Male mating preference of two cryptic species of the herbivorous insect Eccritotarsus catarinensis

When using biological control against pest populations, more than one biocontrol agent might be introduced simultaneously. This could be counterproductive in the event of negative interactions between the biocontrol agents. Within and between species interactions have a strong impact on mating behaviour and reproduction, and can have an impact on the effectiveness of biological control. We studied the reproductive compatibility between two geographically isolated strains (Brazil and Peru) of Eccritotarsus catarinensis (Heteroptera: Miridae), a biocontrol agent against the invasive aquatic weed, Eichhornia crassipes. By performing inter- and intra-species mating experiments, we investigated whether or not males from each of the cryptic species would be able to distinguish between partners from either species, and if they would mate with partners from the opposite species. Our results showed the decrease in lifetime fecundity, and most importantly, the lack of production of offspring from eggs resulting from forced hybridisation. We showed that Peruvian males mated only with females from their own species, and did not mate with females from the Brazilian species. In contrast, Brazilian males mated equally with females from both species, but needed significantly more time in order to commence a mating, and no offspring were produced from eggs resulting from hybridisation. Although future studies demand more rigorous controls, our results indicate asymmetrical sexual isolation between the two species. We speculate on mechanisms involved in reproductive isolation in the two cryptic species, and the possible implications for effective biocontrol, and include some morphological measurements that might support our assumptions.     

A Pathogenic Mosaic TP53 Mutation in Two Germ Layers Detected by Next Generation Sequencing

Background

Li-Fraumeni syndrome is caused by germline TP53 mutations and is clinically characterized by a predisposition to a range of cancers, most commonly sarcoma, brain tumours and leukemia. Pathogenic mosaic TP53 mutations have only rarely been described.

Methods and Findings

We describe a 2 years old child presenting with three separate cancers over a 6 month period; two soft tissue mesenchymal tumors and an aggressive metastatic neuroblastoma. As conventional testing of blood DNA by Sanger sequencing for mutations in TP53, ALK, and SDH was negative, whole exome sequencing of the blood DNA of the patient and both parents was performed to screen more widely for cancer predisposing mutations. In the patient''s but not the parents'' DNA we found a c.743 G>A, p.Arg248Gln (CCDS11118.1) TP53 mutation in 3–20% of sequencing reads, a level that would not generally be detectable by Sanger sequencing. Homozygosity for this mutation was detected in all tumor samples analyzed, and germline mosaicism was demonstrated by analysis of the child''s newborn blood spot DNA. The occurrence of separate tumors derived from different germ layers suggests that this de novo mutation occurred early in embryogenesis, prior to gastrulation.

Conclusion

The case demonstrates pathogenic mosaicim, detected by next generation deep sequencing, that arose in the early stages of embryogenesis.     

Measurement of Free Intracellular Calcium in the Brain by 19F-Nuclear Magnetic Resonance Spectroscopy

We report the first measurement of the free intracellular calcium level in an actively metabolising intact cerebral tissue preparation. To this end, we applied the recently developed 19F-nuclear magnetic resonance calcium chelator, 5,5'-F2-1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (5FBAPTA), in superfused cerebral cortical slices to give values for the intracellular Ca2+ concentration of 350 and 480 nM, at external calcium concentrations of 1.2 and 2.4 mM, respectively. Under both conditions, the intracellular Ca2+ concentration was increased by depolarisation using a high external K+ concentration. Interleaved 31P spectra showed that the presence of the 5FBAPTA had a deleterious effect on the metabolic state of the tissue with an external Ca2+ concentration of 1.2 mM, but normal viability was maintained using 2.4 mM.     

Molecular characterization of PoGT8D and PoGT43B, two secondary wall-associated glycosyltransferases in poplar

Dicot wood is mainly composed of cellulose, lignin and glucuronoxylan (GX). Although the biosynthetic genes for cellulose and lignin have been studied intensively, little is known about the genes involved in the biosynthesis of GX during wood formation. Here, we report the molecular characterization of two genes, PoGT8D and PoGT43B, which encode putative glycosyltransferases, in the hybrid poplar Populus alba x tremula. The predicted amino acid sequences of PoGT8D and PoGT43B exhibit 89 and 75% similarity to the Arabidopsis thaliana IRREGULAR XYLEM8 (IRX8) and IRX9, respectively, both of which have been shown to be required for GX biosynthesis. The PoGT8D and PoGT43B genes were found to be expressed in cells undergoing secondary wall thickening, including the primary xylem, secondary xylem and phloem fibers in stems, and the secondary xylem in roots. Both PoGT8D and PoGT43B are predicted to be type II membrane proteins and shown to be targeted to Golgi. Overexpression of PoGT43B in the irx9 mutant was able to rescue the defects in plant size and secondary wall thickness and partially restore the xylose content. Taken together, our results demonstrate that PoGT8D and PoGT43B are Golgi-localized, secondary wall-associated proteins, and PoGT43B is a functional ortholog of IRX9 involved in GX biosynthesis during wood formation.     

Sequence specificity of the P1 modification methylase (M.Eco P1) and the DNA methylase (M.Eco dam) controlled by the Escherichia coli dam gene.

