Strategies for the biological control of invasive willows (Salix spp.) in Australia

Abstract  Willows ( Salix spp.) are Weeds of National Significance in Australia where a large number of taxa are naturalised in temperate regions and can cause serious environmental degradation of riparian and wetland habitats. Several species are of economic or ornamental value and conflicts of interest could arise with planning their suppression. Biological control of six willow species ( S. alba L., S. cinerea L . , S. fragilis L., S. nigra Marshall, S. viminalis L., S . x  rubens Schrank) is under consideration in Australia using three approaches based on targeting different morphological structures. A rich phytophagous biota associated with Salix in their regions of origin indicate a range of organisms are available for development as biological control agents. Implementation strategies will depend on the host species targeted for biological control, their reproductive strategies, the level of control required to reduce environmental impacts and the level of conflict of interest associated with a biological control program.     

Patterns of interspecific associations of stem gallers on willows

Abstract.  The pattern of interspecific associations of three stem-galling sawfly species ( Euura atra , E. elaeagnos , E. purpureae ) and three stem-galling gallmidge species ( Rabdophaga sp. 3–5) was investigated on five willow taxa ( Salix alba , S. fragilis , S.  ×  rubens , S. elaeagnos , S. purpurea ) at five natural sites in Central Europe. The willow species harboured specific species associations of two stem gallers, each pair consisting of one Euura and one Rabdophaga species. The stem gallers were patchily distributed and their densities varied significantly among willow host plant species, host plant individuals, and host plant sexes. Four of the six species showed a significant increase in galling rate with shoot length. The other two species were the sawfly and cecidomyiid pair that induce galls on S. purpurea . The preference of stem gallers to longer shoots was generally not related to higher larval performance in terms of survival. Only one species, Rabdophaga sp. 5, was found to be more abundant on male plants. The correlation of densities of the species pairs of stem gallers was independent of willow sexes. Species pairs of stem gallers co-occurring on the same willow species tended to attack different shoots within the same host plant individual. When species pairs co-occurred on shoots they were usually found in similar densities as when occurring alone on shoots. The stem-galling sawflies usually formed galls at the basal part of a shoot, whereas the gallmidge R . sp. 5 ( R . sp. 3 and R . sp. 4 showed no clear tendency) preferred the middle or distal part of a shoot. This is interpreted with differences of their phenology and oviposition period.     

PERMANENT GENETIC RESOURCES: Consensus primers of cyp73 genes discriminate willow species and hybrids (Salix, Salicaceae)

Consensus primers, based on exon sequences of the cyp73 gene family coding for cinnamate 4-hydroxylase (C4H) of the lignin biosynthesis pathway, were designed for the tetraploid willow species Salix alba and Salix fragilis. Diagnostic alleles at species level were observed among introns of three cyp73 genes and allowed unambiguous detection of the first generation and introgressed hybrids in populations. Progeny analysis of a female S. alba with a male introgressed hybrid confirmed the codominant inheritance of each intron. Sequences of the diagnostic alleles of both species were similar to those found in the hybrids.     

Camouflage and selective predation in caterpillars of the poplar and eyed hawkmoths (Laothoe populi and Smerinthus ocellata)

Final instar caterpillars of the poplar hawkmoth ( Laothoe populi ) are usually yellow-green when they have been feeding on Salix fragilis but white when on Populus alba. Similarly final instar caterpillars of the eyed hawkmoth (Smerinthus ocellata) vary in colour from yellow-green when fed on Salix cinerea or S. fragilis to grey-green when fed on Salix ripens or S. alba. Most caterpillars of these two moths are thus very cryptic to the human eye. It has already been shown that colour matching is brought about by young caterpillars changing colour depending on the colour of the substrate perceived by their eyes. Evidence is presented here for selective predation, probably by birds. Yellow-green poplar hawk caterpillars disappeared from P. alba more rapidly than did white caterpillars, but there was no significant difference in the survival of the two colour forms on S. fragilis. Yellow-green caterpillars survived better on S. fragilis than on P. alba , but there was no significant difference in the survival of white caterpillars on the two plants. Reasons why selective predation did not occur in all of the experiments are discussed.     

