Unraveling key biological aspects of Delottococcus aberiae (Hemiptera: Pseudococcidae) to enhance pest management

Abstract

Effective pest management relies on understanding how environmental factors, particularly temperature, influence insect development and population dynamics. The invasive mealybug Delottococcus aberiae (De Lotto) has caused significant damage to citrus crops since its detection in 2009 in eastern Spain; despite this, knowledge of its biology remains limited. To address this gap, we examined key biological traits of D. aberiae across a range of temperatures from 15 to 30 °C. The developmental thresholds of D. aberiae were estimated at 9.4 and 30 °C, with the species performing best under moderate conditions but declining at higher temperatures. D. aberiae required a thermal constant of 730 degree-days to complete 1 generation, corresponding to 4 to 5 generations per year under the climatic conditions of eastern Spain, where the species is established. D. aberiae reproduces exclusively sexually, with peak fertility occurring at 22.5 °C, when females laid an average of 500 eggs and exhibited the largest body sizes. Overall, D. aberiae thrives within the temperature range typical of the Mediterranean basin. Although it shows low tolerance to high temperatures, the developmental thresholds and thermal constant determined in this study indicate that D. aberiae can withstand both winter and summer conditions, allowing populations to persist throughout the year. These findings can be used to predict population peaks, prevent spread to other citrus-producing regions with similar climates, and identify critical developmental stages for intervention, thereby supporting more targeted and effective control strategies.

Graphical Abstract

Graphical Abstract.

Introduction

The South African mealybug Delottococcus aberiae (De Lotto) (Hemiptera: Pseudococcidae) is an invasive citrus pest, detected for the first time in citrus orchards in the region of Les Valls in Valencia, eastern Spain, in 2009 (Beltrà et al. 2012, 2013, 2015, García-Marí 2012). Since then, its populations have spread across neighboring regions, posing a threat to other citrus-producing regions in the Mediterranean basin.

Originally from South Africa, D. aberiae is commonly found on wild olive trees and on the roots of Chrysanthemoides monilifera (L.). In the Limpopo Province of northern South Africa, the mealybug also occurs sporadically in citrus orchards, where it is not considered a significant pest (Miller and Giliomee 2011). Spanish populations originate from this region, and its detection in eastern Spain represents the first record of D. aberiae as a major citrus pest (Beltrà et al. 2012, 2013, 2015, García-Marí 2012).

Delottococcus aberiae is a polyphagous species reported on plants from 25 botanical families, including tropical and subtropical crops such as citrus, coffee, guava, pear, persimmon, and olive (De Lotto 1961, Miller and Giliomee 2011, Pérez-Hedo et al. 2018). In Spain, it has been recorded on persimmon, loquat, pomegranate, and several citrus species, where it causes the greatest damage (MAPA 2024).

Unlike other citrus mealybug species, D. aberiae causes direct damage to fruit by distorting and reducing its size, ultimately leading to significant production losses (Beltrà et al. 2013, Soto et al. 2016a, Pérez-Rodríguez et al. 2017, Martínez-Blay et al. 2018a). Delottococcus aberiae produces several generations per year, with the first being the most damaging to citrus (Martínez-Blay et al. 2018b). The critical period for fruit distortion occurs between flowering until fruit reaches a diameter of 25 to 30 mm, typically from March–April to July under the climatic conditions in eastern Spain (Pérez-Rodríguez et al. 2017, Martínez-Blay et al. 2018b). In addition, the secretion of honeydew causes the growth of black sooty mold fungi, which interferes with plant photosynthesis (Franco et al. 2000).

Previous studies indicate that, like other mealybugs, D. aberiae exhibits cryptic behavior, congregating in protected areas of plants where it feeds and reproduces (Martínez-Blay et al. 2018b). Reproduction in D. aberiae has been considered sexual (Martínez-Blay et al. 2018b). However, facultative parthenogenesis, where unmated females can occasionally produce viable offspring, has been reported in other mealybug species considered to reproduce sexually (Lloyd 1952, da Silva et al. 2010, Tabata et al. 2016), and therefore cannot be excluded in D. aberiae.

