Variation in herbivore density among host plants and its consequences for community structure

Summary The densities of four species of gall-forming sawflies were found to vary significantly among willow host plant clones. Two of the speices varied among host plants at four sites in each of three years. The other two species varied in density among host plants at most of the sites in two of the three years. Total sawfly density also varied significantly among clones. Individual species densities on willow clones were significantly positively correlated between years when all sites were combined and frequently when sites were considered separately. Most pairwise species combinations were independent in density between years, but some negative correlations existed between the stem galler and the leaf galler. Gall-former densities also were largely independent among clones within years with all sites combined and with sites considered separately. The significant correlations were nearly all positive. At all four sites the combination of significant variation in sawfly densities among willow clones in the field and independence of species densities among clones resulted in significantly different communities (relative abundance of species) among willow clones in three years. Although sawfly abundances differed substantially among the four sites, this remained true. It is argued that the pattern of community structure among clones is the result of variation in host plant quality of clones. We propose an hypothesis to account for patterns of herbivore species associations based on intrapopulation host plant variation.     

Microsatellite DNA fingerprinting, differentiation, and genetic relationships of clones, cultivars, and varieties of six poplar species from three sections of the genus Populus.

Accurate identification of Populus clones and cultivars is essential for effective selection, breeding, and genetic resource management programs. The unit of cultivation and breeding in poplars is a clone, and individual cultivars are normally represented by a single clone. Microsatellite DNA markers of 10 simple sequence repeat loci were used for genetic fingerprinting and differentiation of 96 clones/cultivars and varieties belonging to six Populus species (P. deltoides, P. nigra, P. balsamifera, P. trichocarpa, P. grandidentata, and P maximowiczii) from three sections of the genus. All 96 clones/cultivars could be uniquely fingerprinted based on their single- or multilocus microsatellite genotypes. The five P. grandidentata clones could be differentiated based on their single-locus genotypes, while six clones of P. trichocarpa and 11 clones of P. maximowiczii could be identified by their two-locus genotypes. Twenty clones of P. deltoides and 25 clones of P. nigra could be differentiated by their multilocus genotypes employing three loci, and 29 clones of P. balsamifera required the use of multilocus genotypes at five loci for their genetic fingerprinting and differentiation. The loci PTR3, PTR5, and PTR7 were found to be the most informative for genetic fingerprinting and differentiation of the clones. The mean number of alleles per locus ranged from 2.9 in P. trichocarpa or P. grandidentata to 6.0 in P. balsamifera and 11.2 in 96 clones of the six species. The mean number of observed genotypes per locus ranged from 2.4 in P. grandidentata to 7.4 in P. balsamifera and 19.6 in 96 clones of the six species. The mean number of unique genotypes per locus ranged from 1.3 in P. grandidentata to 3.9 in P. deltoides and 8.8 in 96 clones of the six species. The power of discrimination of the microsatellite DNA markers in the 96 clones ranged from 0.726 for PTR4 to 0.939 for PTR7, with a mean of 0.832 over the 10 simple sequence repeat loci. Clones/cultivars from the same species showed higher microsatellite DNA similarities than the clones from the different species. A UPGMA cluster plot constructed from the microsatellite genotypic similarities separated the 96 clones into six major groups corresponding to their species. Populus nigra var. italica clones were genetically differentiated from the P. nigra var. nigra clones. Microsatellite DNA markers could be useful in genetic fingerprinting, identification, classification, certification, and registration of clones, clultivars, and varieties as well as genetic resource management and protection of plant breeders' rights in Populus.     

Evidence for strong host clone-parasite species interactions in the Daphnia microparasite system

Abstract Organisms are often confronted with multiple enemy species. Defenses against different parasite species may be traded off against each other. However, if resistance is based on (potentially costly) general defense mechanisms, it may be positively correlated among parasites. In an experimental study, we confronted 19 clones from one Daphnia magna population with two bacterial and three microsporidian parasite species. All parasites were isolated from the same pond as the hosts. Host clones were specific in their susceptibility towards different parasite species, and parasite species were host-clone specific in their infectivity, spore production, and virulence, resulting in highly significant host-parasite interactions. Since the Daphnia 's resistance to different parasite species showed no obvious correlation, neither general defense mechanisms nor trade-offs in resistance explain our findings. None of the Daphnia clones were resistant to all parasite species, and the average level of resistance was quite similar among clones. This may reflect a cost of defense, so that the cumulative cost of being resistant to all parasite species might be too high.     

