[Study on primary screening technique for children with autism spectrum disorder].

To explore screening tools for children with autism spectrum disorder (ASD), which are convenient for primary hospitals, it can provide basic data for formulating ASD prevention policies. This was a cross-sectional study by cluster sampling. Huyi District and Xincheng District were extracted for investigation in Xi'an City. From July 2021 to September 2022, all children aged from 3 months to 36 months who live in the two districts were subjected to primary screening. The child care physician used the routine screening tool "warning signs checklist for screening psychological, behavioral and developmental problems of children" and cartoon pictures of "early high-risk warning signs of autism", the children who were positive in the initial screening were referred to the district level maternal and child health hospital for re-screening, and those who were positive in the re-screening were referred to Xi 'an Children's Hospital for diagnosis. The results showed that a total of 17 905 children aged from 3 months to 36 months were initially screened in the two districts, including 10 588 children aged from 18 months to 36 months, 50 children who were positive in the initial screening and 50 children who were re-screened. 23 children (18 boys and 5 girls) were diagnosed with ASD. The prevalence rate of ASD in children was 2.17‰ (95% confidence interval:1.29‰-3.06‰). 42 children were positive for "warning signs checklist" at the preliminary screening, and 19 were confirmed as ASD. 27 children were positive for "cartoon pictures" in the preliminary screening, and 23 were confirmed with ASD. The "cartoon pictures" in the preliminary screening and diagnosis of consistent rate was higher than the "warning signs checklist", two kinds of screening methods comparison were statistically significant difference in the odds of consistent (χ2=11.01, P=0.001). In conclusion, relying on the three-level network of maternal and child health care, it is conducive to the whole process management of screening and diagnosis of children with ASD, and to guide the formulation of prevention policies. The cartoon pictures of "early high-risk warning signs of autism" can assist the identification of children with ASD based on the "warning signs checklist", which is simple, effective and suitable for promotion in the community health care.

[Relationship between blood glucose variation, collateral circulation and Batman score and prognosis after mechanical thrombectomy with Solitaire stent in acute posterior vascular occlusive stroke].

Objective: To explore the relationship between blood glucose variability, collateral circulation and basilar artery computed scan angiography score (Batman) and prognosis of mechanical thrombectomy with Solitaire stent in patients with large vascular occlusive APCI. Methods: A retrospective study was conducted on 113 patients with large vessel occlusive APCI who underwent Solitaire stent mechanical thrombectomy in the Department of Neurology of Nanyang Central Hospital from March 2021 to July 2022. According to the prognosis, they were divided into outcome group (46 cases) and adverse group (67 cases). Evaluate the prognosis based on the Modified Rankin Scale three months after the surgery. The differences in collateral circulation, GV and Batman score between the two groups were compared, and the related factors affecting the prognosis of large vessel occlusive APCI patients treated with Solitaire stent mechanical thrombectomy were analyzed by multivariate logistic regression model. Results: The age of 113 patients with acute large vessel occlusive APCI was (65.3±8.9) years old. The proportion of female was 34.5% (39 cases). Compared with the outcome group, the adverse group had a lower proportion of collateral circulation [40 cases (87.0%) vs 47 cases (70.2%)], higher GV score [(25.19±3.54) vs (30.36±4.11) points], lower Batman score [(7.49±1.52) vs (6.65±1.33) points], higher proportion of atrial fibrillation history [16 cases (23.9%) vs 4 cases (8.7%)], higher National Institutes of Health Stroke Scale (NIHSS) score at admission [(8.33±0.74) vs (7.25±0.92) points], larger core infarct volume [(32.57±4.87) vs (29.54±5.14) ml], and longer time from admission to vascular recanalization [(123.52±31.17) vs (102.47±29.54) min] (all P<0.05). Atrial fibrillation history, core infarct volume, NIHSS score at admission, time from admission to vascular recanalization, glycemic variability, collateral circulation, and Batman score were related factors for the prognosis of large vessel occlusive APCI patients treated with Solitaire stent mechanical thrombectomy, with ORvalues (95%CI) of 1.383 (1.124-1.641), 1.166 (1.007-1.350), 4.777 (1.856-12.297), 3.068 (2.379-3.757), 1.477 (1.209-1.806), 0.742 (0.654-0.831), and 0.717 (0.214-1.221), respectively (all P<0.05). Conclusion: Blood glucose variation is a risk factor for prognosis of mechanical thrombectomy with Solitaire stent in patients with large vascular occlusive APCI, and collateral circulation and Batman score are protective factors.