Linkage mapping in tetraploid willows: segregation of molecular markers and estimation of linkage phases support an allotetraploid structure for Salix alba x Salix fragilis interspecific hybrids

Salix alba-Salix fragilis complex includes closely related dioecious polyploid species, which are obligate outcrossers. Natural populations of these willows and their hybrids are represented by a mixture of highly heterozygous genotypes sharing a common gene pool. Since nothing is known about their genomic constitution, tetraploidy (2n=4x=76) in willow species makes basic and applied genetic studies difficult. We have used a two-way pseudotestcross strategy and single-dose markers (SDMs) to construct the first linkage maps for both pistillate and staminate willows. A total of 242 amplified fragment length polymorphisms (AFLPs) and 50 selective amplifications of microsatellite polymorphic loci (SAMPL) markers, which showed 1:1 segregation in the F(1) mapping populations, were used in linkage analysis. In S. alba, 73 maternal and 48 paternal SDMs were mapped to 19 and 16 linkage groups covering 708 and 339 cM, respectively. In S. fragilis, 13 maternal and 33 paternal SDMs were mapped in six and 14 linkage groups covering 98 and 321 cM, respectively. For most cosegregation groups, a comparable number of markers linked in coupling and repulsion was identified. This finding suggests that most of chromosomes pair preferentially as occurs in allotetraploid species exhibiting disomic inheritance. The detection of 10 pairs of marker alleles from single parents showing codominant inheritance strengthens this hypothesis. The fact that, of the 1122 marker loci identified in the two male and female parents, the vast majority (77.5%) were polymorphic and as few as 22.5% were shared between parental species highlight that S. alba and S. fragilis genotypes are differentiated. The highly difference between S. alba- and S. fragilis-specific markers found in both parental combinations (on average, 65.3 vs 34.7%, respectively) supports the (phylogenetic) hypothesis that S. fragilis is derived from S. alba-like progenitors.     

Susceptibility trending of blood isolates of the Bacteroides fragilis group over a 12-year period to clindamycin, ampicillin-sulbactam, cefoxitin, imipenem, and metronidazole

Numerous reports have described a steady overall increase in resistance among clinical isolates of the Bacteroides fragilis group to several antimicrobial agents, particularly clindamycin. Determination of resistance rates is significantly influenced by the number of isolates of each species within the B. fragilis group tested. Historically, the B. fragilis species has remained the most susceptible to most antimicrobials when compared to non-B. fragilis species. This study compares the effect of a gradually changing ratio of blood isolates of B. fragilis to non-B. fragilis species tested by broth micro-dilution over a 12-year period on selected antimicrobial agents. In 1987, the ratio of blood isolates of B. fragilis to non-B. fragilis was 68% to 32%; in 1991 it was 59% to 41%; and in 1999 it was 51% to 49%. Both metronidazole and imipenem showed the least changes because of their inherent high activity against all species. For clindamycin, decreases in susceptibility ranged from 84% to 64% for B. fragilis compared to 58% to 67% for non-B. fragilis species. Ampicillin-sulbactam showed a decrease in susceptibility in B. fragilis and non-B. fragilis species, but was highest in 1999 when the ratio of non-B. fragilis species was the highest. Overall resistance rates to cefoxitin varied from 8% to 25% during the testing years and was consistently higher among the non-B. fragilis species. These comparisons indicate that the ratio of B. fragilis group species isolated from the blood has changed over the last 12 years and has appreciably affected the resistance rates to some commonly used anti-anaerobic agents. Whether the noted changes in species isolation rates are a result of selective antibiotic pressure or other factors is yet to be determined.     

Dasineura ingeris sp.n. (Diptera: Cecidomyiidae) on Salix viminalis in Sweden, including comparisons with some other Dasineura species on Salix

Infestations of an undescribed gall midge species were discovered in Southern Sweden in biomass plantations and nurseries of Salix viminalis L. Terminal leaf buds are damaged and side shoots subsequently develop. This midge, Dasineura ingeris sp.n., closely resembles three other species occurring on Salix: D.terminalis (H. Loew) on S.alba L. and S.fragilis L.; D.iteobia (Kieffer) recorded from S.caprea L. and S.cinerea L.; and D.schreiteri (Stelter) (= comb.n. for Rabdophaga schreiteri Stelter) originally found as an inquiline in the galls of another gall midge, Dasineura rosaria (H. Loew), on S.repens L. No qualitative differences in morphology were observed between these four gall midge species, but results of morphometric analyses show significant differences between all of them. In oviposition preference trials, which included host plants of all four midge species, D.ingeris laid eggs mainly on S. viminalis, D.schreiteri preferably on S.repens , and D.iteobia exclusively on S.caprea. In larval performance trials D.ingeris produced many galls on S. viminalis , one gall also on S.caprea , but no galls on either S.alba or S.repens.     