Intraspecific variation among clones of a naïve rare grass affects competition with a nonnative,invasive forb

Intraspecific variation can have a major impact on plant community composition yet there is little information available on the extent that such variation by an already established species affects interspecific interactions of an invading species. The current research examined the competitiveness of clones of a globally rare but locally common native grass, Calamagrostis porteri ssp. insperata to invasion by Alliaria petiolata, a non‐native invasive species. A greenhouse experiment was conducted twice over consecutive years in which 15 clones from three populations of Calamagrostis were paired with rosettes of Alliaria in pots containing native forest soil previously uninvaded by Alliaria. Both species showed a negative response to the presence of the other species, although Alliaria more so than Calamagrostis. Moreover, the effect of Calamagrostis depended upon population, and, to a lesser extent, the individual clone paired with Alliaria. Competitive effects were stronger in the first experiment compared with when the experiment was repeated in the second year. The influence of Calamagrostis clones on the outcome of the experiment varied among populations and among clones, but also between years. Clones from one of the three populations were more influential than clones from the other two populations. Only one of 15 clones, both from the same population, was influential in both experiments. This research supports a growing literature indicating that intraspecific variability among clones of a dominant species can affect interspecific interactions and that such variability in a native species can affect performance of an invading species.     

Culture-independent phylogenetic analysis of the faecal flora of the rat

The dominant faecal flora of the rat was determined using randomly cloned 16S rDNA comparative sequence analysis. A total of 109 near full-length 16S rDNA clones were sequenced, representing 69 unique 16S rRNA phylotypes or operational taxonomic units (OTUs). Estimates of species richness indicated that approximately 338 species were present in the faeces, suggesting that only 20% of species were identified. Only two of 39 Gram-negative clones aligned with previously cultured species, the remainder fell into a separate lineage within the Bacteroides-Cytophaga phylum. Several clones within this new group were related to 16S rDNA sequences previously identified from mouse faeces. Lactobacilli were the most abundant Gram-positive species, representing 23% of the total clones but only 7% of OTUs. The remaining Gram-positive clones were distributed among the Clostridium coccoides group (9%), the Clostridium leptum subgroup (18%), and throughout the low GC Gram-positive bacteria (13%). The majority of OTUs (63/69 or 91%) were less than 97% homologous to previously cultured bacteria. Faecal samples were also cultured using a variety of anaerobic media. With the exception of the lactobacilli, the cultured isolates demonstrated low species diversity and poorly reflected the population, as defined through comparative sequence analysis.     

Clonal diversity and host distribution in Bordetella bronchiseptica.

A total of 303 isolates of Bordetella bronchiseptica recovered from 11 host species were characterized by the electrophoretic mobilities of 15 metabolic enzymes, and 21 distinctive multilocus genotypes (electrophoretic types) were distinguished on the basis of allele profiles at the enzyme loci. The population structure of B. bronchiseptica is clonal, and its genetic diversity is limited in comparison with most other pathogenic bacteria, perhaps reflecting a relatively recent origin of the species. Electrophoretic types mark clones which are, in many cases, nonrandomly associated with host species. Clones differing only slightly in overall chromosomal genetic character may have pronounced differences in virulence potential. There was considerable variation among individual clones and clone families in degree of host specificity and among various species of hosts in the diversity of clones causing disease. The diversity of clones infecting dogs was an order of magnitude greater than that of clones infecting pigs. Most bordetellosis in pigs in the United States and Japan was found to be caused by strains of a single multilocus genotype.     

