Complete genome sequence of bovine polyomavirus type 1 from aborted cattle, isolated in Belgium in 2014

&lt;p&gt;The complete and fully annotated genome sequence of a bovine polyomavirus type 1 (BPyV/BEL/1/2014) from aborted cattle was assembled from a metagenomics data set. The 4,697-bp circular dsDNA genome contains 6 protein-coding genes. Bovine polyomavirus is unlikely to be causally related to the abortion cases. &lt;/p&gt;</p

Complete genome sequences of three African foot-and-mouth disease viruses from clinical samples isolated in 2009 and 2010

&lt;p&gt;The complete genome sequences of three foot-and-mouth disease viruses (one virus of each serotype SAT1, SAT2 and O) were directly sequenced from RNA extracted from clinical bovine samples, demonstrating the feasibility of full-genome sequencing from strong positive samples taken from symptomatic animals. &lt;/p&gt;</p

Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies

<p>Abstract</p> <p>Background</p> <p>During a wildlife screening program for avian influenza A viruses (AIV) and avian paramyxoviruses (APMV) in Belgium, we isolated two hemagglutinating agents from pools of cloacal swabs of wild mallards (<it>Anas platyrhynchos</it>) caught in a single sampling site at two different times. AIV and APMV1 were excluded using hemagglutination inhibition (HI) testing and specific real-time RT-PCR tests.</p> <p>Methods</p> <p>To refine the virological identification of APMV2-10 realized by HI subtyping tests and in lack of validated molecular tests for APMV2-10, random access amplification was used in combination with next generation sequencing for the sequence independent identification of the viruses and the determination of their genomes.</p> <p>Results</p> <p>Three different APMVs were identified. From one pooled sample, the complete genome sequence (15054 nucleotides) of an APMV4 was assembled from the random sequences. From the second pooled sample, the nearly complete genome sequence of an APMV6 (genome size of 16236 nucleotides) was determined, as well as a partial sequence for an APMV4. This APMV4 was closely related but not identical to the APMV4 isolated from the first sample. Although a cross-reactivity with other APMV subtypes did not allow formal identification, the HI subtyping revealed APMV4 and APMV6 in the respective pooled samples but failed to identify the co-infecting APMV4 in the APMV6 infected pool.</p> <p>Conclusions</p> <p>These data further contribute to the knowledge about the genetic diversity within the serotypes APMV4 and 6, and confirm the limited sensitivity of the HI subtyping test. Moreover, this study demonstrates the value of a random access nucleic acid amplification method in combination with massive parallel sequencing. Using only a moderate and economical sequencing effort, the characterization and full genome sequencing of APMVs can be obtained, including the identification of viruses in mixed infections.</p

DNase SISPA-next generation sequencing confirms Schmallenberg virus in Belgian field samples and identifies genetic variation in Europe

In 2011, a novel Orthobunyavirus was identified in cattle and sheep in Germany and the Netherlands. This virus was named Schmallenberg virus (SBV). Later, presence of the virus was confirmed using real time RT-PCR in cases of congenital malformations of bovines and ovines in several European countries, including Belgium. In the absence of specific sequencing protocols for this novel virus we confirmed its presence in RT-qPCR positive field samples using DNase SISPA-next generation sequencing (NGS), a virus discovery method based on random amplification and next generation sequencing. An in vitro transcribed RNA was used to construct a standard curve allowing the quantification of viral RNA in the field samples. Two field samples of aborted lambs containing 7.66 and 7.64 log(10) RNA copies per mu L total RNA allowed unambiguous identification of SBV. One sample yielded 192 SBV reads covering about 81% of the L segment, 56% of the M segment and 13% of the S segment. The other sample resulted in 8 reads distributed over the L and M segments. Three weak positive field samples (one from an aborted calf, two from aborted lambs) containing virus quantities equivalent to 4.27-4.89 log(10) RNA copies per mu L did not allow identification using DNase SISPA-NGS. This partial sequence information was compared to the whole genome sequence of SBV isolated from bovines in Germany, identifying several sequence differences. The applied viral discovery method allowed the confirmation of SBV in RT-qPCR positive brain samples. However, the failure to confirm SBV in weak PCR-positive samples illustrates the importance of the selection of properly targeted and fresh field samples in any virus discovery method. The partial sequences derived from the field samples showed several differences compared to the sequences from bovines in Germany, indicating sequence divergence within the epidemic

The origin of biased sequence depth in sequence-independent nucleic acid amplification and optimization for efficient massive parallel sequencing

Sequence Independent Single Primer Amplification is one of the most widely used random amplification approaches in virology for sequencing template preparation. This technique relies on oligonucleotides consisting of a 39 random part used to prime complementary DNA synthesis and a 59 defined tag sequence for subsequent amplification. Recently, this amplification method was combined with next generation sequencing to obtain viral sequences. However, these studies showed a biased distribution of the resulting sequence reads over the analyzed genomes. The aim of this study was to elucidate the mechanisms that lead to biased sequence depth when using random amplification. Avian paramyxovirus type 8 was used as a model RNA virus to investigate these mechanisms. We showed, based on in silico analysis of the sequence depth in relation to GC-content, predicted RNA secondary structure and sequence complementarity to the 39 part of the tag sequence, that the tag sequence has the main contribution to the observed bias in sequence depth. We confirmed this finding experimentally using both fragmented and non-fragmented viral RNAs as well as primers differing in random oligomer length (6 or 12 nucleotides) and in the sequence of the amplification tag. The observed oligonucleotide annealing bias can be reduced by extending the random oligomer sequence and by in silico combining sequence data from SISPA experiments using different 59 defined tag sequences. These findings contribute to the optimization of random nucleic acid amplification protocols that are currently required for downstream applications such as viral metagenomics and microarray analysis

Risk of adverse pregnancy outcomes of late- and postterm pregnancies in advanced maternal age : A national cohort study

© 2020 The Authors. Acta Obstetricia et Gynecologica Scandinavica published by John Wiley & Sons Ltd on behalf of Nordic Federation of Societies of Obstetrics and Gynecology (NFOG).Peer reviewedPublisher PD

Bluetongue Virus in Wild Deer, Belgium, 2005–2008

To investigate bluetongue virus serotype 8 infection in Belgium, we conducted a virologic and serologic survey on 2,416 free-ranging cervids during 2005–2008. Infection emerged in 2006 and spread over the study area in red deer, but not in roe deer

Perinatal death beyond 41 weeks pregnancy : An evaluation of causes and substandard care factors as identified in perinatal audit in the Netherlands

Peer reviewedPublisher PD

The most likely time and place of introduction of BTV8 into belgian ruminants

Peer reviewe