[Explorations about the correlation between biological changes of meninges in periodontitis mice and cognitive impairment via single-cell RNA sequencing].

Objective: To clarify the potential correlation between biological changes of meninges in periodontitis mice and cognitive impairment by analyzing the biological changes of meninges in periodontitis mice using single-cell RNA sequencing. Methods: Thirty C57BL/6 mice were divided into two groups by using random number table method (15 mice in each group). Mice in the control group were locally administered 2% carboxyl methyl cellulose (CMC) without Porphyromonas gingivalis (Pg) on both buccal sides. A mixture of Pg W83 and 2% CMC was applied on both buccal sides in the experimental group mice three times a week, lasting for 16 weeks in total. The absorption of alveolar bone, locomotor activity and cognitive function, the activation of microglia and astrocytes in the cortex were observed and assessed. The mRNA expression levels of Occludin in meninges and brain were detected in two groups. Single-cell RNA sequencing data of meninges were processed by uniform manifold approximation and projection (UMAP). Differential genes expressions of endothelial cells were processed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. In addition, real-time fluorescence quantitative PCR (RT-qPCR) was used to verify the expressions of transcription activating factor 3 (Atf3) and apolpoprotein L domain-containing 1 (Apold 1). Results: Methylene blue staining found the distances of buccal and palatal cement-enamel junction-alveolar bone crest in experimental mice [(185.60±17.60), (206.90±13.37) µm] increased significantly compared with the control group [(135.33±9.57), (163.05±14.98) µm] (t=5.02, P=0.002; t=4.37, P=0.005). Open field experiment showed the total distance and average speed of mice in the experimental group [(971.88±164.57) cm, (3.25±0.55) cm/s] were not statistically significant compared with the control group [(914.24±278.81) cm, (3.05±0.93) cm/s] (t=0.65, P=0.525; t=0.65, P=0.520). The recognition index of the experimental group [(48.02±16.92) %] was lower than the control group [(66.27±17.90) %] (t=2.40, P=0.027) by novel object recognition tests. Compared with the control group [(63.56±11.88) %], the alternation of experimental group [(50.99±14.17) %] was significantly decreased in Y maze tests (t=2.33, P=0.030). Immunohistochemistry results showed microglia and astrocytes were activated in the cortex of experimental mice. Compared with the control group (1.02±0.25, 1.04±0.31), the relative mRNA expressions of Occludin decreased significantly in the meninges and brain of periodontitis mice, respectively (0.61±0.10, 0.64±0.20) (t=3.47, P=0.010; t=2.66, P=0.024). By single-cell RNA sequencing, meninges cells were divided into 11 types, such as endothelial cells, fibroblasts, immune cells and so on. Endothelial cells were the main cell types in meninges [the control group: 26.47% (1 589/6 004), the experimental group: 26.26% (807/3 073)]. Compared with the control group [5.56% (334/6 004)], the percentage of granulocytes increased in the periodontitis mice [11.65% (358/3 073)]. Using clustering analysis to further focus on endothelial cells, GO enrichment analysis revealed differential genes were mainly related to angiogenesis, cell adhesion, apoptosis and so on. KEGG enrichment analysis revealed that differential genes were related to signaling pathways of interleukin-17, relaxin and so on. The relative mRNA expressions of Atf3 and Apold1 in meninges of periodontitis mice (0.42±0.24, 0.54±0.27) were significantly lower than the control group (1.03±0.26, 1.02±0.23) (t=3.88, P=0.005; t=3.02, P=0.017). Conclusions: The mice chronically infected with Pg W83 occurred memory impairment, neuroinflammation and changes of barrier function. In the meninges of periodontitis mice, there were infiltration of immune cells and down-regulation expressions of Atf3 and Apold1 by single-cell RNA sequencing. Meningeal immunity and changes of barrier function may play an important role in the cognitive impairment caused by periodontitis.

Combined approaches to reveal genes associated with litter size in Yunshang black goats.

