Swine viral detection by adapted Next-Generation Sequencing (NGS) for RNA and DNA species reveals first detection of porcine circovirus type 3 (PCV3) in Chile

AbstractThe outbreak and propagation of COVID-19 have posed a significant challenge to our society, highlighting the relevance of epidemiological monitoring of animal pathogens closely related to humans due to potential zoonosis. To find previously undetected viruses inside Chilean swine farms, we collected non-invasive samples from animals suspected to undergo any viral disease. After screening by Next Generation Sequencing (NGS) adapted to identify viral species with RNA and DNA genomes simultaneously, different viral species were successfully detected. Among viruses with an RNA genome, Porcine Rotavirus A (RVA), Porcine Astrovirus type 5 (PAstV-5) and Porcine Feces-Associated IASV-like virus (PfaIV) were identified; whereas among viruses with DNA genome, Porcine Parvovirus 1 (PPV1), Porcine Circovirus type 1 (PCV1) and Porcine Circovirus type 3 (PCV3) were found. PCV3, a recently described pathogenic virus, was detected for the first time in Chile in different samples from mummified tissue of stillbirths. The whole genome sequence of PCV3 was completed using conventional PCR and Sanger sequencing and compared with previously reported genomes. To the best of our knowledge, this is the first time that an adapted NGS system is used for the screening of DNA and RNA viral species simultaneously inside the Chilean swine industry. This adapted NGS system might be useful for the detection of swine viral emerging diseases worldwide.

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Next-Generation Sequencing Combined With Conventional Sanger Sequencing Reveals High Molecular Diversity in Actinidia Virus 1 Populations From Kiwifruit Grown in China

Frontiers in Microbiology ◽

10.3389/fmicb.2020.602039 ◽

2020 ◽

Vol 11 ◽

Author(s):

Shaohua Wen ◽

Guoping Wang ◽

Zuokun Yang ◽

Yanxiang Wang ◽

Min Rao ◽

...

Keyword(s):

Next Generation Sequencing ◽

Sanger Sequencing ◽

Molecular Diversity ◽

Genome Structure ◽

Viral Disease ◽

Open Reading Frames ◽

Next Generation ◽

Next Generation Sequencing Ngs ◽

First Time ◽

Generation Sequencing

Kiwifruit (Actinidia spp.) is native to China. Viral disease–like symptoms are common on kiwifruit plants. In this study, six libraries prepared from total RNA of leaf samples from 69 kiwifruit plants were subjected to next-generation sequencing (NGS). Actinidia virus 1 (AcV-1), a tentative species in the family Closteroviridae, was discovered in the six libraries. Two full-length and two near-full genome sequences of AcV-1 variants were determined by Sanger sequencing. The genome structure of these Chinese AcV-1 variants was identical to that of isolate K75 and consisted of 12 open reading frames (ORFs). Analyses of these sequences together with the NGS-derived contig sequences revealed high molecular diversity in AcV-1 populations, with the highest sequence variation occurring at ORF1a, ORF2, and ORF3, and the available variants clustered into three phylogenetic clades. For the first time, our study revealed different domain compositions in the viral ORF1a and molecular recombination events among AcV-1 variants. Specific reverse transcriptase–polymerase chain reaction assays disclosed the presence of AcV-1 in plants of four kiwifruit species and unknown Actinidia spp. in seven provinces and one city.

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Identification of tick-borne pathogens by metagenomic next-generation sequencing in Dermacentor nuttalli and Ixodes persulcatus in Inner Mongolia, China

Parasites & Vectors ◽

10.1186/s13071-021-04740-3 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Jun Jiao ◽

Zhiyu Lu ◽

Yonghui Yu ◽

Yangxuan Ou ◽

Mengjiao Fu ◽

...

Keyword(s):

