Selection of suitable predatory mites against, Panonychus citri (McGregor) (Acari: Tetranychidae) using relative control potential metrics and functional response

Abstract

Background

Selection of suitable predators for the control of the spider mite, Panonychus citri is always a challenge due to the lack of their reliable predicted efficacy. In this laboratory study, biological control potential of Neoseiulus californicus, N. cucumeris and Scapulaseius newsami, against P. citri using the “Relative Control Potential” (RCP) metric to compare their efficacies was evaluated. The functional response type was determined at different prey densities (after 24 h).

Results

The type II functional responses were observed from all the tested predators feeding on 3 different life stages of P. citri, except N. cucumeris, which was fed only on eggs. All predatory mites preferred eggs of P. citri with a higher attack rate (1.397) in N. cucumeris, followed by N. californicus (1.238) and S. newsami (0.975). Handling time was also lower in N. cucumeris than other mites, with a trend as: for N. cucumeris, eggs > adults > immature stages; for N. californicus, immature stages > eggs > adults, while for S. newsami, eggs > immature stages > adults. The integration of greenhouse abundance and fecundity (%) data resulted in a higher RCP for N. cucumeris than other predators.

Conclusion

Neoseiulus cucumeris was selected as the most suitable predatory mite for the control of P. citri with a higher RCP and potential to maximum reproduction. S. newsami, which was used for the first time, gave almost similar results compared to N. californicus but with lower efficacy.

Background

Most of the insect pest population is controlled by natural predators as a biological control strategy (Symondson et al. 2002). The selection of a suitable predatory species depends on its generalist characteristics (ability to feed on alternative prey/food sources) and break the pest population during colonization (Messelink et al. 2012). This selection of suitable predators was done by using functional response on prey densities and consumption (Solomon 1949), which allows for the estimation of maximum daily prey consumed or attacked by a predator by determining its stabilizing factor in the predator–prey population (Murdoch and Oaten 1975). The functional response resulted in a linear increase, decelerating increase to a plateau, sigmoid increase, and dome-shaped curves for type I, type II and type III (Trexler et al. 1988) and type IV, respectively based on the number of prey killed by a predator against total available preys (Pervez and Omkar 2005). Functional responses gave the predicted impact of the predator on the prey densities (Dick et al. 2017). However, comparing two or more predators requires an estimation of their relative control potential (RCP) by using their attack rate as a functional response measure and fecundity as a numerical response measure to choose an efficient predator (Cuthbert et al. 2018).

Panonychus citri (McGregor), a citrus red mite, is a major pest in Asia. (Kennett et al. 1999) including southern China (Li et al. 1997). The motile stages (immature stages) and adults feed on cell content which leads to stippling symptoms. Severe infestation of P. citri leads to defoliation, fruit dropping, and dieback (Keetch 1968). Due to extensive chemical control, P. citri developed resistance and changes in its behavior against many acaricides (Qayyoum et al. 2021a, b). In addition to chemical control of P. citri, management cost is higher and difficult (hilly areas), which also may eliminate the role of its natural enemies. Recently, mainly Phytoseiidae and some other mite families used as biological control agents of different phytophagous mites (Cakmak et al. 2009). Among the phytoseiids, Neoseiulus californicus (McGregor) and N. cucumeris (Oudemans) are commonly used as an augmented biological control against different mite and insect pests (Mendel and Schausberger 2011). At the same time, Scapulaseius newsami (Evans) is commonly observed in the citrus or lemon fields of southern China (Song et al. 2019). N. californicus is a selective predator of tetranychid mites with type II functional response, while N. cucumeris and S. newsami are placed in the generalist phytoseiid mites with type III preferring other prey than spider mites (McMurtry et al. 2013). However, these predatory mites have different lifestyles from each other. For evaluating a predator’s effectiveness, the functional response is widely used to predict potential biological control agents. For a better understanding the effectiveness of N. californicus, N. cucumeris, and S. newsami, the functional responses against different life stages of P. citri with their relative control potential (RCP) for comparison were evaluated under laboratory studies.

