Progress 10/01/09 to 09/30/14

Outputs
Target Audience: Agricultural producers, organic farmers, natural resource managers, entomologists Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Oral presentations, posters, and peer-reviewed publications What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? IMPACT: Establishment of the Honduran parasitic fly Lixadmontia franki will save populations of threatened and endangered native Florida bromeliad species from the Mexican bromeliad weevil. Establishment of Phaenochilus kashaya will potentially control populations of the cycad aulacaspis scale (CAS) on ornamental cycads and Guam’s native cycads. Augmentation biological control technology with field releases of insectary-reared predatory stinkbugs and green lacewings give organic growers a management tactic for controlling pests on crucifer crops. Research on entomopathogenic fungi determines those that are effective against numerous pests. Biological studies provide insight into biological control agent/pest interactions. Discovery and descriptions of new species enhance our knowledge of biodiversity for biological control and protection of unique environments. ACCOMPLISHMENTS: 1) A survey for parasitoids of CAS in Florida detected the presence of Arrhenophagus chionaspidis in several localities. A survey of predators and parasitoids of the red bay ambrosia beetle (RAB) detected 3 possible predators and 2 possible parasitoids. 2) A tachinid fly and an ichneumonid wasp were reared from larvae of the yellowmargined leaf beetle (YMLB) in Argentina, but these have not been identified to genus. No new parasitoids of bromeliad-eating weevils were found during exploration in the Neotropics. 3) Development of the Mexican bromeliad weevil (MBW) occurred at 18-30C and is significantly reduced outside this range. Larval and adult weevils are unable to survive 4 days at 0C. Oviposition rate in pineapple leaves was greater than in Tillandsia species; oviposition rate was greater in Florida Tillandsia utriculata than in Tillandsia fasciculata. No larvae fed Central American T. utriculata or T. fasciculata completed development. Female L. franki showed strong antennal responses to a-pinene, beta-pinene, limonene plus ocimene, and ethylanisole and ethylphenol from weevil larvae on chewed pineapple leaves. Larval developmental time of the lady beetle Phaenochilus kashaya was about 47, 32, and 22 days at 20, 25, and 30C, respectively. The degree-day requirements are 119 for eggs, 435 for larvae, 110 for pupae, and 714 from egg to adult. Females at 20C laid about 339 eggs. Daily consumption of CAS by P. kashaya larvae was greater at 25 than at 30C. Eggs of P. kashaya suffered 100% mortality at 0C for more than 1 day. Larvae, pupae, and adults cannot tolerate 0C for more than a few days and cannot tolerate subfreezing temperatures. Phaenochilus kashaya ate other armored scales but not soft scales, large aphids and mealybugs, psyllids, thrips, caterpillars, herbivorous beetle larvae, or mites. It consumed a few butterfly eggs, very small aphids, very small mealybugs, and very small whitefly nymphs when armored scales were not available. Predation was noted when P. kashaya adults were offered larvae of other predatory beetles, parasitized CAS, and green lacewing eggs. First instars of CAS treated with the entomopathogenic fungus Isaria fumosorosea suffered 73% mortality at 30C and 84% at 20C 8 days post-application. The lowest molting rate was obtained with at 20C. These results indicate that I. fumosorosea may be a new biological control weapon for suppressing CAS infestations. Developmental time of YMLB was longest at 15C and shortest at 30C. Leaf area consumed by the 4th instar was 7-fold lower at 30C. The lower developmental threshold varied 7.3-9.8C, and the total degree-days required to complete development from egg to adult was 333. Eggs were most cold tolerant, followed by pupae and adults. The predicted northern distribution of YMLB extends to Kansas, Illinois, Kentucky, and Virginia. Nymphal development of the spined soldier bug (SSB) increased from 23 days at 25C to 99 days at 15C. Nymphs of SSB can develop successfully on a diet of eggs or YMLB larvae, but predation and development rates will be less during cool months when YMLB is a pest in Florida. Larval green lacewings killed 496-634 eggs and 218-225 YMLB larvae, depending on temperature. They showed a 5.2-fold preference for aphid nymphs over YMLB eggs and larvae. On plants, green lacewings killed more YMLB larvae than eggs due to differences in the location of the prey. Larvae of YMLB were more susceptible to infection by I. fumosorosea than other stages. Damage by YMLB on plants sprayed with I. fumosorosea was less than on untreated plants. Adult female RAB were killed in a few days by I. fumosorosea, Metarhizium brunneum, and Beauveria bassiana. Beauveria bassiana and M. brunneum killed females faster. Females dipped in B. bassiana and M. brunneum had more viable spores attached to their bodies. No significant differences were observed in the mortality of beetles that were dipped in fungus and offered avocado logs as substrate, compared to beetles that walked on logs treated with fungi. Survival of the ambrosia beetles Xylosandrus crassiusculus and Xyleborus volvulus was 5.5 and 5.8 days and 17.1 and 7.8 days after exposure to B. bassiana and I. fumosorosea, respectively; B. bassiana killed beetles faster. Xylosandrus crassiusculus females dipped in B. bassiana had more viable spores attached to their bodies, while X. volvulus females dipped in I. fumosorosea had more viable spores attached. 4) A new rearing method for L. franki was developed by which egg-laying females are held in vials and provided pineapple leaves in which to oviposit. The new method produces rates of parasitism similar to the old method. The new method allows for better control of the egg-laying population, reduces the variability of instars within a top, and allows calculation of the number of larvae per top with much greater certainty. Wild material was collected in Honduras to invigorate the gene pool of the laboratory colony. 5) Field releases of 97 L. franki females were made at 4 sites in Florida on 6 dates in 2010-2013. Post-release monitoring gave no indication of establishment of feral populations of this natural enemy. A proposal for field release of P. kashaya was resubmitted to APHIS; a decision on the petition is pending. 6) A study to investigate the effect of straw mulch on the abundance of YMLB and ground predators in turnips in Florida, and subsequent influence on crop damage and yield, concluded that the use of straw mulch as a pest management option for YMLB in crucifer crops on organic farms is not recommended. Higher numbers of YMLB were found in plots with mulch compared to plots with no mulch, whereas greater numbers of ground predators were obtained with no mulch. Turnip leaf biomass was greater in plots with straw mulch, but plants in these plots suffered more herbivory. The predation by SSB on YMLB was evaluated at 3 release rates in field cages. The mean number of YMLB larvae declined during the 1st 4 sampling dates. On the final sampling date, there were no significant differences among treatments with predators. Recommendations for growers are: 1) release 10 SSB nymphs per 6 plants if the plants are expected to have more than 7 leaves; 2) release 4 SSB nymphs per 6 plants if the plants are expected to have less than 6 leaves. In trials on two organic farms during two years, three predator release rates were tested: 0, 3, and 6 nymphs per plant at the first sign of pest adults. The number of YMLB larvae and adults per plant in plots receiving SSB was significantly less than control plots in some weeks. Releases of SSB lowered plant damage by the pest and plants with SSB bugs suffered less proportional leaf area loss compared to control plants. Timely releases of first- and second-instar SSB are capable of reducing larval populations of YMLB. This control tactic will result in less leaf damage and better commodity product for organic growers. The field release of green lacewing larvae did not cause an important effect on YMLB populations.