Although extensive field studies have been conducted on the dynamics, structure, and distribution of D. aberiae populations (Beltrà et al. 2013, Soto et al. 2016a, Pérez-Rodríguez et al. 2017, Martínez-Blay et al. 2018b,c), studies on its biology are scarce. Due to the absence of natural enemies (Soto et al. 2016b, Tena et al. 2017), control of D. aberiae has relied on broad-spectrum insecticides. However, the mealybug’s cryptic behavior and restrictions on pesticide use during flowering have led to alternative strategies, including the introduction of a parasitoid from its native range (Beltrà et al. 2015), pheromone-based monitoring (Vacas et al. 2019), and preventive sanitation measures to limit pest spread (Martínez-Blay et al. 2018b). Given the limitations of conventional control methods, understanding the pest’s fundamental biological traits is crucial for developing more effective management strategies (Chong et al. 2003, Martínez-Blay et al. 2018b, Rebaudo and Rabhi 2018). Environmental factors play a central role in shaping insect life history traits, such as growth, adult longevity, and oviposition. Among these, temperature is one of the most critical, as it regulates development and survival (Uvarov 1931, Davidson 1944, Romoser 1981, Trudgill et al. 2005). Regression models have been applied to describe the relationship between temperature and insect development, providing valuable insights into thermal requirements that can be used to optimize pest management (Davidson 1944, Wagner et al. 1984). For example, developmental thresholds derived from these models have been used to guide the timing of insecticide applications and the release of natural enemies against mealybugs, such as Planococcus ficus (Signoret) and Maconellicoccus hirsutus (Green), as well as to predict the distribution of Paracoccus marginatus (Williams & Granara de Willink) and the population peaks of Dysmicoccus brevipes (Cockerell) (Chong et al. 2003, Amarasekare et al. 2008, Varikou et al. 2010, Bertin et al. 2019).

This study aims to provide valuable insights into the pest’s biology and seasonal occurrence by identifying its developmental thresholds, thermal constant, survival, and reproductive patterns. Such knowledge is essential to develop and implement targeted and sustainable approaches, thereby enhancing the effectiveness of integrated pest management. Ultimately, this research will contribute to the development of science-based strategies to mitigate the spread and impact of D. aberiae in citrus-growing regions.

Materials and Methods

Insect and Plant Materials

A laboratory colony was initiated from individuals collected on infested citrus branches in the region of Les Valls (Valencia, Spain) and identified as D. aberiae. The colony was then established in an insectary at the Instituto Agroforestal Mediterráneo of the Universitat Politècnica de València. The mealybugs were reared on pumpkin fruits, Cucurbita maxima, which were treated with a 2% bleach solution to prevent the growth of mold and other organisms. The infested pumpkins were placed inside rearing cages (47.5 × 47.5 × 47.5 cm) containing between 3 and 5 squashes each. To maintain the colony, 3 or 4 fresh pumpkins were introduced every 2 wk to renew the old, dried ones. The mealybug colony was kept in darkness in climate-controlled chambers at a temperature of 24 °C and 70% ± 10 RH.

Effect of Mating Status on the Reproductive Biology of D. aberiae

To assess the reproductive biology and potential parthenogenetic capacity of D. aberiae, experiments were conducted under mated and unmated female conditions. Newly molted adult females, collected from the laboratory colony before any male contact, were individually transferred to green lemon fruits and placed inside 1-L plastic containers. Containers were randomly assigned to each of these 2 treatments. In the non-mating treatment, females were isolated without male exposure. In the mating treatment, 3 males were introduced into each container and replaced every 3 d to ensure continuous mating. Females from both treatments were observed daily for survival and ovisac formation, and once oviposition began, the number of eggs laid was recorded until female death. Offspring sex ratio was then determined by rearing the laid eggs until the third instar, when sexual differentiation was possible. Survival and fecundity curves were constructed for each treatment using the observed survival (l_x) and fecundity (m_x) rates, where x is female age (d), l_x is the proportion of females surviving to age x and m_x is the mean number of eggs produced per female at age x multiplied by the proportion of females in the sex ratio. Survivorship and fecundity were used to estimate the following demographic parameters (Birch 1948): net reproductive rate (R₀ = ∑(l_x·m_x)), which is to the rate of population multiplication per generation; intrinsic rate of increase (r_m = ∑e^−rmx·l_x·m_x = 1), representing the potential population growth; finite rate of increase (𝜆 = e^rm), which corresponds to the daily population multiplication factor; mean generation time (T = ∑(x·l_x·m_x)/R₀), which is the average female age at reproduction; and the doubling time (DT = ln(2)/r_m) which represents the time required by a population to double in size. The experiments were conducted in climate-controlled chambers at 24 °C with 70% ± 10 RH and darkness, with 30 female replicates per treatment.

