High concordance in preimplantation genetic testing for aneuploidy between automatic identification via Ion S5 and manual identification via Miseq

The Ion S5 (Thermo Fisher Scientific) and Miseq (Illumina) NGS systems are both widely used in the clinical laboratories conducting PGT-A. Each system employs discrepant library preparation steps, sequencing principles, and data processing algorithms. The automatic interpretation via Ion Reporter software (Thermo Fisher Scientific) and the manual interpretation via BlueFuse Multi software (Illumina) for chromosomal copy number variation (CNV) represent very different reporting approaches. Thus, it is intriguing to compare their ability of ploidy detection as PGT-A/NGS system. In the present study, four aneuploid cell lines were individually mixed with a diploid cell line at different aneuploid ratios of 0% (0:5), 10% (1:9), 20% (1:4), 40% (2:3), 50% (3:3), 60% (3:2), 80% (4:1) and 100% (5:0) to assess the sensitivity and specificity for whole chromosomal and segmental aneuploidy detection. The clinical biopsies of 107 blastocysts from 46 IVF/PGT-A cycles recruited between December 2019 and February 2020 were used to calculate the concordance. Initially, the pre-amplified products were divided into two aliquots for different library preparation procedures of each system. Applying the same calling criteria, automatic identification was achieved through the Ion Reporter, while well-trained technicians manually identified each sample through the BlueFuse Multi. The results displayed that both systems reliably distinguished chromosomal CNV of the mixtures with at least 10% aneuploidy from karyotypically normal samples ([Ion S5] whole-chromosomal duplication: 2.14 vs. 2.05, p value = 0.009, segmental deletion: 1.88 vs. 2.05, p value = 0.003; [Miseq] whole-chromosomal duplication: 2.12 vs. 2.03, p value = 0.047, segmental deletion: 1.82 vs. 2.03, p value = 0.002). The sensitivity and specificity were comparable between the Ion S5 and Miseq ([sensitivity] 93% vs. 90%, p = 0.78; [specificity] 100% vs. 100%, p value = 1.0). In the 107 clinical biopsies, three displayed chaotic patterns (2.8%), which could not be interpreted for the ploidy. The ploidy concordance was 99.04% (103/104) per embryo and 99.47% (2265/2277) per chromosome pair. Since their ability of detection were proven to be similar, the automatic identification in Ion S5 system presents comparatively faster and more standardized performance.

Transcriptional and Post-Transcriptional Regulations of Amyloid-β Precursor Protein (APP) mRNA

Alzheimer’s disease (AD) is an age-associated neurodegenerative disorder characterized by progressive impairment of memory, thinking, behavior, and dementia. Based on ample evidence showing neurotoxicity of amyloid-β (Aβ) aggregates in AD, proteolytically derived from amyloid precursor protein (APP), it has been assumed that misfolding of Aβ plays a crucial role in the AD pathogenesis. Additionally, extra copies of the APP gene caused by chromosomal duplication in patients with Down syndrome can promote AD pathogenesis, indicating the pathological involvement of the APP gene dose in AD. Furthermore, increased APP expression due to locus duplication and promoter mutation of APP has been found in familial AD. Given this background, we aimed to summarize the mechanism underlying the upregulation of APP expression levels from a cutting-edge perspective. We first reviewed the literature relevant to this issue, specifically focusing on the transcriptional regulation of APP by transcription factors that bind to the promoter/enhancer regions. APP expression is also regulated by growth factors, cytokines, and hormone, such as androgen. We further evaluated the possible involvement of post-transcriptional regulators of APP in AD pathogenesis, such as RNA splicing factors. Indeed, alternative splicing isoforms of APP are proposed to be involved in the increased production of Aβ. Moreover, non-coding RNAs, including microRNAs, post-transcriptionally regulate the APP expression. Collectively, elucidation of the novel mechanisms underlying the upregulation of APP would lead to the development of clinical diagnosis and treatment of AD.

Arresting chromosome replication upon energy starvation in Escherichia coli

Most organisms possess several cell cycle checkpoints to preserve genome stability in periods of stress. Upon starvation, the absence of chromosomal duplication in the bacterium Escherichia coli is ensured by holding off commencement of replication. During normal growth, accumulation of the initiator protein DnaA along with cell cycle changes in its activity, ensure that DNA replication starts only once per cell cycle. Upon nutrient starvation, the prevailing model is that an arrest in DnaA protein synthesis is responsible for the absence of initiation. Recent indications now suggest that DnaA degradation may also play a role. Here we comment on the implications of this potential new layer of regulation.