[Factors influencing poor prognosis of mechanical thrombectomy in time window of acute ischemic stroke].

Objective: Investigate the causes of poor prognosis of mechanical thrombectomy in the time window of acute ischemic stroke (AIS) with anterior circulation. Methods: A retrospective analysis was made on the data of 78 patients with anterior circulation AIS who underwent mechanical thrombectomy in the time window from January 2017 to December 2017 in the Department of Vascular Neurosurgery of Liaocheng Brain Hospital. The modified Rankin scale (mRS) was used to evaluate the prognosis of the patients 3 months after operation. According to the prognosis,the patients were divided into the group with good prognosis (42 cases, mRS<2 points) and the group with poor prognosis (36 cases, mRS<3 points). Univariate and multivariate Logistic regression analysis was used to analyze the related factors of poor prognosis. Results: (1) Univariate analysis showed that the prognosis of patients with good combination and primary stenosis of diabetes mellitus and atherosclerosis was lower than that of patients with poor prognosis (P<0.05). The collateral circulation compensation rate and vascular recanalization rate of patients with good prognosis were higher than those of patients with poor prognosis (P<0.05). Learning significance (P<0.05). (2) Multivariate analysis showed that diabetes mellitus (P=0.035), collateral circulation compensation (P=0.011) and primary atherosclerotic stenosis (P=0.042) were independent risk factors for poor prognosis. Conclusion: Perfect preoperative evaluation and strict screening of patients, good collateral circulation compensation,individualized treatment for patients with primary atherosclerotic stenosis,and strict control of postoperative hyperglycemia can improve the clinical prognosis of endovascular therapy.

Development, characterization and virus susceptibility of a continuous cell line from the caudal fin of marbled eel (Anguilla marmorata).

A continuous cell line consisting mostly of epithelioid cells was established from the caudal fin of marbled eels (Anguilla marmorata) and designated as marbled eel caudal fin (MECF)-1. The cells multiplied well in Leibovitz's L-15 medium containing 2% to 15% foetal bovine serum at temperatures of 20°C to 35°C and were subcultured for >90 passages during a 5-year period from 2012 to 2017. Transcripts of ictacalcin, keratin 13, cd146, nestin, ncam1 and myod1 were demonstrated in the cells using reverse transcription polymerase chain reaction. The results indicated that MECF-1 was composed of epidermal and mesenchyme stem and progenitor cells including myoblasts. MECF-1 was susceptible to Japanese eel herpesvirus HVA980811, marbled eel polyoma-like virus (MEPyV), aquabirnavirus MEIPNV1310 and aquareovirus CSV. By contrast, MECF-1 was noted refractory to megalocytiviruses RSIV-Ku and GSIV-K1 infection. Moreover, the cells were resistant to betanodavirus infection. In conclusion, MECF-1 derived from marbled eel is suitable for studies on anguillid viruses and interaction with host cells.

Characterization and virus susceptibility of a continuous cell line derived from the brain of Aequidens rivulatus (Günther).

