Effects of sublethal dose of chlorfluazuron on testicular development and spermatogenesis in the common cutworm, Spodoptera litura

This paper describes the physiological mechanism of action of chlorfluazuron on testicular development and spermatogenesis when sublethal doses (LD₁₀: 1.00 ng/larva or LD₃₀: 3.75 ng/larva) are applied topically to the cuticle of newly moulted fifth instars of the common cutworm Spodoptera litura (F.) (Lepidoptera, Noctuidae). These doses disrupt the growth and development of testes by decreasing the volume and weight of testes and thickness of testes sheath as compared with that of the controls. Sublethal doses of chlorfluazuron also significantly reduce the protein content of the testis, but do not affect the carbohydrate and lipid contents in newly emerged treated males when measured in μg/mg of testis as compared with that of the controls. Additionally, such doses disrupt spermatogenesis by reducing the number and size of eupyrene and apyrene sperm bundles in the testis. Very few or no eupyrene sperm bundles are observed in vas deferens of pre‐ and newly moulted adults compared with controls. This result shows that the transfer of sperm bundles from testes to vas deferens is delayed in treated males. The effects of chlorfluazuron on testicular development and spermatogenesis is thought to be one of the factors responsible for the reduction in fecundity, fertility and hatchability caused by sublethal doses of chlorfluazuron.     

斜纹夜蛾抗药性及其防治对策的研究进展

斜纹夜蛾Spodopteralitura是多种作物的重要害虫 ,对有机氯、有机磷、氨基甲酸酯类、拟除虫菊酯类以及Bt制剂等杀虫剂均产生抗药性。本文对斜纹夜蛾抗药性的形成与发展、抗性机理及其防治对策等方面进行了综述     

Effects of residual chlorfluazuron on haemolymph-borne oviposition-stimulating factors in Spodoptera litura

Acylurea insecticides can have various effects on insect reproduction and may even interfere with haemolymph‐borne oviposition‐stimulating factors (OSFs). This study describes the effects of injecting haemolymph into females of the common cutworm, Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae), the effects of chlorfluazuron on the activities of OSFs, when sublethal doses (LD₁₀: 1.00 ng larva^?1 or LD₃₀: 3.75 ng larva^?1) were applied topically to newly ecdysed fifth instars under laboratory conditions. The haemolymph of moths of varying age, sex, and mating status resulted in the following increases in oviposition rates within the first 24 h, listed in decreasing order: the haemolymph of mated females injected into mated females (56%), mated females into newly emerged virgin females (53%), virgin males into virgin females (49%), and virgin females into mated females (29%), compared with uninjected mated or virgin females. Two factors are involved in the activation of ovipositon: OSF‐I is found in virgin females, whereas OSF‐II may be formed or received by males during mating. By contrast, the haemolymph taken from chlorfluazuron‐treated adults (treated as fifth instars with sublethal doses) injected into females resulted in the following percentage decreases in oviposition rates within the first 24 h, listed in decreasing order: the haemolymph of LD₁₀‐ and LD₃₀‐treated virgin males injected into untreated virgin females (77 and 84%, respectively), LD₁₀ and LD₃₀‐treated virgin females into untreated mated females (70 and 80%, respectively), LD₁₀ and LD₃₀‐treated mated females into untreated mated females (61 and 69%, respectively), LD₁₀ and LD₃₀‐treated mated females into untreated virgin females (59 or 68%, respectively), compared with untreated ones. Hence, residual chlorfluazuron decreases the activities of OSFs by significantly decreasing the oviposition rates. Moreover, virgin females’ or males’ OSFs are more sensitive to chlorfluazuron than those of mated cutworms. Both OSFs are most likely proteins.     

宁波地区斜纹夜蛾幼虫寄生蜂调查研究

2007—2008年的6～11月对宁波地区重要害虫斜纹夜蛾的寄生性天敌进行了调查,发现寄生蜂有18种,主要种类有：马尼拉陡胸茧蜂Snellenius manilae、侧沟茧蜂Microplitis sp、棉铃虫齿唇姬蜂Carnpoletis chlorideae和斑痣悬茧蜂Meteorus pulchricornis,并对主要寄生蜂与斜纹夜蛾幼虫的田间消长动态进行了系统观察。     

Pathogenicity of entomopathogenic fungi to eggs and larvae of Spodoptera litura,the common cutworm

