WO2011117444A1 - Method for rearing predator sphaerophoria rueppellii - Google Patents

Abstract

The invention relates to methods for rearing the dipterous predator Sphaerophoria rueppelli for possible use in the biological control of aphids in Mediterranean crops. The invention relates to the method, processes, environmental conditions and diets of both the immature stages and the adults of this species. In all cases, the conditions selected are those that ensure greater reproductive potential and greater rates of survival.

Description

 BREEDING METHOD OF THE PREDATOR SDhaerophoria rueppellii

DESCRICPION

Method of breeding of the predator Sphaerophoria rueppellii.

FIELD OF THE INVENTION

The present invention relates to the breeding method of Sphaerophoria rueppellii (Diptera, Syrphidae), an entomophageal insect that, due to its specificity and predatory efficacy, has been selected for the biological control of aphid agricultural pests.

STATE OF THE PREVIOUS TECHNIQUE

Aphids (aphids) are phytophagous insects that feed on the sap of various parts of plants: leaves, tender stems, roots; preferring the young and developing organs. The high growth rate of aphids, their ability to extract sap with great yield and their character as virus vectors, often leads their populations to cause serious damage to crops, which result in significant economic losses and therefore , to reach the status of plague. The group of useful fauna associated with aphids naturally exercises an effective biological control, especially preventive. When for some reason the action of natural enemies is prevented (usually by the disproportionate use of chemical agents), population peaks occur above the threshold of economic damage and the use of control strategies that reduce the population of the pest species to tolerable levels is necessary, while considering economic interests, of public health and environmental.

Biological control is presented as an effective and risk-free alternative to the numerous and growing problems arising from the use of biocidal chemicals. Biological control consists in the application of techniques compatible with the conservation of the Environment, through the use of natural enemies of pests that control the population level of pest species without causing pollution or waste problems. Biological control agents are released into the environment inoculatively or in a flood, and may be mainly parasitoids or predators.

Due to the complex biology of the controlling agents (mostly insects), the success of their use is not assured if it is not preceded by detailed and rigorous studies on the interrelations established between them, on their specificity and on the characteristics environmental and biological that govern their life cycles.

Sylphs (Diptera, Syrphidae) are a group commonly used as biological control agents for pests. It is a diverse family within the dipterans that have the appearance of hymenoptera such as bees and wasps, with which they are easily confused. To date, about 200 genera and about 5400 species have been described. They are very frequent on flowers, which they feed when they are adults, consuming nectar and pollen, being able in some cases also to be good pollinating agents.

The larvae of the siphids have a biology very different from that of adults and greatly varied within the group. Some are avid predators of aphids and other small phytophagous insects, which are hunted on vegetation, so some species are used in the biological control of pests. Their specificity for soft body homoptera insects makes them suitable candidates for the control of aphid pests. Its life cycle includes the following phases: the egg (H), the larvae, presenting three larval stages (L1, L2 and L3) being the larvae that prey on the aphids, the pupa (P) (immobile and aghagous) and the adult (A) or magician with trophic requirements different from the larvae. The biological cycle scheme is summarized as follows:

Episyphus balteatus es la única especie de sírfido que actualmente se comercializa a nivel mundial como agente biológico de control de plagas de pulgones. H Adult

Episyphus balteatus is the only species of siphon that is currently marketed worldwide as a biological agent for controlling aphid pests.

However, this species, which is especially abundant in central and northern Europe, is not the most abundant or the best adapted to the environmental conditions to which the crops of Mediterranean areas are subjected. There are studies that indicate a high mortality of this species at temperatures above 25 ° C, conditions that are easily reached in Mediterranean agrosystems during the crop cycle.

Bibliographic references

1. Chambers, R. J. 1988. Syrphidae. In: A. K. Minks & P. Harrewijn (eds), Aphid: Their Biology, Natural Enemies and Control, Vol. 2B. Elsevier, Amsterdam, 259-270. 2. Gilbert, F. 1993. Hoverílies. Richmond Press, Surrey.

3. Hart, A. J., Bale, J. S. and Fenlon, J. S. 1997. Developmental threshold, day-degree requirements and voltinism of the aphid predator Episyrphus balteatus (Diptera: Syrphidae). Annals of Applied Biology, 130: 427-437.

4. Pineda A & Marcos-Garcia MA (2006) "First data of the population dynamics of aphidophagous syrphids in Mediterranean pepper greenhouses and reviewing a monitoring trap". lOBC / wprs Bulletin 29: 169-174.

5. Pineda A & Marcos-Garcia MA (2008) "Seasonal abundance of aphidophagous hoverflies (Diptera: Syrphidae) and their population levéis in and outside Mediterranean sweet pepper greenhouses." Annals of the Entomological Society of America 101: 384-391.

