Development of a core set of single-locus SSR markers for allotetraploid rapeseed (Brassica napus L.)

Brassica napus (AACC) is a recent allotetraploid species evolved through hybridization between two diploids, B. rapa (AA) and B. oleracea (CC). Due to extensive genome duplication and homoeology within and between the A and C genomes of B. napus, most SSR markers display multiple fragments or loci, which limit their application in genetics and breeding studies of this economically important crop. In this study, we collected 3,890 SSR markers from previous studies and also developed 5,968 SSR markers from genomic sequences of B. rapa, B. oleracea and B. napus. Of these, 2,701 markers that produced single amplicons were putative single-locus markers in the B. napus genome. Finally, a set of 230 high-quality single-locus SSR markers were established and assigned to the 19 linkage groups of B. napus using a segregating population with 154 DH individuals. A subset of 78 selected single-locus SSR markers was proved to be highly stable and could successfully discriminate each of the 45 inbred lines and hybrids. In addition, most of the 230 SSR markers showed the single-locus nature in at least one of the Brassica species of the U’s triangle besides B. napus. These results indicated that this set of single-locus SSR markers has a wide range of coverage with excellent stability and would be useful for gene tagging, sequence scaffold assignment, comparative mapping, diversity analysis, variety identification and association mapping in Brassica species.     

Modelling for rearrangement of fusiform initials during radial growth of the vascular cambium in Pinus sylvestris L.

In contrast to common belief, recent studies have confirmed that intrusive growth of fusiform cambial initials has a significant role in the rearrangement of the initials, but does not contribute to the cambial circumference increment. We observed a rapid rearrangement of cambial initials on a long series of transverse sections of the vascular cambium and the wood of a 50-year-old pine (Pinus sylvestris L.) tree. A comparison of cell arrangement in consecutive sections, as well as a critical analysis of tangential reconstructions, has confirmed that changes in cell locations in a group of cells on the tangential surface caused no change in the total tangential width of the whole group. Models illustrating changes in locations of the initials have been proposed, assuming that intrusive growth, which makes the growing initials intrude between the neighbouring initials and their immediate derivatives, is localized on the longitudinal edges of cells. We infer that intrusive growth of the cambial initials in P. sylvestris is not involved in the cambial circumference increment, but plays a significant role in the rearrangement of the initials, probably allowing for a relaxation of shearing strains generated during radial growth. The relationship of intrusive growth with the elimination of initials has been discussed with reference to the frequency of anticlinal divisions. It has been proposed that the occurrence of anticlinal divisions in excess over the actual requirement for increase in the cambial circumference could be due to internal shearing strains.     

Diversity of protists and bacteria determines predation performance and stability

Predation influences prey diversity and productivity while it effectuates the flux and reallocation of organic nutrients into biomass at higher trophic levels. However, it is unknown how bacterivorous protists are influenced by the diversity of their bacterial prey. Using 456 microcosms, in which different bacterial mixtures with equal initial cell numbers were exposed to single or multiple predators (Tetrahymena sp., Poterioochromonas sp. and Acanthamoeba sp.), we showed that increasing prey richness enhanced production of single predators. The extent of the response depended, however, on predator identity. Bacterial prey richness had a stabilizing effect on predator performance in that it reduced variability in predator production. Further, prey richness tended to enhance predator evenness in the predation experiment including all three protists predators (multiple predation experiment). However, we also observed a negative relationship between prey richness and predator production in multiple predation experiments. Mathematical analysis of potential ecological mechanisms of positive predator diversity—functioning relationships revealed predator complementarity as a factor responsible for both enhanced predator production and prey reduction. We suggest that the diversity at both trophic levels interactively determines protistan performance and might have implications in microbial ecosystem processes and services.     