Publications

• Type: Journal Articles Status: Published Year Published: 2014 Citation: Cave, R. D., and H. H. Hespenheide. 2014. An annotated list of the Buprestidae (Coleoptera) known to occur in Honduras. The Coleopterists Bulletin 68(1): 53-60.
• Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Cave, R. D. 2014. Four against one: Biological control of the cycad aulacaspis scale. Antenna Special ECE Edition: 24-25.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Ratcliffe, B. C., and R. D. Cave. 2014. Los escarabajos rinocerontes gigantes de El Salvador (Coleoptera: Scarabaeidae: Dynastinae) [El Salvadors Giant Rhinoceros Beetles (Coleoptera: Scarabaeidae: Dynastinae)]. BIOMA 16: 7-27.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Ratcliffe, B. C., and R. D. Cave. 2014. Un inventaire de la biodiversit� des Dynastinae (Coleoptera, Scarabaeidae) des Antilles: pr�sentation du projet. ACOREP?France: Col�opt�res de Petites Antilles. Tome II: 3-9.
• Type: Journal Articles Status: Accepted Year Published: 2015 Citation: Carrillo, D., C. A. Dunlap, P. B. Avery, J. B. Navarrete, R. E. Duncan, M. A. Jackson, R. W. Behle, R. D. Cave, J. H. Crane, A. P. Rooney, and J. E Pe�a. 2015. Entomopathogenic fungi as biological control agents for the vector of laurel wilt disease, the redbay ambrosia beetle, Xyleborus glabratus (Coleoptera: Curculionidae). Biological Control.

