Survival of a specialist natural enemy experiencing resource competition with an omnivorous predator when sharing the invasive prey Tuta absoluta

Can specialist natural enemies persist in ecosystems when competing with omnivorous natural enemies for their shared prey? The consequences of omnivory have been studied theoretically, but empirical studies are still lacking. Omnivory is nevertheless common in nature and omnivorous predators coexist with specialists in many ecosystems, even when they are intraguild predators. This type of association is also common in agroecosystems in which biological control strategies are used. Our study provides an example of the outcome of such an association in the context of biological control of the invasive pest Tuta absoluta (Lepidoptera) in a tomato agroecosystem. The two natural enemies involved, that is, a specialist (Stenomesius japonicus (Hymenoptera) parasitoid) and an omnivore (Macrolophus pygmaeus (Hemiptera) predator), were able to coexist for 3 months in our experimental cages in the absence of metacommunity mechanisms (i.e., emigration and recolonization), contrary to theoretical expectations. However, they negatively affected each other's population dynamics. We found that spatial resource segregation was not a mechanism that promoted their coexistence. Regarding pest control, the specialist and omnivorous natural enemies were found to exhibit complementary functional traits, leading to the best control when together. Mechanisms that may have promoted the coexistence of the two species as well as consequences with regard to the inoculative biological control program are discussed.     

重大潜在入侵害虫番茄潜叶蛾的SS-CoI快速检测技术

【背景】番茄潜叶蛾是源自南美洲的一种最具毁灭性的世界性入侵害虫,其适生区范围广,且与其他潜叶类昆虫难以准确、快速识别。【方法】以田间常见的其他6种潜叶类害虫为对照,采用基于mtDNACOl基因的种特异性SS—COI方法,研究番茄潜叶蛾快速分子检测技术。【结果】种特异性检测结果显示,SS-COl引物只对番茄潜叶蛾mtDNA具有扩增能力,对美洲斑潜蝇、南美斑潜蝇、三叶草斑潜蝇、葱斑潜蝇、豌豆潜叶蝇、桃潜叶蛾等其他潜叶类害虫不具有扩增效果。该引物不仅对成虫和单粒卵具有很好的扩增效能,对单根触角、头部、胸部、腹部以及翅和足等成虫残体亦具有同样的扩增能力,其最低检出阈值为312．5Pg·μL^-1。【结论与意义】该方法在有效防范番茄潜叶蛾侵入我国,保障农业生产安全和生态环境安全中意义重大。     

Development of sequence amplified characterized region (SCAR) markers of helicoverpa armigera: a new polymerase chain reaction-based technique for predator gut analysis

A method is described for the development of DNA markers for detection of Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) in predator gut analysis, based on sequence characterized amplified regions (SCARs) derived from a randomly amplified polymorphic DNA (RAPD) band. A 1200-bp DNA fragment of H. armigera, absent in the predator band pattern and in other closely related prey species, was identified by RAPD analysis. This fragment was cloned and its extremes sequenced to design extended strand-specific 20-mer oligonucleotide primers. Three pairs of SCAR primers, which amplified three different DNA fragments, were used to study the effect of fragment length on detection of prey in the predator gut. Using the pair of primers that amplified the longest fragment of H. armigera DNA, a single band of 1100 bp was obtained, but its detection was not possible in the predator gut. Detection of the ingested prey was possible with the other two pairs of SCAR primers, obtaining bands of 600 and 254 bp, respectively. Detection of H. armigera DNA in the gut of the predator Dicyphus tamaninii was evaluated immediately after ingestion (t = 0) and after 4 h. Detection of H. armigera DNA after 4 h was only possible using the pair of primers that amplified the shortest fragment (254 bp). The test for specificity, using these last pair of primers, showed that H. armigera was the only species detected. The detection threshold was defined at a 1:8192 dilution of a H. armigera whole egg in all samples.     