Cell cultures derived from the brain tissues of Aequidens rivulatus (Günther) have been characterized previously. In this study, a continuous cell line ARB8 was further established, and its growth characteristics, transcription and susceptibility to fish viruses-including chum salmon reovirus (CSV), marbled eel infectious pancreative necrosis virus (MEIPNV), grouper nervous necrosis virus (GNNV), giant seaperch iridovirus (GSIV), red seabream iridovirus (RSIV), koi herpesvirus (KHV), herpesvirus anguilla (HVA) and marbled eel polyoma-like virus (MEPyV)-were examined. ARB8 cells that showed epithelioid morphology and were passaged >80 times grew well at temperatures ranging from 25°C to 30°C in L-15 medium containing 5%-15% foetal bovine serum. The cells constitutively transcribed connexion 43, glutamine synthetase, nestin and nkx6-2, which are markers for neural progenitor cells. The cells were highly susceptible to CSV, MEIPNV, GSIV and RSIV and showed the typical cytopathic effect (CPE). However, the cells were resistant to GNNV, KHV, HVA and MEPyV because no significant CPE was noted after infection. Optimal temperatures for virus production ranged from 25°C to 30°C. The results revealed that the neural progenitor cell line ARB8 can potentially serve as a useful tool for investigating fish viruses and isolating new viruses in ornamental cichlid fishes.

Immunocytochemical characterisation of neural stem-progenitor cells from green terror cichlid Aequidens rivulatus.

In this study, cultures of neural stem-progenitor cells (NSPC) from the brain of green terror cichlid Aequidens rivulatus were established and various NSPCs were demonstrated using immunocytochemistry. All of the NSPCs expressed brain lipid-binding protein, dopamine- and cAMP-regulated neuronal phosphoprotein 32 (DARPP-32), oligodendrocyte transcription factor 2, paired box 6 and sex determining region Y-box 2. The intensity and localisation of these proteins, however, varied among the different NSPCs. Despite being intermediate cells, NSPCs can be divided into radial glial cells, oligodendrocyte progenitor cells (OPC) and neuroblasts by expressing the astrocyte marker glial fibrillary acidic protein (GFAP), OPC marker A2B5 and neuronal markers, including acetyl-tubulin, ßIII-tubulin, microtubule-associated protein 2 and neurofilament protein. Nevertheless, astrocytes were polymorphic and were the most dominant cells in the NSPC cultures. By using Matrigel, radial glia exhibiting a long GFAP+ or DARPP-32+ fibre and neurons exhibiting a significant acetyl-tubulin+ process were obtained. The results confirmed that NSPCs obtained from A. rivulatus brains can proliferate and differentiate into neurons in vitro. Clonal culture can be useful for further studying the distinct NSPCs.

[Effect of continuous subcutaneous insulin infusion on the evaluation of ß cell function].

OBJECTIVE: To evaluate the impact of continuous subcutaneous insulin infusion (CSII) on ß-cell function assessment. METHODS: One hundred and twenty-five patients with type 2 diabetes (T2DM) admitted to Third Affiliated Hospital of Sun Yat-sen University treated with CSII were enrolled from May to December 2015. Blood samples were collected to measure their fasting blood glucose, haemoglobin A1c, blood lipids and plasma C peptide levels (fasting, 30 min and 120 min after a mixed meal) on the next day of their admission before CSII started. When patients achieved the target of fasting capillary glucose ≤ 7.0 mmol/L, C-peptide levels (0 min, 30 min and 120 min after a mixed meal) were measured. Then CSII were stopped at 10 pm with the same tests repeated on the next day. RESULTS: Compared with those measured before CSII [0 min: (0.35±0.20) nmol/L, 30 min: (0.57±0.31) nmol/L, 120 min: (0.84±0.54) nmol/L], C-peptide levels after stopping CSII at all time points [0 min: (0.41±0.16) nmol/L, 30 min: (0.71±0.33) nmol/L, 120 min: (1.37±0.75) nmol/L] increased (P=0.015, P=0.005, P<0.001) even glucose control was achieved, but significantly decreased immediately before CSII was stopped [0 min: (0.23±0.13)nmol/L, 30 min: (0.39±0.26) nmol/L, 120 min: (0.67± 0.50) nmol/L] (P<0.001, P<0.001, P=0.023). The C-peptide after stopping CSII increased to 141% (0 min), 127% (30 min) and 219% (120 min) respectively compared to those before stopping CSII. CONCLUSION: CSII therapy should be stopped for accurate evaluation of ß-cell function due to its"ß-cell rest"effect in T2DM.

Characterization and viral susceptibility of a brain cell line from brown-marbled grouper Epinephelus fuscoguttatus (Forsskål) with persistent betanodavirus infection.