Effect of Temperature on the Biology of D. aberiae

Development, Longevity, and Oviposition

The development time of D. aberiae from egg to adult and the longevity of adult females until death were assessed at 7 constant temperatures: 15, 18, 21, 22.5, 24, 27, and 30 °C. Females from the laboratory colony, presumed to be gravid because of their large size, were incubated on bean pods for oviposition. Bean pods were selected as a suitable substrate for observing oviposition, as newly laid eggs were easily visible and could be readily transferred to experimental containers, facilitating uniform egg distribution. Eggs deposited within 24 h were collected, transferred to green lemon fruits, and placed inside 1-L plastic containers, as described above. Containers were then transferred to each of the 7 experimental temperatures. After hatching, mealybug nymphs were individualized and examined daily. The duration of each developmental stage from egg to adult, as well as adult female longevity, was recorded. Molting from one instar to the next was confirmed by the presence of the exuviae and the absence of body wax. Since adult female mealybugs do not molt, the beginning of egg sac formation was used to distinguish between preovipositional and ovipositional females. Because sex differentiation was not possible before the third instar, data from eggs to second-instar nymphs were pooled regardless of sex, while only female data were recorded from the third instar to adulthood. However, given the biological relevance of 24 °C, data on male prepupa, pupa and adults were also recorded at this temperature.

When females reached adulthood, each was provided with 3 newly emerged males, which were replaced every 3 d to ensure mating until oviposition began. The total number of eggs laid by ovipositing females was recorded at all temperatures tested. Survival curves were constructed for each temperature based on the observed survival rates (l_x), where x represents female age (d) and l_x is the proportion of females surviving to age x. A minimum of 25 observations were obtained for each developmental and adult stage of D. aberiae at each temperature. All tests were conducted in environmental chambers with a relative humidity of 70% ± 10% and complete darkness to facilitate mealybug establishment.

Body Size

The body size of D. aberiae individuals was measured at the same 7 constant temperatures. Length and width were recorded using a stereoscope fitted with an ocular micrometer. The same individuals used in the developmental experiment were measured within 24 h after each molt and at the beginning of ovisac formation. At least 20 individuals were measured per developmental stage at each temperature.

Thermal Thresholds and Phenology of D. aberiae

To evaluate the relationship between mealybug developmental rate and temperature, the development time of D. aberiae from egg to adult stage at 7 temperatures was used to build linear and nonlinear regression models. The accuracy of the models was evaluated based on the adjusted coefficient of determination ($R_{\text{adj}}^2$), the residual sum of squares (RSS), and the goodness of fit to the observed development rates, as well as the thermal threshold estimations.

The linear approximation equation is useful to extrapolate the relationship between temperature and developmental rate within a moderate temperature range, such as 15 to 22.5 °C (Davidson 1944). Hence, a linear regression model was used to calculate the lower developmental threshold (ie the base temperature below which the insect is unable to develop), and the thermal constant (ie the number of accumulated degree-days above T_min needed to complete a developmental stage). The linear regression equation is:

$$D_{\mathrm{r}}=a+bT,$$

where D_r is the developmental rate (1/d), T is the temperature, and a (intercept) and b (slope) are constants to be fitted from the data. The lower development threshold and the thermal constant were calculated as T_min = −a/b and k = 1/b, respectively (Campbell et al. 1974). K is the thermal constant defining the accumulated degree-days needed for insect development.

The upper developmental threshold (T_max) and the optimal temperature (T_opt) for development were estimated using nonlinear regression models. Several models commonly used to describe temperature-dependent development were evaluated, including Logan et al. (1976), Ratkowsky et al. (1983), Rosso et al. (1993), Lactin et al. (1995), and Briere et al. (1999). Among these, only the Logan-6 and Lactin-2 models fit the data adequately. Model performance was compared using the Akaike Information Criterion (AIC) (Akaike 1974), with lower AIC values indicating a better fit. Based on this criterion, the Lactin-2 was selected as the best-fitting model, as it produced lower AIC values than the Logan-6. The Lactin-2 model is given by the expression:

$$D_{\mathrm{r}}=\lambda +{\mathrm{e}}^{\rho T}-{{\mathrm{e}}^{(\rho T_{\mathrm{u}}-(T_{\mathrm{u}}-T)/\mathrm{\Delta }T)}}^{},$$

where λ forces the curve to intercept the x axis below zero, allowing for an estimation of the lower developmental threshold, T_u represents the upper lethal temperature, which is higher than the conceptual upper developmental threshold, and ρ is a constant for the biochemical reaction rate as temperature increases to T_opt.

