Interactions between<Emphasis Type="Italic"> Trichoderma pseudokoningii</Emphasis> strains and the arbuscular mycorrhizal fungi<Emphasis Type="Italic"> Glomus mosseae</Emphasis> and<Emphasis Type="Italic"> Gigaspora rosea</Emphasis>

The interaction between Trichoderma pseudokoningii (Rifai) 511, 2212, 741A, 741B and 453 and the arbuscular mycorrhizal fungi Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe BEG12 and Gigaspora rosea Nicolson & Schenck BEG9 were studied in vitro and in greenhouse experiments. All T. pseudokoningii strains inhibited the germination of G. mosseae and Gi. rosea except the strain 453, which did not affect the germination of Gi. rosea. Soluble exudates and volatile substances produced by all T. pseudokoningii strains inhibited the spore germination of G. mosseae. The germination of Gi. rosea spores was inhibited by the soluble exudates produced by T. pseudokoningii 2212 and 511, whereas T. pseudokoningii 714A and 714B inhibited the germination of Gi. rosea spores by the production of volatile substances. The strains of T. pseudokoningii did not affect dry matter and percentage of root length colonization of soybean inoculated with G. mosseae, except T. pseudokoningii 2212, which inhibited both parameters. However, all T. pseudokoningii strains decreased the shoot dry matter and the percentage of AM root length colonization of soybean inoculated with Gi. rosea. The saprotrophic fungi tested seem to affect AM colonization of root by effects on the presymbiotic phase of the AM fungi. No influence of AM fungi on the number of CFUs of T. pseudokoningii was found. The effect of saprotrophic fungi on AM fungal development and function varied with the strain of the saprotrophic species tested.     

Effect of Mannich bases on some plant pathogenic fungi

3-(2-Pyridyl)-3-iminoisatin, 1-piperidinomethyl-3-(2-pyridyl)-3-iminoisatin, and 1-acetyl-3-(2-pyridyl)-3-iminoisatin affect spore germination ofAlternaria alternata, A. carthemi, Curvularia lunata, Fusarium oxysporum f.sp.cieri andF. udum and influence the development of powdery mildew (Erysiphe pisi) on pea under glasshouse condition as well as conidial germination ofE. pisi on excised pea leaves. Spore germination was inhibited in the sequence 1-acetyl-3-(2-pyridyl)-3-iminoisatin > 1-piperidinomethyl-3-(2-pyridyl)-3-iminoisatin > 3-(2-pyridyl)-3-iminoisatin followed the order accordingly. The powdery mildew development and conidial germination ofE. pisi 1-piperidinomethyl-3-(2-pyridyl)-3-iminoisatin > 1-acetyl-3-(2-pyridyl)-3-iminoisatin > 3-(2-pyridyl)-3-iminoisatin. The chemicals were compared with commonly used antifungal fungicides.     

Effect of copper on germination and seedling growth ofMinuartia,Silene, Alyssum andThlaspi

The influence of increasing copper concentrations on seed germination, seedling survival and radicle length ofMinuartia hirsuta, Silene compacta, Alyssum montanum andThlaspi ochroleucum was studied. Seed germination was highly affected by the higher Cu²⁺ concentrations (80 and 160 μM), while lower Cu²⁺ concentrations seemed to be necessary for seed germination, even for the plants originated from non-Cu²⁺-rich soils (i.e. A. montanum). Nevertheless, plants originated from Cu²⁺-rich soils (M. hirsuta, S. compacta) showed a higher demand of Cu²⁺ for rapid seed germination. Cu²⁺ at higher concentrations severely reduced growth rate of radicle, especially inA. montanum andT. ochroleucum. These data clearly indicate the reduced suitability of the above mentioned plant species for reclamation on Cu²⁺ soils. Lower Cu²⁺-concentrations had no influence on seedling survival inM. hirsuta andS. compacta, but a progressive reduction of a number of survived seedlings with increasing Cu²⁺ concentration was found, that was more pronounced inA. montanum andT. ochroleucum.     

