Evaluation of SHOX defects in the era of next‐generation sequencing

Short stature homeobox (SHOX) haploinsufficiency is a frequent cause of short stature. Despite advances in sequencing technologies, the identification of SHOX mutations continues to be performed using standard methods, including multiplex ligation‐dependent probe amplification (MLPA) followed by Sanger sequencing. We designed a targeted panel of genes associated with growth impairment, including SHOX genomic and enhancer regions, to improve the resolution of next‐generation sequencing for SHOX analysis. We used two software packages, CONTRA and Nexus Copy Number, in addition to visual analysis to investigate the presence of copy number variants (CNVs). We evaluated 15 patients with previously known SHOX defects, including point mutations, deletions and a duplication, and 77 patients with idiopathic short stature (ISS). The panel was able to confirm all known defects in the validation analysis. During the prospective evaluation, we identified two new partial SHOX deletions (one detected only by visual analysis), including an intragenic deletion not detected by MLPA. Additionally, we were able to determine the breakpoints in four cases. Our results show that the designed panel can be used for the molecular investigation of patients with ISS, and it may even detect CNVs in SHOX and its enhancers, which may be present in a significant fraction of patients.Copy number variants analyses and Sanger sequencing of breakpoint regions in Case 11, which has a heterozygous deletions involving exons 4, 5, and 6a of short stature homeobox (SHOX).Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/151254/1/cge13587.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151254/2/CGE_13587-sup-0001-Supinfo.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151254/3/cge13587_am.pd

First report of anthracnose caused by Colletotrichum spaethianum on Allium fistulosum in Brazil

The Welch onion (Allium fistulosum L.) is widely grown in temperate and subtropical regions worldwide and has many important culinary uses. The occurrence of anthracnose on Welch onion has been reported in Korea and is caused by Colletotrichum circinans (Kim et al. 2008). Since 2012, symptoms typical of anthracnose have been observed on Welch onions in a vegetable garden located in the Japanese Colony of Manaus, Amazonas, Brazil (59°59′06″ W; 03°04′16″ S). This disease occurred in 50% of the seedlings and the symptoms consist of brown necrotic spots that extend along the entire leaf. Acervuli collected directly from the leaves were plated onto potato dextrose agar (PDA) and then incubated at 25°C for three to four days. Single-spore cultures were obtained from three isolates from different plants. On PDA medium, the isolates initially produced white colonies, which then turned gray and had an orange-colored conidial mass. On Spezieller Nährstoffarmer Agar (SNA) medium (Leslie and Summerell, 2006), they formed numerous black structures such as sclerotia, setae, and acervuli. Conidia on SNA are hyaline, aseptate, curved or slightly curved, with a rounded apex and truncated base that is 13.1 to 20.2 μm long and 3.3 to 4.0 μm wide. The appressoria are solitary or in loose groups, dark brown, irregularly shaped, sometimes partially lobed, smooth-walled, and from 5.6 to 10.8 μm long and 4.3 to 8.2 μm wide. An alignment of the actin (ACT) and chitin synthase (CHS-1) partial gene sequences showed 100% identity with Colletotrichum spaethianum (Allesch.) Damm, P. F. Cannon & Crous (CBS 167.49). Maximum likelihood analysis was done using the published sequences of the ACT and CHS-1 genes from C. spaethianum and other Colletotrichum species that have curved conidia (Damm et al. 2009; Vieira et al. 2014). The individual data sets were combined using the web tool FaBox (1.41) and analysis with PAUP (1000 bootstrap replicates). Based on morphological characteristics and phylogenetic analysis, the isolates were identified as C. spaethianum. The sequences for the isolates obtained in the present study were deposited in GenBank (ACT Accession Nos. KT184300 to KT184302; CHS-1 Accession Nos. KT184303 to KT184305). The cultures were deposited in the Culture Collection of Microorganisms of the National Institute of the Amazonian Research (Instituto Nacional de Pesquisas da Amazônia, INPA) (INPA 2615, 2770, and 2774). Five Welch onion seedlings were sprayed with a conidial suspension (106 conidia/ml) for each isolate and control seedlings were sprayed with sterile water. Plants were covered with plastic bags and maintained at 28°C ± 2°C in a greenhouse and a 12-h photoperiod. Symptoms typical of anthracnose were induced five days postinoculation, and signs of the pathogen were observed at 12 days postinoculation. No symptoms were observed in the control plants. C. spaethianum was reisolated from symptomatic plants, completing Koch’s postulates. C. spaethianum has been described on Hosta sieboldiana in Germany, Lilium sp. in South Korea, Hemerocallis sp. in New Zealand (Damm et al. 2012), Hemerocallis fulva, Hemerocallis citrine, and Peucedanum praeruptorum in China (Yang et al. 2012; Guo et al. 2013), and Hemerocallis flava in Brazil (Vieira et al. 2014). To our knowledge, this is the first report of C. spaethianum on A. fistulosum. © 2016, American Phytopathological Society. All rights reserved