Response of the zoophytophagous predators Macrolophus pygmaeus and Nesidiocoris tenuis to volatiles of uninfested plants and to plants infested by prey or conspecifics

Knowledge about the orientation mechanisms used by two important predaceous mirids (Macrolophus pygmaeus Rambour and Nesidiocoris tenuis (Reuter)) in finding their prey (whitefly Bemisia tabaci (Gennadius) and the tomato borer Tuta absoluta (Meyrick)) is limited. In a Y-tube olfactometer, we tested the behavioral responses of naïve and experienced predators to uninfested plants, herbivore-induced plant volatiles (HIPVs) from plants infested with T. absoluta and/or B. tabaci, the sex pheromone of T. absoluta, and volatiles produced by plants injured by the predators. Nesidiocoris tenuis responds to volatiles produced by uninfested plants only after experience with the plant, whereas naïve and experienced M. pygmaeus show positive chemotaxis. Both predators are attracted to volatiles from prey-infested plants, and we provide the first evidence that experience affects this response in M. pygmaeus. Infestation of the same plant by both prey species elicited similar responses by the two predators as plants infested by either herbivore singly. Neither predator responded to sex pheromones of T. absoluta. Macrolophus pygmaeus avoided plants injured by conspecifics, while N. tenuis females were attracted by such plants. The implications of these results for augmentative biological control are discussed.     

Detection of secondary predation by PCR analyses of the gut contents of invertebrate generalist predators

Predation by generalist predators is difficult to study in the field because of the complex effects of positive and negative interactions within and between predator species and guilds. Predation can be monitored by molecular means, through identification of prey DNA within predators. However, polymerase chain reaction (PCR) amplification of prey DNA from predators cannot discriminate between primary and secondary predation (hyperpredation), in which one predator feeds on another that has recently eaten the target prey. Here we quantify, for the first time, the potential error caused by detection of prey DNA following secondary predation, using an aphid-spider-carabid model. First, the aphid Sitobion avenae was fed to the spider Tenuiphantes tenuis and the carabid Pterostichus melanarius, and the postconsumption detection periods, for prey DNA within predators, were calculated. Aphids were then fed to spiders and the spiders to carabids. Aphid DNA was detected in the predators using primers that amplified 245- and 110-bp fragments of the mitochondrial cytochrome oxidase I gene. Fragment size and predator sex had no significant effect on detection periods. Secondary predation could be detected for up to 8 h, when carabids fed on spiders immediately after the latter had consumed aphids. Beetles tested positive up to 4 h after eating spiders that had digested their aphid prey for 4 h. Clearly, the extreme sensitivity of PCR makes detection of secondary predation more likely, and the only reliable answer in future may be to use PCR to identify, in parallel, instances of intraguild predation.     

Discrimination of the closely related biocontrol agents Macrolophus melanotoma (Hemiptera: Miridae) and M. pygmaeus using mitochondrial DNA analysis

The separation of the closely related predatory species Macrolophus melanotoma Costa (= M. caliginosus Wagner) and Macrolophus pygmaeus (Rambur) based exclusively on the different colour pattern of the first antennal segment (white central band in M. melanotoma and entirely black in M. pygmaeus) is rather precarious and their taxonomic status is still in doubt. In the present study their taxonomic status was evaluated by DNA confirmatory analysis and hybridization experiments between M. pygmaeus and a Macrolophus taxon, resembling M. melanotoma, with a first antennal segment entirely black or with a white central band collected from Dittrichia viscosa (L.) W. Greuter (Asteraceae) in southern Greece. Adult females from Dittrichia plants hybridized with males of M. pygmaeus and vice versa did not produce viable eggs. The Macrolophus species from Dittrichia irrespective of the first antennal segment coloration differed from M. pygmaeusin digestive patterns generated by AseI, XbaI, and MseI on 16S rRNA and in RAPD profiles produced by the primers OPA-18 and OPA-20. These results demonstrate that on Dittrichia plants there is a distinct dimorphic taxon, M. melanotoma, as it is the only species of the genus Macrolophus bearing a first antennal segment with a central white band. Given the limitation of the coloration pattern, the mtDNA genetic markers are the appropriate method for the identification of M. melanotomaand M. pygmaeus.     

Nectarivory by the plant-tissue feeding predator Macrolophus pygmaeus Rambur (Heteroptera: Miridae): Nutritional redundancy or nutritional benefit?

