Evaluation of the strains of Acinetobacter and Enterobacter as potential biocontrol agents against Ralstonia wilt of tomato

Bacterial wilt (Ralstonia solanacearum) of tomato, Lycopersicon esculentum, causes a considerable amount of damage to tomato in Southern China. Biological control is one of the more promising approaches to reduce the disease incidence and yield losses caused by this disease. Based on antagonistic activity against R. solanacearum and three soil-borne fungal pathogens as well as biocontrol efficacy in the greenhouse, two bacterial strains Xa6 (Acinetobacter sp.) and Xy3 (Enterobacter sp.) were selected out of fourteen candidates as potential biocontrol agents. In order to find a suitable antagonist inoculation method, we compared the methods of root-dipping with soil-drenching in the aspects including rhizocompetence, biocontrol efficacy, and effect of promoting plant growth under greenhouse conditions. The drenching treatment resulted in a higher biocontrol efficacy and plant-yield increase, and this method was also easier to operate in the field on a large scale. Field trials were conducted for further evaluation of these two antagonistic strains. In both greenhouse and field experiments, the strain Xy3 had a better control effect against bacterial wilt than Xa6 did, while Xa6 caused higher biomass or yield increases. As recorded on the 75th day after treatment in two field experiments, biocontrol efficacy of Xy3 was about 65% in both field trials, and the yield increases caused by Xa6 were 32.4 and 40.7%, respectively, in the two trials. This is the first report of an Acinetobacter sp. strain used as a BCA against Ralstonia wilt of tomato.     

Evaluation ofHadena perplexa [Lepidoptera: Phalaenidae] as a biocontrol agent of bladder campionSilene vulgaris [Caryophyllaceae] in Canada: Rearing and host specificity

Rearing techniques and results of preliminary host range tests are reported forHadena perplexa (Denis & Schiffermuller) (Lep.: Phalaenidae) a candidate biocontrol agent against the weed bladder campion,Silene vulgaris (Moench) Garcke, in Manitoba, Canada. In the laboratory, it was necessary to pipette a 15% honey solution in water into the flowers as food for the adult moths. When reared singly to avoid cannibalism, 56% of the 1^st instar larvae developed to pupae. Larvae fed on a natural diet for 10 days can then be reared on either one of 2 artificial diets. Choice oviposition tests and no-choice larval feeding tests were conducted with plant species closely related toS. vulgaris in the generaSilene, Dianthus, Gypsophila, Lychnis, Saponaria. Species in 4 of 5 of these genera were accepted for oviposition, and species in all 5 genera supported the development of 1^st instar larvae to the pupal stage.H. perplexa should not be introduced into Canada.      

Insect natural enemies of coffee green scales [Hemiptera: Coccidae] in Kenya and their potential for biological control ofCoccus celatus andC. viridis in Papua New Guinea

Coffee green scales,Coccus celatus De Lotto andC. viridis (Green), are major pests of coffee in Papua New Guinea (PNG). Surveys for insect natural enemies of coffee green scales were conducted in Kenya to find species suitable for introduction to PNG. Natural enemy communities in Kenya were found to be diverse, particularly those located in the central highlands where 3 primary parasitoids,Metaphycus stanleyi Compere,M. baruensis Noyes andDiversinervus stramineus Compere (Encyrtidae), were dominant onC. celatus andC. alpinus De Lotto. It is suggested that these 3 parasitoids may be potential biological control agents forC. celatus. Several insect predators were found associated withC. celatus in the central highlands but most were only common in ant-free scale populations. An aphelinid primary parasitoid,Coccophagus rusti Compere, was the only natural enemy ofC. viridis located. This species, however, could be of potential as a biological control agent, especially in view of its previous use in biological control against soft scales.      