The eggs and second instar larvae of Spodoptera litura were treated with different concentrations of conidial suspensions of six isolates of fungi belonging to five species, Metarhizium anisopliae var. anisopliae (Metschnikov) Sorokin (ARSEF 7487), Lecanicillium muscarium (Petch) Zare & W Gams (ARSEF 7037 and ARSEF 6118), Cordyceps cardinalis (ARSEF 7193), Fusarium lateritium Nees (ARSEF 8291/MTCC 9050) and Aspergillus sp. (ARSEF 8519). In bioassay with unscaled eggs, M. anisopliae, C. cardinalis, F. lateritium and Aspergillus sp. resulted in 100% mortality above 10⁶ conidia/mL. However, with scaled egg masses, the highest hatching rate (56%) was observed with L. muscarium (ARSEF 6118) whereas the lowest hatchings were observed in the case of M. anisopliae followed by L. muscarium (ARSEF 7037), Aspergillus sp., F. lateritium, and C. cardinalis. The larvae were also found susceptible to all isolates in a dose dependent manner. Three promising isolates against larvae, viz. M. anisopliae, F. lateritium and L. muscarium (ARSEF 7037) resulted in average percent mortalities of 88, 89 and 77%, respectively at ≈10⁸ conidia/mL. When both larvae and the leaves (provided as food) were treated with ≈10⁸ conidia/mL, mortality further increased for these isolates. Based on the mortality data and the LC₅₀ values, we suggest that M. anisopliae, F. lateritium and L. muscarium (ARSEF 7037) could be developed as potential biocontrol agents against rice cutworm in IPM programs.     

Sperm transfer during mating, movement of sperm in the female reproductive tract, and sperm precedence in the common cutworm Spodoptera litura

Abstract. Mating behaviour, sperm transfer and sperm precedence were studied in the moth Spodoptera litura (Fabr.) (Lepidoptera: Noctuidae). There existed a rhythmic, diel pattern of mating behaviour of this moth during the scotophase, presumably set with respect to an endogenous activity rhythm. Approximately 30 min after copulation had started, the formation of the corpus of the spermatophore began in the bursa copulatrix of the female moth, but full inflation of the corpus was not completed until 45–60 min after mating had started. The mature spermatophore contained about 350 eupyrene sperm bundles and a large number of individual (loose) apyrene spermatozoa. The mating status and the age of the male insect influenced the number of sperm transferred to the female within the spermatophore, and also affected the consequent fertility. There was no evidence of sperm reflux within the male tract. Within the female, dissociation of eupyrene sperm bundles was evident within the spermatophore less than 15 min after the completion of mating. Spermatozoa began to move from the bursa (in which the spermatophore is lodged) into the spermatheca 30–45 min after the end of the copulation, and the quantity of sperm in the spermatheca reached a plateau at 90 min after mating. Apyrene sperm reached the spermatheca first, followed by eupyrene sperm. Examination of total (apyrene plus eupyrene) sperm in the female tract showed that 86% of mated females received an apparently normal amount of total sperm from the male. Examination of eupyrene sperm alone showed that 81% of matings resulted in an apparently normal transfer of eupyrene sperm. A small proportion (approximately 8%) of the matings, however, were identified as transferring a clearly subnormal quantity of eupyrene sperm to the spermatheca. The eggs produced as a result of such pairings displayed much reduced fertility (about 43%) compared to those from matings confirmed to have transferred normal quantities of sperm, which showed about 92% fertility. This shows that the availability of eupyrene sperm in the spermatheca may be an important constraint on fertility in normal populations of insects. In the laboratory, S. litura females exhibited multiple matings. Of the females, 93% mated, and the mean frequency of mating was 1.69. Mating with a fertile male led to the oviposition of an increased number of eggs. This effect continued even when the female subsequently mated with an infertile male. Displacement of sperm from previous matings is known to be an important factor in the evolution of multiple mating strategies. Our results on sperm utilization by S. litura indicated that after a second mating, the sperm utilized for subsequent fertilization were almost exclusively from the last mating with little mixing. The proportion of eggs fertilized by sperm from the second mating (P₂) was calculated as 0.95, indicating almost complete sperm precedence from the last mating.     

