Rodrigo Lasa

Efficacy of Spodoptera exigua multiple nucleopolyhedrovirus as a biological insecticide for beet armyworm control in greenhouses of southern Spain

Abstract Chemical control measures targeted at Spodoptera exigua in greenhouse sweet pepper crops in Spain have resulted in pest resistance to virtually all commercially available insecticidal products. A multicapsid nucleopolyhedrovirus (SeMNPV), isolated from diseased S. exigua in Spain, was produced in laboratory reared larvae, tested for insecticidal activity in a laboratory bioassay, and was then applied in eleven commercial greenhouses planted with sweet pepper. Virus occlusion bodies (OBs) were applied on two occasions, at an interval of ∼7 days, at a rate of 5×108 OBs/L of spray in a volume of ∼600 L/ha, depending on crop phenology and greenhouse area. The percentage of plants showing recent (<48 h old) feeding damage fell dramatically in greenhouses with high infestations of S. exigua; the same pattern was observed, although less dramatically, in greenhouses with low infestations. Average mortality of larvae collected from treated plants at 4 days after each application, and reared in the laboratory until death, was high (70–89%) and was not significantly affected by the degree of crop infestation. In a separate trial, the rate of acquisition of infection was examined in larvae that fed on plants treated with 1×108 or 5×108 OBs/L of spray. Of the 27 and 60% of larvae, respectively, that acquired infection in the 48 h period after spraying, about half became infected in the first 6 h post-application, irrespective of application rate. Acquisition of infection proceeded more slowly during the night-time compared to the daytime period, underlining the advantages of early morning applications of the virus. We conclude that the Spanish SeMNPV isolate merits registration as a biological insecticide for use in greenhouse crops in this region.

Efficacy of Commercial Traps and Food Odor Attractants for Mass Trapping of Anastrepha ludens (Diptera: Tephritidae)

ABSTRACT One of the most important factors for the success of a mass trapping strategy to control a fruit fly involves the selection of an effective trap—lure combination. Because different species of fruit flies respond differently to the physical characteristics of a trap and to bait volatiles, the evaluation of commercial traps and lures that have proved useful against other tephtrids is necessary to determine their efficacy for mass trapping of Anastrepha ludens (Loew) (Diptera: Tephritidae). Under caged conditions, a commercial hemispherical trap with lateral holes (Maxitrap Plus) proved more attractive to A. ludens (both sexes) than five other commercial traps that were all baited with hydrolyzed protein. Among these traps, bottom invaginated traps and traps with invaginated lateral holes constructed with transparent cylinders had the best physical retention properties. When evaluated under field conditions, the lure was critical for the efficacy of the trap, and one of the traps that performed poorly in attraction and retention cage tests (MS2) resulted as one of the most effective traps when baited with CeraTrap lure. Considering the use of different trap models under field conditions, CeraTrap liquid bait was more effective in A. ludens capture than Biolure dry synthetic bait, but both lures were not replaced during the entire course of the experiment. The percentage of captured females was also slightly higher using CeraTrap lure (67.2%) than using Biolure baits (54.5–58.8%). In field tests, 75–81% of females were mated and no significant differences were observed among trap—lure combinations. Trap selectivity against nontarget adult lacewings also differed among trap—lure combinations.

Insecticidal Properties and Microbial Contaminants in a Spodoptera exigua Multiple Nucleopolyhedrovirus (Baculoviridae) Formulation Stored at Different Temperatures

Abstract The Spodoptera exigua (Hübner) multiple nucleopolyhedrovirus (SeMNPV) is currently being tested as a biological insecticide for use in greenhouse crops in southern Spain. We performed a study in which semipurified SeMNPV occlusion bodies (OBs) were formulated in phosphate-buffered saline, pH 6.5, with 5% (vol:vol) glycerol and 0.15% (wt:vol) sorbic acid, and they were stored at −20, 4, or 25°C during 18 mo. Initial aerobic counts (±SE) averaged 1.4 (±0.17) × 107 colony-forming units/ml after 17-h incubation at 37°C. Aerobic counts of microorganisms that contaminated OB formulations stored at 25°C decreased markedly over the period of the study, whereas only small decreases were observed in counts from OBs stored at 4 or −20°C. The principal microbial contaminants of OB suspensions were Enterococcus spp., Enterobacteriaceae, and yeasts. Potential human pathogens (Salmonella, Shigella, and Vibrio species) were not detected, and populations of Staphylococcus aureus and Bacillus cereus were extremely low. Compared with newly formulated OBs, the estimated LD50 values of OBs stored at 25°C increased by >16,666-fold over the 18 mo of storage, whereas LD50 values were not greatly affected by storage at 4 or −20°C. Significant changes over time in OB concentrations were only observed in the 25°C treatment. Complete degradation of viral DNA was observed at 25°C but not in refrigerated or frozen OBs. We conclude that OB formulation with bacteriostatic or antioxidant additives, together with storage and distribution in refrigerated conditions, will likely result in an SeMNPV biopesticide shelf life that exceeds 18 mo.