Methods

Mites’ cultures

Neoseiulus californicus and N. cucumeris were obtained from a laboratory culture rearing on Tetranychus urticae Koch. and Tyrophagus putrescentiae. S. newsami and P. citri were collected from citrus/lemon orchards in 2019 and cultured for 6 months before starting the experiment. The predator–prey synchronizations were gained by rearing N. californicus, N. cucumeris, and S. newsami on P. citri. The eggs of predators were separated into new leaf (lemon) discs with enough prey mites to get same-aged adult females. All this rearing process was carried out under controlled conditions with a temperature of 26 ± 1 °C, photoperiod of 16: 8 h (Light: Dark), and relative humidity of 75 ± 1%.

Experimental conditions and methodology

The experiment was performed using a lemon leaf disc (lower leaf surface up) (3.5 cm diameter) by placing it on a water-saturated sponge. All leaf discs were stripped with absorbent paper on their edges to prevent mites from escaping and retain moisture. Mites attached to the absorbent paper were not included in the study data. Each 4 tested units were considered one test group by placing into a plastic box (15 cm × 15 cm × 6.5 cm) and replicated 10 times (10 × 4). All plastic boxes were filled with water just below leaf discs and water was maintained regularly. All experimental procedure was done under controlled conditions as described above. Adult females (> 24 h old) were collected from the stock cultures and isolated from the colony before gaining maturity for uniform age. All collected females were allowed to mate with males for a period of 24 h. After mating, females were starved for 24 h before transferring to the experimental units. Injured or less moved females were excluded from the experiments.

Different densities of P. citri eggs were obtained from placing the 10–20 adults female on the lemon leaf discs and removed after 24 h under controlled conditions. The number of eggs was counted as per density by removing excess eggs. Immature densities were obtained by placing more than 50 adult females on the lemon leaf and allowing them to lay eggs for 5–7 h. before removing the adults. For further growth, the lemon leaf with eggs was placed under controlled conditions for further development. As soon as the immature stage (> 90% proto-nymph) was obtained, they were shifted to the experimental unit.

The selection of prey densities against the 3 predators (N. californicus, N. cucumeris, and S. newsami), a pre-consumption of immatures and adults of P. citri was carried out. The pre-densities selection test revealed that offering predators more than 200 and less than 4 motile stages of P. citri reduced their searching performance and reduced fecundity rate. A maximum of 160 and a lowest of 5 density of each stage was offered to the predator based on the pre-test data, as endorsed by Xiao and Fadamiro (2010).

A functional response experiment was performed using 6 densities (5, 10, 20, 40, 80, and 160) of eggs, immatures, and adults of P. citri by offering to a single female of N. californicus, N. cucumeris, and S. newsami. The prey eggs, immatures, and adults consumed by the predators were replaced by new ones during the experiment. Data of prey eaten and eggs laid by all predators were recorded after 24 h.

Statistical analysis

Data from functional responses are analyzed in 2 steps (Juliano 2001). In the first step, maximum likelihood estimates (intercept (P₀), linear (P₁), quadratic (P₂), and cubic (P₃) coefficients) were calculated, using a logistic regression/polynomial function by comparing the proportions of prey eaten to the initial density and resulted in functional response type.

$$N_{e} {/}N_{0} {\text{ = exp(}}P_{0} { + }P_{1} N_{0} { + }P_{2} N_{0}^{2} { + }P_{3} N_{0}^{3} {\text{)/[1 + exp(}}P_{0} { + }P_{1} N_{0} { + }P_{2} N_{0}^{2} { + }P_{3} N_{0}^{3} {)]}$$

where “N_e/N_o” is the prey consumption probability and “intercept (P₀), linear (P₁), quadratic (P₂)_, and cubic (P₃)” are regression coefficients. A significant functional negative and positive value of P₁, the predator gave functional response types II and III curve, respectively (Juliano 2001).