The number of generations of D. aberiae was estimated by dividing the annual degree-days accumulated above its lower developmental threshold (T_min) by the thermal constant derived from the linear model. Temperature records from 2018 to 2024 (MAPAMA 2025) for Les Valls (eastern Spain) were used to estimate the number of generations in the region.

Statistics

The development, longevity, and body size of each mealybug developmental stage were compared across temperatures using an analysis of variance (ANOVA) with treatment means separated using Tukey’s HSD test at a significance threshold of 0.05. When necessary, variables were transformed using a log, square-root, or an arcsine transformation before analysis to normalize the data and adjust the variances. When data could not be adjusted to a normal distribution, a Kruskal–Wallis 1-way ANOVA test, followed by Dunn’s post hoc test using Bonferroni’s correction (P < 0.05) (STATGRAPHICS Centurion 2022) was used. Fecundity under different mating treatments was assessed using a Wilcoxon rank-sum test (P < 0.05). The effect of temperature and mating treatment on mealybug survival was compared using a Log-Rank test, with P-values adjusted by the Benjamini–Hochberg method to control multiple comparisons (P < 0.05) (STATGRAPHICS Centurion 2022). Demographic parameters (R₀, r_m, λ, T, and DT) were calculated from the life table data, and standard errors were estimated using the jackknife procedure (Hulting et al. 1990). We used regression analysis to describe the relationship between temperature and developmental rate, and to calculate temperature thresholds and thermal constant with STATGRAPHICS Centurion 2022.

Results

Effect of Mating Status on the Reproductive Biology of D. aberiae

Delottococcus aberiae reproduces sexually, with reproductive behavior observed exclusively in female individuals that were exposed to males. Mated females laid an average of 257 eggs throughout their lifespan at 24 °C, whereas female mealybugs that were deprived of male exposure did not produce an ovisac or lay eggs (W = 900, P < 0.001; Fig. 1a). In the mating treatment, the peak oviposition period occurred between the third and sixth days after oviposition began, with females laying an average of 149 eggs, accounting for nearly 60% of the total egg production. Female lifespan differed significantly between mating treatments (χ² = 67.3, df = 1, P < 0.001), with non-mated females surviving longer than mated ones. On average, mated females lived 32.1 d from the last molt to death, compared with 79.1 d for non-mated females (Fig. 1b).

Fig. 1.

Number of eggs laid (a) and age-specific survival (lx) (b) of mated (dashed black line) and unmated (straight blue line) D. aberiae females at 24 °C.

The average net reproduction rate (R₀) of D. aberiae was 160.21 female offspring per female, with an intrinsic rate of increase (r_m) of 0.0754 d⁻¹ and a generation time (T₀) of 67 d. The sex ratio was 1:1.6 (male:female) (Table 1).

Table 1.

Demographic parameters of D. aberiae at 24 °C

Parameter	Mean ± SE
R  o (♀/♀)	160.21 ± 14.42
r  m (♀/♀/d)	0.075 ± 0.0012
λ (♀/♀/d)	1.078 ± 0.0013
T (d)	67.31 ± 0.22
DT (d)	9.19 ± 0.148
Sex ratio (♂:♀)	1:1.6

Effect of Temperature on the Development and Survival of D. aberiae

The development of D. aberiae was significantly affected by temperature (H = 118.6, df = 5, P < 0.001; Table 2). Delottococcus aberiae completed development within a temperature range of 15 to 27 °C, while no development was observed at 30 °C. At 30 °C, the eggs successfully hatched; however, the first instar nymphs failed to develop further and died before molting into the next instar. The development time for each life stage decreased with increasing temperature up to 22.5 °C, after which it rose again until reaching 27 °C. The mean time for D. aberiae to complete development (egg to adult) ranged from 94.92 d at 15 °C to 42.27 d at 22.5 °C, while the generation time (egg to egg) ranged from 120.51 at 15 °C to 54.19 d at 22.5 °C, respectively (Table 2). At 24 °C, the prepupal and pupal stage of males averaged 11.3 ± 0.04 d, and adult males lived a mean of 2.7 ± 0.07 d (Table 2).

Table 2.