A continuous cell line designated BMGB (brown-marbled grouper brain) was established from the brain tissues of the brown-marbled grouper Epinephelus fuscoguttatus and characterized. BMGB cells were identified as astroglial progenitor cells because they expressed glial fibrillary acidic protein and keratin and were persistently infected by betanodavirus, as confirmed through immunocytochemistry, polymerase chain reaction and immunoblot analyses. Because few intact virions were present in the BMGB cell culture fluid, the cytopathic effect (CPE) was not observed when the culture fluid was inoculated with GBC1 cells. However, BMGB cells displayed typical CPE after infection with additional betanodavirus, megalocytivirus and chum salmon reovirus. BMGB cells showed low myxovirus resistance (Mx) protein expression, which increased following betanodavirus and reovirus infection. Because the cells contained several unusual or degraded viral proteins, the persistent infection of betanodavirus in the BMGB cells may have resulted from a mechanism that destroys the viral proteins rather than the result of Mx protein expression. Despite the persistent betanodavirus infection, BMGB cells proliferated in a manner similar to other normal tropic fish cells and supported the propagation of several piscine viruses; however, the yield was lower than that of normal cells. The BMGB cells will be useful for investigating virus and host cell interaction.

Isolation of a novel polyomavirus, related to Japanese eel endothelial cell-infecting virus, from marbled eels, Anguilla marmorata (Quoy & Gaimard).

Marbled eels, Anguilla marmorata (Quoy & Gaimard), cultured in Taiwan exhibited haemorrhage and mortality in January 2012. The severely diseased eels bled from the gills and showed congestion of the central venous sinus of the gill filaments and haemorrhage throughout the body similar to viral endothelial cell necrosis of eel. In this study, a novel polyomavirus (AmPyV) was isolated from the diseased eels using the AMPF cell line established from the pectoral fin of healthy marbled eels. AmPyV was found to encode a long T-antigen orthologous gene. Phylogenetic analysis showed that AmPyV was closely related to Japanese eel endothelial cell-infecting virus. PCR assays revealed AmPyV infection throughout the systemic organs. AmPyV proliferated in the AMPF, EK-1 and EO-2 cells at temperatures 25-30 °C, and the progeny virus yields were 10(7.0) , 10(7.4) and 10(7.7) TCID50  mL(-1) , respectively. The purified virions were icosahedral particles, 70-80 nm in diameter. No clinical signs or mortality was observed among the eels injected with the virus; however, the virus was reisolated from the brain, eyes, kidneys, fins and gills of infected eels 2 month after injection. Our results suggest that AmPyV exhibits a latent infection. Pathogen of the disease needs to study further.

Developing immunological methods for detecting Macrobrachium rosenbergii nodavirus and extra small virus using a recombinant protein preparation.

Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV) have been identified as the causative agents for white tail disease (WTD) of M. rosenbergii. In this study, the gene sequences encoding MrNV and XSV capsid proteins were separately ligated into the pGEX-4T-3 expression vector and transformed into Escherichia coli. After induction, glutathione-S-transferase (GST)-tagged MrNV and XSV fusion proteins were obtained with molecular masses of 68 and 43 kDa, respectively. Specific polyclonal antibodies for MrNV and XSV against viral recombinant proteins and infected prawn tissues were verified using Western blotting. According to immunodot blot assay results, the detection sensitivities of antibodies were approximately 5 ng µL(-1) for both recombinant proteins GST-MrNV and GST-XSV. In additional, MrNV and XSV were detected at dilution levels of 1:2560 and 1:640 in the infected prawn tissues, respectively. No cross-reactions with white spot syndrome virus or grouper nervous necrosis virus were observed using immunodot blot assays. MrNV and XSV in infected muscle tissues were detected using immunohistochemistry. Although the detection limit of the immunodot blot assay was lower than that of nested reverse transcription polymerase chain reaction, these polyclonal antibodies can be useful for confirming MrNV and XSV infections in field tests.

The prevalence of actinic keratosis in patients visiting dermatologists in two hospitals in China.