Identification of chloroplast genome loci suitable for high‐resolution phylogeographic studies of Colocasia esculenta (L.) Schott (Araceae) and closely related taxa

Recently, we reported the chloroplast genome‐wide association of oligonucleotide repeats, indels and nucleotide substitutions in aroid chloroplast genomes. We hypothesized that the distribution of oligonucleotide repeat sequences in a single representative genome can be used to identify mutational hotspots and loci suitable for population genetic, phylogenetic and phylogeographic studies. Using information on the location of oligonucleotide repeats in the chloroplast genome of taro (Colocasia esculenta), we designed 30 primer pairs to amplify and sequence polymorphic loci. The primers have been tested in a range of intra‐specific to intergeneric comparisons, including ten taro samples (Colocasia esculenta) from diverse geographical locations, four other Colocasia species (C. affinis, C. fallax, C. formosana, C. gigantea) and three other aroid genera (represented by Remusatia vivipara, Alocasia brisbanensis and Amorphophallus konjac). Multiple sequence alignments for the intra‐specific comparison revealed nucleotide substitutions (point mutations) at all 30 loci and microsatellite polymorphisms at 14 loci. The primer pairs reported here reveal levels of genetic variation suitable for high‐resolution phylogeographic and evolutionary studies of taro and other closely related aroids. Our results confirm that information on repeat distribution can be used to identify loci suitable for such studies, and we expect that this approach can be used in other plant groups.     

Oral Ingestion of Transgenic RIDL Ae. aegypti Larvae Has No Negative Effect on Two Predator Toxorhynchites Species

Dengue is the most important mosquito-borne viral disease. No specific treatment or vaccine is currently available; traditional vector control methods can rarely achieve adequate control. Recently, the RIDL (Release of Insect carrying Dominant Lethality) approach has been developed, based on the sterile insect technique, in which genetically engineered ‘sterile’ homozygous RIDL male insects are released to mate wild females; the offspring inherit a copy of the RIDL construct and die. A RIDL strain of the dengue mosquito, Aedes aegypti, OX513A, expresses a fluorescent marker gene for identification (DsRed2) and a protein (tTAV) that causes the offspring to die. We examined whether these proteins could adversely affect predators that may feed on the insect. Aedes aegypti is a peri-domestic mosquito that typically breeds in small, rain-water-filled containers and has no specific predators. Toxorhynchites larvae feed on small aquatic organisms and are easily reared in the laboratory where they can be fed exclusively on mosquito larvae. To evaluate the effect of a predator feeding on a diet of RIDL insects, OX513A Ae. aegypti larvae were fed to two different species of Toxorhynchites (Tx. splendens and Tx. amboinensis) and effects on life table parameters of all life stages were compared to being fed on wild type larvae. No significant negative effect was observed on any life table parameter studied; this outcome and the benign nature of the expressed proteins (tTAV and DsRed2) indicate that Ae. aegypti OX513A RIDL strain is unlikely to have any adverse effects on predators in the environment.     

Bacteriophage Receptor Binding Protein Based Assays for the Simultaneous Detection of Campylobacter jejuni and Campylobacter coli

Campylobacter jejuni and Campylobacter coli are the most common bacterial causes of foodborne gastroenteritis which is occasionally followed by a debilitating neuropathy known as Guillain-Barré syndrome. Rapid and specific detection of these pathogens is very important for effective control and quick treatment of infection. Most of the diagnostics available for these organisms are time consuming and require technical expertise with expensive instruments and reagents to perform. Bacteriophages bind to their host specifically through their receptor binding proteins (RBPs), which can be exploited for pathogen detection. We recently sequenced the genome of C. jejuni phage NCTC12673 and identified its putative host receptor binding protein, Gp047. In the current study, we localized the receptor binding domain to the C-terminal quarter of Gp047. CC-Gp047 could be produced recombinantly and was capable of agglutinating both C. jejuni and C. coli cells unlike the host range of the parent phage which is limited to a subset of C. jejuni isolates. The agglutination procedure could be performed within minutes on a glass slide at room temperature and was not hindered by the presence of buffers or nutrient media. This agglutination assay showed 100% specificity and the sensitivity was 95% for C. jejuni (n = 40) and 90% for C. coli (n = 19). CC-Gp047 was also expressed as a fusion with enhanced green fluorescent protein (EGFP). Chimeric EGFP_CC-Gp047 was able to specifically label C. jejuni and C. coli cells in mixed cultures allowing for the detection of these pathogens by fluorescent microscopy. This study describes a simple and rapid method for the detection of C. jejuni and C. coli using engineered phage RBPs and offers a promising new diagnostics platform for healthcare and surveillance laboratories.