Retroperitoneal paraganglioma with loss of heterozygosity of the von Hippel-Lindau gene: a case report and review of the literature.

Von Hippel-Lindau (VHL) disease is an autosomal dominant disease related to germline mutations in VHL. In VHL disease, pheochromocytoma develops in 10%-20% of patients because of germline mutations and loss of heterozygosity of VHL. However, the rate of paraganglioma associated with VHL is low compared with that of pheochromocytoma, and the reason is unknown. In this study, we performed germline and somatic mutation analyses of retroperitoneal paraganglioma that developed in a patient with clinically diagnosed VHL disease and investigated the tumorigenic mechanism of paraganglioma. The patient was a 25-year-old woman who was considered to have VHL disease on the basis of her family history. She was referred to our clinic to investigate a tumor at the bifurcation of the common iliac artery. The tumor was diagnosed as retroperitoneal paraganglioma by clinical evaluations. A left renal cell carcinoma was also suspected. Polymerase chain reaction direct sequencing analysis and polymorphic microsatellite analysis within the VHL locus suggested that loss of heterozygosity of VHL was associated with paraganglioma and renal cell carcinoma. Multiplex ligation-dependent probe amplification analysis showed a loss of the copy number of VHL exons in paraganglioma. These results suggest that VHL disease contributes to the development of paraganglioma. A literature review showed no reported common missense variants involved in the progression of paraganglioma. The loss of heterozygosity of VHL can be a tumorigenic mechanism of retroperitoneal paraganglioma in VHL disease. However, the low rate of paraganglioma compared with pheochromocytoma is not explained by their genetic background alone.

Development and characterization of microsatellite markers in Rosy-faced and other lovebirds (Agapornis spp.) using next-generation sequencing.

Agapornis are a group of small African parrots that are heavily traded around the world. They are invasive species in many places, but some of them are listed as Vulnerable or Near Threatened. However, the genetic tools for assessing inter-individual relationships, population structure, and genetic diversity of these birds are very limited. Therefore, we developed polymorphic microsatellite markers in A. roseicollis and tested the transferability on 5 lovebird species including A. personatus, A. nigrigenis, A. fischeri, A. pullarius, and A. canus, and two closely related outgroups (i.e. Bolbopsittacus lunulatus and Loriculus galgulus). We first performed whole-genome re-sequencing on five individuals of A. roseicollis to identify potential polymorphic loci. Out of 37 loci tested in 11 A. roseicollis, 27 loci were demonstrated to be polymorphic, with the number of the alleles ranging from 2 to 7 (mean = 3.963). The observed heterozygosity ranged from 0 to 0.875 (mean = 0.481) and expected heterozygosity ranged from 0.233 to 0.842 (mean = 0.642). Five loci (Agro-A13, p < 0.01; Agro-A15, p < 0.05; Agro-A43, p < 0.05, Agro-A65, p < 0.05; Agro-A67, p < 0.05) were detected to deviate from Hardy-Weinberg equilibrium, with the presence of null alleles suggested in locus Agro-A13 and Agro-A77. The exclusion powers for PE1 and PE2 are 0.997 and 0.999, respectively. The 27 novel polymorphic markers developed here will be useful for parentage and kinship assignment and population genetics study in Agapornis, and provide a tool for scientific research, captive breeding industry, and invasion and conservation management of these species.

Development of polymorphic microsatellite markers by using de novo transcriptome assembly of Calanthe masuca and C. sinica (Orchidaceae).

BACKGROUND: Calanthe masuca and C. sinica are two genetically closely related species in Orchidaceae. C. masuca is widely distributed in Asia, whereas C. sinica is restricted to Yunnan and Guangxi Provinces in southwest China. Both play important roles in horticulture and are under the pressure of population decline. Understanding their genetic background can greatly help us develop effective conservation strategies for these species. Simple sequence repeats (SSRs) are useful for genetic diversity analysis, presumably providing key information for the study and preservation of the wild populations of the two species we are interested in. RESULTS: In this study, we performed RNA-seq analysis on the leaves of C. masuca and C. sinica, obtaining 40,916 and 71,618 unigenes for each species, respectively. In total, 2,019/3,865 primer pairs were successfully designed from 3,764/7,189 putative SSRs, among which 197 polymorphic SSRs were screened out according to orthologous gene pairs. After mononucleotide exclusion, a subset of 129 SSR primers were analysed, and 13 of them were found to have high polymorphism levels. Further analysis demonstrated that they were feasible and effective against C. masuca and C. sinica as well as transferable to another species in Calanthe. Molecular evolutionary analysis revealed functional pathways commonly enriched in unigenes with similar evolutionary rates in the two species, as well as pathways specific to each species, implicating species-specific adaptation. The divergence time between the two closely related species was tentatively determined to be 3.42 ± 1.86 Mya. CONCLUSIONS: We completed and analysed the transcriptomes of C. masuca and C. sinica, assembling large numbers of unigenes and generating effective polymorphic SSR markers. This is the first report of the development of expressed sequence tag (EST)-SSR markers for Calanthe. In addition, our study could enable further genetic diversity analysis and functional and comparative genomic studies on Calanthe.

De novo generation of adipocytes from circulating progenitor cells in mouse and human adipose tissue.