Absence of geographical structure of chloroplast DNA variation in sallow, Salix caprea L

In the present study, we have used PCR-RFLP markers to investigate the chloroplast DNA variation in 24 European populations of Salix caprea L. A subset of these populations has also been analysed with chloroplast microsatellites. The main feature of both markers is the absence of a clear geographic structure (G(ST(PCR-RFLP))=0.090, G(ST(microsatellites))=-0.017) and high levels of variation within populations. This lack of phylogeographic structure in S. caprea is suggested to be the consequence of the joint action of several factors: (i) presence of intermediate latitude refugia with large population sizes during the last glacial maximum, (ii) a high speed of recolonisation and dispersal ability, (iii) a high mutation rate and (iv) extensive hybridisation with other willow species. In addition to the S. caprea samples, a limited number of individuals from several other Salix species were also analysed with PCR-RFLP: S. cinerea, S. aurita, S. purpurea, S. atrocinerea, S. appendiculata, S. elaeagnos, S. fragilis and S. alba. Many of the haplotypes found in Salix caprea were also detected in S. cinerea, S. aurita, S. purpurea, S. atrocinerea and/ or S. appendiculata but not in S. alba, S. elaeagnos or S. fragilis. Our data suggest that hybridisation and gene flow have occurred within these two groups but not between them.     

Microscopic fracture aspects of impact tested human bones

The fracture aspects of impact twisted human femurs and tibias have been studied in their wet condition. The main tool used to correlate fracture behaviour with bone structure was the scanning electron microscope. Supplementary data were obtained from macroscopic fracture pattern analyses and stress coat experiments on slowly tested dried bone. The interest of the data reported concerns two major points. Firstly the analysis of the fractographic results leads to the conclusion that the fracture has presumably been initiated by shearing parallel with the torsion axis. The fracture propagates along so called spiral and oblique fracture lines. Secondly, the correlation of the macroscopic fracture pattern with microscopic fracture aspects indicates that the fracture is initiated along the cement lines, which can be considered as the weak matrix in an osteon-cement lining composite. The fracture propagates along the cement lines as well as through osteons and lamellae. Throughout the impact fracture surfaces ductile and brittle fracture aspects were observed, although the brittle fracture mode was predominant during propagation.     

Effect of seed size on seedling growth response to elevated CO2 in Picea abies and Picea rubens

Several previous studies have observed that species and individuals with large seeds respond more positively to elevated CO (2) than those with small seeds. We explored the reasons for this pattern by examining the relationship between seed size and CO (2) response in Picea abies and P. rubens using growth analysis. The large seeded species (P. abies) responded more positively to elevated CO (2) than the small seeded species (P. rubens). At the intraspecific level, P. abies individuals from large seeds responded more positively to elevated CO (2) than individuals from small seeds, however, there was no significant intraspecific variation in CO (2) response in P. rubens. The greater CO (2) response of plants from large seeds was not simply the result of a larger starting capital compounded at the same rate as in plants from small seeds. Elevated CO (2) increased relative growth rate to a greater extent in individuals from large seeds. This effect appears to be related to differences in time of establishment, source to sink ratio and nutrient availability with seed size. These results are significant not only in understanding the potential effect of rising atmospheric CO (2) concentrations on plant populations, but also in understanding the factors affecting plant success at current atmospheric CO (2) levels due to the elevation of CO (2) within the litter layer that occurs at many germination sites.     

Yield reduction in Brassica napus, B. rapa, B. juncea, and Sinapis alba caused by flea beetle (Phyllotreta cruciferae (Goeze) (Coleoptera: Chrysomelidae)) infestation in northern Idaho

Phyllotreta cruciferae is an important insect pest of spring-planted Brassica crops, especially during the seedling stage. To determine the effect of early season P. cruciferae infestation on seed yield, 10 genotypes from each of two canola species (Brassica napus L. and Brassica rapa L.) and two mustard species (Brassica juncea L. and Sinapis alba L.) were grown in 2 yr under three different P. cruciferae treatments: (1) no insecticide control; (2) foliar applications of endosulfan; and (3) carbofuran with seed at planting plus foliar application of carbaryl. Averaged over 10 genotypes, B. rapa showed most visible P. cruciferae injury and showed greatest yield reduction without insecticide application. Mustard species (S. alba and B. juncea) showed least visible injury and higher yield without insecticide compared with canola species (B. napus and B. rapa). Indeed, average seed yield of S. alba without insecticide was higher than either B. napus or B. rapa with most effective P. cruciferae control. Significant variation occurred within each species. A number of lines from B. napus, B. juncea, anid S. alba showed less feeding injury and yield reduction as a result of P. cruciferae infestation compared with other lines from the same species examined, thus having potential genetic background for developing resistant cultivars.     