Effects of sublethal doses of chlorfluazuron on insemination and number of inseminated sperm in the common cutworm, Spodoptera litura (F.) (Lepidoptera: Noctuidae)

Sublethal doses (LD₁₀: 1.00 ng larva^?1; or LD₃₀: 3.75 ng larva^?1) of chlorfluazuron were applied topically to the cuticles of newly moulted fifth instars of the common cutworm, Spodoptera litura (F.) (Lepidoptera, Noctuidae). During mating, the treated males transferred spermatophores of a significantly lower weight into females. The weight of spermatophores transferred by LD₃₀‐treated males was significantly lower than the weight of spermatophores transferred by LD₁₀‐treated males, which was in turn significantly lower than the weight of spermatophores transferred by untreated males. The transfer of spermatophores was delayed by 5–15 min in LD₁₀‐ and LD₃₀‐treated males. However, mating duration was not affected by chlorfluazuron treatment. The transfer of spermatozoa was also delayed by 5–10 min in LD₁₀‐ and LD₃₀‐treated females. Polygynous male adults mated an average of ten times during their lifespan of 11–13 days when paired every day with a new virgin female of the same age. The number of matings per polygynous male was not affected by chlorfluazuron treatment, but the first mating was delayed by 1 day. The number of inseminated sperm found in the spermatophore averaged 10.3 (± 2.1) × 10⁵ over the lifespan of a male, in which the number of eupyrene sperm was 5.4 (± 1.1) × 10⁵. The number of inseminated eupyrene sperm decreased by 66% and 88%, respectively, in LD₁₀‐ and LD₃₀‐treated males. No significant reduction in the number of inseminated eupyrene sperm was observed when females were treated with LD₁₀ or LD₃₀ doses, nor was there a significant reduction when both sexes were treated with the LD₁₀ or LD₃₀ doses relative to treatment of males with the same doses. The ratio of inseminated eupyrene to apyrene sperm was not affected by chlorfluazuron treatment.     

幼虫密度对斜纹夜蛾抗病能力的影响

为了阐明斜纹夜蛾Spodoptera litura Fabricius幼虫密度对其抗病能力的影响,在室内条件下(温度23℃±1℃,相对湿度75%)对不同幼虫密度(1、2、5、10、15头/皿(直径为12cm))饲养的斜纹夜蛾幼虫抵抗斜纹夜蛾核型多角体病毒侵染的能力及其免疫指标进行了研究。结果表明:幼虫密度对斜纹夜蛾幼虫接种核型多角体病毒后的存活率、存活时间及血淋巴酚氧化酶活性影响显著。随着幼虫密度的增加,接种核型多角体病毒后幼虫的存活率降低,存活时间缩短。当幼虫密度达到15头/皿时,幼虫存活率显著低于其它幼虫密度。不同幼虫密度幼虫的存活时间以1头/皿的最高,15头/皿的最低,且二者之间差异显著。幼虫血淋巴中酚氧化酶活性随幼虫密度的增加而明显降低,当幼虫密度达到5头/皿时,幼虫酚氧化酶活性显著低于1头/皿的。另外,幼虫溶菌酶活性和血细胞总数受幼虫密度影响不显著。不同密度幼虫抗病性的变化与其血淋巴中酚氧化酶活性的变化趋势较为一致。所以斜纹夜蛾幼虫抗病能力的降低可能与幼虫酚氧化酶活性的下降有关。因此,幼虫密度是影响斜纹夜蛾幼虫抗病性变化的重要因子。     

Cloning and expression analysis of six small heat shock protein genes in the common cutworm, Spodoptera litura

Small heat shock proteins (sHsps) are probably the most diverse in structure and function among the various superfamilies of stress proteins. To explore the diverse functions of insect sHsps, six sHsp cDNAs were cloned from the midgut cDNA library of Spodoptera litura, and a phylogenetic tree was constructed based on the conserved α-crystalline domains. The expression patterns in different developmental stages and tissues, as well as in response to both thermal and 20-hydroxyecdysone (20E) induction, were studied by real-time quantitative PCR. Based on sequence characteristics and phylogenetic relationships, the six SlHsps were classified into three independent groups: BmHsp20.4 like proteins (SlHsp19.7, 20.4, 20.7, 20.8), BmHsp26.6 like protein (SlHsp20), and BmHsp21.4 like protein (SlHsp21.4). All the SlHsps showed highest expression in the Malpighian tubules. The four BmHsp20.4 like protein genes were up-regulated by thermal stress and showed expression variation with development. SlHsp20 exhibited lower expression levels in both egg and larval stages than in pupal and adult stages. SlHsp21.4 retained a constant expression level during all life stages. The expression of both SlHsp20.4 and SlHsp20.8 was significantly up-regulated by 20E. These results indicate that sHsps play diverse functions in S. litura: the BmHsp20.4 like proteins are involved in both thermal adaptation and development; SlHsp20 does not respond to temperature stress but possibly plays a role in metamorphosis; SlHsp21.4 may have no direct relationship with either thermal response or development.     