Mean development time (±SE, d) and longevity (±SE, d) for each developmental stage of D. aberiae reared under 7 constant temperatures

T (°C)	Immature stages				Adult stage		Development (egg to adult)	Generation (egg to egg)
	Egg	First instar	Second instar	Third instar	Preoviposition	Oviposition
15	26.6 ± 0.4a	29.1 ± 0.4a	19.1 ± 0.4a	20.3 ± 0.4a	25.6 ± 0.8a	39.0 ± 2.7a	94.9 ± 0.4a	120.5 ± 0.5a
18	16.4 ± 0.3ab	22.7 ± 1.0 b	13.9 ± 0.6 b	15.9 ± 0.7 b	21.5 ± 1.2 b	31.7 ± 2.4a	68.8 ± 0.7ab	90.3 ± 0.8ab
21	12.6 ± 0.2 b	14.4 ± 0.4d	9.6 ± 0.5c	11.5 ± 0.3c	14.6 ± 0.5c	16.6 ± 0.9c	48.1 ± 0.3 cd	62.6 ± 0.4 cd
22.5	10.2 ± 0.2c	13.1 ± 0.2 cd	9.2 ± 0.4c	9.9 ± 0.5d	11.9 ± 0.6d	21.3 ± 1.2 b	42.3 ± 0.3d	54.2 ± 0.4d
24	9.3 ± 0.2 cd	14.9 ± 0.5d	11.0 ± 0.8c	11.8 ± 0.5c	16.2 ± 1.3c	15.9 ± 1.7c	47.1 ± 0.5 cd	63.3 ± 0.7 cd
				11.3 ± 0.1 (♂)a	2.7 ± 0.1 (♂)		46.6 ± 0.1 (♂)	–
27	8.5 ± 0.1d	17.2 ± 0.7c	16.0 ± 1.5 b	15.3 ± 1.7 b	21.4 ± 1.4 b	11.2 ± 0.8d	56.9 ± 1.0bc	78.4 ± 1.1bc
30	6.9 ± 0.1e	–	–	–	–	–	–	–
H	460.2	281.4	88.3	91.9	89.9	87.1	118.6	122.2
P-value	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001

Means within a column followed by the same lowercase letter are not significantly different among temperature treatments (Dunn’s test, P < 0.05).

^aMale prepupa and pupa data combined.

Similarly, the adult lifespan of both preovipositional and ovipositional females significantly shortened with increasing temperature (H = 89.9 and H = 87.1, respectively; df = 5, P < 0.001; Table 2). The age-specific survival was highest for adult females reared at temperatures of 15 and 18 °C, doubling that of those held at higher temperatures (χ² = 118, df = 5, P < 0.001; Fig. 2).

Fig. 2.

Survival of female D. aberiae reared under 7 constant temperatures. Survival curves followed by the same lowercase letter are not significantly different (pairwise Log-Rank test, P < 0.05).

Effect of Temperature on the Oviposition and Body Size of D. aberiae

Temperature significantly affected oviposition (H = 73.9, df = 5, P < 0.001), with the highest production observed between 15 and 22.5 °C, peaking at 504.5 eggs per female at 22.5 °C (Table 3). Egg production declined at higher temperatures, with females laying 213.93 and 73.7 eggs at 24 and 27 °C, respectively. The proportion of hatched eggs remained above 80% at 22.5 and 24 °C, dropping to around 60% at extreme temperatures. However, these differences were not statistically significant (H = 10.9, df = 5, P = 0.08959).

Table 3.

Mean oviposition (±SE) and proportion of hatched eggs (%) of female D. aberiae reared under 7 constant temperatures

T (°C)	Oviposition (number of eggs)	Hatched eggs (%)
15	439.03 ± 42.7a	77.0
18	490.35 ± 56.5a	65.4
21	434.3 ± 29.3a	63.5
22.5	504.5 ± 46.4a	82.5
24	213.93 ± 31.9 b	84.0
27	73.7 ± 7.56 b	70.0
H	73.9	10.9
P-value	<0.001	0.08959

Means within a column followed by the same lowercase letter are not significantly different among treatments (Dunn’s test, P < 0.05).

Temperature also significantly affected body size, particularly during the adult stage in both preovipositional (length, H = 44.9, df = 5, P < 0.001) and ovipositional phases (length, H = 45.9, df = 5, P < 0.001). The largest body sizes were observed between 15 and 22.5 °C, coinciding with the temperatures that resulted in the highest oviposition, whereas both length and width were smallest at higher temperatures (Table 4).

Table 4.