Progress 10/01/13 to 09/30/14

Outputs
Target Audience: Agricultural producers, organic farmers, natural resource managers, entomologists Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? presentations and posters at professional meetings, peer-reviewed publications What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? IMPACT: Establishment of the Honduran parasitic fly Lixadmontia franki will save populations of threatened and endangered native Florida bromeliad species from the Mexican bromeliad weevil. Establishment of Phaenochilus kashaya will potentially control populations of the cycad aulacaspis scale (CAS) on ornamental cycads and Guam’s native cycads. Augmentation biological control technology with field releases of insectary-reared predatory stinkbugs and green lacewings give organic growers a management tactic for controlling pests on crucifer crops. Research on entomopathogenic fungi determines those that are effective against numerous pests. Biological studies provide insight into biological control agent/pest interactions. Discovery and descriptions of new species enhance our knowledge of biodiversity for biological control and protection of unique environments. ACCOMPLISHMENTS: 1) No activity. 2) No new parasitoids of bromeliad-eating weevils were found during exploration in Honduras, Puerto Rico, Lesser Antilles, and Belize. 3) Development of the Mexican bromeliad weevil (MBW), Metamasius callizona, occurred at 18-30C. At 15C, there was no survival beyond second instar. At 35C, no eggs hatched. The number of eggs collected at 22-33C were statistically similar at 0.3-0.5 eggs per day per female. At 15 and 18C, MBW oviposited infrequently; at 35C, it oviposited infrequently and died prematurely. In a cold tolerance study, half of the test cohort adults held at 0C for 1 day survived, 20% survived 2 days at 0C, and no adults survived 4 days at 0C. Of the test cohort larvae, 60% held at 0C for 1 day survived, 20% survived 2 days at 0C, and no larvae survived 4 days at 0C. Leaves from four host plants were compared for MBW oviposition rate, development time from egg to pupa, and adult size. The oviposition rate in pineapple leaves was greater than all other host types. The oviposition rate in Central American Tillandsia utriculata was similar to that in Florida T. utriculata and Tillandsia fasciculata, but the oviposition rate was greater in Florida T. utriculata than in T. fasciculata. None of the larvae that were fed leaves from Central American T. utriculata or T. fasciculata developed beyond third instar. Larvae developed almost twice as fast on pineapple leaves than on leaves from Florida T. utriculata; development times were similar from pupation to adult emergence. Adult MBW reared on pineapple leaves were significantly larger than those raised on Florida T. utriculata leaves. These studies allow us to understand the demographics of MBW in Florida and hypothesize why it is a pest in Florida but not in Central America. Eggs of the lady beetle Phaenochilus kashaya suffered 100% mortality at 0C for more than 1 day. Larvae, pupae, and adults cannot tolerate 0C for more than a few days and cannot tolerate subfreezing temperatures. Predation by P. kashaya adults on green lacewing eggs was very high in both no-choice and choice tests, but the green lacewing is not a predator of CAS. A laboratory study evaluated the effect of four concentrations (0.1, 0.5, 1.0, and 2.0 g per 100 ml of distilled water) of the entomopathogenic fungus Isaria fumosorosea on adult yellowmargined leaf beetle (YMLB), Microtheca ochroloma, mortality and leaf consumption by the beetle. There were no significant differences in adult mortality among the treatments. However, damage on plants sprayed with 0.5 and 1.0 g per 100 ml of distilled water was significantly less than that on the control plants. Control plants had on average 3.7% more damage than the treated plants. Adult female redbay ambrosia beetles (RAB), Xyleborus glabratus, were exposed separately to two strains of I. fumosorosea, one strain of Metarhizium brunneum, and one strain of Beauveria bassiana. Contact with any of the entomopathogenic fungi resulted in death of all RAB females within a few days. Beauveria bassiana and M. brunneum killed RAB females faster. Female RAB dipped in B. bassiana and M. brunneum had the highest number of viable spores attached to their bodies. No significant differences were observed in the mortality of beetles that were dipped in fungal spore solutions and then offered avocado logs as substrate, compared to beetles that were allowed to walk on logs previously treated separately with the different fungi. This is the first study to demonstrate that entomopathogenic fungi are potential biological control agents against adult RAB. Two sets of bioassays were conducted to determine the time that each of three commercial formulations of B. bassiana, I. fumosorosea, and M. brunneum require to cause mortality of the ambrosia beetles Xylosandrus crassiusculus and Xyleborus volvulus. Spore densities of the treatments were all 2,400,000 spores per ml. Contact with all of the entomopathogenic fungi resulted in death of all females. Mean survival ranged from 5.5 and 5.8 days (B. bassiana) to 17.1 and 7.8 days (I. fumosorosea) for X. crassiusculus and X. volvulus, respectively. Beauveria bassiana killed the beetle females faster, followed by M. brunneum, and then I. fumosorosea. Xylosandrus crassiusculus females dipped in B. bassiana had the highest number of viable spores attached to their bodies, with no differences between M. brunneum and I. fumosorosea. Xyleborus volvulus females dipped in I. fumosorosea had significantly more viable spores attached to their bodies, followed by M. brunneum and then B. bassiana. This is the first study to demonstrate that entomopathogenic fungi are potential biological control agents of X. crassiusculus and X. volvulus. 4) No activity. 5) Monitoring a release sites for L. franki has provided no evidence of the parasitic fly’s establishment. A proposal for field release of P. kashaya was resubmitted to APHIS; a decision on the petition is pending. 6) An open-field study evaluated guidelines for releasing nymphs of the spined soldier bug (SSB), Podisus maculiventris, on organically grown bok choy. In three trials on two organic farms during two years, three predator release rates were tested: 0, 3, and 6 nymphs per plant at the first sign of pest adults. The number of YMLB larvae and adults per plant in plots receiving SSB was significantly less than control plots in some weeks. Releases of SSB lowered plant damage by the pest and plants with SSB bugs suffered less proportional leaf area loss compared to control plants. Timely releases of first- and second-instar SSB are capable of reducing larval populations of YMLB. This control tactic will result in less leaf damage and better commodity product for organic growers. The predators should be purchased at the first sign of adult YMLB and released within five days of receipt.

Publications

• Type: Journal Articles Status: Published Year Published: 2014 Citation: Cave, R. D., and H. H. Hespenheide. 2014. An annotated list of the Buprestidae (Coleoptera) known to occur in Honduras. The Coleopterists Bulletin 68(1): 53-60.
• Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Cave, R. D. 2014. Four against one: Biological control of the cycad aulacaspis scale. Antenna Special ECE Edition: 24-25.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Ratcliffe, B. C., and R. D. Cave. 2014. Los escarabajos rinocerontes gigantes de El Salvador (Coleoptera: Scarabaeidae: Dynastinae) [El Salvadors giant rhinoceros beetles (Coleoptera: Scarabaeidae: Dynastinae)]. BIOMA 16: 7-27.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: Ratcliffe, B. C., and R. D. Cave. 2014. Un inventaire de la biodiversit� des Dynastinae (Coleoptera, Scarabaeidae) des Antilles: pr�sentation du projet. ACOREP?France: Col�opt�res de Petites Antilles. Tome II: 3-9.
• Type: Journal Articles Status: Accepted Year Published: 2015 Citation: Carrillo, D., C. A. Dunlap, P. B. Avery, J. B. Navarrete, R. E. Duncan, M. A. Jackson, R. W. Behle, R. D. Cave, J. H. Crane, A. P. Rooney, and J. E Pe�a. 2015. Entomopathogenic fungi as biological control agents for the vector of laurel wilt disease, the redbay ambrosia beetle, Xyleborus glabratus (Coleoptera: Curculionidae). Biological Control.

