Tsai, 2001;Kuo et al., 2006).In the present research, the highest and the lowest r m values were observed under 35 °C and 20 °C, respectively.Mehrnejad (1998) investigated the r m of A. pistaciae under 20 °C to 35 °C and obtained the lowest value at 20 °C, which is in agreement with our results.However, this researcher determined the highest r m of A. pistaciae at 30 °C (vs.35 °C in our study).e longest and shortest mean generation time of B. germari also occurred at 20 °C and 35 °C, respectively.Mehrnejad (1998) obtained the mean generation time of A. pistaciae at 20 °C and 35 °C and the lowest value of this parameter was observed at 20 °C, similar to our results.It also have been documented that the mean generation time decreases with increasing temperature in the range of 20 °C to 30 °C (Mehrnejad, 1998;Basiratet al., 2016).e highest r m and R 0 were observed under 35 °C and 20 °C, respectively.is could imply the highest population growth under this temperature range.
Our results showed that the temperature has significative impact on growth, survival and reproductive rate of pistachio green stink-bug, playing an important role in phenology regulation, seasonal abundance and population growth of this pest.Although insects in nature are not always exposed to constant temperatures, our studies under controlled conditions provided useful information about dynamics of B. germari that could be used in forecasting populations growth and evaluate integrated pest management (IPM) programs.
L x : number of individuals life at time x (age specific survival); m x : number of female offspring per female at time x (age specific fertility); x: age interval in days; r: per capita rate of population change; e: natural base = 2.71878 (constant), α: age of first laying; β: age of last laying Data analysis Statistical analysis was carried out using SAS version 9.1 soware (SAS Institute, 1998) and the means were compared by Tukey's test (P<0.05).
20 and 25 °C compared to the other temperatures (F=47.04,df=346, P<0.0001).e longest and the shortest pre-oviposition, oviposition and post-oviposition periods were observed at 20 and 35 °C, respectively.e age-specific fecundity of B. germari under five experimental temperatures is presented in Figure 2. e first oviposition was observed on day 78, 63, 54, 44 and 29 at 20, 25, 27.5, 30 and 35 °C, respectively.Table I.Immature mortality rate (%) of Brachynema germari under five constant temperatures.Table II.Immature developmental time, adult longevity and reproduction period values (days) of Brachynema germari under five constant temperatures.Mean of values that followed by the same letter in each row are not significantly different (Tukey test, P<0.05).