Most predators and parasitoids feed on plant-provided food (nectar, pollen) or engage in herbivory during at least part of their life stages. Plant feeding by these insects plays an important role in driving predator-herbivore dynamics. Thus, understanding the effects of plant feeding on omnivores could be an important element in improving biological control strategies. The mirid Macrolophus pygmaeus is an omnivorous heteropteran predator of whitefly and other pests. Unlike other predators that need to seek out accessible nectar to meet their carbohydrate requirements, mirid bugs can access the plant's carbohydrate resources by feeding directly on plant tissues. Leaf and stem feeding could be seen as a nutritional surrogate that allows mirids to become independent of nectar availability. However, to date feeding experiments have not yet considered nectar feeding by these mirid predators. In this study we demonstrate that M. pygmaeus survival is prolonged on broad bean plants featuring extrafloral nectar as compared to broad bean with extrafloral nectaries removed, irrespective of the presence of cattail pollen. Survival on extrafloral nectar was comparable to the survival by individuals kept on broad bean provided with eggs of Ephestia kuehniella as prey. Also, a greater proportion of mirid females laid eggs when extrafloral nectar was available as compared to those confined on nectariless plants without supplemental food.     

Intraguild predation and sublethal interactions between two zoophytophagous mirids,Macrolophus pygmaeus and Nesidiocoris tenuis

The omnivorous predators Macrolophus pygmaeus and Nesidiocoris tenuis (Hemiptera: Miridae) are important biological control agents of pests on tomato crops. In this study, potential intraguild predation (IGP) interactions between the two species were investigated on tomato. We examined: (a) the within plant distribution of both species in the field, (b) the within plant distribution of each predatory species when co-occurred at high densities on tomato caged plants, (c) their behavioral interactions when enclosed in experimental arenas and (d) the development young and old nymphs of M. pygmaeus when enclosed together with N. tenuis adults. Results revealed that the two predators showed a different distribution pattern on the plants, with N. tenuis exploiting mostly the upper part, whereas M. pygmaeus were mostly observed on the 5th to the 7th leaf from the top. However, when the predators co-occurred, N. tenuis or M. pygmaeus individuals were recorded with increased numbers on the lower or the higher part of the plant, respectively. In the presence of N. tenuis adult young nymphs of M. pygmaeus completed their development to the adult stage, when alternative prey (lepidopteran eggs) was present on the plant, however failed to reach adulthood in the absence of alternative prey. A high percentage of the dead nymphs found with their body fluids totally sucked indicating predation by N. tenuis. However, large 4th instar nymphs of M. pygmaeus were much less vulnerable to N. tenuis than younger. The behavior of N. tenuis was affected by the presence of M. pygmaeus, but at a rate similar to that when two individuals of N. tenuis were enclosed together. Contacts between the predators were recorded in a similar frequency in mono- and heterospecific treatments, whereas aggressive behavior was not observed. This study shows that intraguild interactions between M. pygmaeus and N. tenuis occur but are not intensive. The potential implications of the outcomes for biological control are discussed.     

Dispersal of Podisus nigrispinus (Het., Pentatomidae) nymphs preying on tomato leafminer: effect of predator release time, density and satiation level

Abstract:   The dispersal potential of nymphal stinkbug, Podisus nigrispinus (Dallas, 1851) (Het., Pentatomidae) preying on larvae of the tomato leafminer Tuta absoluta (Meyrick, 1917) (Lep., Gelechiidae) was studied in an open-sided greenhouse. The parameters investigated were (1) the density of nymphs released per plant (one or five); (2) the release time (0600 and 1800 h); and (3) predator satiation level (satiated and 24-h-starved nymphs). Tomato plants were infested with larvae of the tomato leafminer (one third or fourth instar per leaf). The evaluations started 30 min after the release of predators second instar and hourly evaluations were carried out over 36 and 24 h for release times of 0600 and 1800 h, respectively. Starved nymphs released in the morning, either alone or in groups of five, dispersed faster than satiated nymphs. All of the starved individuals had left the plant by the end of the observation period, whereas 25 and 36% of the satiated nymphs released alone and in groups, respectively, stayed on the plant until the end of the observation period. Both satiated and starved nymphs showed slower rates of dispersal when released at 1800 h. Satiated nymphs delayed prey attack up to 9 h, whereas starved individuals started to attack T. absoluta caterpillars 1 h after their release at both densities. Our findings suggest more effective biological control of T. absoluta is possible with the release of second instar nymphs of P. nigrispinus when starved for 24 h prior to release and then released either in the morning or in the evening.     