Interspecific and intraspecific variation in the performance of three pest aphid species on five grain legume hosts

The suitability of five grain legume species (narrow-leafed lupin, chickpea, faba bean, field pea, lentil) as hosts for three aphid species (green peach aphid, cowpea aphid, bluegreen aphid) was evaluated by measuring the mean relative growth rate (MRGR) and survivorship of nymphs over a 5 day period. For each aphid species, intraspecific (interclonal) variation was also determined by independently measuring the performance of 30 clones collected from a variety of hosts and from different parts of the Western Australia (WA) wheatbelt. The suitability of the grain legumes varied among aphid species. Chickpea was not a suitable host for any of the aphids tested. Averaged over all clones, lentil and faba bean were the most suitable hosts for cowpea aphid, and narrow-leafed lupin was the most suitable host for green peach aphid. Field pea was a suitable host for all three species, but only at a suboptimal level. Cowpea aphid showed the greatest amount of intraspecific variation, with significant variation in MRGR among clones on all hosts except chickpea and significant variation in survivorship on chickpea and lupin. For green peach aphid, there was significant variation in MRGR among clones on field pea and lupin, but in survivorship on lupin only. Bluegreen aphid clones showed significant variation only for MRGR on faba bean and lupin. There were positive correlations in performance of green peach aphid clones on faba bean and lentil, and of cowpea aphid clones on faba bean and lentil. Bluegreen aphid clones showed a negative correlation in performance on field pea and faba bean. These results show the importance of screening cultivars against a wide variety of aphid clones when assessing aphid susceptibility in breeding programmes. The implications of these results on the adaptability of parthenogenetic aphids are discussed.     

Isolation and Characterization of Ty1/copia-like Reverse Transcriptase Sequences from Mung Bean (Vigna radiata)

Ty1/copia-like sequences were amplified from mung bean (Vigna radiata (L.) Wilczek) genomic DNA, by PCR with degenerate oligonucleotide primers corresponding to highly conserved domains in the Ty1/copia-like retrotransposons. PCR fragments of roughly 270 bp were isolated and cloned, and forty clones were sequenced. Thirty-six of the forty clones had unique nucleotide sequences, and eighteen clones had a frameshift, a stop codon, or both. Alignment of the nucleotide sequences indicated that these clones, denoted Tvr, fell into nine subgroups and nine ungrouped sequences. The nucleotide sequence similarity between these elements ranged from 8% to 100%, which indicates high level of sequence heterogeneity among these clones. A phylogenetic analysis comparing these clones with corresponding sequences from other plant species showed that some of the Tvr clones are more closely related to Ty1/copia-like retrotransposons from other species than to other Tvr clones. Dot blot analysis revealed that Ty1/copia-like retrotransposons comprise about 9.3% of the mung bean genome.     

Control of laminin synthesis during differentiation of F9 embryonal carcinoma cells

Molecular clones complementary to the mRNA species for the A, B1 and B2 chains of murine laminin were identified by hybrid-selection and in vitro translation. Northern blot analysis demonstrated that the three clones, p59 (A), p2 (B1) and p16 (B2) hybridized to mRNA species 9.8, 6.0, and 8.0 kb in length, respectively. The three clones were used as probes to monitor the steady-state levels of laminin mRNA species during differentiation of F9 embryonal carcinoma cells induced by treatment with retinoic acid and dibutyryl cyclic AMP. The steady-state levels of the three mRNA species appeared to increase in a coordinate manner. Undetectable levels at the beginning of induction were followed by a dramatic increase in the levels of the three mRNA species between 48 and 72 h. The kinetics parallel the increase in laminin synthesis and the striking morphological changes previously reported.     

Changes in the species composition of a thermotolerant community of acidophilic chemolithotrophic microorganisms upon switching to the oxidation of a new energy substrate

Construction and analysis of the 16S rDNA clone libraries was used to investigate the species composition of two thermotolerant communities of acidophilic chemolithotrophic microorganisms (ACM) isolated from the pulp of laboratory reactors used for oxidation of different gold-containing ore concentrates. The first community was formed during oxidation of the pyrite-arsenopyrite ore concentrate from the Kyuchus deposit. The clones of the bacterial component of this community belonged to the genera Sulfobacillus (32 clones) and Leptospirillum (33 clones). The Sulfobacillus clones belonged to three groups: Sb. thermosulfidooxidans, Sb. benefaciens, and Sb. thermotolerans. All Leptospirillum clones were closely related to L. ferriphilum. All clones of the archaeal component belonged to Ferroplasma acidiphilum. The microorganisms of this community were used as inoculum for biooxidation of a different mineral concentrate, the pyrrhotite-containing pyrite-arsenopyrite ore concentrate from the Olympiadinskoe deposit, and the structure of the community formed in the process was investigated. The clones of the bacterial component of the second community also belonged to the genera Sulfobacillus (14 clones) and Leptospirillum (48 clones). The Sulfobacillus clones belonged to the species Sb. thermosulfidooxidans (13 clones) and Sb. thermotolerans (1 clone). All Leptospirillum clones were closely related to L. ferriphilum. All clones of the archaeal component belonged to Ferroplasma acidiphilum. During the adaptation of the community to a new oxidized mineral substrate, both the composition and the ratio of the constituent microbial species changed.     