BACKGROUND: Actinic keratosis (AK) is one of the most common conditions treated by dermatologists in western countries. Studies have shown that AK prevalence in Europe, the U.S.A. and Australia is 4·5-60%. No data of AK prevalence in China has been reported. OBJECTIVES: This study aimed to explore the prevalence of AK in patients visiting dermatologists in two hospitals in China. METHODS: This study was conducted in the dermatology departments of two teaching hospitals (Peking University First Hospital, Beijing, and Fourth Military Medical University, Xi'an). All records for 5 years between 2008 and 2012 with clinically or pathologically diagnosed AKs were collected from the pathological databases of both hospitals. Data from these records were used to calculate the prevalence of AKs among patients who were seen by dermatologists in these hospitals. To estimate the reliability of data from the previous database, a cross-sectional study was conducted simultaneously in the two hospitals from 15 October to 8 December in 2012 after all dermatologists in the two departments were retrained through intensive courses on recognizing AK clinically. RESULTS: The prevalence of total clinical AKs through 2008-2012 was 0·52% in 1 590 817 patient visits in the two hospitals. The yearly prevalence of clinical AKs was 0·30-1·20%. In the cross-sectional study, 72 437 clinical patients were screened and 76 patients (1·05%) were identified to have clinically recognized AK. CONCLUSIONS: The overall prevalence of AKs in patients visiting dermatologists in the two hospitals in China was 0·52%, which is much lower than the prevalence in western countries.

Characterization of a novel cell line from the caudal fin of koi carp Cyprinus carpio.

A continuous cell line (KF-101) derived from the caudal fin of the koi carp Cyprinus carpio was established and characterized. The KF-101 cell line multiplied abundantly in Leibovitz's L-15 medium containing 10% foetal bovine serum at 25° C, and was subcultured for >90 passages over a period of 3 years. Immunocytochemistry revealed that the KF-101 cells contain keratin, junction proteins connexin-43 and occludin, and ectodermal stem-cell marker Pax-6, but not vimentin. Furthermore, the KF-101 cells reacted with anti-human DARPP-32 and anti-human GATA-4 antibodies, and the labelling was regulated according to the cell cycle. The labels of the DARPP-32 and GATA-4 antibodies in the KF-101 cells were the suggested phosphatase-1 inhibitor-1 and GATA-3, respectively. In addition, the KF-101 cells were susceptible to koi herpesvirus but were resistant to eel herpesvirus, iridovirus, grouper nodavirus and chum salmon (Oncorhynchus keta) virus. The results indicate that the KF-101 cells are suitable materials for investigating biological and virological development.

Viral susceptibility, transfection and growth of SPB--a fish neural progenitor cell line from the brain of snubnose pompano, Trachinotus blochii (Lacépède).

This study investigates the susceptibilities of the SPB cell line to fish viruses including giant seaperch iridovirus (GSIV-K1), red sea bream iridovirus (RSIV-Ku), grouper nervous necrosis virus (GNNV-K1), chum salmon reovirus (CSV) and eel herpesvirus (HVA). GSIV-K1, RSIV-Ku and CSV replicated well in SPB cells, with a significant cytopathic effect and virus production. However, the cells were HVA and GNNV refractory. To examine the ability of SPB cells to stably express foreign protein, expression vectors encoding GNNV B1 and B2 fused to enhanced green fluorescent protein (EGFP) and GSIV ORF35L fused to DsRed were constructed and introduced by transfection into SPB cells. Stable transfectants displayed different morphologies compared with SPB and with each other. EGFP-B1 was predominantly localized in the nuclei, EFPF-B2 was distributed throughout the cytoplasm and nucleus, and granular 35L-DsRed was localized with secreted vesicles. The expression of EFPF-B2 in SPB cells produced blebs on the surface, but the cells showing stable expression of EGFP, EGFP-B1 or 35L-DsRed showed normal morphologies. Results show the SPB cells and the transfected cells grow well at temperatures between 20 and 35 °C and with serum-dependent growth. SPB cells are suitable for studies on foreign protein expression and virology.