Since it accounts for prey loss per density, the non-linear least square regression was used in the second step to estimate handling time and attack rate coefficients.

$$N_{e} = N_{0} \left\{ {1 - \exp \left[ {\alpha \left( {T_{h} N_{e} - T} \right)} \right]} \right\}$$

where “Ne” stands for the number of prey killed, “N_o” stands for initial prey density, “T_h” stands for the handling time, and “T” stands for the total time given to the predator (24 h). The attack rate “α” and handling time “T_h” parameters were calculated using a nonlinear least-squares regression procedure.

Curve fitting using the polynomial equation and Holling’s disc equation was performed using R (RCore-devlopment-team 2019). ANOVA analyzed the prey density-dependent effects on consumption and number of eggs laid by predators, and then the Tukey test was used to compare the results (P = 0.05). The abundance data of the 3 predatory mites were obtained from a greenhouse augmented biological control experiment by releasing the predatory mites (n = more than 50/plant) at the highest infestation of P. citri and other insect pests. Abundance data (Mean ± SE) from highest (predators released) to almost zero P. citri was used. The fecundity (%) of predators (per adult female) was calculated by using the below formula by feeding on different life stages of P. citri;

$${\text{Fecundity}} = \left( {{\raise0.7ex\hbox{${{\text{Eggs}}\;{\text{production}}}$} \!\mathord{\left/ {\vphantom {{{\text{Eggs}}\;{\text{production}}} {{\text{prey}}\;{\text{consumed}}}}}\right.\kern-\nulldelimiterspace} \!\lower0.7ex\hbox{${{\text{prey}}\;{\text{consumed}}}$}}} \right) \times 100$$

The relative control potential (RCP) was calculated to compare their efficacies based on a maximum feeding rate (1/h), as a result of their functional response and abundance of predator in greenhouse or fecundity (%) as calculated above. The RCP was calculated as below formula;

$${\text{RCP}} = \left( {\frac{{{\text{FR}}_{{(1/{\text{h}})}} {\text{predator}} - A}}{{{\text{FR}}_{{(1/{\text{h}})}} {\text{predator}} - B}}} \right) \times \left( {\frac{{X\;{\text{predator}} - A}}{{X\;{\text{predator}} - B}}} \right)$$

where FR is the maximum feeding rate (1/h), and “X” is proxies for the responses (a measure of predator greenhouse abundance or fecundity of predator). If the value of RCP = 1, no difference between the 2 predators; for RCP < 1, predicted predator-A less efficient than predator-B, and RCP > 1, predator-A is predicted as greater efficacy than predator-B. RCP indicating the predicted increase or decrease relative efficacy of predator-A compared to predator-B (Cuthbert et al. 2018).

Results

The prey's (Panonychus citri) survival rate varies depending on predator species, prey life stage, and density. Prey survival was a maximum in their adult stage at the highest density (160 prey) when fed by N. californicus and S. newsami, while for N. cucumeris, prey survival rate was higher at the immature stage (86.1%) (Table 1). Daily consumption rate of N. cucumeris was significantly higher than N. californicus and S. newsami. The prey consumption rate increased with an increase in prey density then decreased after attaining maximum consumption at the density of 80 prey. The daily consumption rate of all tested predators among densities and within the specific densities of all predatory mites were also observed highly significantly (Table 1). The consumption rate of all predatory mites against P. citri depended on density, life stage of prey, and interaction, as presented in Table 1. Fecundity rate of female N. californicus and N. cucumeris was increased with the increase of gradual level of prey density rather than prey consumption, except for S. newsami by feeding on immature stages and adults as presented in relationship expression between the rate of oviposition and density of prey (Table 2). Daily eggs laid by a female of N. californicus and S. newsami depended on prey density only, while N. cucumeris number of eggs laid in a day depended on prey-stage and density (Table 2).