Invasive drosophilid pests Drosophila suzukii and Zaprionus indianus (Diptera: Drosophilidae) in Veracruz, Mexico

Summary The invasive drosophilid pests Drosophila suzukii (Matsumura) and Zaprionus indianus Gupta (Diptera: Drosophilidae) were captured in traps baited with hydrolyzed protein lures during a trial aimed at monitoring Anastrepha flies infesting guava, Psidium guajava L. (Myrtales: Myrtaceae) in Xico, Veracruz, Mexico, during Sep 2014. This is the first record of these species in Veracruz State. These exotic species represented over 80% of the total drosophilid flies captured. Studies on the commercial impact of these pests in guava and in the production of other fruit species in the region should be considered a priority.

The attractiveness of phagostimulant formulations of a nucleopolyhedrovirus-based insecticide depends on prior insect diet

The multiply-enveloped nucleopolyhedrovirus (SeMNPV) of Spodoptera exigua (Hübner) is being employed on an increasing scale as the basis for bioinsecticidal products for control of this pest in greenhouse crops in Europe. The mortality of diet-reared S. exigua larvae was determined after feeding on lettuce leaf discs contaminated by mixtures of SeMNPV occlusion bodies and 1 of 13 substances reported to have phagostimulant properties. Of the substances tested, wheatgerm and soya flour resulted in significantly increased mortality compared to the virus alone. However, these preferences disappeared when larvae were reared on lettuce. We conclude that (1) the activity of potential phagostimulant substances depends on the prior feeding experience of S. exigua and, (2) laboratory tests using diet-reared insects require validation using plant-reared larvae before being developed for field testing.

Fruit firmness, superficial damage, and location modulate infestation by Drosophila suzukii and Zaprionus indianus: the case of guava in Veracruz, Mexico

The exotic pestiferous flies Drosophila suzukii (Matsumura) and Zaprionus indianus Gupta (Diptera: Drosophilidae) were recently identified in traps used for monitoring tephritid pests of guava, Psidium guajava L. (Myrtaceae), in Veracruz, Mexico. To determine whether both drosophilids were infesting guava, a previously unreported host, samples were taken from fruits on trees and fallen fruits on the ground. Overall, 74 and 36% of visually intact fruits attached to the tree were infested by D. suzukii and Z. indianus, respectively. Under laboratory conditions, small artificial punctures on the surface of ripe guavas did not result in increased oviposition by D. suzukii compared to undamaged fruit, whereas Z. indianus almost completely avoided oviposition, or were not capable of developing in fruit. Females of D. suzukii were capable of ovipositing in early ripe guavas in laboratory tests (23% of fruits were used for oviposition), although a high penetration force is required to pierce fruit (mean ± SEM = 89.0 ± 3.0 cN). Fully ripe and overripe guavas were softer (52.2–53.5 cN penetration force) and were more frequently infested (ca. 60% infestation). Numbers of females that developed in guavas were not influenced by ripeness/firmness, whereas male development was reduced in early ripe fruit compared to ripe and overripe fruit. In laboratory choice experiments with crushed fruits, D. suzukii adults were equally attracted to guava and blueberry, independent of gender and age. However, raspberry was more attractive than guava. This study demonstrates that D. suzukii is attracted to guava, is capable of ovipositing in fruit and, under field conditions, is more abundant in fruits still attached to the tree compared to fallen fruit that remain intact. In contrast, Z. indianus was not capable of developing in intact guavas and, although present in fruits attached to the tree, was most abundant in fallen damaged fruits.

Baculovirus-Induced Climbing Behavior Favors Intraspecific Necrophagy and Efficient Disease Transmission in Spodoptera exigua

Shortly prior to death, many species of Lepidoptera infected with nucleopolyhedrovirus climb upwards on the host plant. This results in improved dissemination of viral occlusion bodies over plant foliage and an increased probability of transmission to healthy conspecific larvae. Following applications of Spodoptera exigua multiple nucleopolyhedrovirus for control of Spodoptera exigua on greenhouse-grown sweet pepper crops, necrophagy was observed by healthy S. exigua larvae that fed on virus-killed conspecifics. We examined whether this risky behavior was induced by olfactory or phagostimulant compounds associated with infected cadavers. Laboratory choice tests and olfactometer studies, involving infected and non-infected cadavers placed on spinach leaf discs, revealed no evidence for greater attraction of healthy larvae to virus-killed over non-infected cadavers. Physical contact or feeding on infected cadavers resulted in a very high incidence of transmission (82–93% lethal disease). Observations on the behavior of S. exigua larvae on pepper plants revealed that infected insects died on the uppermost 10% of foliage and closer to the plant stem than healthy conspecifics of the same stage, which we considered clear evidence of baculovirus-induced climbing behavior. Healthy larvae that subsequently foraged on the plant were more frequently observed closer to the infected than the non-infected cadaver. Healthy larvae also encountered and fed on infected cadavers significantly more frequently and more rapidly than larvae that fed on non-infected cadavers. Intraspecific necrophagy on infected cadavers invariably resulted in virus transmission and death of the necrophagous insect. We conclude that, in addition to improving the dissemination of virus particles over plant foliage, baculovirus-induced climbing behavior increases the incidence of intraspecific necrophagy in S. exigua, which is the most efficient mechanism of transmission of this lethal pathogen.