Molecular diversity of methanogens in feedlot cattle from Ontario and Prince Edward Island, Canada

The molecular diversity of rumen methanogens in feedlot cattle and the composition of the methanogen populations in these animals from two geographic locations were investigated using 16S rRNA gene libraries prepared from pooled PCR products from 10 animals in Ontario (127 clones) and 10 animals from Prince Edward Island (114 clones). A total of 241 clones were examined, with Methanobrevibacter ruminantium accounting for more than one-third (85 clones) of the clones identified. From these 241 clones, 23 different 16S rRNA phylotypes were identified. Feedlot cattle from Ontario, which were fed a corn-based diet, revealed 11 phylotypes (38 clones) not found in feedlot cattle from Prince Edward Island, whereas the Prince Edward Island cattle, which were fed potato by-products as a finishing diet, had 7 phylotypes (42 clones) not found in cattle from Ontario. Five sequences, representing the remaining 161 clones (67% of the clones), were common in both herds. Of the 23 different sequences, 10 sequences (136 clones) were 89.8 to 100% similar to those from cultivated methanogens belonging to the orders Methanobacteriales, Methanomicrobiales, and Methanosarcinales, and the remaining 13 sequences (105 clones) were 74.1 to 75.8% similar to those from Thermoplasma volcanium and Thermoplasma acidophilum. Overall, nine possible new species were identified from the two clone libraries, including two new species belonging to the order Methanobacteriales and a new genus/species within the order Methanosarcinales. From the present survey, it is difficult to conclude whether the geographical isolation between these two herds or differences between the two finishing diets directly influenced community structure in the rumen. Further studies are warranted to properly assess the differences between these two finishing diets.     

Emergence of brown tides caused by Aureococcus anophagefferens Hargraves et Sieburth in China

Large-scale blooms suspected to be “brown tides” occurred in early summer for three consecutive years from 2009 to 2011 in the coastal waters of Qinhuangdao, China, and had significant negative impacts on the shellfish mariculture industry. To identify the causative species of the blooms, phytoplankton samples were collected from regions with and without bloom in the coastal waters of Qinhuangdao in 2011, and clone libraries were built using eukaryote-specific 18S ribosomal RNA gene (18S rDNA). Altogether 50 clones, including 17 clones from bloom area and 33 clones from nearby regions without bloom were amplified. Blasted in GenBank, 17 clones amplified from the bloom area were assigned to Pelagophyceae (8 clones), Mediophyceae (2 clones), Cryptophyta (2 clones), Dinophyceae (2 clones) and unidentified eukaryotic species (3 clones). Those from the non-bloom site were assigned to Cryptophyta, Eustigmatophyceae, Prasinophyceae, Coscinodiscophyceae, Mediophyceae, Raphidophyceae and Dinophyceae, but not Pelagophyceae. All 8 pelagophyte clones from the bloom area were 99.7–100% similar to a single species, Aureococcus anophagefferens Hargraves et Sieburth, the causative species of brown tides on the east coast of USA. For nearly the entire length of the 18S rDNA, there were 0–6 base pair differences between the 8 amplicons and those of A. anophagefferens from USA. Furthermore, all of the 8 clones were clustered into the same well-supported clade with A. anophagefferens (posterior probability = 0.99) in a phylogenetic tree established for pelagophytes and other related microalgae. In our previous studies, the causative species of the bloom was tentatively identified as a pelagophyte, haptophyte or silicoflagellate, based on the pigment profile of the size-fractioned phytoplankton samples. Based on this study, we conclude that blooms in the coastal waters of Qinhuangdao of the Bohai Sea were brown tides caused by A. anophagefferens. As far as we know, this is the first report of brown tide events caused by A. anophagefferens in China, which is the third country in the world reporting A. anophagefferens blooms in addition to USA and South Africa.     