Distribution of similar self-incompatibility (<Emphasis Type="Italic">S</Emphasis>) haplotypes in different genera,<Emphasis Type="Italic"> Raphanus</Emphasis> and<Emphasis Type="Italic"> Brassica</Emphasis>

The nucleotide sequences of ten SP11 and nine SRK alleles in Raphanus sativus were determined, and deduced amino acid sequences were compared with those of Brassica SP11 and SRK. The amino acid sequence identity of class-I SP11s in R. sativus was about 30% on average, the highest being 52.2%, while that of the S domain of class-I SRK was 77.0% on average and ranged from 70.8% to 83.9%. These values were comparable to those of SP11 and SRK in Brassica oleracea and B. rapa. SP11 of R. sativus S-21 was found to be highly similar to SP11 of B. rapa S-9 (89.5% amino acid identity), and SRK of R. sativus S-21 was similar to SRK of B. rapa S-9 (91.0%). SP11 and SRK of R. sativus S-19 were also similar to SP11 and SRK of B. oleracea S-20, respectively. These similarities of both SP11 and SRK alleles between R. sativus and Brassica suggest that these S haplotype pairs originated from the same ancestral S haplotypes.     

Effect of<Emphasis Type="Italic">Trichoderma</Emphasis> species on growth of Fusarium<Emphasis Type="Italic">proliferatiom</Emphasis> and production of fumonisins,fusaproliferin and beauvericin

Trichoderma species has been suggested as potential biocontrol agent forFusarium verticillioides on maize. In this cereal,F. verticillioides and F. proliferatum contributed to fumonisin accumulation. In addition,F. proliferatum could produce beauvericin and fusaproliferin. The aim of this work was to evaluate the effect ofTrichoderma spp. on growth and fumonisin B¹ fusaproliferin and beauvericin production byF. proliferatum. Dual cultures of F.proliferatum andT. harzianum ITEM 3636 andT. longibrachiatum ITEM 3635 on maize meal agar at 0.995 aw were done. The effect ofTrichoderma spp. on the lineal growth ofF. proliferatum was determined. The effect ofTrichoderma species on fumonisin B¹, fusaproliferin and beauvericin production byF. proliferatum was determined on co-inoculated maize kernels by HPLC.T. harzianum suppressedF. proliferatum growth once contact between the colonies occurred.T. longibrachiatum showed a less antagonistic effect againstF. proliferatum. A reduction on fumonisin B¹ production of 98% and 88% was observed in the co-incubation ofF. proliferatum withT. harzianum andT. longibrachiatum, respectively. The decrease of FB¹ production was significant even in maize kernels on whichF. proliferatum had been growing 7 days prior to the addition ofTrichoderma spp. The concentration of beauvericin and fusaproliferin produced during 30 days coincubation ofF. proliferatum with bothTrichoderma spp. did not differ to those produced byF. proliferatum alone. These mycotoxins might enter the food chain causing so far unknown consequences to the health of domestic animals and humans. For this reason it is important, when a potential biocontrol agent is under study, to test the effect on the fungal growth and on the putative mycotoxin produced. Part of the information was presented at the Mycotoxin Prevention Cluster Dissemination Day and Mycoglobe Launch Conference, Brussels, Belgium, Oct 20–21, 2004 Financial support: Agenda Córdoba Ciencia, grant N^o 0279–000431/00     

Specificity of a strain ofNeozygites sp. (Zygomycetes: Entomophthorales) toMononychellus tanajoa (Acari: Tetranychidae)

Neozygites sp. is commonly found infecting the cassava green mite,Mononychellus tanajoa (Bondar), in parts of northeast Brazil. The introduction of this fungus into other regions requires the knowledge of its specificity, especially in relation to natural enemies of different cassava pests. Laboratory tests indicated the development of germination tubes ofNeozygites in some females ofTetranychus bastosi Tuttle et al. andT. urticae Koch, with subsequent formation of a reduced number of hyphal bodies in someT. bastosi. No females of the phytoseiid predatorsAmblyseius idaeus (Denmark & Muma) andAmblyseius limonicus Garman & McGregor s.l. were infected byNeozygites sp.     