Progress 10/01/12 to 09/30/13

Outputs
Target Audience: Scientific community, Florida Council of Bromeliad Societies,horticulture community, organic growers Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Presentations at annual meetings of the Entomological Society of America, Florida Academy of Sciences, and Florida Entomological Society. How have the results been disseminated to communities of interest? Website, oral presentations, written reports, peer-reviewed publications What do you plan to do during the next reporting period to accomplish the goals? Increase laboratory colony of L. franki for for adequate amount of material for biological studies and field releases.Release adult L. franki at weevil-infested sites in south Florida and monitor for establishment.Complete research on the fertility of M. callizona.Researchhost plant effects on the growth and development of M. callizona.Continue research on the chemical ecology of L. franki with USDA scientists.Increase a viable colony of P. kashaya in quarantine.Publishresults ofcomparative studies of development and consumption rates of Rhyzobius lophanthae and Cryptolaemus montrouzieri with cycad aulacaspis scale as prey.Release P. kashaya in the field if approved byFDACS and APHIS PPQ.Publish resultson killing rates and development of C. rufilabris preying on yellowmargined leaf beetle eggs and larvae.Continue on-farm evaluation of the effectiveness of two release rates of P. maculiventris for managing populations of the yellowmargined leaf beetle. Completecold tolerance studies on adults of the Sri Lankan weevil.Use historical collection data and cold tolerance data for the Sri Lanka weevil to determine its potential distribution within Florida and spread to comparable climate zones in the United States. Prepare and submit manuscript on potential distribution of the Sri Lanka weevil in North America. Conductbioassays to assess the infectivity and efficacy for controlling the Sri Lanka weevil life stages by three entomopathogenic fungi, Isaria fumosorosea, Beauveria bassiana, and Metarhizium anisopliae and three OMRI- approved insecticides.Complete research on the efficacy of I. fumosorosea used as a dip treatment for biocontrol of P.madeirensis pre- and post-shipping using leaf bioassays.Begin research on the efficacy of I. fumosorosea and insect growth regulators alone and in combination as a dip treatment for biocontrol of P. madeirensis pre- and post-shipping.Perform laboratory bioassyason the practical application of various entomopathogenic fungi in infested natural areas and commercial avocado groves for managing and mitigating the spread of ambrosia beetles.

Impacts
What was accomplished under these goals? Establishment of the Honduran parasitic fly L. franki will save populations of native Florida bromeliad species which are currently threatened and endangered due to the presence of the Mexican bromeliad weevil in the state. Information from biological studies of the weevil will help us better understand the ecology of the pest, as well as improve our method for mass-rearing L. franki. Introduction and establishment of Phaenochilus kashaya will potentially control populations of the cycad aulacaspis scale on ornamental cycads in Florida and native cycads in Guam. Development of augmentation biological control technology with field releases of insectary-reared predatory stinkbugs and green lacewings will provide organic growers a needed management tactic for controlling the yellowmargined leaf beetle on crucifer crops. Initial on-farm experiments with preliminary results from data analysis indicate that augmentative releases of the predatory spined soldier bug can have a significant effect in the control of yellowmargined leaf beetle populations in organically grown crucifers. The research on entomopathogenic fungal strains will determine those that are infective against the red bay ambrosia beetle under field conditions, cause epizootics, and reduce brood production in infested natural areas and commercial avocado groves. Biological studies will provide greater insight into biological control agent/pest interactions. Discovery and descriptions of new species will enhance our knowledge of biodiversity for biological control and protection of unique environments. Objective 1) Carry out surveys and assessments of natural enemies already attacking selected invasive pest insects in Florida. This research was done previously and no new research for this objective was conducted during the reporting period. Objective 2) Conduct and coordinate overseas exploration for exotic natural enemies with potential as biological control agents of adventive herbivorous insect pests in Florida. This was not conducted due to lack of funding. Objective3) Study the natural history, climatic tolerance, behavior, and host specificity of the potential biological control agents in a quarantine facility.Temperature-dependent studies were completed on a Mexican and a Belizeanpopulation of the Mexican bromeliad weevil.The study evaluated development time at nine temperatures, oviposition rate at seven temperatures, temperature tolerance of the egg, third instar, pupa, and adult, and the longevity of adults at three temperatures.The Belizean population had a mean oviposition rate significantly lower than that of the Mexican population (0.26 eggs per day for the Belizean weevils versus 0.44 eggs per day for the Mexican population; P = 0.0102) and a significantly longer development time from egg to pupa (53 days for the Belizean weevils and 43 days for the Mexican weevils; P < 0.001). We will continue collecting data on the Belizean weevils, including temperature tolerance for eggs, larvae, pupae, and adults, and longevity at high, optimal, and low temperatures.Host range testing on Phaenochilus kashaya was performed by exposing adult beetles to a variety of insects in different stages in no-choice trials. Lesser snow scale, small nymphs of the Madeira mealybug and aphids, and green lacewing eggs and larvae, and eggs of the cabbage butterfly were consumed. In a cold tolerance study, half of the test cohort adults held at 0°C for 1 day survived, 20% survived 2 days at 0°C, and no adults survived 4 days at 0°C. Of the test cohort larvae, 60% held at 0°C for 1 day survived, 20% survived 2 days at 0°C, and no larvae survived 4 days at 0°C. A study quantified the consumption rate of Chrysoperla rufilabris when provided with immature stages of the yellowmargined leaf beetle and measured the developmental time of the predator at different temperatures under laboratory conditions. Eggs or first instars of M. ochroloma were offered to the predator, and they were kept at 10, 15, 20, or 25oC. The predator was able to complete its life cycle feeding on eggs and first instars of the beetle. The predator was not able to develop at the lowest temperature and there was an inverse interaction between developmental time and temperature. The consumption rate of the predator varied among stadia. Chrysoperla rufilabris has the potential to reduce populations of the yellowmargined leaf beetle when temperatures are above 10°C. The predator showed a 5.2-fold preference for aphids over M. ochroloma eggs and larvae. When only eggs and larvae of M. ochroloma were offered in Petri dishes, the predator did not show any preference. However, when the immature stages of M. ochroloma were offered to the predator on bok choy plants, C. rufilabris killed more first instars than eggs due to differences in the location of the prey. The entomopathogenic fungus Isaria fumosrosea infected 1st and 2nd instars after they emerged from the contaminated ovisac. Adult males were susceptible to infection when in contact with an infected female on the leaf surface or in the ovisac. The LC50 was 103.7 ± 0.56 and LC90 was 107.7 ± 0.75 blastospores/mL when sprayed on the ovisacs. Fewer nymphs emerged from the eggs as dosage increased. A cold tolerance study on the Sri Lankan weevil determined that the adults can tolerate 0°C for a day. Objective 4) Develop rearing methods for production of high-quality imported parasitoids and predators in quantities sufficient for release in the field.The rearing of Lixadmontia franki for biocontrol of the Mexican bromeliad weevil has been difficult due to the fly’s fragility to variations in environmental conditions. To improve the colony, we imported wild flies from Honduras to add to our colony. Weevil larvae (Metamasius quadrilineatus) were collected in the field in Honduras and the fly puparia that emerged from these larvae were collected and shipped to our facility. Seven shipments were made from 9 October to 5 December 2012, and a total of 635 puparia were received). Of those 635 puparia, 477 emerged as adult flies and 158 died, for 75% emergence. Objective 5) Release parasitoids and predators with a reasonable probability of establishing viable populations in the field, and monitor their post-release establishment and dispersal. One fly release was made on 25 April 2013. Because our fly population has declined so drastically, we have been unable to make any other releases and have no releases planned for the near future.Objective 6) Evaluate the ecological impacts of the released biological control agents on pest populations and elucidate interactions with other natural enemies. A field experiment in winter 2013 assessed augmentative releases of the predatory spined soldier bug (SSB), Podisus maculiventris, on populations of the yellowmargined leaf beetle on two organic farms. Each trial had three treatments: 1) six SSB per plant; 2) 12 SSB per plant; 3) no SSB. Preliminary data analysis shows that augmentative releases of the predator can lower the incidence of yellowmargined leaf beetle larvae compared to no release in mustard and turnips. Plants that received predators suffered significantly less damage than plants without predators. The performance of C. rufilabris feeding on M. ochroloma larvae under field conditions on an organic farm was evaluated. All possible combinations of cage-uncaged bok choy plants and with-without predator were evaluated. The release of the predator did not cause an important effect on M. ochroloma populations. Differences in the number of prey on the last sampling period were more related to the use of cages.