Osmotic stress adaptation, compatible solutes accumulation and biocontrol efficacy of two potential biocontrol agents on Fusarium head blight in wheat

Fusarium head blight (FHB) caused by Gibberella zeae (anamorph = Fusarium graminearum) is a devastating disease that causes extensive yield and quality losses to wheat in humid and semi-humid regions of the world. Biological control has been demonstrated to be effective under laboratory conditions but a few biocontrol products have been effective under field conditions. The improvement in the physiological quality of biocontrol agents may improve survival under field conditions, and therefore, enhance biocontrol activity. Bacillus subtilis RC 218 and Brevibacillus sp. RC 263 were isolated from wheat anthers and showed significant effect on control of FHB under greenhouse assays. This study showed the effect of water availability measured as water activity (a_W) using a growth medium modified with NaCl, glycerol and glucose on: (i) osmotic stress tolerance, (ii) viability in modified liquid medium, (iii) quantitative intracellular accumulation of betaine and ectoine and (iv) the biocontrol efficacy of the physiologically improved agents. Viability of B. subtilis RC 218 in NaCl modified media was similar to the control. Brevibacillus sp. RC 263 showed a limited adaptation to growth in osmotic stress. Betaine was detected in high levels in modified cells but ectoine accumulation was similar to the control cells. Biocontrol activity was studied in greenhouse assays on wheat inoculated at anthesis period with F. graminearum RC 276. Treatments with modified bacteria reduced disease severity from 60% for the control to below 20%. The physiological improvement of biocontrol agents could be an effective strategy to enhance stress tolerance and biocontrol activity under fluctuating environmental conditions.     

Investigations on the parasitism ofOtiorrhynchus salicicola andO. Sulcatus [Col.: Curculionidae] byHeterorhabditis sp. [Nematoda]

Experiments were conducted to study the efficacy of the insect parasitic nematodeHeterorhabditis sp. (HW79) as a biological control agent ofOtiorrhynchus salicicola. This weevil species is reported as a pest of ornamental plants in Switzerland and Italy. Dipping plastic boxes containing heavily infested cuttings of laurel (Prunus laurocerasus) in a nematode suspension resulted in approximately 100% parasitisation of full-grown larvae, pupae and non-emerged young adults. The average dose resulting from dipping varied between 56,000 and 62,000 nematodes per liter soil. This experiment was run under natural outdoor conditions. In a further outdoor experiment, pottedLigustrum plants were inoculated with eggs ofO. salicicola and later 20,000 infective juvenile nematodes per liter soil were added to the soil surface. The resulting weevil mortality in the treated pots was 78%. In seven greenhouse tests using the same nematode dose in pots with horticultural soil to which weevil larvae had been added, weevil mortality varied between 76% and 100%, the arithmetic average being 90%. These results indicate that Heterorhabditid nematodes may provide an effective means of controllingO. salicicola. In an other experiment usingO. sulcatus larvae, the influence of application time on nematode efficacy was investigated. When nematodes were added a few days before weevil larvae had hatched from the eggs, no parasitic effect was obtained. Nematode applications done shortly after larval hatching however, resulted in complete weevil control. These results are of significance in timing nematode applications in practice.      

Notes on natural enemies attackingLepidosaphes species [Homoptera: Diaspididae] associated withCitrus in Costa Rica

Aphytis lepidosaphes Compere (Hymenoptera: Aphelinidae) parasitize two species ofLepidosaphes onCitrus in Costa Rica.Zagloba beaumonti andPentilia discors (Col.: Coccinellidae) also feed on both species.      

Trichilogaster sp. [Hymenoptera: Pteromalidae], a potential biocontrol agent for the weedAcacia pycnantha [Fabaceae]

The gall wasp,Trichilogaster sp., was imported from Australia to assess its potential as an agent for the control of the invasive shrub/treeAcacia pycnantha Benth. in South Africa. Host specificity tests indicate safety for release; of 19 tree/shrub species tested, including 16 species closely related toA. pycnantha, galls developed only onA. pycnantha. However, galling intensity remained consistently low on the host plant; only 21–29% of the branches exposed to the wasp were galled during 3 years of rearing. Neither the prolonged presence of males in test cages (someTrichilogaster species are thelytokous) nor the stage of maturity of reproductive buds exposed to oviposition affected the percentage of branches galled. It is not recommended thatTrichilogaster sp. be released before the possibility of insect-plant homeostasis or mis-matching of wasp and host plant populations/strains/subspecies is investigated, especially since galling intensities of 30% were ineffective in reducing seed production of a relatedTrichilogaster species/Acacia association.      