Seed dispersal by the Florida box turtle (<Emphasis Type="Italic">Terrapene carolina bauri</Emphasis>) in pine rockland forests of the lower Florida Keys,United States

Seed dispersal by animals is one of the most important plant-animal mutualisms, but saurochory, the dispersal of seeds by reptiles, has received little attention. We investigated the role of the Florida box turtle (Terrapene carolina bauri) as a seed dispersal agent in pine rockland forests of the lower Florida Keys and examined the effect of turtle digestion on seed germination. We obtained seeds of 11 species with fleshy fruits and 2 species with non-fleshy fruits (a grass and legume) from the feces of 145 box turtles collected on Key Deer National Wildlife Refuge from 1999 to 2000. We planted the seeds of nine species and germination percentage (percentage of seeds that germinated during the experiment) varied from 10% to 80%. Comparative germination experiments were conducted with Thrinax morrissii, Serenoa repens, and Byrsonima lucida. We compared the germination percentage and germination rate (number of days from planting to seedling emergence) of seeds from three treatments (seeds recovered from feces, control seeds with pulp, and control seeds without pulp) and continued these experiments for up to 2 years. Passage through the box turtle digestive tract greatly enhanced the germination percentage and germination rate of S. repens, but decreased the germination percentage of B. lucida and T. morrissii, and decreased germination rate for T. morrissii. Subsequent destructive seed viability tests revealed that many ungerminated T. morrissii seeds remained viable, suggesting long-term seed dormancy may occur, even after passage through the turtle digestive system. In addition, the proportion of ungerminated seeds which remained viable was greater for seeds recovered from turtle feces than from control seeds with pulp. Furthermore, removal of fleshy pulp either manually or by the turtle digestive system may allow T. morrissii to escape insect predation.     

Stereoselective Degradation and Microbial Epimerization of Triadimenol in Soils

Triadimenol is a widely used triazole fungicide and consists of four stereoisomers with 1R,2S, 1S,2R, 1R,2R, and 1S,2S configurations. The trans‐enantiomeric pair (1R,2S‐isomer and 1S,2R‐isomer) is also called triadimenol‐A and the cis‐enantiomeric pair (1R,2R‐isomer and 1S,2S‐isomer) triadimenol‐B. In this study, the stereoselective degradation and chiral stability of triadimenol in two soils were investigated in details. The dissipation of technical triadimenol, a 6:1 mixture of triadimenol‐A and triadimenol‐B, showed significant epimerization from triadimenol‐A to triadimenol‐B occurred along with the dissipation process. The degradation exhibited some stereoselectivity, resulting in a concentration order of 1S,2S > 1R,2R > 1R,2S > 1S,2R or 1S,2S > 1R,2R > 1S,2R > 1R,2S at the end of the 100 days incubation for Baoding soil or Wuhan soil, respectively. Further incubation of triadimenol‐B revealed no epimerization, i.e. triadimenol‐B was configurationally stable in soil, and 1R,2R‐triadimenol degraded slightly slower in the former part and slightly faster in the later part of the incubation than 1S,2S‐triadimenol. Moreover, by incubation of enantiopure 1S,2R‐triadimenol and 1R,2S‐triadimenol, the results documented the epimerization for each enantiomer occurred at both C‐1 and C‐2 positions. Finally, the present work also documented that the enantiomerization reaction for all the four stereoisomers was nearly negligible in the soils. Chirality 25:355‐360:, 2013. © 2013 Wiley Periodicals, Inc.     

Germination and growth ofErythrorchis ochobinesis (Orchidaceae) accelerated by monokaryons and dikaryons ofLenzites betulinus andTrametes hirsuta

Four sib-monokaryons and two reconstituted dikaryons of two basidiomycetes,Lenzites betulinus andTrametes hiruta, accelerated the seed germination ofErythrorchis ochobiensis, an achlorophyllous orchid. All isolates ofL. betulinus and three isolates ofT. hirsuta induced the development of plants from germinated seeds. Although three monokaryotic isolates ofT. hirsuta failed to induce the development of plants, the reconstituted dikaryons induced the development.     