Immunoglobulin production by a human-mouse somatic cell hybrid

The fusion of human lymphocytes and TEPC-15 mouse myeloma cells, which had not been adapted to culture, resulted in the establishment of in vitro hybrid cell cultures. Ten clones of this somatic cell hybrid were examined. There was preferential exclusion of human chromosomes: between two and five human chromosomes were identified in the hybrid clones by Giemsa banding. All of the clones had the mouse parental histocompatibility antigens, but only four clones also retained the human parental histocompatibility antigens. Secretion of parental immunoglobulin was determined by SDS-gel electrophoresis of species-specific immune precipitates. Synthesis of parental immunoglobulin by individual hybrid cells was determined by double label fluorescent antibody staining. Individual cells from six of the clones secreted and synthesized both human and mouse parental immunoglobulins. Three clones secreted only one parental immunoglobulin. Cells from one of these clones secreted and synthesized only human immunoglobulin. Cells from the remaining two clones secreted only one parental species of immunoglobulin but synthesized both human and mouse immunoglobulins. Finally, one clone did not secrete immunoglobulin, yet the individual cells synthesized both human and mouse parental species of immunoglobulin.     

Cloning and identification of lupin nodule specific cDNA sequences

Yellow lupin nodule specific sequences were selected by screening of cDNA library prepared from lupin nodule poly(A)+RNA. From about 3,000 clones containing fragments of lupin DNA 150-1,500 base pair long, 7% of clones carrying nodule specific sequences were identified. Among them the most abundant sequence species, represented by 32% clones, encodes leghemoglobin. Another abundant species designated pLN13 is represented by 13% clones. The Northern blot analysis of lupin mRNA confirmed nodule specificity of the cloned sequences. The nucleotide sequence of one clone, pLN281 of 225 bp, is presented.     

Uptake of amino acids by uncultivated bacterioplankton of a eutrophic water reservoir

Water bacterioplankton species (clones) feeding on lysine and glycine were revealed in experimental microecosystems using nonsterile cultivation. The structure of the community was analyzed by separation of amplified fragments of the bacterial 16S rRNA gene of by denaturing gradient gel electrophoresis. The dominant species (clones) of the bacterioplankton were identified by molecular cloning and determining the nucleotide sequence of part of the 16S gene of ribosomal RNA. It was found that glycine, one of the prevalent amino acids of a eutrophic reservoir, and lysine, whose content in natural water is lower, are nutrient substrates for various subdominant bacterioplankton species (clones). Thus, free-living bacterial species are highly specialized for the uptake of individual amino acids.     

Metagenomic profiling: microarray analysis of an environmental genomic library

Genomic libraries derived from environmental DNA (metagenomic libraries) are useful for characterizing uncultured microorganisms. However, conventional library-screening techniques permit characterization of relatively few environmental clones. Here we describe a novel approach for characterization of a metagenomic library by hybridizing the library with DNA from a set of groundwater isolates, reference strains, and communities. A cosmid library derived from a microcosm of groundwater microorganisms was used to construct a microarray (COSMO) containing approximately 1-kb PCR products amplified from the inserts of 672 cosmids plus a set of 16S ribosomal DNA controls. COSMO was hybridized with Cy5-labeled genomic DNA from each bacterial strain, and the results were compared with the results for a common Cy3-labeled reference DNA sample consisting of a composite of genomic DNA from multiple species. The accuracy of the results was confirmed by the preferential hybridization of each strain to its corresponding rDNA probe. Cosmid clones were identified that hybridized specifically to each of 10 microcosm isolates, and other clones produced positive results with multiple related species, which is indicative of conserved genes. Many clones did not hybridize to any microcosm isolate; however, some of these clones hybridized to community genomic DNA, suggesting that they were derived from microbes that we failed to isolate in pure culture. Based on identification of genes by end sequencing of 17 such clones, DNA could be assigned to functions that have potential ecological importance, including hydrogen oxidation, nitrate reduction, and transposition. Metagenomic profiling offers an effective approach for rapidly characterizing many clones and identifying the clones corresponding to unidentified species of microorganisms.