The biology and host specificity of the stem-boring weevilListronotus setosipennis [Col.: Curculionidae] A potential biocontrol agent forParthenium hysterophorus [Asteraceae]

The stem-boring weevilListronotus setosipennis (Hustache) is widespread and damaging to its hostParthenium hysterophorus L. in northern Argentina and southern Brazil. In detailed host-testing it was shown to have a restricted host-range and, despite some feeding and development on sunflower in tests, to be a safe biological control agent againstP. hysterophorus. Field releases in Queensland, Australia took place from 1982 to 1986 and the weevil has established over several thousand hectares at numerous sites. Spread is however slow and the effect on the plant still negligible.      

Potential of an antagonistic bacterium isolate obtained fromLentinus lepideus basidiospores as a biocontrol agent

TheBurkholderia sp. isolate 87-11 obtained from basidiospores ofLentinus lepideus was antagonistic against severalPythium andRhizoctonia isolates. The bacterium was tested against soilborne diseases of five plants caused byP. aphanidermatum andR. solani by soil and seed application, and its potential as a biocontrol agent is discussed.     

Isolation and identification of entomopathogenic nematodes from citrus orchards in South Africa and their biocontrol potential against false codling moth

A survey was conducted to determine the diversity and frequency of endemic entomopathogenic nematodes (EPN) in citrus orchards in the Western Cape, Eastern Cape and Mpumalanga provinces of South Africa. The main aim of the survey was to obtain nematodes as biological control agents against false codling moth (FCM), Thaumatotibia leucotreta, a key pest of citrus in South Africa. From a total of 202 samples, 35 (17%) tested positive for the presence of EPN. Of these, four isolates (11%) were found to be steinernematids, while 31 (89%) were heterorhabditids. Sequencing and characterisation of the internal transcribed spacer (ITS) region was used to identify all nematode isolates to species level. Morphometrics, morphology and biology of the infective juvenile (IJ) and the first-generation male were used to support molecular identification and characterisation. The Steinernema spp. identified were Steinernema khoisanae, Steinernema yirgalemense and Steinernema citrae. This is the first report of S. yirgalemense in South Africa, while for S. citrae it is the second new steinernematid to be identified from South Africa. Heterorhabditis species identified include Heterorhabditis bacteriophora, Heterorhabditis zealandica and an unknown species of Heterorhabditis. Laboratory bioassays, using 24-well bioassay disks, have shown isolates of all six species found during the survey, to be highly virulent against the last instar of FCM larvae. S. yirgalemense, at a concentration of 50 IJs/FCM larva caused 100% mortality and 74% at a concentration of 200 IJs/pupa. Using a sand bioassay, S. yirgalemense gave 93% control of cocooned pupae and emerging moths at a concentration of 20 IJs/cm². This is the first report on the potential use of EPN to control the soil-borne life stages of FCM, which includes larvae, pupae and emerging moths. It was shown that emerging moths were infected with nematodes, which may aid in control and dispersal.     

A survey of black scale,Saissetia oleae [Hom.: Coccidae] parasitoids [Hym.: Chalcidoidea] in southern California

A survey of black scaleSaissetia oleae (Olivier) parasitoids present in southern California was conducted between September 1987 and September 1989. From 308 collections of black scale-infested citrus, olive, and oleander twigs from 19 sites in southern California, 1,610 specimens were collected. Nine primary and six secondary parasitoid species were identified. Four primary species were abundant in southern California:Metaphycus bartletti Annecke & Mynhardt,M. helvolus (Compere),Scutellista caerulea (Fonscolombe) (=S. cyanea Motschulsky), andDiversinervus elegans Silvestri. The most common secondary parasitoids wereMarietta mexicana (Howard),Cheiloneurus noxius Compere, andTetrastichus minutus (Howard). In the coastal region of southern California,M. bartletti was the most abundant parasitoid, followed in order byD. elegans, S. Caerulea, andM. helvolus. In the intermediate and interior regions,M. helvolus was most abundant.D. elegans was second most abundant in the intermediate region, but was rare in the interior region.M. bartletti was second in abundance in the interior region and third in the intermediate region.      

