Bio-guided isolation of alpha-glucosidase inhibitory compounds from Vietnamese liverwort Marchantia polymorpha: in vitro and in silico studies.

Bio-guided isolation was applied to Vietnamese Marchantia polymorpha L. to find alpha-glucosidase inhibition. Fifteen compounds were isolated and structurally determined, including two new compounds, marchatoside (7) and marchanol (8), along with thirteen known compounds: marchantin A (1), isoriccardin C (2), riccardin C (3), marchantin K (4), lunularin (5), 3R-(3,4-dimethoxybenzyl)-5,7-dimethoxyphthalide (6), vitexilactone (9), 12-oleanene-3-one (10), 3,11-dioxoursolic acid (11), ursolic acid (12), artemetin (13), kaempferol (14), and quercetin (15). The structures of these compounds were determined through extensive spectroscopic analyses (1D and 2D NMR, HRESIMS, and ECD) and by comparisons to the existing literature. There are five types of carbon skeleton, including bibenzyl (1-5), 3-benzylphthalide (6 and 7), diterpenoid (8 and 9), triterpenoid (10-12), and flavonoid (13-15). Compounds 6-12 were reported for the first time within the genus Marchantia. Compounds 1-12 were evaluated for their alpha-glucosidase inhibition. Among them, 1-5 and 10-12 displayed potent inhibition, with IC50 values ranging from 28.9 to 130.6 µM, compared to the positive control acarbose 330.9 µM. A kinetic study and molecular docking were also performed to understand the mechanism.

Structural and functional analysis of natural capsid variants suggests sialic acid-independent entry of BK polyomavirus.

BK polyomavirus (BKPyV) is an opportunistic pathogen that uses the b-series gangliosides GD1b and GT1b as entry receptors. Here, we characterize the impact of naturally occurring VP1 mutations on ganglioside binding, VP1 protein structure, and virus tropism. Infectious entry of single mutants E73Q and E73A and the triple mutant A72V-E73Q-E82Q (VQQ) remains sialic acid dependent, and all three variants acquire binding to a-series gangliosides, including GD1a. However, the E73A and VQQ variants lose the ability to infect ganglioside-complemented cells, and this correlates with a clear shift of the BC2 loop in the crystal structures of E73A and VQQ. On the other hand, the K69N mutation in the K69N-E82Q variant leads to a steric clash that precludes sialic acid binding. Nevertheless, this mutant retains significant infectivity in 293TT cells, which is not dependent on heparan sulfate proteoglycans, implying that an unknown sialic acid-independent entry receptor for BKPyV exists.

A cluster of broadly neutralizing IgG against BK polyomavirus in a repertoire dominated by IgM.

The BK polyomavirus (BKPyV) is an opportunistic pathogen, which is only pathogenic in immunosuppressed individuals, such as kidney transplant recipients, in whom BKPyV can cause significant morbidity. To identify broadly neutralizing antibodies against this virus, we used fluorescence-labeled BKPyV virus-like particles to sort BKPyV-specific B cells from the PBMC of KTx recipients, then single-cell RNAseq to obtain paired heavy- and light-chain antibody sequences from 2,106 sorted B cells. The BKPyV-specific repertoire was highly diverse in terms of both V-gene usage and clonotype diversity and included most of the IgM B cells, including many with extensive somatic hypermutation. In two patients where sufficient data were available, IgM B cells in the BKPyV-specific dataset had significant differences in V-gene usage compared with IgG B cells from the same patient. CDR3 sequence-based clustering allowed us to identify and characterize three broadly neutralizing "41F17-like" clonotypes that were predominantly IgG, suggesting that some specific BKPyV capsid epitopes are preferentially targeted by IgG.

Quantification of APOBEC3 Mutation Rates Affecting the VP1 Gene of BK Polyomavirus In Vivo.

Mutations in the BK polyomavirus (BKPyV) capsid accumulate in kidney transplant (KTx) recipients with persistent virus replication. They are associated with neutralization escape and appear to arise as a result of cytosine deamination by host cell APOBEC3A/B enzymes. To study the mutagenic processes occurring in patients, we amplified the typing region of the VP1 gene, sequenced the amplicons to a depth of 5000-10,000×, and identified rare mutations, which were fitted to COSMIC mutational signatures. Background mutations were identified in amplicons from plasmids carrying the BKPyV genome and compared to mutations observed in 148 samples from 23 KTx recipients in France and in Vietnam. Three mutational signatures were consistently observed in urine, serum, and kidney biopsy samples, two of which, SBS2 and SBS13, corresponded to APOBEC3A/B activity. In addition, a third signature with no known etiology, SBS89, was detected both in patient samples, and in cells infected in vitro with BKPyV. Quantitatively, APOBEC3A/B mutation rates in urine samples were strongly correlated with urine viral load, and also appeared to vary between individuals. These results confirm that APOBEC3A/B is a major, but not the only, source of BKPyV genome mutations in patients.

Identification of three novel mutations in PCNT in vietnamese patients with microcephalic osteodysplastic primordial dwarfism type II.

BACKGROUND: Primordial dwarfism (PD) is a group of genetically heterogeneous disorders related to developmental disabilities occurring in the uterus and prolongs during all stages of life, resulting in short stature, facial deformities and abnormal brain. OBJECTIVE: To determine the exact cause of the disease in two Vietnamese patients priory diagnosed with PD by severe pre-and postnatal growth retardation with marked microcephaly and some bone abnormalities. METHODS: Whole-exome sequencing was performed for the two patients and mutations in genes related to PD were screened. Sanger sequencing was applied to examine the mutations in the patients of their families. RESULTS: Three novel mutations in the PCNT gene which have not been reported previously were identified in the two patients. Of which, two frameshift mutations (p.Thr479Profs*6 and p.Glu2742Alafs*8) were detected in patient I and one stop-gained mutation (p.Gln1907*) was detected in the patient II. These mutations may result in a truncated PCNT protein, leading to an inactivated PACT domain corresponding to residue His3138-Trp3216 of PCNT protein. Therefore, the three mutations may cause a deficiency of protein functional activity and result in the phenotypes of primordial dwarfism in the two patients. CONCLUSIONS: Clinical presentations in combination with genetic analyses supported an accurate diagnosis of the two patients with microcephalic osteodysplastic primordial dwarfism type II (MOPD II). In addition, these results have important implications for prenatal genetic screening and genetic counseling for the families.