Improved capture of Drosophila suzukii by a trap baited with two attractants in the same device

The improvement of trap-lure combinations is an important part of integrated pest management programs that involve monitoring pests for timely insecticide applications, or for their use in control strategies such as mass trapping or bait stations. In this study improvements in the capture of Drosophila suzukii were not observed following the inclusion of different color stimuli with respect to a red-black stripe cup trap. This red-black stripe trap with a hemispherical dome-shaped lid had a significantly improved physical retention of flies compared to traps fitted with a flat lid. Retention was further improved when an additional tube device, which could be baited with a supplemental attractant, was introduced through the dome-shaped lid. Under laboratory conditions, this trap, in which apple cider vinegar + 10% ethanol was present as the drowning solution and the additional tube device was baited with a fermenting mixture of sugar and yeast, was significantly more effective in catching D. suzukii flies than other conventional attractants or a commercial lure. The capture rate of this trap-lure combination remained higher than that of a commercial lure, even after 20 days of use under laboratory conditions. In a guava orchard this trap was 15-fold more effective in catching D. suzukii flies than similar traps baited with apple cider vinegar alone, 4 to 7 fold more effective than similar traps baited with a commercial lure, and 1.7-fold more effective than a fermenting mixture of yeasts and wheat flour. In commercial blackberry orchards, this trap was 6-fold more effective in trapping D. suzukii flies than the clear trap baited with apple cider vinegar used by growers. The efficacy of this trap presents a promising line of future research for monitoring and control of D. suzukii and likely other drosophilid pests.

Experimental hybridization and reproductive isolation between two sympatric species of tephritid fruit flies in the Anastrepha fraterculus species group

Among tephritid fruit flies, hybridization has been found to produce local adaptation and speciation, and in the case of pest species, induce behavioral and ecological alterations that can adversely impact efficient pest management. The fraterculus species group within Anastrepha (Diptera: Tephritidae), is a rapidly radiating aggregate, which includes cryptic species complexes, numerous sister species, and several pest species. Molecular studies have highlighted the possibility of introgression between A. fraterculus and A. obliqua. Reproductive isolation has been studied among morphotypes of the A. fraterculus species complex as a tool for species delimitation. Here we examined the existence and strength of prezygotic and postzygotic isolation between sympatric populations of two closely related species within the highly derived fraterculus group (A. fraterculus and A. obliqua), coexisting in nature. Although adults of both species showed a strong tendency for assortative mating, a small proportion of hybrid pairings in both directions were observed. We also observed asymmetric postzygotic isolation, with one hybrid cross displaying a strong reduction in fecundity and F1 egg fertility. Survival was greater for the progeny of homotypic and hybrid crosses in the maternal host. There was a marked female biased sex ratio distortion for both F1 hybrid adults. Hybridization between A. fraterculus and A. obliqua in nature may be difficult but possible; these two species display stronger reproductive isolation than all pairs of species previously examined in the A. fraterculus species complex. Asymmetric postzygotic isolation is suggestive of Wolbachia mediated cytoplasmic incompatibilities that may be exploited in area‐wide pest management.

Anticarsia gemmatalis Nucleopolyhedrovirus from Soybean Crops in Tamaulipas, Mexico: Diversity and Insecticidal Characteristics of Individual Variants and their Co-Occluded Mixtures

Abstract In 1999, Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) was introduced into a major soybean-growing region in Tamaulipas, Mexico, for control of its lepidopteran host, Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae). The virus introduction proved to be highly successful in controlling this agronomically important pest. In order to determine the genotypic diversity and insecticidal traits of Mexican AgMNPVs, we obtained 30 field-collected isolates from Tamaulipas State. Five distinct variants (genotypes 1–5) were identified from plaques replicated in A. gemmatalis larvae by examination of restriction profiles using HindIII. Initial screening indicated that none of the variants, or co-occluded mixtures of variants in different proportions, was more pathogenic than the 30 field isolates mixture or a reference variant from Brazil (AgMNPV-2D). Mean occlusion body production also was similar among genotype variants, the mixture of 30 field isolates and AgMNPV-2D treatments, but was significantly reduced in 1 co-occluded mixture. Speed of kill also was similar among variants (except genotype 1) and their mixtures. Lethal concentration metrics indicated that these results were unlikely due to selection of variants with reduced pathogenicity during the plaque purification process. We conclude that the mixture of 30 field isolates most likely would prove suitable for use as a biological insecticide in the soybean-growing region of Mexico.