Phenology of the black bean aphid,Aphis fabae,on organic crops and effect of parasitoid communities

1. Parasitoids represent the third trophic level in plant-insect food webs. They develop in or on herbivorous insects at the second trophic level, which in turn feed on plants that constitute the first trophic level. This food web system might be affected by agricultural landscape and practices.
2. We studied the phenology and population density of Aphis fabae (Hemiptera: Aphididae) and determined the impact of parasitoids in organic fields of beans and beet plants.
3. Twenty leaves per field were collected weekly. For each leaf, the total number of aphids, apterous and alate morphs, the number of mummies, and mortality of aphids were recorded.
4. Our results showed that the phenology of A. fabae varied throughout the season with maximal numbers observed at the beginning of summer. In addition, the production of alate morphs and the proportion of parasitism were positively correlated to an increase in aphid densities.
5. Six parasitoid species were recorded attacking A. fabae in the field, with Lysiphlebus fabarum and Binodoxys angelicae (Hymenoptera: Aphidiinae) being the most abundant parasitoids.
6. Field characteristics and availability of host plants in the surroundings affected A. fabae density. Lysiphlebus fabarum and B. angelicae appeared to be good biocontrol candidates against A. fabae.

     

Selective antimicrobial action of chitosan against spoilage yeasts in mixed culture fermentations

The effect of chitosan on Saccharomyces cerevisiae (the yeast that carries out alcohol fermentation), Brettanomyces bruxellensis and Brettanomyces intermedius (contaminants of alcohol fermentations), was investigated. The effect of chitosan was tested on each yeast, as well as on mixed cultivations of S. cerevisiae + B. bruxellensis and S. cerevisiae + B. intermedius. Chitosan enhanced the lag period of both strains of Brettanomyces (80 h for B. bruxellensis and 170 h for B. intermedius with 6 and 2 g/l chitosan, respectively). The growth rate of S. cerevisiae was inversely proportional to the chitosan concentration; the former was 50% when 6 g/l polysaccharide was used. Moreover, in mixed cultivations of S. cerevisiae and Brettanomyces strains, it was found that both B. bruxellensis and B. intermedius failed to grow while growth of S. cerevisiae was not affected (using 3 and 6 g/l chitosan, respectively). An interesting collateral result was that the presence of chitosan accelerated the consumption of glucose in the mixed cultivations (60 h instead of 120 h).     

Molecular cloning,functional expression and characterization of two serine/threonine-specific protein kinases from<Emphasis Type="Italic"> Nicotiana tabacum</Emphasis> pollen

The existence of different kinds of kinases in pollen and pollen tubes suggests that kinase-mediated signaling pathways are likely involved in regulating pollen germination and pollen tube growth during the life cycle of higher plants. We have used RT-PCR and RACE to isolate full-length cDNAs for two pollen-expressed kinases, named NtPK1 and NtPK2, of Nicotiana tabacum. NtPK1 and NtPK2 encode proteins of 365 and 369 amino acids with calculated molecular masses of 39.2 kDa and 39.5 kDa, respectively, and both proteins possess the 12 sub-domains that are conserved among protein kinases. The nucleotide and deduced amino acid sequences of NtPK1 and NtPK2 share 88% and 91% identity, respectively, with the C-terminal region being the most conserved. RT-PCR analysis revealed that NtPK1 was specifically expressed in pollen and pollen tubes, and that NtPK2 was also expressed in pistil and petal. Immunoblot analysis using anti-NtPK1 and anti-NtPK2 antibodies confirmed that both NtPK1 and NtPK2 were produced in pollen and pollen tubes, and that NtPK2 was also produced in developing male gametophytes and other floral tissues. Biochemical fractionation experiments showed that, in all the tissues examined, NtPK1 and NtPK2 were present in the cytosolic fraction and not in the microsomal fraction. NtPK1 and NtPK2 were found to autophosphorylate on threonine and, for NtPK2, on serine as well. All the results taken together suggest that NtPK1 and NtPK2 are novel receptor-like cytosolic serine/threonine kinases, and could mediate signaling pathways required for pollen germination and/or pollen tube growth.The nucleotide sequence data of NtPK1 and NtPK2 reported in this paper will appear in the EMBL/GenBank/DDBJ nucleotide sequence databases under the accession numbers AJ608156 and AJ608157, respectively     