Publications

• Type: Journal Articles Status: Published Year Published: 2013 Citation: Cooper, T. M., J. H. Frank, R. D. Cave. 2013. Loss of phytotelmata due to an invasive bromeliad-eating weevil and its potential effects on faunal diversity and biogeochemical cycles. Acta Oecologcia http://dx.doi.org/10.1016/j.actao.2013.01.016
• Type: Books Status: Published Year Published: 2013 Citation: Ratcliffe, B.C., R.D. Cave, and E. B. Cano. 2013. The Dynastine Scarab Beetles of Mexico, Guatemala, and Belize. Bulletin of the University of Nebraska State Museum Volume 27. University of Nebraska State Museum, Lincoln, NE. 666 pp.
• Type: Theses/Dissertations Status: Published Year Published: 2013 Citation: Nino Beltran, A. A. 2013.Evaluation of the green lacewing Chrysoperla rufilabris Burmeister(Neuroptera: Chrysopidae) as a biological control agent of the yellowmargined leaf beetle Microtheca ochroloma Stal (Coleoptera: Chrysomelidae)
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Van Ekert, E., D. Borovsky, C. A. Powell, R. D. Cave, R. T. Alessandro, and R. G. Shatters. Targeting juvenile hormone metabolic genes in the Asian citrus psyllid (Diaphorina citri) as a strategy to reduce the spread of citrus greening disease. Presentation at the annual meeting of the Entomological Society of America, Knoxville, TN, 11-14 November 2012.
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Ben-Mahmoud, S., D. Borovsky, J. Ramos, C. A. Powell, R. D. Cave, D. G. Hall, S. L. Lapointe, and R. G. Shatters. Transgenic citrus strategies for control of the citrus root weevil, Diaprepes abbreviatus (Coleoptera: Curculionidae). Presentation at the annual meeting of the Entomological Society of America, Knoxville, TN, 11-14 November 2012.
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Cooper, T. M., R. D. Cave, and J. H. Frank. Growth and development of Metamasius callizona on four host plants. Presentation at the annual meeting of the Entomological Society of America, Knoxville, TN, 11-14 November 2012.
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Hawkings, C., J. K. Morgan, L. Shaffer, C. A. Powell, D. Borovsky, R. D. Cave, W. Dawson, S. Gowda, and R. G. Shatters. RNAi-based strategy for Asian cirus pyllid (Diaphorina citri) control: A method to reduce the spread of citrus greening disease. Presentation at the annual meeting of the Entomological Society of America, Knoxville, TN, 11-14 November 2012.
• Type: Conference Papers and Presentations Status: Other Year Published: 2012 Citation: Ni�o, A., and R. D. Cave. Development time and consumption rate of Chrysoperla rufilabris Burmeister (Neuroptera: Chrysopidae) feeding on Microtheca ochroloma St�l (Coleoptera: Chrysomelidae). Presentation at the annual meeting of the Entomological Society of America, Knoxville, TN, 11-14 November 2012.
• Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Ni�o Beltran, A. A., and R. D. Cave. Development time and consumption rate of Chrysoperla rufilabris Burmeister (Neuroptera: Chrysopidae) feeding on Microtheca ochroloma St�l (Coleoptera: Chrysomelidae). Florida Academy of Sciences, Miami Shores, FL. March 2013
• Type: Conference Papers and Presentations Status: Other Year Published: 2013 Citation: Richardson, C., P. B. Avery, and R. Cave. 2013. Exposure of yellowmargined leaf beetle larvae to entomopathogenic fungus, Isaria fumosorosea Wize, and its effects on the predatory behavior of Podisus maculiventris. Florida Academy of Sciences annual meeting. Miami Shores, FL. March, 2013