Labeled oligonucleotides have been fractionated from pancreatic DNase digests of DNA that had been methylated in vitro with the P1 modification enzyme (M·Eco P1) or with the DNA-adenine methylase (M·Eco dam) controlled by the Escherichia coli dam gene. The sequences of methylated oligonucleotides were established for M·Eco dam modification of calf thymus DNA. The results show that M·Eco dam inethylates adenine residues contained in the twofold symmetrical sequence, 5′ … G-A-T-C … 3′. The sequence for the site methylated by M·Eco P1 has also been deduced; we propose that M·Eco P1 modification produces the following methylated pentameric sequence: 5′ … A-G-A¹-C-Py … 3′ (where $\text{A}{}^1\text{= N}{}^6$ methyladenine and Py is C or T).     

Host location and selection by the symbiotic sargassum crab <Emphasis Type="Italic">Portunus sayi</Emphasis>: the role of chemical,visual and tactile cues

The Sargassum community consists of a unique and diverse assemblage of symbiotic fauna critical to pelagic food chains. Associated symbionts presumably have adaptations to assist in finding Sargassum. In situ scattered Sargassum patches accumulate as they are pushed toward the shoreline (via wind, waves, currents or tides) and are frequently less than 1 m apart and in depths of 10 cm or less as the patches approach the shoreline Crabs, and other symbiotic fauna, must relocate to another patch that is seaward in direction or likely perish as their current patch will likely become beached. This study investigated sensory cues used for host location and selection by the Sargassum crab, Portunus sayi. Chemical detection trials were conducted with a two-chamber choice apparatus with Sargassum spp. and Thalassia testudinum as habitat source odors. Visual detection trials (devoid of chemical cues) and habitat selection trials were conducted in which crabs were given a choice between hosts. Results showed that P. sayi responded to chemicals from Sargassum spp. Crabs visually located host habitats but did not visually distinguish between different hosts. In host selection trials, crabs selected Sargassum spp. over artificial Sargassum and T. testudinum. These results suggest that crabs isolated from Sargassum likely use chemoreception; within visual proximity of a potential patch, crabs likely use both chemical and visual information.     

Characterization of an analphoid, neocentromere-positive inv dup 8p marker chromosome using multiplex whole chromosome and sub-telomere FISH analyses

A 30-year-old male patient with mild mental retardation was found to have a small supernumerary marker chromosome (SMC) in 90% of his peripheral blood cells and in 100% of his fibroblast cells. Multiplex whole chromosome and sub-telomere FISH analyses were used to determine that this SMC is an inverted duplicated distal chromosome 8p fragment. Although it was negative for alpha-DNA sequences, this marker had a functional kinetochore (neocentromere) demonstrated by a positive signal with a CENP-C antibody. Apparently intact 8p telomeres at the marker's ends were demonstrated by using a telomere repeat FISH probe. The patient's phenotypically normal mother on G-banding analysis had a small marker chromosome in 8% of her peripheral blood cells in two cultures of the first specimen studied. The marker was not seen in any subsequent maternal peripheral blood or fibroblast specimens. Although it was impossible to further characterize the maternal SMC, it was suggested that the mother had the same marker as the one seen in the proband. Inverted duplicated chromosomal fragments are the most frequent type of analphoid markers. Stable inverted duplicated 8p marker chromosomes were previously reported in three other patients. They all apparently occurred de novo and were found to be positive for kinetochore-associated proteins. Evidence for the possible inheritance of an inverted-duplicated, analphoid SMC was not shown to-date. This study also demonstrates a practical, straightforward approach for analphoid marker characterization in clinical laboratory settings, using whole chromosome multiplex and subtelomere-specific FISH analyses. FISH probes for all sub-telomere chromosomal regions are commercially available and the large majority of analphoid marker chromosomes involve telomere regions.     

A PCR-RFLP for KIT associated with tobiano spotting pattern in horses

An MspI polymorphism was identified in intron 13 of the equine homologue of proto-oncogene c-kit (KIT) by comparing DNA sequences from horses with solid coat colour and horses homozygous for the tobiano spotting (To) gene. The allele associated with solid coat colour was designated KM0, while the allele associated with the tobiano pattern created an additional MspI restriction site and was designated KM1. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) studies using DNA from hair follicles demonstrated that all 129 of 129 tobiano patterned horses possessed the KM1 allele. However, three of 104 solid-coloured thoroughbred horses also possessed the KM1 allele. Therefore, while KM1 is strongly associated with the gene for To, the association is not absolute. However, this test appears more efficacious to identify putative homozygotes for To than current biochemical testing methods using albumin (Alb) and vitamin D binding protein (Gc) haplotypes.     

Effect of latent human immunodeficiency virus infection on cell surface phenotype.

Highly active antiretroviral therapy has succeeded in many cases in suppressing virus production in patients infected with human immunodeficiency virus (HIV); however, once treatment is discontinued, virus replication is rekindled. One reservoir capable of harboring HIV in a latent state and igniting renewed infection once therapy is terminated is a resting T cell. Due to the sparsity of T cells latently infected with HIV in vivo, it has been difficult to study viral and cellular interactions during latency. The SCID-hu (Thy/Liv) mouse model of HIV latency, however, provides high percentages of latently infected cells, allowing a detailed analysis of phenotype. Herein we show that latently infected cells appear phenotypically normal. Following cellular stimulation, the virus completes its life cycle and induces phenotypic changes, such as CD4 and major histocompatibility complex class I down-regulation, in the infected cell. In addition, HIV expression following activation did not correlate with expression of the cellular activation marker CD25. The apparently normal phenotype and lack of HIV expression in latently infected cells could prevent recognition by the immune response and contribute to the long-lived nature of this reservoir.