Length and width (mm) of developmental stages of D. aberiae reared under 6 constant temperatures

T (°C)	Second instar nymph		Third instar nymph		Preovipositional adult female		Ovipositional adult female
	Length	Width	Length	Width	Length	Width	Length	Width
15	0.68 ± 0.01a	0.31 ± 0.01a	1.1 ± 0.02a	0.52 ± 0.01a	1.64 ± 0.04bc	0.81 ± 0.02bc	3.47 ± 0.13a	1.89 ± 0.07a
18	0.69 ± 0.02a	0.33 ± 0.01a	1.1 ± 0.02a	0.52 ± 0.01a	1.83 ± 0.05a	0.9 ± 0.02a	3.43 ± 0.14a	1.8 ± 0.08a
21	0.66 ± 0.01ab	0.3 ± 0.01a	1.03 ± 0.02ab	0.48 ± 0.01ab	1.68 ± 0.03abc	0.81 ± 0.02abc	3.49 ± 0.08a	1.78 ± 0.05a
22.5	0.67 ± 0.01a	0.31 ± 0a	1.05 ± 0.02ab	0.49 ± 0.01ab	1.72 ± 0.03ab	0.83 ± 0.02ab	3.47 ± 0.09a	1.79 ± 0.05a
24	0.64 ± 0.02ab	0.3 ± 0.01a	0.99 ± 0.03 b	0.46 ± 0.02 b	1.54 ± 0.04c	0.8 ± 0.02bc	2.93 ± 0.15ab	1.55 ± 0.09ab
27	0.60 ± 0.01 b	0.3 ± 0a	1.01 ± 0.01 b	0.48 ± 0.01ab	1.49 ± 0.03c	0.73 ± 0.02c	2.22 ± 0.11 b	1.14 ± 0.05 b
H	28.4	18.5	24.9	20.9	44.9	34.3	45.9	45.6
P-value	<0.001	0.00233	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001

Means within a column followed by the same letters are not significantly different among temperature treatments (Dunn’s test, P < 0.05).

Thermal Thresholds and Phenology of D. aberiae

The linear model fit the developmental rates of each life stage, as well as the total development (egg to adult) and complete generation time (egg to egg) of D. aberiae. The high values of R² indicate a good fit of the model, accurately describing the relationship between temperature and the developmental rate of D. aberiae at the linear part of the curve (Table 5). The estimated lower temperature threshold (T_min) ranged from 8.3 °C for third instar nymphs to 11.2 °C for ovipositional females. For total development, T_min was 9.4 °C, with a thermal constant (k) of 561.7 DD above the base temperature being required to complete development from egg to adult (Table 5; Fig. 3). Completing 1 generation (egg to egg) required a T_min of 9.3 °C and a thermal constant (k) of 730 DD (Table 5).

Table 5.

Linear regression equations, lower thermal threshold (T_min) and thermal constant (k) of developmental stages of D. aberiae

Developmental stage	Linear regression model	R  2	T  min (°C)	Thermal constant (degree-days)	Accumulated degree-days
Egg	D  r = −0.06559 + 0.00702T	0.986	9.3	142.4	142.4
First instar	D  r = −0.05728 + 0.00593T	0.9652	9.6	168.5	310.9
Second instar	D  r = −0.06869 + 0.00806T	0.9794	8.6	124	434.9
Third instar	D  r = −0.05731 + 0.00692T	0.9779	8.3	144.5	579.4
Preovipositional female	D  r = −0.05453 + 0.00596T	0.938	9.1	167.7	747.1
Ovipositional female	D  r = −0.06437 + 0.00575T	0.874	11.2	173.9	921
Total development (egg to adult)	D  r = −0.01667 + 0.00178T	0.9902	9.4	561.7	–
Generation (egg to egg)	D  r = −0.01278 + 0.00137T	0.9829	9.3	730	–

Fig. 3.

Relationship between temperature and developmental rate for 1 generation of D. aberiae fitted to a linear (dashed blue line) and a Lactin 2 nonlinear (black) development regression model.

The Logan et al. (1976) nonlinear model, as modified by Lactin et al. (1995) (Lactin 2), provided a good fit to the data, with high R² and low RSS values. The upper temperature threshold coincided for total development and for 1 full generation of D. aberiae, both estimated at 30 °C, whereas the optimal temperatures differed slightly, at 23.1 and 22.9 °C, respectively (Table 6; Fig. 3). The model also estimated a lower temperature threshold at 12.1 °C for the total development and 11.8 °C to complete 1 generation (Table 6; Fig. 3). In the Les Valls region, D. aberiae was estimated to complete 4.4 generations per year between 2018 and 2024.

Table 6.