Intensive artificial selection has been imposed in Yunshang black goats, the first black specialist mutton goat breed in China, with a breeding object of improving reproductive performance, which has contributed to reshaping of the genome including the characterization of SNP, ROH and haplotype. However, variation in reproductive ability exists in the present population. A WGS was implemented in two subpopulations (polytocous group, PG, and monotocous group, MG) with evident differences of litter size. Following the mapping to reference genome, and SNP calling and pruning, three approaches - GWAS, ROH analysis and detection of signatures of selection - were employed to unveil candidate genes responsible for litter size. Consequently, 12 candidate genes containing OSBPL8 with the minimum P-value were uncovered by GWAS. Differences were observed in the pattern of ROH between two subpopulations that shared similar low inbreeding coefficients. Two ROH hotspots and 12 corresponding genes emerged from ROH pool association analysis. Based on the nSL statistic, 15 and 61 promising genes were disclosed under selection for MG and PG respectively. Of them, some promising genes participate in ovarian function (PPP2R5C, CDC25A, ESR1, RPS26 and SERPINBs), seasonal reproduction (DIO3, BTG1 and CRYM) and metabolism (OSBPL8, SLC39A5 and SERPINBs). Our study pinpointed some novel promising genes influencing litter size, provided a comprehensive insight into genetic makeup of litter size and might facilitate selective breeding in goats.

Isolation and sequence analysis of the complete VP2 gene of canine parvovirus from Chinese domestic pets and determination of the pathogenesis of these circulating strains in beagles.

Canine parvovirus (CPV) causes acute gastroenteritis in domestic dogs, cats, and several wild carnivore species. In this study, the full-length VP2 gene of 36 CPV isolates from dogs and cats infected between 2016 and 2017 in Beijing was sequenced and analyzed. The results showed that, in dogs, the new CPV-2a strain was the predominant variant (n = 18; 50%), followed by the new CPV-2b (n = 6; 16.7%) and CPV-2c (n = 3; 8.3%) strains, whereas, among cats, the predominant strain was still CPV-2 (n = 9; 25%). One new CPV-2a strain, 20170320-BJ-11, and two CPV-2c strains, 20160810-BJ-81 and 20170322-BJ-26, were isolated and used to perform experimental infections. Multiple organs of beagles that died tested PCR positive for CPV, and characteristic histopathological lesions were observed in organs, including the liver, spleen, lungs, kidneys, small intestines, and lymph nodes. Experimental infections showed that the isolates from the epidemic caused high morbidity in beagles, indicating their virulence in animals and suggesting the need to further monitor evolution of CPV in China.

Molecular identification and biological characteristic analysis of an Apiotrichum mycotoxinivorans (formerly Trichosporon mycotoxinivorans) strain isolated from sputum specimens of a pediatric patient with pneumonia.

Apiotrichum mycotoxinivorans (formerly Trichosporon mycotoxinivorans) has long been used to degrade fungal toxins in livestock feed. However, clinic reports about this type of fungus are rare. In this study, we report the morphology, biochemistry, and molecular characteristics of an A. mycotoxinivorans strain isolated from a pediatric patient with congenital ventricular septal defect and pneumonia. A female patient, 26 months old, presented with congenital ventricular septal defect. Pulmonary infection symptoms were observed after the patient received cardiac repair surgery. Sputum bacterial and fungal cultures were positive for Elizabethkingia anophelis and a fungus, which was not readily identifiable using biochemical identification, or MALDI-TOF MS analysis. The strain was finally identified as A. mycotoxinivorans using amplification and sequencing of the D1/D2 region of 26S rDNA, ITS, and IGS1. Antifungal susceptibility test results suggested that fluconazole or voriconazole may be an appropriate choice for antifungal therapy. A biodegradability of ochratoxin A was considered as a characteristic of the fungal strain. Our results support the existing evidence that A. mycotoxinivorans is an opportunistic pathogen for human beings. Nucleic acid analysis allows for the accurate identification of the species in instances where conventional identification methods such as biochemical testing and MALDI-TOF MS may be unsuccessful.

[Gender difference in clinical manifestations and comorbidities in the patients with spondyloarthritis].