Phylogeny of the GenusTrichodermaBased on Sequence Analysis of the Internal Transcribed Spacer Region 1 of the rDNA Cluster

Sequences of the internal transcribed spacer region 1 (ITS1) of the ribosomal DNA were used to determine the phylogenetic relationships of species ofTrichodermasect.Pachybasium.To this end, 85 strains—including all the availableex-type strains—were analyzed. Parsimony analysis demonstrated that the section is nonmonophyletic, distributing the 85 strains among three main groups that were supported by bootstrap values. Group A comprises two clades (A1 and A2), with A1 includingT. polysporum, T. piluliferum,andT. minutisporum,while A2 includedT. hamatum, T. pubescens,andT. strigosumin addition to species previously included in sect.Trichoderma(i.e.,T. viride, T. atroviride,andT. koningii). Theex-type strain ofT. fasciculatumformed a separate branch basal to clade A. Clade B contained the sect.PachybasiummembersT. harzianum, T. fertile, T. croceum, T. longipile, T. strictipile, T. tomentosum, T. oblongisporum, T. flavofuscum, T. spirale,and the anamorphs ofHypocrea semiorbisandH. cf. gelatinosa.Sequence differences among clades A1, A2, and B were in the same order of magnitude as between each of them andT. longibrachiatum,which was used as an outgroup in these analyses. Sequence differences within clades A1, A2, and B were considerably smaller: in some cases (i.e.,T. virensandT. flavofuscum; T. strictipileandH. cf. gelatinosa), the ITS1-sequences were identical, suggesting conspecifity. In other cases (e.g.,T. crassumandT. longipile; T. harzianum, T. inhamatum, T. croceum, T. fertile,andH. semiorbis; T. hamatumandT. pubescens;andT. viride, T. atroviride,andT. koningii) differences were in the range of 1–3 nt only, suggesting a very close phylogenetic relationship. The sequence of a previously described aggressive mushroom competitor group ofT. harzianumstrains (Th2) was strikingly different from that of theex-type strain ofT. harzianumand closely related species and is likely to be a separate species.     

Karyological and taxonomical notes on three species of the genus<Emphasis Type="Italic">Epipactis (Neottioideae,Orchidaceae)</Emphasis> in the central-western Iberian Peninsula

Studies on the karyotypes and chromosome numbers of species ofEpipactis from the central-western Iberian Peninsula show that the species harbour enormous chromosome variability, have very asymmetric karyotypes and possess extraordinary diversity of aneuploidy. This paper provides the first report of a chromosome number forE. fageticola (2n=36, 40 + 0–2 B), as well as the first counts for Portuguese populations ofE. helleborine (2n=18, 32, 38) and first counts for Iberian populations ofE. tremolsii (n=20, 30, 2n=16, 24, 32, 34, 36, 38 + 1B, 40 + 1B, 52, 60). Among populations ofE. tremolsii there is a significant differentiation in ecology and somatic chromosome number, suggesting that there may be two different taxa in the region studied. Chromosomes are large to small, ranging in length from 10.8 μm to 1.8 μm. Karyotype asymmetry is of type 3C inE. fageticola andE. tremolsii and 2C inE. helleborine andE. tremolsii.     

Effects of Bacteria on Cadmium Bioaccumulation in the Cadmium Hyperaccumulator Plant Beta Vulgaris Var. Cicla L.

To investigate the effects of two cadmium-tolerant bacteria, Staphylococcus pasteuri (S. pasteuri X1) and Agrobacterium tumefaciens (A. tumefaciens X2), on cadmium uptake by the cadmium hyperaccumulator plant Beta vulgaris var. cicla L., a pot experiment with artificially contaminated soil was conducted. The results demonstrated that both cadmium-tolerant bacteria enhanced the dry weight of Beta vulgaris var. cicla L. The total dry weights of plants in the control CK20, S. pasteuri X1 and A. tumefaciens X2 treatments were 0.85, 1.13, and 1.38 g/pot, respectively. Compared with the control CK20 findings, the total dry weight of plants was increased by 32.8 and 61.1% after inoculation with S. pasteuri X1 and A. tumefaciens X2, respectively, indicating that A. tumefaciens X2 more strongly promoted the growth of Beta vulgaris var. cicla L. than S. pasteuri X1. In addition, inoculation with S. pasteuri X1 and A. tumefaciens X2 significantly (p < 0.05) promoted cadmium uptake by plants and improved the bioaccumulation of cadmium by the plants from the soil. Moreover, the inoculation of S. pasteuri X1 and A. tumefaciens X2 effectively facilitated the transfer of cadmium in the soil from the Fe-Mn oxide and residual fractions to the soluble plus exchangeable and weakly specially adsorbed fractions in the rhizosphere soils of plants. The bacterial enhancement of cadmium phytoavailability might provide a potential and promising method to increase the efficiency of phytoextraction.     