White adipocytes in adults are typically derived from tissue resident mesenchymal progenitors. The recent identification of de novo production of adipocytes from bone marrow progenitor-derived cells in mice challenges this paradigm and indicates an alternative lineage specification that adipocytes exist. We hypothesized that alternative lineage specification of white adipocytes is also present in human adipose tissue. Bone marrow from transgenic mice in which luciferase expression is governed by the adipocyte-restricted adiponectin gene promoter was adoptively transferred to wild-type recipient mice. Light emission was quantitated in recipients by in vivo imaging and direct enzyme assay. Adipocytes were also obtained from human recipients of hematopoietic stem cell transplantation. DNA was isolated, and microsatellite polymorphisms were exploited to quantify donor/recipient chimerism. Luciferase emission was detected from major fat depots of transplanted mice. No light emission was observed from intestines, liver, or lungs. Up to 35% of adipocytes in humans were generated from donor marrow cells in the absence of cell fusion. Nontransplanted mice and stromal-vascular fraction samples were used as negative and positive controls for the mouse and human experiments, respectively. This study provides evidence for a nontissue resident origin of an adipocyte subpopulation in both mice and humans.

The Correlation Between Candida Colonization of Distinct Body Sites and Invasive Candidiasis in Emergency Intensive Care Units: Statistical and Molecular Biological Analysis.

Both statistical and molecular biological methods were used to evaluate the association between Candida colonization of different body sites and invasive candidiasis (IC) and analyse the potential infection sources of IC. Candida surveillance cultures from the urine, sputum, rectum and skin were performed on patients admitted to an emergency intensive care units (EICU) of a tertiary care hospital in Shanghai, China, from February 2014 to January 2015. Specimens were collected once a week at admission and thereafter. The patients' clinical data were collected, and Candida isolates were genotyped using polymorphic microsatellite markers. A total of 111 patients were enrolled. Patients with positive urine (23.3 vs. 2.5 %, p = 0.001) and rectal swab (13.6 vs. 0 %, p = 0.010) cultures were more likely to develop IC. However, the risk for IC was not significantly different among patients with and without respiratory (10.0 vs. 5.8 %, p = 0.503) and skin (33.3 vs. 6.5 %, p = 0.056) colonization. Gene microevolution frequently occurred at rectal swab and urine sites, and IC with possible source of infection was caused by rectal isolates (2/7), urine isolates (4/7) and sputum isolate (1/7).The colonization of gut and urinary tract maybe more relevant indicators of IC, which should be taken into consideration when selecting practical body sites for Candida surveillance cultures.

Genome-wide development of polymorphic microsatellite markers and their application in peanut breeding program

BACKGROUND: Cultivated peanut (Arachis hypogaea. L) represents one of the most important oil crops in the world. Although much effort has been expended to characterize microsatellites or Simple Sequence Repeats (SSRs) in peanut, the quantity and quality of the markers in breeding applications remain limited. Here, genome-wide SSR characterization and marker development were performed using the recently assembled genome of the cultivar Tifrunner. RESULTS: In total, 512,900 microsatellites were identified from 2556.9-Mb genomic sequences. Based on the flanking sequences of the identified microsatellites, 7757 primer pairs (markers) were designed, and further evaluated in the assembled genomic sequences of the tetraploid Arachis cultivars, Tifrunner and Shitouqi, and the diploid ancestral species, A. duranensis and A. ipaensis. In silico PCR analysis showed that the SSR markers had high amplification efficiency and polymorphism in four Arachis genotypes. Notably, nearly 60% of these markers were single-locus SSRs in tetraploid Arachis species, indicating they are more specific in distinguishing the alleles of the A and B sub-genomes of peanut. In addition, two markers closely related with purple testa color and 27 markers near to FAD2 genes were identified, which could be used for breeding varieties with purple testa and high-oleic acid content, respectively. Moreover, the potential application of these SSR markers in tracking introgressions from Arachis wild relatives was discussed. CONCLUSIONS: This study reported the development of genomic SSRs from assembled genomic sequences of the tetraploid Arachis Tifrunner, which will be useful for diversity analysis, genetic mapping and functional genomics studies in peanut

Development of 12 novel polymorphic microsatellite markers using a next generation sequencing approach for Spiculopteragia spiculoptera, a nematode parasite of deer.

Twelve novel polymorphic microsatellite markers were produced and characterized for Spiculopteragia spiculoptera (Nematoda, Trichostrongyloidae) a common parasite of abomasum of Roe and Red deer, using next generation sequencing approach, and two multiplexes PCR were developed with these markers. Polymorphism of each locus was tested in 40 individuals of this species from diverse wild populations of cervids, and was tested for crossed-amplification on four other species of nematodes, close to S. spiculoptera among the Trichostrongyloidea: 20 Spiculopteragia houdemeri, 34 Ostertagia leptospicularis, 16 Ashworthius sidemi, and 25 Trichostrongylus spp. Our new microsatellite markers seem to be specific to Spiculopteragia spiculoptera since no amplifications were obtained for the four other species. The number of alleles per locus ranged from 2 to 12, the average observed and expected heterozygosity per locus ranged from 0.025 to 0.641 and from 0.049 to 0.664, respectively. Four of the 12 microsatellite loci showed significant deviations from Hardy-Weinberg equilibrium (which two slightly significant). One locus pair showed significant linkage disequilibrium (Sspi4 vs. Sspi8). Neither evidence of scoring error due to stuttering nor evidence of large allele dropout was found at all of the 12 loci, but evidence of null alleles was indicated at three loci because of general excess of homozygotes for most allele size classes. These polymorphic loci will be useful markers to study population genetics structure of Spiculopteragia spiculoptera in order to understand transfer and to explain the relationships between deer populations.