Reassortment and genomic analysis of a G9P[8]-E2 rotavirus isolated in China.

OBJECTIVE: To isolate a prevalent G9P[8] group A rotavirus (RVA) (N4006) in China and investigate its genomic and evolutionary characteristics, with the goal of facilitating the development of a new rotavirus vaccine. METHODS: The RVA G9P[8] genotype from a diarrhea sample was passaged in MA104 cells. The virus was evaluated by TEM, polyacrylamide gel electrophoresis, and indirect immunofluorescence assay. The complete genome of virus was obtained by RT-PCR and sequencing. The genomic and evolutionary characteristics of the virus were evaluated by nucleic acid sequence analysis with MEGA ver. 5.0.5 and DNASTAR software. The neutralizing epitopes of VP7 and VP4 (VP5* and VP8*) were analyzed using BioEdit ver. 7.0.9.0 and PyMOL ver. 2.5.2. RESULTS: The RVA N4006 (G9P[8] genotype) was adapted in MA104 cells with a high titer (105.5 PFU/mL). Whole-genome sequence analysis showed N4006 to be a reassortant rotavirus of Wa-like G9P[8] RVA and the NSP4 gene of DS-1-like G2P[4] RVA, with the genotype constellation G9-P[8]-I1-R1-C1-M1-A1-N1-T1-E2-H1 (G9P[8]-E2). Phylogenetic analysis indicated that N4006 had a common ancestor with Japanese G9P[8]-E2 rotavirus. Neutralizing epitope analysis showed that VP7, VP5*, and VP8* of N4006 had low homology with vaccine viruses of the same genotype and marked differences with vaccine viruses of other genotypes. CONCLUSION: The RVA G9P[8] genotype with the G9-P[8]-I1-R1-C1-M1-A1-N1-T1-E2-H1 (G9P[8]-E2) constellation predominates in China and may originate from reassortment between Japanese G9P[8] with Japanese DS-1-like G2P[4] rotaviruses. The antigenic variation of N4006 with the vaccine virus necessitates an evaluation of the effect of the rotavirus vaccine on G9P[8]-E2 genotype rotavirus.

Efficacy of high-intensity focused ultrasound combined with LNG-IUS for adenomyosis: a systematic review and meta-analysis.

OBJECTIVE: To evaluate the efficacy and safety of high-intensity focused ultrasound (HIFU) combined with the levonorgestrel intrauterine system (LNG-IUS) for adenomyosis. METHODS: We searched PubMed, Embase, Cochrane Library, Web of Science, CNKI, SinoMed, Wanfang, and VIP databases from their inception to Nov 20, 2021 for relevant articles that compared HIFU combined with LNG-IUS vs. HIFU alone in patients with adenomyosis. RevMan5.4 software was used for the data analysis. The primary outcome was changes in volume of the uterine. Secondary outcomes included visual analog scale (VAS) scores for dysmenorrhea, serum CA125 level, recurrence rate, changes in volume of the adenomyotic lesion, menstrual volume scores, and adverse reactions. Data synthesis was conducted using a random-effects model with significant heterogeneity (I2 > 50%), and using a fixed-effects model otherwise. This study is registered on the PROSPERO platform (CRD42021295214). RESULTS: The final analysis included 13 studies, with a total of 1861 patients. Results of analysis revealed that there was no significant difference in uterine volume reduction between the HIFU control group and the HIFU/LNG-IUS group at 3 months after procedure (MD:30.63). Compared with the HIFU control group, the HIFU/LNG-IUS group had more pronounced reduction in uterine volume at 6 (MD:29.04) and 12 months (MD:22.10) after procedure. The HIFU/LNG-IUS group has lower VAS scores for dysmenorrhea than the HIFU control group at 3 (MD:1.68), 6 (MD:1.69), and 12 months (MD:1.30) after procedure. Serum CA125 level in the HIFU/LNG-IUS group decreased more significantly than the HIFU control group at 6 (MD:18.34) and 12 months (MD:18.49) after procedure. The recurrence rate in the HIFU/LNG-IUS group was lower than that in the HIFU control group (RR:0.20). CONCLUSIONS: Compared to HIFU control group, HIFU/LNG-IUS group for the management of adenomyosis had more advantages in alleviating symptoms and decreasing the volumes of the uterine and adenomyotic lesions. However, since the number of the included studies was too small and some of them were not RCT, this conclusion needs to be referenced with caution.

Comparison of immune response to human rhinovirus C and respiratory syncytial virus in highly differentiated human airway epithelial cells.

BACKGROUND: Human rhinovirus C (HRV-C) accounts for a large proportion of HRV-related illnesses, but the immune response to HRV-C infection has not been elucidated. Our objective was to assess the effect of HRV-C on cytokine secretion in human bronchial epithelial (HBE) cells grown at air-liquid interface (ALI) and compare it with that of respiratory syncytial virus (RSV). METHODS: HBE cells were differentiated at ALI culture and the full-length cDNA clones of HRV-C651 and HRV-C15, clinical isolates of HRV-C79 and HRV-C101, and two RSV isolates were inoculated in the HBE cells. The effect of HRV-C on cytokine secretion was assessed and compared with that of RSV. RESULTS: HRV-Cs infect and propagate in fully differentiated HBE cells and significantly increase the secretion of IFN-λ1, CCL5, IP10, IL-6, IL-8, and MCP-1. The virus loads positively correlated with the levels of the cytokines. HRV-C induced lower secretion of CCL5 (P = 0.048), IL-6 (P = 0.016), MCP-1 (P = 0.008), and IL-8 (P = 0.032), and similar secretion of IP10 (P = 0.214) and IFN-λ1 (P = 0.214) when compared with RSV. CONCLUSION: HBE ALI culture system supported HRV-C infection and propagation and HRV-C induced relatively weaker cytokine expression than RSV.