Estimates of thermal thresholds and parameters of developmental stages of D. aberiae with Lactin 2 nonlinear regression model

Developmental stage	Upper threshold (T  max) (°C)	Optimum threshold (T  o) (°C)	Lower threshold (T  min) (°C)	λ	ΔT	R  2	RSS
Total development (egg to adult)	30	23.1	12.1	−0.0928	6.905	0.9732	0.000011
Generation (egg to egg)	30	22.9	11.8	−0.0907	7.085	0.9647	0.0000081

Discussion

In citrus orchards of central-eastern Valencia, where D. aberiae was first detected, the average annual minimum and maximum temperatures between 2018 and 2024 were 12.2 and 23.5 °C, respectively. Based on the estimated lower and upper developmental thresholds of D. aberiae, these conditions allow the species to survive throughout the year. Both the linear and the nonlinear (Lactin-2) models estimted T_min. Although nonlinear models describe the full thermal response, T_min is typically derived from linear approaches, which perform well within the lower-mid temperature range and provide more reliable estimates of the lower developmental threshold (Uvarov 1931, Davidson 1944). Its lower threshold is comparable to other mealybug species commonly found in Mediterranean citrus, such as Planococcus citri (Green) (8 to 9 °C), Paracoccus burnerae (Brain) (8.7 °C), and Pseudococcus longispinus (Targioni Tozzzetti) (8 °C) (Bodenheimer 1951, Arai 1996, Martínez Ferrer 2003, Johnson and Giliomee 2011, Santa-Cecilia et al. 2011). In contrast, D. aberiae shows a lower upper thermal threshold than these species, all of which tolerate temperatures above 30 °C, and a lower optimal developmental temperature, whereas most mealybugs develop optimally above 27 °C (Bodenheimer 1951, Arai 1996, Martínez Ferrer 2003, Johnson and Giliomee 2011, Santa-Cecilia et al. 2011). This suggests a narrower tolerance to high temperatures and may help explain its distribution and impact within the Valencian Community, where temperatures are typically Mediterranean and the pest is well established and damaging. In areas south of this region, where temperatures are slightly higher and heat events are more frequent, the pest remains less problematic so far. Similarly, in South African citrus orchards, with generally higher temperatures (mean annual minimum and maximum of 16 and 29.7 °C, respectively, for 2010–2018; Shikwambana et al. 2021), D. aberiae rarely reaches damaging levels (Miller and Giliomee 2011), although natural enemies and management practices may also contribute to population regulation.

Degree-day accumulation is closely correlated with phenology and population dynamics. Based on the thermal constant of D. aberiae (730 DD), the mealybug is expected to complete 4 to 5 generations annually in eastern Spain, 3 of which occur between April and August. These estimates are consistent with field observations reported by Martínez-Blay et al. 2018c, who recorded the highest population densities during these months. In comparison, P. citri completes at least 5 generations per year due to its lower thermal constant (562 DD) and greater heat tolerance (Martínez Ferrer 2003), whereas P. longispinus has a higher thermal constant (956.3 DD) and completes only 3 generations (Plata et al. 2023).

Temperature also influences physiological traits such as body size and fecundity. In most oviparous species, these traits vary in response to environmental factors like food availability, population density and temperature (Honěk 1993). In our study, D. aberiae ovipositional females reared at lower temperatures developed larger bodies and exhibited higher fecundity, exceeding that reported for other mealybug species under similar conditions, such as P. citri (186 eggs at 20 °C) (Arai 1996) and P. marginatus (300 eggs at 25 °C) (Amarasekare et al. 2008). A positive relationship between body size and fecundity has been documented in mealybugs (Cocco et al. 2018) and was also observed in D. aberiae. The largest females and highest egg production occurred under moderate spring temperatures (up to 22.5 °C), coinciding with citrus phenological stages most vulnerable to mealybug attack.

Another key factor influencing insect populations is reproductive biology. Our study confirms that D. aberiae reproduces exclusively sexually, with no evidence of facultative parthenogenesis. Although both obligate and facultative parthenogenetic reproduction have been reported in some species (Lloyd 1952, Tabata et al. 2016), obligate sexual reproduction predominates among mealybugs (da Silva et al. 2010, Prasad et al. 2012). Extended longevity in unmated females, documented in other mealybugs (James 1937, Waterworth et al. 2011, Lentini et al. 2018), may increase mating opportunities when males are scarce. In our study, unmated females survived significantly longer than mated females.