A POLYPLOID SPECIES COMPLEX OF SPIROGYRA COMMUNIS (CHLOROPHYTA) OCCURRING IN NATURE 1

Investigations were conducted to determine whether ploidal changes found in laboratory cultures of Spirogyra also occur in nature. In an earlier study filament types identifiable as three different species (Spirogyra singu-laris Nordstedt, S. communis (Hassall)Kütz., S. fragilis Jao) arose from an original clonal culture through vegetative growth and sexual reproduction. These three “species” or filament groups differed in filament width, chloroplast number, zygospore size, and chromosome number. The differences in chromosome number represented a polyploid series of diploid (S. communis), triploid (S. fragilis), and tetraploid (S. singularis) forms in which width increased with ploidal level. The three width groups constituted a “species complex.” Five years after isolation of the original strain in this species complex, filaments corresponding to two of the width groups (S. singularis and S. communis) were found at the original collection site in the Santa Catalina Mountains in southern Arizona. The two field-collected groups were indistinguishable from the laboratory species complex in morphology and chromosome number. Homothallic conjugation within the two field width groups yielded progeny similar to those from homothallic conjugation of groups in the laboratory species complex. Filament widths of progeny were generally within the width limits of respective parental groups. The four possible intergroup crosses between the two laboratory and two field width groups yielded progeny similar to the wider parent (S. singularis) or the parent of intermediate width (S. fragilis). Progeny characteristics were determined by the width groups of parents, regardless of whether parents came from the laboratory or field. The similarities in morphology, chromosome numbers, and reproductive behavior of laboratory and field width groups imply that the laboratory species complex of S. communis has a natural counterpart in the field.     

Transformation of Kluyveromyces fragilis

For the transformation of the yeast species Kluyveromyces fragilis, we have constructed a vector containing a bacterial kanamycin resistance (Kmr) gene, the TRP1 gene of Saccharomyces cerevisiae, and an autonomously replicating sequence of Kluyveromyces lactis called KARS2 . By utilizing the method based on treatment by alkali cations and with the Kmr gene as the selective marker, a wild-type strain of K. fragilis was transformed to resistance against the antibiotic G418 . In the transformed cell the plasmid replicates autonomously. The same plasmid could also be used to transform S. cerevisiae trp1 mutant to Trp+. Thus, KARS2 of K. lactis enables the vector to replicate in K. fragilis, K. lactis, and S. cerevisiae, whereas ARS1 of S. cerevisiae allows autonomous replication only in S. cerevisiae.     

Experimental taxonomy of Silene section Elisanthe (Caryophyllaceae): crossing experiments

Silene section Elisanthe is a well-defined group containing (in Europe) the following species: S. alba, S. diclinis, S. dioica, S. heuffelii and S. marizii (dioecious perennials or biennials) and S. noctifloara (a self-compatible hermaphrodite annual). Crosses were attempted among these species, and between these species and members of other Silene sections.
Crosses among the first five species revealed partial cross-incompatibility with moderate hybrid fertility. S. alba proved especially incompatible with S. diclinis. S. noctiflora would not cross at all with other members of the section. It is suggested that S. noctiflora evolved from a dioecious precursor of S. alba , the species to which it is most similar in morphology, distribution and habitat; hybrid sterility, even without incompatibility, would have assured mutual isolation.
Crosses with species from other sections of Silene have usually either failed consistently or revealed high cross-incompatibility with hybrid sterility. Those crosses which were successful have all been within the boundaries of the old genus Melandrium , or with Lychnis species.     