PCR amplification and sequence analysis of the major capsid protein gene of megalocytiviruses isolated in Taiwan.

Viruses belonging to the genus Megalocytivirus in the family Iridoviridae are one of the major agents causing mass mortalities in marine and freshwater fish in Asian countries. Outbreaks of iridovirus disease have been reported among various fish species in Taiwan. However, the genotypes of these iridoviruses have not yet been determined. In this study, seven megalocytivirus isolates from four fish species: king grouper, Epinephelus lanceolatus (Bloch), barramundi perch, Lates calcarifer (Bloch), silver sea bream, Rhabdosargus sarba (Forsskal), and common ponyfish, Leiognathus equulus (Forsskal), cultured in three different regions of Taiwan were collected. The full open reading frame encoding the viral major capsid protein gene was amplified using PCR. The PCR products of approximately 1581 bp were cloned and the nucleotide sequences were phylogenetically analysed. Results showed that all seven PCR products contained a unique open reading frame with 1362 nucleotides and encoded a structural protein with 453 amino acids. Even though the nucleotide sequences were not identical, these seven megalocytiviruses were classified into one cluster and showed very high homology with red sea bream iridovirus (RSIV) with more than 97% identity. Thus, the seven iridovirus strains isolated from cultured marine fish in Taiwan were closer to the RSIV genotype than the infectious spleen and kidney necrosis virus genotype.

Macrobrachium rosenbergii nodavirus infection in M. rosenbergii (de Man) with white tail disease cultured in Taiwan.

White tail disease (WTD) is a serious problem in Macrobrachium rosenbergii hatcheries and nursery ponds in Asia. The causative agents have been identified as M. rosenbergii nodavirus (MrNV) and its associated extra small virus. This is the first report demonstrating MrNV virus in M. rosenbergii displaying WTD signs in Taiwan by reverse transcriptase-polymerase chain reaction (RT-PCR). Amplified fragments of 850 and 425 bp for RNA-1 and RNA-2 of MrNV, respectively, were obtained by RT-PCR. RT-PCR products of about 850 and 1121 bp for RNA-1 and RNA-2 of MrNV were also obtained using different primer pairs. The amplicons were individually cloned into pGEM-T vector and sequenced. Using this recombinant plasmid of MrNV RNA-2 as DNA template, the non-radioactive DNA probes were prepared by PCR amplification with DIG-11-dUTP. The probes were used to successfully detect MrNV infection in the striated muscle tissues of WTD-diseased prawns using in situ hybridization. The 1121 bp genomic fragment of RNA-2 of MrNV consisted of a unique open reading frame with 1116 nucleotides, and it encoded a structural protein with 371 amino acids. The nucleotide sequence of the partial genome of MrNV RNA-2 revealed a 97% identity with an Indian isolate. A phylogenetic tree constructed using the nucleotide sequence of the viral capsid gene from insect and fish nodaviruses revealed that the MrNV Taiwan isolate could be interpreted as a new genus within the family Nodaviridae. However, its position showed more affinity with Alphanodavirus than with Betanodavirus. The study confirmed the presence of MrNV infection in freshwater prawns cultured in Taiwan suffering from WTD.

Effect of oxygen transfer on lipase production by Acinetobacter radioresistens.

The influence of oxygen on alkaline lipase production by Acinetobacter radioresistens was studied under two operating modes: controlled dissolved oxygen (DO) concentration and controlled aeration rate. Compared with cell growth, the lipase production depended more extensively on oxygen. The intrinsic factor determining cell growth and lipase production was oxygen transfer rate (OTR) rather than DO concentration. Improvements in OTR, either by aeration or agitation, resulted in an increase in lipase yield and/or a reduction in fermentation time. The formation of A. radioresistens lipase could be described by a mixed-growth-associated model, and the enzyme was mainly a growth-associated product. The overall productivity for the lipase, which depended more strongly on agitation than aeration, could be related with kLa. DO concentration could not be employed in this correlation, though it has been useful as a criterion for ensuring no oxygen limitation in an aerobic fermentation.