Table 1 Mean (± SE) consumption of three predatory mites at different densities of different stages of P. citri

Table 2 Mean (± SE) eggs laid by a female(three predatory mites) mites at different densities of different stages of P. citri

The functional responses were observed from all predators fed on the 3 different life stages, with significant negative P1 or first-order terms, followed by significant negative P2 or second-order terms, except N. cucumeris fed on eggs of P. citri (weak type-II with non-significant negative P1). Attack rate increased significantly by feeding on adults > immatures > eggs by N. californicus and S. newsami, except for N. cucumeris, which trends as immatures > adults > eggs. The overall attack rate for N. cucumeris was significantly higher than the other two predatory mites (Table 3).

Table 3 Results of logistic regression to denote functional response type across predatory mites by feed on three life stages of P. citri using Holing’s disc equation

The Relative Control Potential (RCP) was calculated using the abundance data of the 3 predatory mites with a max. Feeding rate as presented in Table (4). N. cucumeris was much more abundant than N. californicus and S. newsami, resulting in greater RCP than both predators, as both functional response parameters were used (Table 4). Overall, asymptotes (maximum feeding rate (1/T_h)) of functional response curve observed non- significant difference between N. californicus and S. newsami, but N. cucumeris maximum feeding rate was significantly different from other both species. There was a significant difference between the max. feeding rate in all life stages. The increase in max. feeding rate pattern varied within the species as: for N. californicus, adults > eggs > immatures; for S. newsami, adults > immatures > eggs and for N. cucumeris, immatures > adults > eggs (Table 5). By the change in prey stage, the RCP of predators varied among each other and within prey-stage feeding (Table 5). Overall, N. cucumeris had a greater efficacy than N. californicus and S. newsami. In comparing N. californicus and S. newsami, N. californicus efficacy was greater than S. newsami by feeding on immatures of P. citri, followed by adults and eggs. In a comparison of N. cucumeris and N. californicus, N. cucumeris significantly showed a higher efficacy than N. californicus by feeding on the eggs of P. citri, followed by immatures and adults. While in comparison of N. cucumeris with S. newsami, resulted in higher efficacies than other comparisons, trend as; adults > eggs > immatures (Table 5). A significant difference between the predatory mites to response against prey consumption at different stages, with N. cucumeris exhibiting a higher predation efficacy by feeding on immatures (comparison with S. newsami) and eggs (comparison with N. californicus) was concluded. N. cucumeris was also observed consistently in its prey consumption response to different life stages (increase in movement) of P. citri.

Table 4 Mean Relative Control Potential (RCP) using abundance mean per leaf (n = 6) (Additional file 1: Fig. S1), for N. californicus, S. newsami, and N. cucumeris, that the RCP output exceeds 1 using bootstrapped maximum feeding and attack rates

Table 5 Mean relative control potential (RCP) using fecundity (%) (Additional file 1: Table S1–3) across three life stages of P. citri for N. californicus, S. newsami, and N. cucumeris, that the RCP output exceeds 1 using bootstrapped maximum feeding and attack rates

Discussion

In biological control systems, phytoseiid predatory mites are considered effective biological control agents against different mites’ species and insect pests (Gerson et al. 2003). Many attempts have been made to predict predatory mite efficacy, using functional and numerical responses to various population density levels (Teodoro et al. 2020). However, considering the functional response, these efforts to predict predatory mites/natural enemies' efficacy/consumption are insufficient (per capita effects) (Fernández-arhex and Corley 2003) and numerical response (population responses) (Dick et al. 2017) who used the RCP metric by combining both responses (functional and numerical), which gave more accurate predictive and explanatory results of biological control agents (invasive) with population reduction of target species (prey). Furthermore, RCP also efficiently integrated the dependencies associated with a change in environmental/ habitat conditions (Laverty et al. 2015) and food preference, for strong predator–prey interactions. This study presents the new RCP metric, which compares efficacy among predatory mites using per capita, population responses, and food preferences to predict change in their efficacies.