Overseas migration of the common cutworm, Spodoptera litura (Lepidoptera: Noctuidae), from May to mid-July in East Asia

Spodoptera litura (Fabricius) males were monitored with sex-pheromone traps from spring to early summer in 1999, 2002, 2003, 2009, and 2010 in four regions in East Asia: western Japan, South Korea, China, and Taiwan. The backward trajectory analysis was conducted to investigate whether any air current arrived over the monitoring sites around the dates when distinguished increases in catches of >100 insects per day occurred. The catch data showed abrupt catch increases. Some of the catch increases coincidentally appeared among monitoring sites several hundreds of kilometers apart. These catch increases were found to coincide with the arrival of southwesterly air currents within 24–36 h from southern China and/or Taiwan. Other evidence related to the overseas migration are discussed in detail. The results of this and previous studies suggest that S. litura males immigrate into western Japan and South Korea in spring to early summer using air currents from southern China and/or Taiwan.     

Analysis of expressed sequence tags and characterization of a novel gene, Slmg7, in the midgut of the common cutworm, Spodoptera litura

Background

Out of total 3,081 assembled expressed sequence tags (ESTs) sequences representing 6,815 high-quality ESTs identified in three cDNA libraries constructed with RNA isolated from the midgut of Spodoptera litura, 1,039 ESTs showed significant hits and 1,107 ESTs did not show significant hits in BLAST searches. It is of interest to clarify whether or not these ESTs that did not show hits function in S. Litura.

Results

Twenty “no-hit” ESTs containing at least one putative open reading frame were selected for further expression analysis. The results from northern blot analysis showed that six of the selected ESTs are expressed in the larval midgut of this insect at different levels, suggesting that these ESTs represent true mRNA products, whereas the other 14 ESTs could not be detected. Homologues of the four larval midgut-predominant genes (Slmg2, Slmg7, Slmg9 and Slmg17) were detected in the genomes of other lepidopteran insects but not in Drosophila melanogaster. A novel gene, Slmg7, is expressed at a high level specifically in the midgut during each of the larval stages. Slmg7 is a single copy gene and encodes a 143-amino acids protein. The SLMG7 protein was localized to the cytoplasm of Spli-221 cells.

Conclusions

Six ESTs from the no hit list are transcribed into mRNA and are mainly expressed in the midgut of S. litura. Slmg7 is a novel gene that is localized to the cytoplasm.     

An extremely low frequency magnetic field increases unconditioned larval movement of the common cutworm,Spodoptera litura: A novel model for a magnetoreceptive neurobehavioral study

One of the most interesting features of magnetic field (MF) responsiveness in animals is the signal transduction mechanism from sensing MF to behavior. To develop a model system for understanding the process, we performed a preliminary behavioral assay using an insect, the common cutworm, Spodoptera litura (Fabricius). In contrast to sham exposure, a continuous 60 Hz oscillating MF (0.2 mT) induced a significant increase in movement in third instar larvae: the MF‐induced movement was longer in total duration and had an earlier onset than movement from sham exposure. Not surprisingly, the movement duration of the group test was notably higher than that of the individual test, suggesting that the individual test is more favorable for assessing the MF effect on movement. Considering the simplicity of the experimental operation and obvious non‐conditioned responsiveness to the MF, this model might be suitable for studying magnetoreception and following the signal transduction mechanism between neurons and behavior.     

Control of reproduction and a survival cost to mating in female Bicyclus anynana butterflies

Abstract.  1. Given that sexual conflict is all pervasive, investigating potential costs to mating and the control of female reproduction are important issues.
2. Here, female reproductive output and longevity are investigated in relation to mating status (virgin vs. once-mated females) and host-plant availability in the tropical butterfly Bicyclus anynana .
3. Both factors significantly affected realised fecundity in a manner demonstrating that female reproduction is not entirely under control of the female herself, but is rather subject to additive contributions of the female, her male partner, and intrinsic physiological processes. As evident from the deposition of significant egg numbers by virgin, host-deprived females, B. anynana is effectively unable to completely inhibit oogenesis.
4. Mated females suffered a reduction in adult life span, which cannot be explained as a side-effect of variation in egg size, lifetime, or early fecundity.
5. Such detrimental effects of mating per se are indicative of the cooperation–conflict balance between sexes being shifted towards conflict in B. anynana .     