6. Rojo S, Gilbert F, Marcos-Garcia MA, Nieto JM & Mier MP (2003) "A world review of predatory hoverflies (Diptera, Syrphidae: Syrphinae) and their prey." CIBIO, Alicante. 7. Speight MCD (2005) "Species accounts of European Syrphidae (Diptera)" 2005, Vol. 49: Syrph the Net, the datábase of European Syrphidae (ed. By MCD Speight, E Castella, JP Sarthou & C Monteil) Syrph the Net publications, Dublin, p. 242

8. Weems, H. V. 1971. A hoverfly, Allograpta obliqua (Say) (Diptera: Syrphidae). Entomology Circular 106. Division of Plant Industry, Florida Department of Agriculture and Consumer Services. Gainesville, FL.

EXPLANATION OF THE INVENTION

It is necessary in the light of the above, to look for an alternative to the use of E. bateatus, such as a species that, while retaining its characteristics as a good biological agent, develops in optimal conditions in Mediterranean agricultural ecosystems.

Sphaerophoria rueppellii is shown as an ideal biological agent in pest control as it is a much more abundant species in arid regions naturally than E. balteatus, which supports its greater adaptation to these types of environments.

Sphaerophoria rueppellii is an insect belonging to the group of syphilids (Diptera, Syrphidea), some of whose species are important aphid control agents. Sphaerophoria rueppellii is a species of wide distribution, especially abundant in crops of the Mediterranean area. Given the geographical distribution of this species, its use in biological control does not have associated problems related to the release of alien species since The environmental conditions allow the optimum of its predatory activity on aphid pests, without causing any damage on biocenosis (as in the case of invasive species), nor any ecological imbalance (due to displacement, competitiveness, etc.).

In addition, Sphaerophoria rueppellii is adapted to the extreme conditions of temperature and humidity present in crops in the Mediterranean area, being the most abundant species of sapphire in both greenhouses and open crops in the driest periods.

On the other hand it should be noted that the larvae of this species prey on more than 40 species of aphids, which cause, most of them, serious pests in agricultural crops. This wide range of prey together with the specific nature of their diet (aphids only), make this species an ideal candidate for the biological control of aphid pests in the Mediterranean area.

However, to date nothing was known about the biology or the life cycle of Sphaerophoria rueppellii, except that its larvae (immature phases) are aphids predators. This lack of knowledge precluded its breeding on an industrial scale and therefore its use as a biological control agent. It is well known in the field of Entomology that trophic requirements, oviposition stimuli, climatic conditions of survival, optimal fertility and fertility conditions, etc., are different in each species of insect and therefore, the data that are known for the only industrially produced species of sapphire {Episyrphus balteatus), cannot be applied successfully in the breeding of other species. Therefore, for the use of Sphaerophoria rueppellii as a biological control agent, it is necessary to develop a method or protocol of breeding of this species that can be used on an industrial scale and that allows obtaining a large number of individuals of this biological agent, in that the appropriate diet is also indicated in each phase of its life cycle, as well as the means and requirements necessary to complete its breeding in optimal conditions.

The present invention is based on studies conducted in laboratory conditions to design a methodological procedure that allows artificial breeding (growth, maturation, reproduction and oviposition) of Sphaerophoria rueppellii in the laboratory. Given the biological characteristics of these holometábolos insects, good flying and with a great capacity of dispersion when they are adults, it is impossible to know what are the biotic and abiotic parameters that in natural conditions influence their biology, both in the immature phases and in the adults Therefore, since it cannot be simulated in the laboratory, a large number of probative experiments must be carried out.

For this reason, it has been necessary for the elaboration of the breeding method that the present invention exposes, the study and determination of the following aspects:

 - Protocol of action and necessary means for breeding in laboratory conditions.

 - The larvae diet.

 - The diet of adults.

- The optimum and humidity ranges, brightness and temperature required in the breeding of this predator, from the egg phase to the emergence of adults. - The optimum humidity, temperature and luminosity necessary during the period of sexual maturation, intercourse, oviposition and longevity of adults.

 - The biological requirements necessary for laying.

Likewise, the determination of the method and breeding protocol of the present invention allows to overcome inconveniences in the aspects listed below, optimizing the culture conditions:

 - Growth of fungi in the nutritional environment.

 - Growth of fungi or bacteria in the immature phases.

- Avoid death by drying out or excessive humidity.

- Avoid mortality due to infections.

 - Avoid lengthening the life cycle by temperatures different from the optimum.

 - Overcome oviposition deficiency due to lack of stimuli (prey, molasses, support plant, etc.).

The present invention thus relates to a breeding method with a specific protocol that allows the development from egg, through the three larval phases and through the physiological processes of pupation and metamorphosis to adults.