Progress 10/01/11 to 09/30/12

Outputs
OUTPUTS: No Lixadmontia franki were released due to poor production from a weak laboratory colony. Wild material was collected in Honduras to invigorate the gene pool of the laboratory colony. Temperature-dependent studies quantified the Mexican bromeliad weevil's development and oviposition rates and cold tolerance. Growth and development occurred at 18-30C. At 15C, there was no survival beyond second instar. At 35C, no eggs hatched. The number of eggs collected at temperatures of 22-33C were statistically similar at 0.3-0.5 eggs per day per female. At 15 and 18C, weevils oviposited infrequently; at 35C, weevils oviposited infrequently and died prematurely. Pupae and eggs of the weevil are the least cold tolerant, whereas the adults are very cold hardy. Preliminary studies tested the hypothesis that the Florida form of Tillandsia utriculata is more susceptible to attack by the weevil than the Central American form. A colony of the predatory lady beetle Phaenochilus kashaya from Thailand continues for host ranging testing and biological studies for finalization of a field release petition to be resubmitted to APHIS PPQ. The predator feeds on lesser snow scale and first instars of cotton aphid and Madeira mealybug, but does not feed on yellowmargined leaf beetle larvae, Diaprepes root weevil eggs, or giant whitefly nymphs. Experiments were done on 2 farms to assess augmentative releases of the predatory spined soldier bug, Podisus maculiventris, on populations of the yellowmargined leaf beetle in organically grown crucifers. In one experiment, 5 treatments were evaluated: 10 predators per 6 plants released once; 16 predators per 6 plants released once; 10 predators per 6 plants released twice; 16 predators per 6 plants released twice; and a no-release control. In a second experiment, 3 treatments were evaluated: 10 predators per 6 plants released once; 16 predators per 6 plants released once; and a no-release control. Preliminary data analysis shows that augmentative releases of the predator can lower the incidence of yellowmargined leaf beetle larvae compared to no release in mustard and turnips. Data on the developmental and killing rates of the green lacewing,Chrysoperla rufilabris, preying on yellowmargined leaf beetle eggs were collected and are being analyzed. Chrysoperla rufilabris is able to complete its development at temperatures above 10C when yellowmargined leaf beetle eggs are the prey. The predator kills more prey at 15C during the larval stage, but at 20 and 25C the predator kills more prey per day. Five entomopathogenic fungi(commercial and experimental) were tested against the red bay ambrosia beetle to determine the infectivity and pathogenicity of each strain by topical application via dipping and passive acquisition of spores via walking on contaminated bark substrates of avocado bolts. All the fungal pathogens tested against the beetle were infective, LT50s varied, and some strains acquired while the beetle was walking hindered the pest's ability to bore completely into the bolt; however, all fungi showed much potential for managing this beetle under laboratory conditions. PARTICIPANTS: Teresa Cooper, Indian River Research & Education Center, Ft. Pierce, FL. Biological studies and field releases of L. franki. Pasco Avery, Indian River Research & Education Center, Ft. Pierce, FL. Entomopathogenic fungi of invasive insects. Angie Nino, Indian River Research & Education Center, Ft. Pierce, FL. Evaluation of C. rufilabris as a biological control agent of the yellowmargined leaf beetle. Anita Neal, Indian River Research & Education Center, Ft. Pierce, FL. Biological control of the Sri Lanka weevil. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: New research is being conducted on the use of commercial and experimental fungal pathogens to control the red bay ambrosia beetle in avocados. New graduate student research will examine the ecology of the Sri Lanka weevil, Myllocerus undecimpustulatus undatus, and biological control methods.

Impacts
Establishment of the Honduran parasitic fly L. franki will save populations of native Florida bromeliad species which are currently threatened and endangered due to the presence of the Mexican bromeliad weevil in the state. Information from biological studies of the weevil will help us better understand the ecology of the pest, as well as improve our method for mass-rearing L. franki. Introduction and establishment of Phaenochilus kashaya will potentially control populations of the cycad aulacaspis scale on ornamental cycads in Florida and native cycads in Guam. Development of augmentation biological control technology with field releases of insectary-reared predatory stinkbugs and green lacewings will provide organic growers a needed management tactic for controlling the yellowmargined leaf beetle on crucifer crops. Initial on-farm experiments with preliminary results from data analysis indicate that augmentative releases of the predatory spined soldier bug can have a significant effect in the control of yellowmargined leaf beetle populations in organically grown crucifers. The research on entomopathogenic fungal strains will determine those that are infective against the red bay ambrosia beetle under field conditions, cause epizootics, and reduce brood production in infested natural areas and commercial avocado groves. Biological studies will provide greater insight into biological control agent/pest interactions. Discovery and descriptions of new species will enhance our knowledge of biodiversity for biological control and protection of unique environments.