The intrinsic rate of increase (r_m) is a key demographic parameter for predicting population growth (Andrewartha and Birch 1954). For D. aberiae, r_m at 24 °C falls within reported ranges for other species at similar temperatures, such as P. citri (0.17 at 25 °C) and P. burneare (0.045 at 25 °C) (Goldasteh et al. 2009, Johnson and Giliomee 2011). However, the r_m of D. aberiae (0.075) is generally lower than that reported for many mealybugs under optimal conditions, likely because the experimental temperature is slightly above the optimum for this species, estimated at ∼23 °C in the present study. Accordingly, a higher r_m would be expected under these conditions. Comparing pest and natural enemy demographic parameters helps assess biological control potential, as higher r_m of natural enemies relative to the pest generally indicates greater effectiveness (Huffaker et al. 1976). Both the introduced parasitoid Anagyrus aberiae (Guerrieri) and coccinellid predators like Cryptolaemus montrouzieri (Mulsant) reduce mealybug populations, particularly in areas difficult to reach with insecticides (Martínez-Blay et al. 2018b, Pérez-Rodríguez et al. 2019, Soto et al. 2021). The r_m estimated for A. aberiae at 24 °C (0.1) (Romero et al. unpublished data) supports its potential as a biocontrol agent. In contrast, optimal r_m values for C. montrouzieri occur at higher temperatures (26 to 28 °C; r_m = 0.07 to 0.14) (Persad and Khan 2002, Elsherif et al. 2010, Saeedi et al. 2015, Özgökçe and Kara 2018), which may explain its greater effectiveness during summer (Pérez-Rodríguez et al. 2019).

The thermal thresholds and constants estimated in this study provide useful tools for improving D. aberiae management. Degree-day accumulation can be used to predict the timing of susceptible developmental stages and optimize control measures. When insecticide applications are necessary, they usually target early nymphal stages, which require 311 to 435 DD. Monitoring degree-day accumulation between peaks of these stages can therefore help determine optimal treatment timing. In addition, the obligate sexual reproduction of D. aberiae highlights the potential of mating disruption as a management strategy, as previously reported for this and other mealybug species (Cocco et al. 2014, Gavara et al. 2024).

Biological control plays a key role in the management of D. aberiae. Because parasitoid development depends on that of their hosts (Hentz et al. 1998), synchrony between host and natural enemy phenology is critical. Larger female mealybugs, typically associated with cooler conditions, are expected to produce more female parasitoid offspring. However, studies on A. aberiae show that lower temperatures (18 °C) result in 79% male offspring, whereas a female-biased sex ratio is achieved at intermediate temperatures (21 to 24 °C) (Romero et al. 2026). These results suggest that host size alone does not determine either the sex ratio or the effectiveness of this natural enemy. Anagyrus aberiae develops within a temperature range similar to that of D. aberiae (Romero et al. 2026) and preferentially parasitizes third-instar nymphs and adult females (Romero et al. unpublished data). Consequently, the peak of adult females (approximately at 561 DD) represents the most suitable period for parasitism. Similarly, early larvae of C. montrouzieri primarily feed on eggs and first-instar mealybugs, whereas older stages attack individuals of any size (Azhar et al. 2012). The development of eggs and first instars between 142 and 311 DD, therefore, represents a suitable period for C. montrouzieri activity.

Delottococcus aberiae can persist year-round under Mediterranean conditions, with optimal development at moderate temperatures and limited heat tolerance, explaining its lower impact in citrus-growing areas with higher temperatures in eastern Spain. The thermal thresholds identified allow prediction of key population dynamics and improve control timing. These results also support biological control, as A. aberiae shows thermal and demographic compatibility with the pest. Overall, these findings provide a basis for a more targeted and effective management of D. aberiae in Mediterranean citrus.

Author Contributions

Elena Romero (Conceptualization [equal], Data curation [equal], Formal analysis [equal], Investigation [equal], Methodology [equal], Writing—original draft [equal]), Josep Sesé (Data curation [equal], Investigation [equal]), Marco Benito (Data curation [equal], Investigation [equal]), Rolando González (Data curation [equal], Investigation [equal]), and Antonia Soto (Conceptualization [equal], Funding acquisition [lead], Methodology [equal], Supervision [equal], Validation [equal], Writing—review & editing [equal])

Funding

Part of the funding for this research was provided by Generalitat Valenciana’s budget program 714.80 through the Conselleria de Agricultura, Desarrollo Rural, Emergencia Climática y Transición Ecológica. Funding for openaccess charge: Universitat Politècnica de València.

Conflicts of Interest

The authors declare no conflict of interest.