In addition, to obtain successive generations, once the adults of Sphaerophoria rueppellii are born, the method of the present invention allows:

- the sexual maturation of adults born through the proportion of an adequate diet. - the intercourse, allowing adults the conditions and sufficient space for the reconnaissance flight and the consummation of the intercourse.

 - oviposition, providing adequate stimuli for laying inside the container.

 - the development of immature phases until the birth of the adult

Finally it has been necessary to determine the duration of the different stages of development and growth (from oviposition to the birth of the larva, the three larval stages to pupation, from the pupa to the adult's emergence, the longevity of the adult, the period of maturation of adults and the duration of the oviposition period). Once these parameters are known, the breeding of this species has been optimized by:

 to. The decrease of the life cycle duration, so that a greater number of generations can be achieved in the minimum possible time.

 b. The optimization of fertility and fertility.

These improvements have been achieved by modifying the relative humidity and ambient temperature under laboratory conditions.

The characteristics of the breeding receptacles for adults (FIGURE 3) and for the immature phases (FIGURE 4) have been selected specifically for this species seeking its optimum efficiency and contemplating space limitation, an important requirement in the production of this large scale species. The breeding of adult specimens was carried out in a receptacle with certain conditions. The breeding method of Sphaerophoria rueppellii begins with the introduction of pupae of Sphaerophoria rueppellii in the receptacle (FIGURE 3), where adult males and females are born. In that receptacle there is the sexual maturation of adults, intercourse and oviposition. Said receptacle also contains inside:

 a) Capsicum plants (Capsicum annuum) grown for six weeks in small pots, infested with colonies of Myzus persicae developing on pepper plants two weeks before exposure to the larvae and which constitute the stimulus of oviposition.

 b) A container to hold water.

 c) Sphaerophoria rueppellii diet containing sugars and pollen.

The eggs laid by adults on the pepper plants in this receptacle are transferred to another receptacle for incubation until the birth and development of the born larvae. This second receptacle is similar to that of adult breeding but the upper part is covered with a layer of plastic film (FIGURE 4) to achieve in its interior conditions of relative humidity greater than in the first receptacle. This second receptacle also contains in its interior some twelve pepper plants (Capsicum annuum) grown in pots and infested with abundant specimens of Myzus persicae that will be the only trophic resource of the larvae until pupation.

Finally, the new pupae obtained in the immature breeding receptacle (FIGURE 4), are collected and transferred to the first adult breeding receptacle (FIGURE 3) to begin the process again.

In both receptacles, the environmental conditions are achieved by means of heated chambers where the following conditions are programmed:

 • temperature at 25 ° C +/- 1 ° C,

 • relative humidity of 60% +/- 10%

 • photoperiod of approximately 12:12 or 14:10 (light hours: dark hours)

BRIEF DESCRIPTION OF THE FIGURES FIGURE 1: Larval mortality rate for different combinations of humidity and temperature. The photoperiod has been 12:12 (12 hours of light and 12 hours of darkness).

FIGURE 2: Maturation period (days) of the eggs until the larvae are born for different combinations of humidity and temperature. The photoperiod has been 12:12 (12 hours of light and 12 hours of darkness).

FIGURE 3: Receptacle where adults are born and lay eggs and their biological requirements.

FIGURE 4: Receptacle where the breeding of immature phases (eggs, larvae and pupae) and biological requirements is developed. DETAILED EXHIBITION OF REALIZATION MODES

The breeding method was carried out in receptacles (1 and 9) of washable, translucent material, provided with a cover that prevents the passage of insects but allows the passage of light and air. It must be provided with a sleeve (3) that allows the maneuver with the breeding material.

 The Sphaerophoria rueppellii breeding method protocol begins with the introduction of Sphaerophoria rueppellii pupae into the adult receptacle (FIGURE 3) from which adult males and females are born. Right there is the sexual maturation of magicians, intercourse and oviposition. The conditions inside this receptacle should be 25 ° C-30 ° C temperature and 60% relative humidity with a variation range of 10%.

For this, this receptacle must contain inside: a) Capsicum plants (5) (Capsicum annuum), grown for six weeks in small pots infested with colonies of Myzus persicae (4) growing on pepper plants two weeks before its exposure to the larvae and that constitute the stimulus of oviposition.

 b) A container to hold water (6).

 c) Sphaerophoria rueppellii diet containing sugars (7) pollen (8).

In the invention the sugars that make up the diet of adults of Sphaerophoria rueppellii were experimentally tested and selected among them sucrose and brown sugar in combination with milflores pollen and water. Eggs laid by adults on pepper plants in the receptacle (1) are transferred to the receptacle (FIGURE 4) for incubation until the birth and development of the born larvae. This receptacle is similar to that of adult breeding but covering the upper part with a layer of plastic film (10) to achieve inside conditions of relative humidity of 80-100%. The rest of the environmental conditions are similar: 25-30 ° C of temperature, and photoperiod of 12:12 or 14:10 that are programmed in the breeding chamber.