Publications

• Manrique, V., R. Diaz, C. Montemayor, D. Serrano, and R. D. Cave. 2012. Temperature-dependent development and cold tolerance of Microtheca ochroloma (Coleoptera: Chrysomelidae), a pest of cruciferous crops in the southeastern United States. Annals of the Entomological Society of America 105(6):859-864.
• Montemayor, C. M., and R. D. Cave. 2012. Evaluation of the predation capacity of Podisus maculiventris (Hemiptera: Pentatomidae) on Microtheca ochroloma (Coleoptera: Chrysomelidae) in field cages. Journal of Economic Entomology 105(5):1719-1725.
• Manrique, V., D. Mancero-Castillo, R. D. Cave, and R. Nguyen. 2012. Effect of temperature on the development and consumption of Phaenochilus kashaya (Coleoptera: Coccinellidae), a predator of the cycad aulacaspis scale, Aulacaspis yasumatsui. Biocontrol Science & Technology 22(11):1245-1253.

Progress 10/01/10 to 09/30/11

Outputs
OUTPUTS: Six female Lixadmontia franki were released the Fakahatchee Strand State Preserve in August. No indication of reproduction in the field was detected. Wild material was collected in Honduras to invigorate the gene pool of the laboratory colony. Temperature-dependent studies tested the Mexican bromeliad weevil's development and oviposition rates, cold tolerance, and survivorship of adults. A new rearing method was developed by which egg-laying females are held in vials and provided pineapple leaves in which to oviposit. Leaves with eggs are collected and reared until larvae emerge. Three to 4 larvae are placed on a pineapple top, which are held for a week and then placed in the cage with gravid L. franki. The new method produces rates of parasitism similar to the old method. The new method allows for better control of the egg-laying population, reduces the variability of instars within a top, and allows calculation of the number of larvae per top with much greater certainty. A colony of the predatory lady beetle Phaenochilus n. sp. from Thailand continue to be increased to complete host ranging testing and biological studies for finalization of a field release petition to be resubmitted to APHIS. Experiments were done on 2 organic farms to assess the augmentative release of the predatory spined soldier bug, Podisus maculiventris, on populations of the yellowmargined leaf beetle. Five treatments were evaluated: 10 predators per 6 plants released once; 16 predators per 6 plants released once; 10 predators per 6 plants released twice; 16 predators per 6 plants released twice; and a no-release control. Spined soldier bug eggs were purchased from Rincon-Vitova, Inc. The first releases took place 5 weeks after planting and the second release 2 weeks after planting. Every 5 days, 6 plants per plot were examined and the numbers of beetle larvae, pupae, and adults, and number of spined soldier bug nymphs were recorded. Data is being analyzed. To survey the insects associated with the red bay ambrosia beetle, 5 beetle-infested red bay logs were placed at each of 4 sites in April and recovered in May, and 5 logs placed at the same sites in September and recovered in October. Logs were placed individually in carton boxes and held in the laboratory for 1 month. Insects were then collected from the logs and sent to collaborators for identification. Data is being analyzed. PARTICIPANTS: : Teresa Cooper, Indian River Research & Education Center, Ft. Pierce, FL. Biological studies and field releases of L. franki. Ru Nguyen, Florida Dept. of Agriculture & Consumer Services, Gainesville, FL. Biological studies and colony maintenance of Phaenochilus. Jorge Pena, Tropical Research & Education Center, Homestead, FL. Insects associated with red bay ambrosia beetle. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Establishment of the Honduran parasitic fly L. franki will save populations of native Florida bromeliad species which are currently threatened and endangered due to the presence of the Mexican bromeliad weevil in the state. Introduction and establishment of Phaenochilus n. sp. will potentially control populations of the cycad aulacaspis scale on ornamental cycads in Florida and native cycads in Guam. Development in the field of augmentation biological control technology with field releases of insectary-reared predatory stinkbugs will provide organic growers a needed management tactic for controlling the yellow-margined leaf beetle on crucifer crops. Knowledge of insects associated with the red bay ambrosia beetle will be useful for assessing the non-target impact of any exotic biological control agents that might be introduced. Biological studies will provide greater insight into parasitoid/host interactions. Discovery and descriptions of new species will enhance our knowledge of biodiversity for biological control and protection of unique environments.

Publications

• Castillo, J. A., P. B. Avery, R. D. Cave, and C. O. Montemayor. 2011. Mortality of the cycad aulacaspis scale (Hemiptera: Diaspididae) by the entomopathogenic fungus Isaria fumosorosea (Wize) under laboratory conditions. Journal of Entomological Science 46(3): 256-264.
• Cooper, T. M., J. H. Frank, R. D. Cave, M. S. Burton, J. S. Dawson, and B. W. Smith. 2011. Release and monitoring of a potential biological control agent, Lixadmontia franki (Diptera: Tachinidae), to control an invasive bromeliad-eating weevil, Metamasius callizona (Coleoptera: Curculionidae), in Florida. Biological Control 59(3): 319-325.
• Montemayor, C. M., and R. D. Cave. 2011. Development time and predation rate of Podisus maculiventris (Hemiptera: Pentatomidae) preying on Microtheca ochroloma (Coleoptera: Chrysomelidae). Environmental Entomology 40(4): 948-954.
• Pena, J. E., J. H. Crane, J. L. Capinera, R. E. Duncan, P. E. Kendra, R. C. Ploetz, S. McLean, G. Brar, M. C. Thomas, and R. D. Cave. 2011. Chemical control of the redbay ambrosia beetle, Xyleborus glabratus, and other Scolytinae (Coleoptera: Curculionidae). Florida Entomologist 94: 882-896.