Taxonomic reappraisal on<Emphasis Type="Italic">Suaeda australis</Emphasis> (Chenopodiaceae) in korea based on the morphological and molecular characteristics

We used morphological and molecular characteristics to perform a taxonomic reappraisal ofSuaeda australis (Brown) Moquin-Tandon from Korea. Populations of this species are dispersed at the bottoms of sand zones, and usually exhibit a depressed habit. Except for their total heights and leaf lengths, the morphology of these plants does not differ from that ofS. maritima. Molecular traits were examined based on ITS and psbB-psbH spacer region sequences. The former region included ITS-1, 5.8S, and ITS-2, which were 629 nucleotide bases long. Pair-wise distances (p-distance) among KoreanSuaeda species ranged from 1.12 to 17.84. The psbB-psbH spacer region sequences were 618 nucleotide bases long, and were conserved in the alignment of KoreanSuaeda species. In our ML and MrBayesian analysis of ITS sequences aligned with other sequences from GenBank, the plants of KoreanSuaeda made three clades: 1)S. japonica;S. australis, andS. maritima;2)S. malacosperma; and 3) S.glauca. However, the psbB-psbH region sequences could not be resolved amongS. japonica, S. maritima, andS. australis from Korea. Molecular and vegetative characteristics indicated that the plants now classified asS. australis from Korea should instead be named as S.maritima (L.) Dumont.     

Potential of genes and gene products fromTrichoderma sp. andGliocladium sp. for the development of biological pesticides

Fungal cell wall degrading enzymes produced by the biocontrol fungiTrichoderma harzianum andGliocladium virens are strong inhibitors of spore germination and hyphal elongation of a number of phytopathogenic fungi. The purified enzymes include chitinolytic enzymes with different modes of action or different substrate specificity and glucanolytic enzymes with exo-activity. A variety of synergistic interactions were found when different enzymes were combined or associated with biotic or abiotic antifungal agents. The levels of inhibition obtained by using enzyme combinations were, in some cases, comparable with commercial fungicides. Moreover, the antifungal interaction between enzymes and common fungicides allowed the reduction of the chemical doses up to 200-fold. Chitinolytic and glucanolytic enzymes fromT. harzianum were able to improve substantially the antifungal ability of a biocontrol strain ofEnterobacter cloacae. DNA fragments containing genes encoding for different chitinolytic enzymes were isolated from a cDNA library ofT. harzianum and cloned for mechanistic studies and biocontrol purposes. Our results provide additional information on the role of lytic enzymes in processes of biocontrol and strongly suggest the use of lytic enzymes and their genes for biological control of plant diseases.     

Phylogenetic analyses of <Emphasis Type="Italic">Uromyces viciae-fabae</Emphasis> and its varieties on <Emphasis Type="Italic">Vicia</Emphasis>, <Emphasis Type="Italic">Lathyrus</Emphasis>, and <Emphasis Type="Italic">Pisum</Emphasis> in Japan

A pea rust fungus, Uromyces viciae-fabae, has been classified into two varieties, var. viciae-fabae and var. orobi, based on differences in urediniospore wall thickness and putative host specificity in Japan. In principal component analyses, morphological features of urediniospores and teliospores of 94 rust specimens from Vicia, Lathyrus, and Pisum did not show definite host-specific morphological groups. In molecular analyses, 23 Uromyces specimens from Vicia, Lathyrus, and Pisum formed a single genetic clade based on D1/D2 and ITS regions. Four isolates of U. viciae-fabae from V. cracca and V. unijuga could infect and sporulate on P. sativum. These results suggest that U. viciae-fabae populations on different host plants are not biologically differentiated into groups that can be recognized as varieties.Contribution no. 184, Laboratory of Plant Parasitic Mycology, Institute of Agriculture and Forestry, University of Tsukuba, Japan     