The receptacle (9) is occupied at its base by pepper plants (Capsicum annuum) grown in pots and infested with abundant specimens of Myzus persicae (1 1) that will be the only trophic resource of the larvae until pupation.

Finally, the new pupae obtained in the immature breeding receptacle (9) are collected and transferred to the adult breeding receptacle (1) to begin the process again.

The oviposition period of mature females is a maximum of 15 days. Sexual maturation is reached between the sixth and seventh day of adult life. The duration of the complete cycle, from egg to adult emergence ranges between 14 and 17 days. The invention will now be illustrated by tests carried out by the inventors, which shows the specificity and effectiveness of the breeding method and protocol. The experimental results demonstrate the successful breeding of successive generations of Sphaerophoria rueppellii. EXAMPLE 1: Results of the breeding of Sphaerophoria rueppellii a. Adult breeding: adult emergence, maturation, intercourse and oviposition (FIGURE 3)

 The pupae of Sphaerophoria rueppellii are introduced into the receptacle (1) where male and female adult individuals emerge, mature, copulate and lay eggs.

The receptacle in this case must contain:

 · Artificial diet of adults:

^■ 6 gr sucrose (7)

^■ 6 gr of brown sugar (7)

^■ 2 gr of pollen (8) milflores

^■ 125 ml of water (6)

The stimulus for oviposition consisting of a pepper plant (5) with aphids of the species Myzus persicae (4). b. Egg maturation, birth and larval rearing (FIGURE 4) Once the larvae of the eggs were born, they developed inside the receptacle (9) of immature phase rearing, with conditions that did not vary since the introduction of the Eggs until pupation.

 This receptacle contains approximately twelve 6-week-old pepper plants in small pots infested with Myzus persicae (11) two weeks before exposure to the larvae.

These conditions were designed for the development of between 75 and 100 eggs. The humidity and temperature conditions were gone in each treatment of the experiment. In all cases the photoperiod was 12:12. The data obtained for the larval mortality rate (FIGURE 1) showed that:

 to. At 30 ° C and a relative humidity range of 80-100%, the highest percentage of larval birth (84%) was obtained. b. Maintaining this humidity between 80 and 100%, high levels of adult emergencies continued to be obtained, although the temperature was below 30 ° C.

 C. At humidities below this range of between 80 and 100%, the number of births decreases dramatically. Data referring to the maturation period of the eggs until the larvae are born (FIGURE 2) showed that:

 to. At 30 ° C and a relative humidity of between 80 and 100%, the shortest period of time from oviposition to the emergence of the larvae (less than 2 days) was obtained. b. At lower temperatures, the cycle was extended although humidity remained in this range,

 C. At humidities below 80-100%, the emergency period was considerably extended (more than 4 days).

Claims

1.- Method of breeding of the predator Sphaerophoria rueppellii that allows the development of the complete cycle, starting with oviposition, the birth and development of the three larval phases, followed by the physiological processes of pupation and metamorphosis to adults and their maturation. This method distinguishes: A- An initial phase where the pupae are introduced inside the receptacle (1) until the emergence of adults (males and females), sexual maturation and oviposition and comprising at least: 1. Pepper plants (Capsicum annuum) infested with colonies of Myzus persicae developing on these plants and constituting the stimulus and place of oviposition.  2. A container with water.  3. Adult diet of Sphaerophoria rueppellii comprising at least sugars and pollen.  All this under certain conditions: 4. Constant temperature between 25-30 ° C.  5. Relative humidity of 60% ± 10%.  6. Photoperiod of 12:12 or 14:10. In this way the females lay the eggs on the pepper leaves from where they are transferred to a second receptacle (9). B- A second breeding phase where the eggs obtained in the previous phase are carefully transferred to a second receptacle (9) where the optimal conditions for the larvae are born, their growth and transformation into pupae. This second receptacle (9) comprises at least: Abundant pepper plants infested with the Myzus persicae aphid on which the transferred eggs are placed. All this under certain conditions: 1. Photoperiod of 12:12 or 14:10.  2. Constant temperature of 25 ° C-30 ° C.  3. Relative humidity inside the 80% - 100% receptacle that is achieved by covering it with a layer of plastic film. C- A third phase where the new pupae obtained in the receptacle (9), are carefully collected and transferred to the first receptacle (1) to begin the process again. 2. Use of the breeding method of the predator Sphaerophoria rueppellii according to claim 1 for its application in the biological control of aphid pests in Mediterranean climate crops.