Persistent BK Polyomavirus Viruria is Associated with Accumulation of VP1 Mutations and Neutralization Escape.

To investigate the relationship between neutralization escape and persistent high-level BK polyomavirus replication after kidney transplant (KTx), VP1 sequences were determined by Sanger and next-generation sequencing in longitudinal samples from KTx recipients with persistent high-level viruria (non-controllers) compared to patients who suppressed viruria (controllers). The infectivity and neutralization resistance of representative VP1 mutants were investigated using pseudotype viruses. In all patients, the virus population was initially dominated by wild-type VP1 sequences, then non-synonymous VP1 mutations accumulated over time in non-controllers. BC-loop mutations resulted in reduced infectivity in 293TT cells and conferred neutralization escape from cognate serum in five out of six non-controller patients studied. When taken as a group, non-controller sera were not more susceptible to neutralization escape than controller sera, so serological profiling cannot predict subsequent control of virus replication. However, at an individual level, in three non-controller patients the VP1 variants that emerged exploited specific "holes" in the patient's humoral response. Persistent high-level BK polyomavirus replication in KTx recipients is therefore associated with the accumulation of VP1 mutations that can confer resistance to neutralization, implying that future BKPyV therapies involving IVIG or monoclonal antibodies may be more effective when used as preventive or pre-emptive, rather than curative, strategies.

Environnement réconfortant et respect de l'intimité. L'organisation du cabinet médical pour la consultation gynécologique.

INTRODUCTION: Social science concepts (intimate distance, personal space) suggest that the gynecological examination environment (GEE) might influence women’s feelings during the exam.Purpose of research: We explore this hypothesis by assessing women’s preferences for the GEE. RESULTS: An opinion poll was conducted, without randomization to explore women’s point of view. In 2017, questionnaires were referred to women by 14 general practitioners in the Lille region. Among 173 answers, 73% granted importance to the medical setting, especially to a comfortable ambiance and an isolated place during an exam (to respect women’s privacy). Women expected at least: a separation of consultation/examination (77%), disposal of sanitary towels (80%), a place to leave clothes and underwear (74%), a changing area (56%). Fifty-five percent judged it unnecessary to cover the lower body. Concerning the examination table: comfort was satisfactory (93%), “calm and peaceful colors” were appreciated, but clamps should be improved. CONCLUSIONS: The GEE is characterized by a balance of natural comfort and a sequentially delimited spatial configuration. The study revealed cultural and subjective dimensions of privacy. Physicians need to apply psychosocial competences to perform a person-centered gynecological exam.

Socioeconomic Inequalities in the HIV Testing during Antenatal Care in Vietnamese Women.

Although HIV (human immunodeficiency virus) testing for all women has been promoted by Vietnam's Ministry of Health since 2000, test acceptance rates in this country were reported to be less than 30% in the community. This country has been facing the barriers to approach the national services towards transmission prevention from mother to child including HIV testing during antenatal care (ANC) towards mothers. Here, we aim to assess the socioeconomic inequalities in HIV testing during ANC among Vietnamese women. This study used available data from the Vietnam Multiple Indicator Cluster Survey 2014. Overall, the prevalence of HIV testing during antenatal care was 30% and the concentrate index (CCI) was 0.1926. There was significant inequality between women classified as poor and rich, and when stratified by social characteristics, inequality was found in women aged 15-49 years (CCI: 0.4), living in rural areas (CCI: 0.3), belonging to ethnic minorities (CCI: 0.5) and having primary or less education (CCI: 0.4). In the multivariate logistic regression analysis, ethnicity and socioeconomic status were significant factors associated with HIV testing during ANC. We found the prevalence of HIV testing during ANC was low, and its inequalities were associated with age, living area, ethnicity, education, and economic status.

Validation of ambiguous MLPA results by targeted next-generation sequencing discloses a nonsense mutation in the DMD gene.

BACKGROUND: Duchenne muscular dystrophy (DMD) is the most common inherited muscular disease and caused by mutations in the DMD gene on the X-chromosome. Multiplex ligation-dependent probe amplification (MLPA) is recognized as a convenient and reliable technique to detect exon deletion/duplication mutations in the DMD gene. Here, we applied targeted semi-conductor next-generation sequencing to clarify the cause of ambiguous MLPA results. METHODS: Targeted semi-conductor next-generation sequencing was carried out using the Inherited Disease Panel (IDP) on the Ion Torrent Personal Genome Machine (PGM). RESULTS: MLPA analysis disclosed unclassifiable relative peak ratio of exon 18 in a DMD boy. His female cousin was indicated to have exon 18 deletion in one allele. To validate these incompatible results, targeted next-generation sequencing was conducted. A nucleotide change, C.2227 C>T creating a premature stop codon, was in exon 18. Concomitantly, both C and T nucleotides were identified in his cousin's genome. Ambiguous values of the relative peak ratio in MLPA were considered due to the one nucleotide mismatch between the genomic sequence and the probe used in MLPA. CONCLUSION: Analysis using IDP on PGM disclosed a nonsense mutation in the DMD gene as a cause of ambiguous results of MLPA.