Development of a real-time PCR assay for the detection of Cladosporium fulvum in tomato leaves

Aims: The aim of this study was to develop a sensitive real-time polymerase chain reaction (PCR) assay for the rapid detection of Cladosporium fulvum in tomato leaves. Methods and Results: Three PCR primer pairs were designed based on the nucleotide sequences of: (i) the internal transcribed spacer regions of ribosomal RNA; (ii) a microsatellite region amplified by the microsatellite primer M13; and (iii) the β-tubulin gene of C. fulvum. Each primer pair amplified the expected target DNA fragment from geographically diverse isolates of C. fulvum. No PCR products were amplified with these primer pairs from DNA of other fungal species. Among the three pairs of primers, the primer pair CfF1/CfR1 developed based on the microsatellite region was the most sensitive. Using this sensitive primer pair, a real-time PCR assay was developed to detect early infection of C. fulvum in tomato leaves. Significance and Impact of the Study: DNA regions amplified by the microsatellite primer M13 have a high potential for developing highly sensitive species-specific PCR primers for the detection of phytopathogenic fungi. The real-time PCR assay developed in this study is useful in monitoring early infection of C. fulvum, and can help growers make timely decisions on fungicide application.     

Infiltration by alien predators into invertebrate food webs in Hawaii: a molecular approach

Abstract Alien invertebrate predators have been introduced to Hawaii to control pests, particularly in lowland areas where most crops are grown. We developed techniques for assessing the impact of these predators on native food webs in relatively pristine upland areas where, it was hypothesized, few lowland predators might be found. Predator densities were assessed along transects within the Alakaii Swamp on Kaua'i. The most numerous alien biocontrol agents found were Halmus chalybeus (Coccinellidae), a species known to feed on Lepidoptera eggs. Laboratory experiments were conducted using two genera of endemic Lepidoptera, Scotorythra and Eupithecia (Geometridae), that are of considerable conservation value, the former because of its recent speciation across Hawaii, the latter for its unique predatory larvae. Techniques were developed for detecting Lepidoptera DNA within the guts of alien predators using prey-specific PCR primers. General primers amplified fragments of the mitochondrial cytochrome oxidase I gene from beetles and Lepidoptera. The sequences were aligned and used successfully to design target-specific primers for general detection of the remains of Geometridae and for particular species, including Scotorythra rara and Eupithecia monticolans. DNA fragments amplified were short [140-170 base pairs (bp)], optimizing detection periods following prey ingestion. Trials using the introduced biocontrol agent Curinus coeruleus (Coccinellidae) demonstrated detection of Lepidoptera DNA fragments = 151 bp in 85-100% of beetles after 24 h digestion of an early instar larva. This study provides a framework for future use of molecular gut analysis in arthropod conservation ecology and food web research with considerable potential for quantifying threats to endemic species in Hawaii and elsewhere.     

Adaptation of indigenous larval parasitoids to Tuta absoluta (Lepidoptera: Gelechiidae) in Italy

Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) is a serious threat to tomato (Solanum lycopersicum L.) crops in South America. In Europe, after its first detection in Spain in 2006, it rapidly spread through the Mediterranean basin, reaching Italy 2 yr later. The aim of our work was to find indigenous effective biological control agents and to evaluate their potential role in the control of larval populations of T. absoluta in controlled conditions. Nine species of larval parasitoids emerged from field-collected tomato leaves infested by T. absoluta. The most abundant, Necremnus near artynes (Walker) and Necremnus near tidius (Walker) (Hymenoptera: Eulophidae), were tested in laboratory parasitism trials. Furthermore, because the species N. artynes and N. tidius are each reported in literature as an ectoparasitoid of Cosmopterix pulchrimella Chambers (Lepidoptera: Cosmopterigidae) on upright pellitory plants, olfactometer bioassays were performed to assess the response of our parasitoids to the odors of tomato and pellitory leaves infested by T absoluta and C. pulchrimella, respectively, compared with healthy ones. Both Necremnus species showed good adaptation to the invasive pest, and we observed a high larval mortality of T. absoluta because of host feeding and parasitism. Even olfactory responses highlighted a preference of both wasps for tomato plants infested by the exotic pest. These preliminary results demonstrated a high suitability of these indigenous natural enemies for controlling T. absoluta. Further investigations are needed to confirm their role as potential biological agents in commercial tomato plantations.     