The functional responses of all predatory mites species exhibited the type-II by giving significant negative P1, which the consumption increased with the increase of prey availability and started to decline after gaining maximum peak as previously reported by Song et al. (2019) for N. cucumeris and N. californicus against Tetranychus urticae. At the same time, N. cucumeris exhibited a weak functional response type-II by offering eggs of P. citri (non-significant negative P1). The predators with functional response type-III are known as effective biological control agents (Pervez and Omkar 2005), but most of the predators showed type-II (Xiao and Fadamiro 2010) wrong arranged. S. newsami was identified as a dominant and efficient predator species from the citrus growing region of Southern China (Guangdong Province) during the highest infestation of P. citri (Song et al. 2019) with type-II functional responses similar to N. californicus (Table 3). In contrast, S. newsami was placed with functional response type-III (living on glabrous leaves) by McMurtry et al. (2013) due to habitat preferences, not prey consumption ability.

Here, RCP resulted in S. newsami with least efficient biological control agent than others which explained that other factors (like: environmental and alternative food source) involved in the expected availability of S. newsami in the citrus or lemon fields in China (Song et al. 2019). The change in prey life stage was shown to mediate changes to functional responses of all predatory mites. N. cucumeris was found more efficient than other predatory mites against P. citri with a higher RCP, integrated with fecundity (%) and field abundance as previously reported by Zheng et al. (2017) against T. urticae. The usages of abundance and fecundity estimate how predator–prey interaction strong (numerical response proxy) and their reproduction ability when introduced, respectively. N. cucumeris gave a higher RCP by feeding on eggs and immatures than N. californicus and S. newsami, respectively. The attack rate parameters revealed the efficacy of predators. The consumption rate of N. cucumeris was significantly higher than N. californicus and S. newsami due to increased metabolic demand. In contrast to Zhang et al. (2001), N. cucumeris by converting P. citri energy to maximum (2 × than Zhang et al. 2001) eggs production in this study. This can explain that even N. cucumeris was not associated with P. citri, but maximum potential against P. citri with a higher reproductive turnover than other predatory mites.

Obtained results on the preference of prey-stage profound effective on pre-predator interactions. All predatory mites preferred eggs of P. citri due to an effortless approach with more nutritional values (Ebrahim et al. 2014). In contrast, previously reported that N. californicus preferred active larval/nymphal stages (immature stages) of T. urticae (Zheng et al. 2017), T. cinnabarinus (T. cinnabarinus is a junior synonym of T. urticae) (Li et al. 2014), and P. citri (Xiao and Fadamiro 2010). These variations in preference within predatory species may be due to genetic diversity, experimental conditions, prey density differences, and rearing technology of both prey and predator. The above comparison of obtained results mostly with Tetranychus spp. as prey (Popov and Kondryakov 2008) suggested that the predacious ability may change with change in prey species. There was a slighter difference in fecundity rate of all 3 predatory mites by feeding on eggs of P. citri compared to immatures and adults (Table 2), which resulted in non-significant dependency on the prey life stage, except for N. cucumeris (Table 1). These results explained that nutritional level had no impact on the N. californicus and S. newsami with the same profitability, while in the case of N. cucumeris, a maximum nutritional intact with a maximum output (reproduction).

Conclusions

In conclusion, using the RCP metric provides a successful explanation and prediction of predators /invasive species for the selection of the best biocontrol agent. Overall, change in prey stage will induce change in predation ability of predatory mites towards P. citri. N. cucumeris was the most suitable predatory mite for the control of P. citri with a higher RCP and potential to maximum reproduction. S. newsami used for the first time gave almost similar results compared to N. californicus with less efficacy. The addition of environmental variations and prey responses will be more helpful to get better RCP for the selection of biocontrol agents as a future consideration.