Objectives: The aim of the study was to determine whether there was gender difference in clinical manifestations and comorbidities in the patients with Spondyloarthritis (SpA) in China. Methods: 346 patients fulfilling ASAS criteria for SpA were recruited from the Third Affiliated Hospital of Sun Yat-sen University, including 280 males and 66 females. A comparison was conducted in terms of age at onset, disease course, family history, HLA-B27 positivity, clinical manifestations, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP), the bath ankylosing spondylitis disease activity index (BASDAI) and AS disease activity score (ASDAS), and comorbidities between male and female patients. Results: Compared with female patients, male patients were younger at disease onset (22±7 vs 27±9, P<0.001),had higher rates of morning stiffness (74.3%), and higher scores of CRP and ASDAS-CRP (P<0.010, P=0.014). However, no significant gender difference was observed in other clinical parameters like clinical manifestations, family history, HLA-B27 positivity, BASDAI, and BASFI and treatment. Male SpA patients had a higher prevalence of Hepatitis B virus (HBV) infection (26.2%) than that of female patients (8.3%), and a higher prevalence of osteoporosis (30.5% vs 14.3%,P<0.01), especially with a lower lumbar T score. Logistic regression analysis reviewed that limited weight (OR=0.94, P<0.001), high ASDAS-CRP (OR=1.58, P=0.006) and male (OR=8.02, P=0.004) are more inclined to have osteoporosis. Conclusion: Compared with female patients, male patients were younger at disease onset and higher scores of CRP and ASDAS-CRP. No significant gender difference was observed in clinical manifestations, family history, HLA-B27 positivity, BASDAI, and BASFI and treatment. Male SpA patients had a higher prevalence of HBV infection and osteoporosis than female patients. Comorbidities should be paid more attention in the patients with SpA.

Saline conditions alter morpho-physiological intensification in purslane (Portulaca oleracea l.).

In this study, primary investigations of selected cultivar of purslane named as Tall Green under articular salinity stress were evaluated to understand the basic concept of different mechanisms of physiological attributes which will play an important role for molecular and proteomic level research. The evaluation of morphological and physiological attributes under 0 mM (without salt addition) 100 mM and 200 mM salt stress changed dramatically. The results showed high salt stress at 200 mM significantly decreasing the morphological attributes and performance of leaves, stems, and roots. At moderate salt stress levels, 100 mM, the ratio of Fv/Fm slightly increased compared to high stress. In addition, salt stress significantly decreased the total chlorophyll content (chl a+b) at 200 mM. The relative water content percentage was high at 0 mM. Moreover, the electrolyte leakage (EL) significantly increased with increasing salinity stress compared to control 0 mM.

Biological changes of Enterococcus faecalis in the viable but nonculturable state.

Enterococcus faecalis may enter a viable but nonculturable (VBNC) state under adverse conditions. E. faecalis, the major bacterial species present in failed root canal treatments, is thought to survive after endodontic treatment by entering a VBNC state. In this study, we characterized the VBNC state of E. faecalis. We designed 3 different protocols to successfully induce the VBNC state. Approximately one-third of bacteria entered a VBNC state after 15-30 days, and all remained viable for at least 2 months. The morphology, glycometabolism, and adhesion capabilities of VBNC cells differed from those of E. faecalis during the exponential growth phase. Specifically, VBNC E. faecalis cells could not decompose lactose, D-mannitol, or D-sorbitol, although they were able to metabolize sucrose. Transmission electron microscopy showed that the morphology of the VBNC E. faecalis cells changed significantly; the cytoplasmic matrix was unevenly condensed and the overall morphology of the cells became irregular, but the cell membranes remained intact. Although the adhesion ability of the bacteria decreased, VBNC E. faecalis could still adhere to collagen fiber type I and tooth dentine. The persistence of this adhesion ability may be important in the virulence of VBNC E. faecalis.

Inheritance of balanced translocation t(17; 22) from a Down syndrome mother to a phenotypically normal daughter.

We report that a 30-year-old woman with mental retardation was referred for prenatal diagnoses during pregnancy. An ultrasound scan showed that the heart structure and function of the fetus were normal. Cytogenetic analysis showed that the female karyotype was 47,XX, t(17; 22) (q21; q11), +21. The woman's husband had a normal male karyotype and was phenotypically normal. During this first pregnancy, an amniocentesis, which was done at 19 weeks, revealed that the fetal karyotype was 46,XX, t(17; 22) (q21; q11). Fluorescence in situ hybridization testing of amniotic fluid gave a normal result for chromosome 21. The child was a phenotypically normal female baby.

Sperm retrieval from patients with nonmosaic Klinefelter's syndrome by semen cytology examination.