Divergência em QTLs e variância genética para teores de proteína e óleo em soja

Resumo: O objetivo deste trabalho foi avaliar a relação entre os parâmetros de divergência em regiões de QTLs e a variância genética em genótipos de soja, quanto aos teores de proteína e óleo nos grãos. Dois grupos de genótipos foram avaliados, em diferentes ambientes, quanto aos teores de proteína e óleo e genotipados com marcadores moleculares de regiões de QTLs. A partir de cada grupo, estabeleceram-se subgrupos por critérios pré-definidos e avaliou-se a relação entre os parâmetros, tendo-se comparado a divergência média e a variância genética entre os subgrupos. Os subgrupos foram definidos com base nos critérios de diferença em divergência média, homogeneidade e heterogeneidade nos subgrupos e proximidade em uma projeção tridimensional da matriz de distância. As percentagens de concordância entre maiores valores de divergência média e de variância genética para o total de subgrupos de cada grupo inicial foram de 72,5 e de 73,4%, respectivamente. Portanto, nestes genótipos, há relação positiva entre as estimativas de divergência em regiões de QTL e variância genética para os teores de proteína e de óleo dos grãos. As distâncias genéticas com base nos marcadores moleculares de regiões de QTLs são eficientes para a predição da variabilidade genética em genótipos de soja para os teores de proteína e de óleo dos grãos

Track A Basic Science

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/138319/1/jia218438.pd

Risk profiles and one-year outcomes of patients with newly diagnosed atrial fibrillation in India: Insights from the GARFIELD-AF Registry.

BACKGROUND: The Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF) is an ongoing prospective noninterventional registry, which is providing important information on the baseline characteristics, treatment patterns, and 1-year outcomes in patients with newly diagnosed non-valvular atrial fibrillation (NVAF). This report describes data from Indian patients recruited in this registry. METHODS AND RESULTS: A total of 52,014 patients with newly diagnosed AF were enrolled globally; of these, 1388 patients were recruited from 26 sites within India (2012-2016). In India, the mean age was 65.8 years at diagnosis of NVAF. Hypertension was the most prevalent risk factor for AF, present in 68.5% of patients from India and in 76.3% of patients globally (P < 0.001). Diabetes and coronary artery disease (CAD) were prevalent in 36.2% and 28.1% of patients as compared with global prevalence of 22.2% and 21.6%, respectively (P < 0.001 for both). Antiplatelet therapy was the most common antithrombotic treatment in India. With increasing stroke risk, however, patients were more likely to receive oral anticoagulant therapy [mainly vitamin K antagonist (VKA)], but average international normalized ratio (INR) was lower among Indian patients [median INR value 1.6 (interquartile range {IQR}: 1.3-2.3) versus 2.3 (IQR 1.8-2.8) (P < 0.001)]. Compared with other countries, patients from India had markedly higher rates of all-cause mortality [7.68 per 100 person-years (95% confidence interval 6.32-9.35) vs 4.34 (4.16-4.53), P < 0.0001], while rates of stroke/systemic embolism and major bleeding were lower after 1 year of follow-up. CONCLUSION: Compared to previously published registries from India, the GARFIELD-AF registry describes clinical profiles and outcomes in Indian patients with AF of a different etiology. The registry data show that compared to the rest of the world, Indian AF patients are younger in age and have more diabetes and CAD. Patients with a higher stroke risk are more likely to receive anticoagulation therapy with VKA but are underdosed compared with the global average in the GARFIELD-AF. CLINICAL TRIAL REGISTRATION-URL: http://www.clinicaltrials.gov. Unique identifier: NCT01090362