Seasonal variation in allelopathic potential of<Emphasis Type="Italic">Artemisia princeps</Emphasis> var.<Emphasis Type="Italic">orientalis</Emphasis> on plants and microbes

We investigated seasonal variations in allelopathic potential ofArtemisia princeps var.orientalis. Aqueous and meth-anol extracts and volatile substances were prepared in the laboratory from samples collected monthly (April through October). Their impacts were then assessed on the germination and seedling growth ofLactuca sativa andAchyranthes japonica. The allelopathic potential varied with the time of sample collection and the concentration tested. For example, germination ofL. sativa was not inhibited by the aqueous extract but seedling growth (shoots and roots) was, with its seasonal effect being significant. ForA. japonica, seed germination was not inhibited at lower concentrations (except for August samples). However, at higher concentrations and in certain months (especially July), germination was more negatively affected. The degree of seedling growth inhibition also differed by month and by extract concentration, with roots being impacted more than shoots. Volatile substances also had a time-dependent influence on the germination and seedling elongation ofA. japonica. In a separate experiment, the ethyl-acetate and water fractions of a crude methanol extract were prepared monthly fromA. princeps var.orientalis. Here, we examined their antimicrobial activities against three gram-positive bacteria (Bacillus cereus, Bacillus subtilis, andStaphylococcus aureus), two gramnegative bacteria (Escherichia coli andPseudomonas fluorescens), and one lactic acid bacterium,Lactobacillus plantar urn. The ethyl-acetate fraction that was sampled in September was remarkably potent againstB. cereus andB. subtilis, whereas the water fraction collected in August and September showed great antimicrobial activity against the grampositive and -negative bacteria. In contrast,L. plantarum was not inhibited by the water fraction, regardless of the sampling month. Likewise, the ethyl-acetate and water fractions collected in April and October had the lowest levels of antimicrobial activity.     

Effect of Two Opiine Parasitoids (Hymenoptera: Braconidae) Introduced for Fruit Fly Control on a Native Hawaiian Tephritid,Trupanea dubautiae(Diptera: Tephritidae)

Diachasmimorpha longicaudata(Ashmead) andPsyttalia fletcheri(Silvestri) are opiine parasitoids introduced into Hawaii for control of the Oriental fruit fly,Bactrocera dorsalis(Hendel) and the melon fly,Bactrocera cucurbitae(Coquillett), respectively. Both species have recently been mass-reared and released for research in augmentative biocontrol programs. Laboratory and field sleeve cage experiments were conducted to investigate the potential impact of mass-producedD. longicaudataandP. fletcherion a native Hawaiian tephritid,Trupanea dubautiae(Bryan), infesting the flowerheads of the native composite shrubDubautia raillardioidesHillebrand. Gravid females ofD. longicaudataandP. fletcheriwere confined with bloomingD. raillardioidesflowerheads infested with late instarT. dubautiae.BothD. longicaudataandP. fletcherilacked positive oviposition responses toT. dubautiaelarvae in infested flowerheads and caused neither parasitism nor mortality to the flies. However, when larvae were removed from the flowerheads and presented in screened dishes containing artificial diet of the parasitoids' normal rearing hosts (B. dorsalisandB. cucurbitae), bothD. longicaudataandP. fletcherireadily oviposited in the test larvae. Oviposition byD. longicaudatadid not significantly affect the percentage pupation ofT. dubautiae,but did reduce the emergence of adult flies. Oviposition byP. fletcherisignificantly reduced both pupation and adult fly emergence. All progeny of both parasitoid species died as eggs or first-instar larvae. Results from our experiments demonstrate that biological control programs targeted against frugivorous tephritid pests byD. longicaudataandP. fletcherihave no harmful impact on flowerhead-infestingT. dubautiae.