Competitiveness and dry matter allocation of oilseed rape (Brassica napus L.) and two mustards (Sinapis alba L. and S. arvensis L.) under water stress conditions

In Morocco, oilseed rape is commonly exposed to mustards competition which are not totally controlled by herbicides. To understand the competitiveness of each species, growth parameters should be studied notably dry matter allocation. The objective of this study was to confirm the competitiveness of oilseed rape with regard to Sinapis alba and S. arvensis and to investigate how the dry matter is allocated. A pot experiment was undertaken with a quartz sand as substrate. Two plant densities were tested (one and two plants). The binary density was either a monoculture or a mixture. Half the pots were maintained at field moisture capacity and the other half was irrigated up to 70% of its water holding capacity. Dry matter allocation of each species at density two was compared to that of the same species at density one. Results of replacement series diagrams and those of the relative crowding coefficient (based on total dry matter) showed that Brassica napus was more competitive than S. alba. S. arvensis was the least competitive. Under competition, B. napus irrigated at water holding capacity allocated more dry matter to stem when compared to its dry matter at density one. Under the same condition, when reducing water supply, B. napus allocated more dry matter to leaves. In case of S. alba, dry matter percent in leaves and roots were respectively low and high in normally irrigated plant and inversely under water shortage. S. arvensis allocated high and low dry matter percent respectively to root than to leaves when sufficiently irrigated. But no clear tendency was noticed under water shortage, for this species.     

16S rRNA PCR鉴定脆弱类杆菌

目的:应用16SrRNA序列设计PCR引物鉴别脆弱类杆菌。方法:通过脆弱类杆菌16SrRNA序列特异性位点设计引物,对4株脆弱类杆菌及大肠杆菌、乳酸杆菌、嗜热链球菌等进行PCR扩增。应用琼脂糖电泳法对PCR扩增产物进行特异性检测。结果:脆弱类杆菌在176bp左右出现特异性条带,而其他细菌均未出现特异性条带。结论:通过16SrRNA序列中特异位点设计引物进行PCR,可特异性鉴定脆弱类杆菌。     

Molecular and phytochemical evidence for the taxonomic integrity of Salix alba, S. fragilis, and their hybrid S. × rubens (Salicaceae) in mixed stands in SE Germany

AFLP fingerprinting, nrDNA sequencing, and HPLC of phenolic compounds from bark samples was used to assess the intensity of hybridisation in two mixed stands of Salix alba, S. fragilis, and their hybrid S. × rubens (Salicaceae) along the Danube in the vicinity of Regensburg (SE Germany). AFLP analyses of 130 individuals from two mixed and two pure strands of the parental species resulted in a matrix of 337 polymorphic loci that was analysed with Bayesian cluster algorithms implemented in the software programmes Structure and NewHybrids. The results, together with sequence polymorphisms of the nrDNA ITS1 + 5.8S region, indicate that the three taxa are distinctly separated and that only a few F2-hybrids and no backcrosses to the parental species are present in the surveyed mixed stands. These findings were corroborated by HPLC studies of the phenolic compounds from bark samples that were carried out for the same individuals. A multivariate statistical analysis of relative peak areas of chromatograms (PCA, principal component analysis) indicates the clear separation of parental and the intermediate position of S. × rubens individuals.     

Weak bases inhibit cleavage and embryogenesis in amphibians and echinoderms

The action of weak bases was studied on the early embryonic development of a number of species. Gastrulation was disrupted in the frog, Xenopus laevis, the newt, Pleurodeles watlii, the sea urchins, Paracentrotus lividus and Sphaerechinus granularis and the starfish, Asterias rubens. This required only submillimolar amounts of either NH+4 (pH 9.0) or procaine (pH 8.2). At higher concentrations even early cell division was inhibited in all the species with furrow regression particularly noticeable in Xenopus eggs. A similar action of the weak bases on early development, the lack of any action at lower extracellular pH, and the counteracting action of NH+4 on acidity-induced disruption of sea urchin development, all implicate an elevation of intracellular pH. However, a more direct intracellular action of the weak bases cannot be ruled out.     

Production of bacteriocin by Bacteriodes fragilis and partial characterization

The ability of Bacteroides fragilis strains, isolated from various sources, to produce bacteriocin was evaluated. All strains isolated from intestinal infections were producers in high levels and less susceptible to the others. Strains from other origins were found to produce bacteriocin at a medium level and they were variably susceptible. Some properties of one bacteriocin produced by the Bact. fragilis 079298-3 strain were analysed, providing evidence of its protein nature, with stability over a wide range of pH and retained inhibitory activity after heating. This variability seems to suggest that bacteriocin typing is a good method for this species.