Effects of male genetic contribution and paternal investment to egg and hatchling size in the cricket,Gryllus firmus

Egg provisions represent the complete energy supply of oviparous organisms from fertilization until hatching, and egg size is generally correlated with initial offspring size and a suite of other early fitness related traits. Since egg size is determined by the mother, little attention has been given to potential sources of paternal effects on either egg size or initial offspring size. This study considers two processes by which the sire can affect the egg size and/or initial body size of his offspring, in the multiply mated cricket, Gryllus firmus. The first is a paternal genetic effect, whereby differences in offspring genotype result in differences in the efficiency of metabolising available resources. The second is a paternal environmental effect, whereby the quality or size of paternal investment varies among male phenotypes and this is correlated with the size of eggs the females subsequently produce. Using a one-locus two-allele recessive mutation for pale eye colour as a marker, a mating experiment was designed which enabled the discrimination between eggs fertilized by two males mated simultaneously to a single female. The results of this experiment suggest that sire effects on egg and initial body size occur through both processes. Eggs fertilised by the two males were significantly different at day ten of development, suggesting that the genetic contribution of the sire is affecting embryonic body size. Further, a negative correlation was found between the head size of the pale eyed male and the size of all the eggs that the female laid, suggesting an effect of male size of the amount of nutrients the female receives from her mates. The results of this study suggest that paternal effects may be both more common and more profound than previously thought, and that studies examining early fitness traits that are correlated to egg size or initial body size, in oviparous animals, should consider the possibility and importance of the paternal contribution.     

Behavioral responses in prey location of the predatory stink bug, Eocanthecona furcellata, to chemical cues in the larvae of Spodoptera litura

Behavioral responses of the predatory stink bug Eocanthecona furcellata (Wolff) (Heteroptera: Pentatomidae) towards Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) larvae were observed in olfactometers. In single-and dual-tube olfactometers, adult bugs were observed to approach and stretch their proboscis toward the odor source. These types of behavior were also observed toward the control odor source in the dual-tube olfactometer. However, the frequency of these phases toward the control odor was significantly lower than that toward the odors from intact or dead larvae. The solvent extracts of larvae elicited these similar behavioral phases significantly more frequently and for significantly longer duration than the control. In a linear track olfactometer, E. furcellata nymphs were observed to be attracted to the fraction eluted with hexane from silica gel chromatography but proboscis-protruding behavior was not observed. The other fraction, 15%-ether-in-hexane from silica gel chromatography, elicited the proboscis-protruding behavior by a simple behavioral assay using a Pasteur pipette. These findings therefore indicate the presence of at least two different chemical cues in prey location.     

Impact of short‐term exposure to low subzero temperatures on egg hatch in the hemlock looper,Lambdina fiscellaria

The frequency of extreme events, such as cold spells, is expected to increase under global warming. Therefore, the ability of insects to survive rapid changes in temperature is an important aspect to investigate in current population ecology. The hemlock looper (HL), Lambdina fiscellaria (Guenée) (Lepidoptera: Geometridae), a defoliator of boreal balsam fir forests in eastern Canada, overwinters at the egg stage on tree trunks and branches where eggs can be exposed to very low subzero air temperatures. Using eggs from the island of Newfoundland (NL) and Quebec mainland (QC), we undertook field and laboratory experiments to determine: (1) their supercooling point (SCP) in mid‐January and mid‐February; (2) overwintering mortality; (3) cold tolerance to various combinations of subzero temperatures (?25, ?30, ?33, ?35, or ?37 °C) and exposure durations (2, 4, 8, 12, or 16 h); and (4) potential causes of death at subzero temperatures above the SCP. Regardless of population or sampling date, eggs supercooled on average at ?40.1 °C. In the field, 59% of eggs from either population that overwintered in Sainte‐Foy (QC) and Corner Brook (NL) hatched successfully, whereas none did in Armagh (QC) or Epaule (QC). In the laboratory, 50% of eggs survived after 4 h at ?34.4 °C or after 14 h at ?32.9 °C. In contrast, regardless of exposure duration, >50% of eggs hatched at temperatures ≥?33 °C, but <50% did so at ≤?35 °C, suggesting high pre‐freeze mortality. However, when eggs were attached to thermocouples and exposed to temperatures ranging from ?25 to ?37 °C for 16 h, 69% froze at temperatures of ?35 to ?37 °C, but only 2% did at ?25 or ?30 °C. Time to freeze decreased as subzero temperatures declined, and this was more evident in island eggs than in mainland eggs. Overall, eggs can freeze after a brief exposure to subzero temperatures higher than the standard SCP, and are thus highly vulnerable to cold spells.     