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time, and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space. While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes, vast areas of the tropics remain understudied. In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity, but it remains among the least known forests in America and is often underrepresented in biodiversity databases. To worsen this situation, human-induced modifications may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge, it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

First report of anthracnose on welsh onion (Allium fistulosum) in Brazil caused by Colletotrichum theobromicola and c. truncatum

Welsh onion (Allium fistulosum L.) is cultivated in temperate and subtropical regions worldwide. Brown, necrotic, anthracnose-like lesions that extended the length of Welsh onion leaves were first observed in the village Paraná do Supiá (60°45′34.8″ W; 3°24′17.11″ S), Manacapuru district, in 2008 and in the village Nossa Senhora de Fátima (60°17′35.2″ W; 3°15′51.5″ S), Iranduba district, in 2014. Both villages are situated in Amazonas state, Brazil. Aggregated orange spores and acervuli collected directly from the necrotic lesions were plated on potato dextrose agar (PDA) and incubated at 25°C. Single spore cultures were deposited in the Microorganisms Culture Collection of the National Institute of Amazonian Research, Manaus, Brazil (INPA 1809 and INPA 2743). For morphological characterization, the strains were grown on SNA. The isolate INPA 1809 formed a colony varying from light to dark gray. Microscopically, conidia were cylindrical to subcylindrical, aseptate, measuring (n = 20) 10.8 to 17.4 × 4.0 to 4.6 μm. Appressoria were solitary, dark brown, irregular, sometimes lobate, measuring 6.3 to 13.6 × 4.0 to 5.6 μm. The isolate INPA 2743 formed light gray colonies. Conidia were hyaline, aseptate, slightly curved with tapered tips and truncate bases, measuring (n = 20) 16 to 22 × 2.5 to 3.5 μm. Appressoria were solitary or in groups, light brown, lobate or rounded, measuring from 9.0 to 19.0 × 4.3 to 5.0 μm. Total DNA was extracted and fragments of actin (ACT), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and chitin synthase (CHS) genes were amplified, sequenced, and deposited in GenBank. Accession numbers of sequences from isolate INPA 1809 were KX702276 (ACT), KX702274 (GADPH), and KX702272 (CHS). Likewise, accession numbers deposited from isolate INPA 2743 were KX702275 (ACT), KX702273 (GADPH), and KX702271 (CHS). Bayesian inference analyses were performed with concatenated genes sequences (ACT, GAPDH, and CHS) and showed that the isolate INPA 1809 clustered with the ex-type specimen of Colletotrichum theobromicola (CBS 124945) in a clade with high support (posterior probability = 1) and the isolate INPA 2743 clustered with the ex-epitype specimen of C. truncatum (CBS 151.35) in a clade with high support (posterior probability = 1). Fifteen Welsh onion seedlings were used to verify pathogenicity and to complete Koch’s postulates. Five of the seedlings were sprayed with 106 conidia/ml spore suspension from INPA 1809, another five seedlings were inoculated with 106 conidia/ml of INPA 2743, and five seedlings were sprayed with sterile water. All seedlings were covered with plastic bags, 24 h after inoculation, and kept at 27°C in a greenhouse and a 12-h photoperiod. Typical symptoms of anthracnose (brown necrotic lesions) were observed 5 days after inoculation with both inoculated strains. The fungal species were recovered from symptomatic plants, while no symptoms were observed on the controls. Anthracnose on Welsh onion caused C. circinans was reported from Korea (Kim et al. 2008). In Brazil, C. spaethianum was reported causing the same disease on Welsh onion in a vegetable garden in Manaus, Amazonas (Santana et al. 2016). To our knowledge, this is the first report of C. theobromicola and C. truncatum causing anthracnose on Welsh onion in Brazil. The identification of these species will contribute to the adoption of correct measures of control and reduction of anthracnose losses in Welsh onion. © 2017, American Phytopathological Society. All rights reserved