Effects of the concurrent exclusion of ants and earwigs on aphid abundance in an organic citrus grove

Based on the well-known mutualism between ants (Hymenoptera: Formicidae) and aphids (Homoptera: Aphididae), we conducted a five-year experiment of ant-exclusion from the canopies of citrus trees as a possible method of biological control of aphids. However, our results showed that the exclusion of ants from the canopies increased, instead of reducing, aphid abundance. To explain this unexpected result, we reasoned that the exclusion of ants from the canopies might also have excluded crawling insects that prey on aphids, such as the European earwig (Forficula auricularia L., Dermaptera: Forficulidae). Such a possibility is supported by the negative relationship between aphid density and the abundance of earwigs, consistent with a top-down control of aphids by earwigs. In contrast, the abundance of other aphid predators (Coleoptera: Coccinellidae, and Heteroptera) had no such negative effect on aphid density but a positive one, suggesting a bottom-up control, and showed no differences between control and ant-excluded trees. Thus, the most likely explanation for the increase in aphid abundance in the ant-excluded trees is the absence of earwigs from the canopies of the experimental trees, providing further evidence of the major role that earwigs play as control agents of aphids in cultivated trees.     

Production of chitinase and its optimization from a novel isolate Alcaligenes xylosoxydans: potential in antifungal biocontrol

A novel, highly chitinolytic strain of Alcaligenes xylosoxydans was isolated which showed potential for use as an antifungal biocontrol agent for the control of two fungal plant pathogens. It could degrade and utilize dead mycelia of Rhizoctonia bataticola and Fusarium sp. (fungal plant pathogens of Cajanus cajan). In vitro it could inhibit the growth of Fusarium sp. and R. bataticola. Chitin at 10–15 g/l was found to be good carbon and nitrogen source. Alcaligenes xylosoxydans showed optimum chitinase production at 72 h, pH optima at 8 and growth peak at 120 h. Yeast extract, arabinose, Tween 20 and several other surfactants enhanced chitinase production.     

Observations on the biology and host specificity ofEpiblema strenuana [Lepidoptera,Tortricidae], a potential biocontrol agent forParthenium hysterophorus [Compositae]

The tortricid mothEpiblema strenuana (Walker), which attacksAmbrosia spp. over most of North America, is shown to have the annual weedParthenium hysterophorus L. [Compositae] as its main host in northern Mexico. The larvae produce galls in the stems and growing-points which can considerably stunt growth when young plants are attacked. In tests it was shown to have a restricted host-range and not to damage any composite of economic importance. Its release in Australia for trial as a biological control agent againstP. hysterophorus has been approved.      

Temperatures lethal for citrus blackfly parasites,Encarsia opulenta andE. Smithi [Hym.: Aphelinidae]

Seasonally acclimatized adult and immature parasites of the citrus blackfly (CBF),Aleurocanthus woglumi Ashby, were exposed to high or low temperature extremes for 3 h periods. Death of all summer adults ofEncarsia opulenta Silvestri andE. smithi Silvestri occurred between 35° and 40°C. Within CBF hosts,E. opulenta were not able to emerge when temperatures reached between 45° and 50°C. In winter experiments adults of bothEncarsia species succumbed between −5° and −10°C. In a comparison of the 2 seasonal tests, a higher percentage ofE. smithi adults were able to survive both higher and lower temperatures thanE. opulenta, but the main interspecific difference was the ability ofE. opulenta within CBF to survive −10° to − 15°C whileE. smithi did not. Limited data forAmitus hesperidum Silvestri [Hym.: Platygasteridae] indicated that the immatures survived better at low, and not as well at high, temperatures as either species ofEncarsia. Florida Agricultural Experiment Station Journal Series # 5549.     