Successful sperm retrieval from ejaculates of nonmosaic Klinefelter's syndrome (KS) patients by using semen cytology examination was described in this report. The clinical parameters of KS patients with sperm compared to patients without sperm were described. One hundred and fifty-one patients were proven to suffer from KS by chromosomal analysis using G-banding. Spermatozoa were obtained from 10 patients (10/151, 6.6%) using semen analysis. After semen cytology examination, 32 patients (32/151, 21.2%) were found to have sperm or germ cell in their ejaculate. The patients with successful sperm retrieval were significantly younger (27.1 ± 3.7 years) than the patients for whom sperm retrieval failed (28.9 ± 4.2 years). The mean serum testosterone level and the mean T/LH ratio of KS patients with successful sperm retrieval were significantly higher in men with sperm than in men without sperm (testosterone: 3.2 ± 2.1 ng/mL vs 2.7 ± 1.5 ng/mL; T/LH ratio: 0.2 ± 0.3 vs 0.1 ± 0.1). In conclusion, semen cytology examination should be performed to identify sperm and germ cells in the ejaculate of KS patients if no sperm can be detected by traditional semen analysis. The serum testosterone level and T/LH ratio revealed an association between impaired Leydig cell function and impaired spermatogenesis in KS males. KS patients should receive earlier diagnosis and treatment.

45,X/46,X,r(Y)/46,X,dic r(Y) karyotype in an azoospermic male: a case report.

Y chromosome abnormalities are frequently associated with male infertility. Men with ring Y chromosomes can present with sexual infantilism, ambiguous genitalia, hypospadias, or azoospermia. AZF microdeletions can result in spermatogenic defects in such patients. Here, we report an unusual karyotype of 45,X/46,X,r(Y)/46,X,dic r(Y) in an azoospermic man. However, the reason for this patient's azoospermia is not an AZF microdeletion but might be the abnormal structure of the r(Y) chromosome, the 45,X cell line, mosaicism of the 3 cell lines, or another unknown cause. In such cases, if the couple wishes to reproduce, cytogenetic, molecular and fluorescent in situ hybridization evaluations should be performed, and preimplantation genetic diagnosis should be used together with assisted reproductive technology.

Impact of chromosomal translocations on male infertility, semen quality, testicular volume and reproductive hormone levels.

OBJECTIVE: To analyse the relationship between male infertility and chromosomal translocations, and the influence of different types of chromosomal translocations on semen quality, testicular volume and hormone levels. METHODS: A retrospective cohort of infertile men was recruited for chromosomal analysis using standard Giemsa stain banding. Physical examinations, semen analysis, hormonal analysis and the detection of azoospermia factor (AZF) microdeletions were carried out. Men with normal fertility were used as controls. RESULTS: Among the 1056 infertile men, 22 had chromosomal translocations (2.1%), including seven with Robertsonian translocations (0.7%), 11 with autosome-autosome reciprocal translocations (1.0%) and four with gonosome-autosome reciprocal translocations (0.4%). Left and right testicular volumes of patients with chromosomal translocations were significantly smaller than those in the fertile control group. There were no significant differences in hormone levels between patients with chromosomal translocations and fertile controls, except for significantly lower testosterone levels in patients with Robertsonian and gonosome-autosome reciprocal translocations compared with the controls. All AZF microdeletion analyses showed normal results. CONCLUSIONS: Chromosomal translocations may cause reductions in testicular volume and testosterone level, which may impact spermatogenesis, resulting in azoospermia or oligozoospermia and male infertility.

Changes in spatial distribution of flexor digitorum superficialis muscle activity is correlated to finger's action.

Multitendoned extrinsic muscles of the human hand can be divided into several neuromuscular compartments (NMCs), each of which contributes to the ability of human finger to produce independent finger movements or force. The aim of this study was to investigate the changes in the spatial activation of flexor digitorum superficialis (FDS) during the fingertip force production with non-invasive multichannel surface electromyography (sEMG) technique. 7 healthy Subjects were instructed to match the target force level for 5s using individual index finger (I), individual middle finger (M) and the combination of the index and middle finger (IM) respectively. Simultaneously, a 2 × 6 electrode array was employed to record multichannel sEMG from FDS as finger force was produced. The entropy and center of gravity of the sEMG root mean square (RMS) map were computed to assess the spatial inhomogeneity in muscle activation and the change in spatial distribution of EMG amplitude related to the force generation of specific task finger. The results showed that the area and intensity of high amplitude region increased with force production, and the entropy increased with force level under the same task finger. The findings indicate that the change of spatial distribution of multitendoned extrinsic hand muscle activation is correlated to specific biomechanical functions.

Preliminary study of the presence and association of bacteria and archaea in teeth with apical periodontitis.