Natural Control of Helicoverpa armigera in Sunflower: Assessment of the Role of Predation

In three field experiments in Kenya , the seasonal population trend of Helicoverpa armigera (Hubner) (= Heliothis armigera) in sunflower was followed in plots from which predators were excluded and in plots from which predators were not excluded . In experiment 1 , complete exclusion of crawling predators (predominantly Pheidole spp . of ants occurring at densities of 25 per plant) resulted in H. armigera densities 3 - 5 times higher than in plots where ants were not excluded . Pheidole sp . had more impact on young H. armigera larvae (instars 2 - 3) than on older larvae (instars 4 - 6) . The results of experiment 2 were less striking , because H. armigera infestation was low , and ant densities were moderate . Here , Myrmicaria spp . and Camponotus spp . were the predominant ants . Exclusion of ants resulted in a 1 . 8 - fold increase in densities of large H. armigera instars . In experiment 3 , the impact of predators on H. armigera was studied under three conditions: exclusion of crawling predators , exclusion of both crawling and flying predators , and the control where no predators were excluded . To evaluate the role of predation in total mortality , the recruitment of H. armigera larvae was determined with Southwood and Jepson ' s graphical method , and recruitment of newly laid eggs was directly measured on trap plants . Because it was difficult to detect older eggs due to colour change , the direct measurement of egg recruitment was superior to the graphical method . Ants and Anthocoridae were the dominant predators . M ortality from egg to older larvae (instars 4 - 6) was 73 - 78% . Exclusion of ants and anthocorids did not affect the densities of H. armigera larvae . Anthocorid predators increased only after the main oviposi tion peak of H. armigera and , therefore , their exclusion had little impact on the pest . Ant density was considerably lower than in experiment 1 , and did not significantly suppress H. armigera.     

Development of species-specific PCR primer sets for the detection of Leptospira

Spirochetes of the genus Leptospira infect animals and humans and are the causative agents for the emerging infectious disease leptospirosis. Rapid and simple assays for the identification of individual Leptospira species are currently not available. For identification of individual Leptospira species, PCR primers that detect the ompL1 gene sequence for the majority of pathogenic leptospires were developed in this study. The primer pairs detect Leptospira interrogans, Leptospira borgpetersenii, Leptospira kirschneri, Leptospira santarosai, Leptospira weilii and Leptospira noguchii, without cross-reacting with other Leptospira species. The development of the primers revealed a divergence of the ompL1 gene within L. interrogans, splitting this species into two separate groups. The species-specific primers will be especially useful in epidemiological studies and disease outbreak investigations for the detection of Leptospira species in human, animal and environmental samples.     

褐飞虱特异引物的设计和评估

研究稻田生态系统中天敌对褐飞虱Nilaparvata lugens( St(a)l)的捕食非常重要,但很困难.本研究设计褐飞虱的PCR特异引物,并对利用从稻田66种稻田常见节肢动物中提取的DNA进行了特异性验证,得到褐飞虱的特异引物( Lug-F1a/Lug-R1),在实验室以拟环纹豹蛛Pardosa pseudoan...     

Tracking the movement of Nesidiocoris tenuis among banker plants and crops in a tomato greenhouse by DNA markers of host plants