Selection of rhizosphere-competent Pseudomonas strains as biocontrol agents in tropical soils

Pseudomonas strains were isolated from the rhizosphere of maize grown in yellow-red latosol from Rio de Janeiro, Brazil, to serve as a delivery system for heterologous genes and for risk assessment studies in tropical soils. Selected strains were modified by insertion of the cryIVB gene from Bacillus thuringiensis and tested for pathogenicity gene expression against larvae of a susceptible model species, Anopheles aquasalis. Modified strains Br8 and Br12 showed similar survival performance to their parental strains, and presented a viable density of 10⁷ c.f.u./g dry soil 30 days after release. A strain of P. fluorescens (Br12) that presented positive results for gene expression and the best survival performance, was selected for risk assessment studies in soil microcosms.     

Predation byCheilomenes vicina [Coleoptera: Coccinellidae] on the cowpea aphid,Aphis craccivora [Homoptera: Aphididae]: Effect of prey stage and density

Consumption of larvae and females ofAphis craccivora Koch by 1st and 4th larvae and adults ofCheilomenes vicina (Muls.) was studied under fluctuating temperature (24–30°C). The early aphid instars were consumed in significantly greater numbers than later instars and females. The feeding rates ofC. vicina were significantly positively correlated with the population density of prey. The number of prey consumed daily by each predator stage tested, increased more steeply at lower than at higher prey densities, exhibiting thus the type 2 functional response

---

Résumé Consommation des larves et des ♀ d'Aphis craccivora par les11^e stades, les 4^e stades et les adultes deCheilomenes vicina a été étudiée à une température variant de 24 à 30°C. Les jeunes stades du puceron sont consommés en nombre plus grand significativement que les derniers stades et les ♂. Les taux d'alimentation deC. vicina manifestent une corrélation hautement significative avec la densité de population de la proie. Le nombre de proies consommées quotidiennement par chaque stade considéré du prédateur augmentait plus brusquement aux faibles qu'aux fortes densités de proies, manifestant ainsi chez le prédateur le type 2 de réponse fonctionnelle.

     

The biology and behaviour ofIphiaulax varipalpis [Hymenoptera: Braconidae] as a parasite ofDirphya nigricornis [Coleoptera: Cerambycidae]

The ability of the braconid parasite,Iphiaulax varipalpis Cary to control the beetleDyrphya nigricornis Olivier was evaluated. Field collected samples and specimens reared in polyethylene paper tubes under laboratory conditions (23.0°C±1.0 at 70% r.h) were used. Field parasitism was low (10.72%) while laboratory parasitism was high (57.66%). The parasite detected the hosts very easily in the laboratory. This ability is minimised in thick canopies ofCoffea arabica L. Host colonization was brisk (90 seconds) and confined to a near absolute area of discovery, measuring 4.18±0.41 cm. The parasite reproduces gregariously with female lifespan being approximately double the male period. The parasites are very active at the age of 10 days. As they age their potential fecundity (232 oocytes) reduces drastically with oviposition.      

Diurnal and spatial patterns of Phytoseiidae in the citrus canopy

Phytoseiid mites were sampled in a grapefruit orchard at various times of the day to study their diurnal and seasonal distributions in the tree canopy. Samples were collected on 14, 20 and 28 October 1999 at 2 h intervals from 0600 to 2200 h. Similar samples were collected in a grapefruit orchard at 3 h intervals from 0600 to 2100 h on 9 and 16 March and on 17 and 24 August 2000 for phytoseiid mites. No differences in numbers of phytoseiid motiles were observed among the hours sampled in any of the three months. However, significant differences were observed in the number of phytoseiids per leaf based on location within the tree (eastern, western sides of the canopy or interior). Interior leaves collected in March and August 2000 had higher numbers of phytoseiids than exterior leaves taken from either the eastern or western sides of the tree canopy. Phytoseiids were more abundant in the March 2000 samples (mean = 1.10 phytoseiids/leaf) than in either October 1999 or August 2000 samples (mean = 0.16 and 0.19 phytoseiids/leaf, respectively). Prevalent phytoseiid species were Typhlodromalus peregrinus (Muma) (42.1%) and Iphiseiodes quadripilis (Banks) (50.4%) in October 1999, Typhlodromalus peregrinus (Muma) (76.2%) in March 2000, and Euseius mesembrinus (Dean) (54%) in August 2000.