AIM: To investigate, by reverse transcription polymerase chain reaction (RT-PCR), the presence and association of bacteria and archaea in primary and secondary root canal infections. METHODOLOGY: A total of 77 root canal samples from 77 Chinese patients, 42 with necrotic pulp tissues (primary infection) and 35 with failed prior conventional root canal treatment (secondary infection), aseptically exposed at the first patient visit, were studied. Total RNA was isolated directly from each sample, and 16S rRNA gene-based RT-PCR assays were used to determine the presence of bacteria and archaea, respectively. RESULTS: Bacteria were detected in 39/42 (93%) of root canal samples from teeth with primary infections, and archaea in 16/42 (38%). In the cases diagnosed as secondary root-infected canals, bacteria were detected in 30/35 (86%), whilst archaea were detected in 6/35 (17%) of cases. Amongst the canals, which were positive for bacteria, archaea were always found in combination with bacteria. The incidence of symptomatic cases positive for both bacteria and archaea (16/22, 73%) were significantly higher than those positive for bacteria alone (21/47, 45%) (P < 0.05). CONCLUSIONS: This study confirms the presence of archaea in root canal infections and further implicates them in an association with clinical symptoms. The nature of this association requires further study.

Prevalence and molecular diversity of Archaea in subgingival pockets of periodontitis patients.

INTRODUCTION: The aim of this study was to investigate the prevalence and molecular diversity of Archaea in the subgingival crevices of patients with chronic periodontitis. METHODS: Subgingival plaque was collected from 41 patients with chronic periodontitis and 15 healthy subjects. The prevalence of Archaea in those plaque samples was tested by polymerase chain reaction with two broad-range archaeal primer sets. Amplicons from eight Archaea-positive plaque samples were cloned and sequenced for molecular diversity analysis using one of these two primer sets and a novel third primer set. RESULTS: Archaea were detected in the subgingival plaque of patients with chronic periodontitis at a prevalence of 70.7-73.2%, but were not detected in healthy subjects. Using one primer set, all sequences of the archaeal amplicons were identified as Methanobrevibacter oralis-like species. With another primer set, the amplicons were also found to be identical to the uncultured M. oralis-like species except one phylotype was found to belong to the class Thermoplasmata. CONCLUSION: Archaea might be correlated with periodontal diseases. The diversity of Archaea associated with periodontitis was limited. Almost all sequenced amplicons fell into the genus Methanobrevibacter of the Euryarcheota phylum. M. oralis-like species was the predominant but non-exclusive archaeon in the subgingival dental plaque of patients with periodontitis.

Association of uncultivated oral phylotypes AU126 and X112 with periodontitis.

OBJECTIVE: To discuss the relationship between uncultivated pathogenic bacteria and periodontitis. SUBJECTS AND METHODS: Specific polymerase chain reaction (PCR) primers were designed for phylotypes AU126 and X112; PCRs were applied to determine the prevalence of these phylotypes in 35 patients with chronic periodontitis, 26 patients with plaque-induced gingivitis and 20 healthy control subjects. RESULTS: The specificity of each primer is validated on the basis of the results from sequence analysis of PCR products. AU126 and X112 were detected in the subgingival plaque samples in all the three groups. The prevalence of AU126 in subgingival plaque in chronic periodontitis (77.1%) and plaque-induced gingivitis (61.5%) is relatively higher than that in the healthy subjects (10.0%), and the difference is statistically significant (P < 0.01). The prevalence of X112 in subgingival plaque in periodontitis patients (85.7%) is higher than that in healthy subjects (30.0%), the difference (P < 0.01) being equally statistically significant. The difference between the chronic periodontitis group and the plaque-induced gingivitis group (50.0%) is statistically significant (P < 0.05). CONCLUSIONS: It might be assumed that the novel uncultivated AU126 phylotype could possibly be related to chronic periodontitis and plaque-induced gingivitis, and that X112 might play a role in the progress of lesion from gingivitis to periodontitis.

[Studies on the chemical constituents from Aralia elata].

Eight compounds have been isolated from the root bark of Aralia elata. Their structures have been identified by means of physico-chemical and spectral analysis. They are (6'-O-palmitoyl)-beta-sitosterol-3-O-beta-D-glucoside (A5), silphiosideA (A9), chikusetusaponin Ib (A11), araloside A (A12), araloside C (A15), acanthoside D (B1). Compound A10 is a new natural product, named as araloside A methyl-ester. 3-O-beta-D-glucopyranosyl (1----3)[beta-D-glucopyranosy (1----4)]-beta-D-glucopyranosyl-oleanolic acid-28-O-beta-D-glucopyranoside (A16) is a new compound, named as araloside G. Compounds A5, A9, A10, A11, A16, and B1 were isolated for the first time from the plant. 13C-NMR chemical shifts of compounds A9 and A15 were assigned for the first time.