Progress 10/01/09 to 09/30/10

Outputs
OUTPUTS: Field releases of 199 adults of the parasitic fly Lixadmontia franki, a biological control agent of the Mexican bromeliad weevil, Metamasius callizona, were made at 2 sites in south Florida on 4 dates. No indication of establishment of feral populations of this natural enemy is yet evident. A collaborative study of the chemical ecology of L. franki is determining what chemicals attract female flies to weevil-infested bromeliads, then to use those chemicals to create an attractive trap for detecting presence of the fly in the field. Testing female fly antennal response to volatiles collected from weevil-infested bromeliads showed clear responses to several chemicals. Several no-choice (without CAS) and choice (with CAS) tests were conducted to evaluate the host range of Phaenochilus n. sp. adults. Results showed that Phaenochilus adults will not feed on spider mites, cactus moth larvae, chinch bugs, or eggs and larvae of the tropical soda apple beetle, Gratiana boliviana. There was some feeding on butterfly eggs in a no-choice test, but no feeding occurred in a choice test. A proposal for field release of Phaenochilus was submitted to APHIS and NAPPO; more data, including a formal name and description of the species, were requested pending approval. A field study determined guidelines for releasing the spined soldier bug, Podisus maculiventris, for control of the yellowmargined leaf beetle, Microtheca ochroloma, in field cages. The release rate of 16 first instar P. maculiventris per 6 large turnip plants significantly reduced the M. ochroloma population over time compared to 4 and 10 predators released per 6 plants. For six small turnip plants, a release rate of 10 P. maculiventris significantly reduced the M. ochroloma population over time compared to the lowest release rate, but its pest population suppression capabilities were not significantly different from the higher release rate. Laboratory experiments evaluated the infectivity and lethal time (LT) of I. fumosorosea on M. ochroloma at the concentration of 1g of the product PFR-97 in 100 ml of distilled water in the laboratory. The larval stage is more susceptible to PFR-97 than eggs, pupae, and adults. Infectivity rates of 17 and 20% were confirmed in the first and third instars of M. ochroloma, respectively. The LT10 for first and third instars of M. ochroloma were 4 and 3 days, respectively. Concentrations of 1, 2, 3, and 4 g of PFR-97 in 100 ml of distilled water were applied to first instars of M. ochroloma to compare infectivity, LT, and lethal concentrations (LC). Confirmed infectivity rates for 1, 2, 3, and 4 g concentrations were 2, 5, 10, and 27%, respectively. The LT10 and LT25 for the 4 g concentration were 3.4 and 5.7 days, respectively. The LC10 and LC25 were 1.4 g and 5.5 g per 100 ml of distilled water, respectively. PARTICIPANTS: Dr. Ru Nguyen, Florida Dept. of Agriculture & Consumer Services, Division of Plant Industry, Gainesville, FL. Biological studies and colony maintenance for Phaenochilus n. sp. Dr. Teresa Cooper, Indian River Research & Education Center, Ft. Pierce, FL. Biological studies, field releases, and colony maintenance for Lixadmontia franki. Mr. Bradley Smith, Indian River Research & Education Center, Ft. Pierce, FL. Biological studies, field releases, and colony maintenance for Lixadmontia franki. Dr. Paul Robbins, USDA Horticultural Research Laboratory, Ft. Pierce, FL. Chemical ecology research on Lixadmontia franki. Ms. Cecil Montemayor, Indian River Research & Education Center, Ft. Pierce, FL. Biological studies and field research on biological control of Microtheca ochroloma. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Establishment of the Honduran parasitic fly will save populations of native Florida bromeliad species which are currently threatened and endangered due to the presence of the Mexican bromeliad weevil in the state. Introduction and establishment of Phaenochilus n. sp. will potentially control populations of the cycad aulacaspis scale on ornamental cycads in Florida and native cycads in Guam. Development in the field of augmentation biological control technology with field releases of insectary-reared predatory stinkbugs will provide organic growers a needed management tactic for controlling the yellow-margined leaf beetle on crucifer crops. Biological studies will provide greater insight into parasitoid/host interactions. Discovery and descriptions of new species will enhance our knowledge of biodiversity for biological control and protection of unique environments.

Publications

• Manrique, V., C. O. Montemayor, R. D. Cave, E. A. Skvarch, and B. W. Smith. 2010. Effect of straw mulch on populations of Microtheca ochroloma (Coleoptera: Chrysomelidae) and ground predators in turnip Brassica rapa in Florida. Florida Entomologist 93(3): 407-411.
• Montemayor, C. O., and R. D. Cave. 2009. Prospects for biological control of the yellow-margined leaf beetle, Microtheca ochroloma Stal. Proceedings of the Florida State Horticultural Society 122: 250-252.
• Ratcliffe, B. C., and R. D. Cave. 2010. New species of Orizabus (Coleoptera: Scarabaeidae: Dynastinae: Pentodontini) from Mexico and Guatemala, with a revised key and checklist of the species in the genus. Insecta Mundi 0127: 1-16.
• Ratcliffe, B. C., and R. D. Cave. 2010. The Dynastinae (Coleoptera: Scarabaeidae) of the Cayman Islands (West Indies), with a description of Tomarus adoceteus, new species, (Pentodontini) and Caymania nitidissima, new genus and species, (Phileurini) from Little Cayman. Insecta Mundi 0139: 1-15.
• Orapa, W., and R. D. Cave. 2010. Coccinellids for combating cycad scale: promising news from Palau and the USA. Biocontrol News & Information 31(1): 2N-3N.
• Vecco, C. D., R. Pinedo, M. Fernandez, and R. D. Cave. 2009. Analisis funcional del nicho ecologico de Syphrea sp. (Coleoptera: Chrysomelidae) asociado a Plukenetia volubilis L. (Euphorbiales: Euphorbiaceae) en la Amazonia peruana. Ceiba 50: 40-46.