Experimentally reducing clutch size reveals a fixed upper limit to egg size in snakes, evidence from the king ratsnake, Elaphe carinata

Snakes are free of the pelvic girdle's constraint on maximum offspring size, and therefore present an opportunity to investigate the upper limit to offspring size without the limit imposed by the pelvic girdle dimension. We used the king ratsnake (Elaphe carinata) as a model animal to examine whether follicle ablation may result in enlargement of egg size in snakes and, if so, whether there is a fixed upper limit to egg size. Females with small sized yolking follicles were assigned to three manipulated, one sham-manipulated and one control treatments in mid-May, and two, four or six yolking follicles in the manipulated females were then ablated. Females undergoing follicle ablation produced fewer, but larger as well as more elongated, eggs than control females primarily by increasing egg length. This finding suggests that follicle ablation may result in enlargement of egg size in E. carinata. Mean values for egg width remained almost unchanged across the five treatments, suggesting that egg width is more likely to be shaped by the morphological feature of the oviduct. Clutch mass dropped dramatically in four- and six-follicle ablated females. The function describing the relationship between size and number of eggs reveals that egg size increases with decreasing clutch size at an ever-decreasing rate, with the tangent slope of the function for the six-follicle ablation treatment being -0.04. According to the function describing instantaneous variation in tangent slope, the maximum value of tangent slope should converge towards zero. This result provides evidence that there is a fixed upper limit to egg size in E. carinata.     

A limit on the extent to which increased egg size can compensate for a poor postnatal environment revealed experimentally in the burying beetle,Nicrophorus vespilloides

It is often assumed that there is a positive relationship between egg size and offspring fitness. However, recent studies have suggested that egg size has a greater effect on offspring fitness in low‐quality environments than in high‐quality environments. Such observations suggest that mothers may compensate for poor posthatching environments by increasing egg size. In this paper we test whether there is a limit on the extent to which increased egg size can compensate for the removal of posthatching parental care in the burying beetle, Nicrophorus vespilloides. Previous experiments with N. vespilloides suggest that an increased egg size can compensate for a relatively poor environment after hatching. Here, we phenotypically engineered female N. vespilloides to produce large or small eggs by varying the amount of time they were allowed to feed on the carcass as larvae. We then tested whether differences between these groups in egg size translated into differences in larval performance in a harsh postnatal environment that excluded parental care. We found that females engineered to produce large eggs did not have higher breeding success, and nor did they produce larger larvae than females engineered to produce small eggs. These results suggest that there is a limit on the extent to which increased maternal investment in egg size can compensate for a poor posthatching environment. We discuss the implication of our results for a recent study showing that experimental N. vespilloides populations can adapt rapidly to the absence of posthatching parental care.     

Sex allocation in Telenomus busseolae, a solitary parasitoid of concealed eggs: the influence of host patch size

Telenomus busseolae Gahan (Hymenoptera: Scelionidae) is an important egg parasitoid of noctuid stem borers of gramineous crops, attacking egg masses of Sesamia spp. Under natural conditions, and whatever the host species attacked, these egg masses are generally concealed under the leaf sheaths or other narrow spaces, and vary greatly in size. In the work presented here, the influence of host patch size (4, 8, 16, 32, 64, or 128 eggs per mass) on the sex ratio and sex sequence pattern of ovipositing T. busseolae was investigated in the laboratory using Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae) as host. The results are similar to those described for other parasitoids of aggregated hosts, and are in accordance with the Local Mate Competition model. With increasing egg mass size, the overall sex ratio (proportion of males) decreased, although additional males were laid at the end of the sequence in the larger masses (64 and 128 eggs). Sex sequence pattern always followed a males‐first strategy, i.e., with a higher proportion of males at the beginning, but the whole sex ratio sequence was influenced by the size of the egg mass. Such results in a parasitoid of concealed eggs are compared to those observed in parasitoids of exposed eggs and discussed in terms of parasitoid reproductive strategies and evolutionary adaptations.