The zoophytophagous predator, Nesidiocoris tenuis (Reuter) (Hemiptera: Miridae), is an important biological control agent. To maintain this insect, several non-crop host plants are used as banker plants in greenhouse crop systems. To optimize the efficiency of the predator-banker plant interaction, it is necessary to investigate how individual predators move between banker plants and crops. However, the movement is difficult to quantify under field conditions. Therefore, we investigated the movement of N. tenuis between tomato plants (Solanum lycopersicum L., Solanales: Solanaceae) and three banker plants (Cleome hassleriana Chod., Brassicales: Cleomaceae; Sesamum indicum L., Lamiales: Pedaliaceae; and Verbena × hybrida Voss, Lamiales: Verbenaceae) in a greenhouse by conducting PCR using plant-species-specific primers. Laboratory analysis results showed that our molecular method could detect N. tenuis activity within a relatively short time (≤ 24 h). In addition, N. tenuis predation on a pest species was unlikely to result in false detection of plant DNA in the pest (suggesting that N. tenuis had been on the plants). Multiple plant species were detected in adult insects collected from the greenhouse plants, indicating that N. tenuis frequently moved across the mentioned plant species. The movement patterns of N. tenuis between plant species varied substantially based on the plant species from which they were collected, which suggested each of the plant species had different functions for N. tenuis. Our findings revealed that planting multiple host plants would stabilize the N. tenuis population in biological control programs.

     

Detecting Cacopsylla pyricola (Hemiptera: Psyllidae) in predator guts using COI mitochondrial markers

Cacopsylla pyricola (F?rster) is one of the most important pests of pear in North America, where several native predators have been considered for integrated pest management (IPM) programmes. Two molecular markers of 271 and 188 bp were developed from C. pyricola cytochrome oxidase I (COI) fragments, in order to study the detection of this species in the gut of arthropod predators. Primer sensitivity and the detection period for pear psylla remains in the guts of Anthocoris tomentosus Pericart were determined. The sensitivity threshold was defined at 10-5 dilution of a C. pyricola fifth-instar nymph in all samples. Predator adults were evaluated immediately after ingestion of one to five C. pyricola nymphs (t = 0) and after 2, 4, 6, 8, 16, 24 and 32 h. Detection of the presence of C. pyricola DNA always lasted longer using the shorter fragment and was observed after 32 h of digestion using both markers. The primers amplifying the 188 bp fragment amplified all four psyllid species tested, whereas the primers designed to amplify the 271 bp fragment did so exclusively for C. pyricola and its close relative, Cacopsylla pyri (Linnaeus). Both primers failed to amplify DNA from representative species of the Coccinellidae, Chrysopidae, Hemerobiidae, Anthocoridae, Miridae, Salticidae, Aphididae, Tetranychidae and the Tortricidae, suggesting their suitability for general trophic studies.     

Macrolophus pygmaeus (Hemiptera: Miridae) population parameters and biological characteristics when feeding on eggplant and tomato without prey

The population parameters and biological characteristics of the predator Macrolophus pygmaeus Rambur when feeding on eggplant, Solanum melongena L., and tomato, Lycopersicon esculentum (Miller) (both Solanaceae), without access to prey, were investigated at several temperatures. The experiments were conducted at 15, 20, 25, and 30 degrees C, 65 +/- 5% RH, and a photoperiod of 16:8 (L:D) h. Females oviposited on both plant species at all temperatures, with the exception of tomato at 30 degrees C. Fecundity was highest at 20 degrees C (21.55 and 8.28 eggs per female on eggplant and tomato, respectively). Adult longevity on both host plants was greatest at 15 degrees C, reaching 38.72 and 34.20 d for females and 92.88 and 62.80 d for males, on eggplant and tomato, respectively. The estimated values of population parameters showed that this predator increased its numbers on eggplant at all temperatures tested, but the highest intrinsic rate of increase occurred at 25 degrees C (0.0401 d(-1)). On tomato, this predator could not increase in number, although it could survive for a relatively long period. Results demonstrated that M. pygmaeus can survive on both host plants in periods of prey scarcity, particularly on eggplant. Therefore, phytophagy can positively contribute to the effectiveness of this biological control agent. The importance of eggplant as a reservoir for the predator in mixed crops that are less suitable host plants is discussed.     

Universal mtDNA primers for species identification of degraded bony fish samples

We developed primers for amplifying and sequencing highly degraded mtDNA from diverse fish species. The primers flank a variable 148-bp fragment within the 12S region of mtDNA. We screened and sequenced 82 samples of bony fishes representing 17 families to confirm cross-species amplification and identification. Salmonid species were analysed and demonstrate 13 species-specific SNPs within this region. Based on alignments of additional deposited sequences, these primers are conserved in many other species, making them useful for species identification using degraded DNA samples such as archaeological specimens.