Duplication of chromosome 16p13.11-p12.3 with different expressions in the same family

The knowledge about genetic involvement in neurodevelopmental disorders, and especially in autism, is currently rising. To date, more than 100 gene mutations related to autistic syndromes have been described. Some disorders that affect multiple family members are caused by gene mutations, which can be inherited. Recently, array comparative genomic hybridization (aCGH) has identified sub microscopic deletions and duplications as a common cause of mental retardation and autism. In this article we report the occurrence of the same genetic finding (chromosome 16p13.11-p12.3 duplication) in a family with four small children, where two older siblings manifested a global neurodevelopmental delay associated with an autism spectrum disorder (ASD), but younger twin brothers with the same mutation, have typical development. Genetic analysis showed that the chromosomal duplication was inherited from the father, in which phenotype and functioning are quite typical. As is known, the duplication can pass from parents to children. The 16p13.11 micro duplication has been implicated in several neurodevelopmental and behavioral disorders and is characterized by variable expressivity and incomplete penetrance.

High Concordance Between Automatic Identification in Ion S5 and Manual Identification in Miseq During Next-Generation Sequencing for Preimplantation Genetic Testing of Aneuploidy

The Ion S5 (Thermo Fisher Scientific) and Miseq (Illumina) NGS systems are both widely used in the clinical laboratories conducting PGT-A. Each system employs discrepant library preparation steps, sequencing principles, and data processing algorithms. The automatic interpretation via Ion Reporter software (Thermo Fisher Scientific) and the manual interpretation via BlueFuse Multi software (Illumina) for chromosomal copy number variation (CNV) represent very different reporting approaches. Thus, it is intriguing to compare their ability of ploidy detection as PGT-A/NGS system. In the present study, four aneuploid cell lines were individually mixed with a diploid cell line at different aneuploid ratios of 0% (0:5), 10% (1:9), 20% (1:4), 40% (2:3), 50% (3:3), 60% (3:2), 80% (4:1) and 100% (5:0) to assess the sensitivity and specificity for whole chromosomal and segmental aneuploidy detection. The clinical biopsies of 107 blastocysts from 46 IVF/PGT-A cycles recruited between December 2019 and February 2020 were used to calculate the concordance. Initially, the pre-amplified products were divided into two aliquots for different library preparation procedures of each system. Applying with the same calling criteria, automatic identification was achieved through the Ion Reporter, while well-trained technicians manually identified each sample through the BlueFuse Multi. The results displayed that both systems reliably distinguished chromosomal CNV of the mixtures with at least 10% aneuploidy from karyotypically normal samples ([Ion S5] whole-chromosomal duplication: 2.14 vs. 2.05, p-value=0.009, segmental deletion: 1.88 vs. 2.05, p-value=0.003; [Miseq] whole-chromosomal duplication: 2.12 vs. 2.03, p-value=0.047, segmental deletion: 1.82 vs. 2.03, p-value=0.002). The sensitivity and specificity were comparable between the Ion S5 and Miseq ([sensitivity] 93% vs. 90%, p=0.78; [specificity] 100% vs. 100%, p-value=1.0). In the 107 clinical biopsies, three displayed chaotic patterns (2.8%), which could not be interpreted for the ploidy. The ploidy concordance was 99.04% (103/104) per embryo and 99.47% (2265/2277) per chromosome pair. Since their ability of detection were proven to be similar, the automatic identification in Ion S5 system presents comparatively faster and more standardized performance.

Pitt-Hopkins Syndrome: A Review of Current Literature, Clinical Approach, and 23-Patient Case Series

Pitt-Hopkins syndrome (PTHS) is a rare, genetic disorder caused by a molecular variant of TCF4 which is involved in embryologic neuronal differentiation. PTHS is characterized by syndromic facies, psychomotor delay, and intellectual disability. Other associated features include early-onset myopia, seizures, constipation, and hyperventilation-apneic spells. Many also meet criteria for autism spectrum disorder. Here the authors present a series of 23 PTHS patients with molecularly confirmed TCF4 variants and describe 3 unique individuals. The first carries a small deletion but does not exhibit the typical facial features nor the typical pattern of developmental delay. The second exhibits typical facial features, but has attained more advanced motor and verbal skills than other reported cases to date. The third displays typical features of PTHS, however inherited a large chromosomal duplication involving TCF4 from his unaffected father with somatic mosaicism. To the authors’ knowledge, this is the first chromosomal duplication case reported to date.