Next Generation Sequencing ◽

Inner Mongolia ◽

Human Infection ◽

Ixodes Persulcatus ◽

Next Generation ◽

Nested Polymerase Chain Reaction ◽

Hard Ticks ◽

Animal Pathogens ◽

First Time ◽

Generation Sequencing

Abstract Background Hard ticks act as arthropod vectors in the transmission of human and animal pathogens and are widely distributed in northern China. The aim of this study is to screen the important tick-borne pathogens (TBPs) carried by hard ticks in Inner Mongolia using metagenomic next-generation sequencing (mNGS) and to estimate the risk of human infection imposed by tick bites. Methods The adult Dermacentor nuttalli (n = 203) and Ixodes persulcatus (n = 36) ticks feeding on cattle were collected. The pooled DNA samples prepared from these ticks were sequenced as the templates for mNGS to survey the presence of TBPs at the genus level. Individual tick DNA samples were detected by genus--specific or group-specific nested polymerase chain reaction (PCR) of these TBPs and combined with DNA sequencing assay to confirm the results of mNGS. Results R. raoultii (45.32%, 92/203), Candidatus R. tarasevichiae (5.42%, 11/203), Anaplasma sp. Mongolia (26.60%, 54/203), Coxiella-like endosymbiont (CLE) (53.69%, 109/203), and Babesia venatorum (7.88%, 16/203) were detected in D. nuttalli, while R. raoultii (30.56%, 11/36), Anaplasma sp. Mongolia (27.80%, 10/36), and CLE (27.80%, 10/36) were detected in I. persulcatus. The double- and triple-pathogen/endosymbiont co-infections were detected in 40.39% of D. nuttalli and 13.89% of I. persulcatus, respectively. The dual co-infection with R. raoultii and CLE (14.29%, 29/203) and triple co-infection with R. raoultii, Anaplasma sp. Mongolia, and CLE (13.79%, 28/203) were most frequent in D. nuttalli. Conclusions This study provides insight into the microbial diversity of D. nuttalli and I. persulcatus in Inner Mongolia, China, reporting for the first time that Candidatus R. tarasevichiae had been found in D. nuttalli in China, and for the first time in the world that Anaplasma sp. Mongolia has been detected in I. persulcatus. This study proves that various vertically transmitted pathogens co-inhabit D. nuttalli and I. persulcatus, and indicates that cattle in Inner Mongolia are exposed to several TBPs. Graphic Abstract

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Mitochondrial m.13513G>A Point Mutation in ND5 in a 16-Year-Old Man with Leber Hereditary Optic Neuropathy Detected by Next-Generation Sequencing

Journal of Pediatric Genetics ◽

10.1055/s-0039-1691812 ◽

2019 ◽

Vol 08 (04) ◽

pp. 231-234

Author(s):

Daniel Vázquez-Justes ◽

Lidia Carreño-Gago ◽

Elena García-Arumi ◽

Alicia Traveset ◽

Julio Montoya ◽

...

Keyword(s):

Next Generation Sequencing ◽

Optic Neuropathy ◽

Nadh Dehydrogenase ◽

Deoxyribonucleic Acid ◽

Next Generation ◽

Leber Hereditary Optic Neuropathy ◽

Next Generation Sequencing Ngs ◽

First Time ◽

Generation Sequencing ◽

Hereditary Optic Neuropathy

AbstractThis article reports a Leber hereditary optic neuropathy (LHON) case associated for the first time with mitochondrial m.13513G>A mutation. We present a 16-year-old man who complained of subacute, painless, visual loss. Ocular examination showed optic nerve atrophy, papillary pseudoedema, and optic disc pallor. Extraocular manifestations included hypertrophic myocardiopathy and myopathy. Initial genetic analysis excluded the three most common LHON mutations. Sanger sequencing of the whole mitochondrial deoxyribonucleic acid showed no mutation. Next-generation sequencing (NGS) revealed m.13513G>A mutation in the NADH dehydrogenase (ND5) subunit gene (MT-ND5). The m.13513G>A mutation has never been associated with LHON phenotype without Leigh/mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes features. NGS techniques should be considered when this diagnosis is strongly suspected.

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Yield of Rare Variants Detected by Targeted Next-Generation Sequencing in a Cohort of Romanian Index Patients with Hypertrophic Cardiomyopathy

Diagnostics ◽

10.3390/diagnostics10121061 ◽

2020 ◽

Vol 10 (12) ◽

pp. 1061

Author(s):

Miruna Mihaela Micheu ◽

Nicoleta-Monica Popa-Fotea ◽

Nicoleta Oprescu ◽

Stefan Bogdan ◽

Monica Dan ◽

...

Keyword(s):

Next Generation Sequencing ◽

Hypertrophic Cardiomyopathy ◽

Rare Variants ◽

Causal Variant ◽

Next Generation ◽

Targeted Next Generation Sequencing ◽

Novel Variants ◽

Next Generation Sequencing Ngs ◽

First Time ◽

Generation Sequencing

Background: The aim of this study was to explore the rare variants in a cohort of Romanian index cases with hypertrophic cardiomyopathy (HCM). Methods: Forty-five unrelated probands with HCM were screened by targeted next generation sequencing (NGS) of 47 core and emerging genes connected with HCM. Results: We identified 95 variants with allele frequency < 0.1% in population databases. MYBPC3 and TTN had the largest number of rare variants (17 variants each). A definite genetic etiology was found in 6 probands (13.3%), while inconclusive results due to either known or novel variants were established in 31 cases (68.9%). All disease-causing variants were detected in sarcomeric genes (MYBPC3 and MYH7 with two cases each, and one case in TNNI3 and TPM1 respectively). Multiple variants were detected in 27 subjects (60%), but no proband carried more than one causal variant. Of note, almost half of the rare variants were novel. Conclusions: Herein we reported for the first time the rare variants identified in core and putative genes associated with HCM in a cohort of Romanian unrelated adult patients. The clinical significance of most detected variants is yet to be established, additional studies based on segregation analysis being required for definite classification.

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