NeiyiKangfu tablets control the progression of endometriosis through inhibiting RAF/MEK/ERK signal pathway by targeting RKIP.

BACKGROUND: NeiyiKangfu tablets (NYKF) are widely used clinically for the treatment of endometriosis (EMS), whose mechanism of action has been extensively studied. Researchers have found that NYKF may control the development of ectopic lesions by inhibiting angiogenesis and inflammatory cytokine secretion. Nevertheless, NYKF's mechanism of action remains unclear. METHODS: In the present study, the function of NYKF in the progression of EMS and the associated underlying mechanism was investigated by in vivo and in vitro experiments. EMS model mice were treated with NYKF and the pro-inflammatory factors and apoptosis of ectopic endometrium as well as RAF/mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling activation were assessed. In addition, human endometriosis-derived immortalized entopic stromal (hEM15A) cells transfected with or without RAF kinase inhibitor protein (RKIP)-small-interfering RNA (siRNA) were also treated with NYKF and the proliferation, migration, apoptosis, and RAF/MEK/ERK signaling activation were measured by Cell Counting Kit-8 (CCK-8), flow cytometry, Transwell, and western blot, respectively. RESULTS: Results showed that NYKF increased the expression of RKIP, inhibited RAF/MEK/ERK signaling activation, and induced apoptosis while inhibiting proliferation and migration both in EMS mice and hEM15A cells. RKIP knockdown could inhibit the effect of NYKF treatment, leading to the activation of RAF/MEK/ERK signaling and the proliferation and migration of hEM15A cells. CONCLUSIONS: In conclusion, these results suggest that NYKF treatment promotes apoptosis and inhibits proliferation and migration in EMS by inhibiting the RAF/MEK/ERK signaling pathway by targeting RKIP.

Strategies for managing the destruction of calcar femorale.

BACKGROUND: The calcar femorale was identified long ago. However, our current understanding of the calcar is insufficient, and its related concepts are sometimes confused. The calcar femoral is an important anatomical structure of the proximal femur, and its function can be overlooked. In trauma, tumors, or other diseases, the calcar femorale can be destroyed or changed pathologically. As a result, the mechanical structure of the proximal femur becomes destroyed, causing pathological fractures. How to address the destruction of the calcar femorale or the damage to the calcar femorale is discussed in this article. MAIN TEXT: Destruction of the calcar femorale is accompanied by many conditions, including trauma, tumors, and other diseases. The types of hip fractures caused by trauma include femoral neck fractures and intertrochanteric fractures. Dynamic hip screws, proximal femoral nail anti-rotation, and multiple parallel cannulate pins can be used in different conditions. When metastatic and primary bone tumors involve the calcar femorale, endoprostheses are widely used. Other diseases, such as fibrous dysplasia and aneurysmal bone cyst are treated differently. CONCLUSIONS: The calcar femorale can redistribute stresses and the destruction of the calcar femorale can lead to an increase in posterior medial stress. Many factors need to be considered when deciding whether to reconstruct the calcar femorale. Effective treatment strategies for managing the destruction of calcar femorale will need first establishing the precise mechanism of the destruction of the calcar and then designing therapies towards these mechanisms. Further investigation to the calcar needs to be carried out.

GII.13/21 Noroviruses Recognize Glycans with a Terminal ß-Galactose via an Unconventional Glycan Binding Site.

Human noroviruses (huNoVs) recognize histo-blood group antigens (HBGAs) as host susceptibility factors. GII.13 and GII.21 huNoVs form a unique genetic lineage that emerged from mainstream GII NoVs via development of a new, nonconventional glycan binding site (GBS) that binds Lea antigen. This previous finding raised the question of whether the new GII.13/21 GBS really has such a narrow glycan binding spectrum. In this study, we provide solid phenotypic and structural evidence indicating that this new GBS recognizes a group of glycans with a common terminal ß-galactose (ß-Gal). First, we found that P domain proteins of GII.13/21 huNoVs circulating at different times bound three glycans sharing a common terminal ß-Gal, including Lec, lactose, and mucin core 2. Second, we solved the crystal structures of the GII.13 P dimers in complex with Lec and mucin core 2, which showed that ß-Gal is the major binding saccharide. Third, nonfat milk and lactose blocked the GII.13/21 P domain-glycan binding, which may explain the low prevalence of GII.13/21 viruses. Our data provide new insight into the host interactions and epidemiology of huNoVs, which would help in the control and prevention of NoV-associated diseases.IMPORTANCE Evidence from both phenotypic binding assay and structural study support the observed interactions of human noroviruses (huNoVs) with histo-blood group antigens (HBGAs) as receptors or attachment factors, affecting their host susceptibility. GII.13 and GII.21 genotypes form a unique genetic lineage that differs from the mainstream GII huNoVs in their unconventional glycan binding site. Unlike the previous findings that GII.13/21 genotypes recognize only Lea antigen, we found in this study that they can interact with a group of glycans with a common terminal ß-Gal, including Lec, lactose, and mucin core 2. However, this wide glycan binding spectrum in a unique binding mode of the GII.13/21 huNoVs appears not to increase their prevalence, probably due to the existence of decoy glycan receptors in human gastrointestinal tract limiting their infection. Our findings shed light on the host interaction and epidemiology of huNoVs, which would impact the strategy of huNoV control and prevention.

An outbreak of gastroenteritis associated with a novel GII.8 sapovirus variant-transmitted by vomit in Shenzhen, China, 2019.

BACKGROUND: Human Sapoviruses (SaVs) has been reported as one of the causative agents of acute gastroenteritis (AGE) worldwide. An outbreak of SaVs affected 482 primary school students during spring activities from February 24 to March 11, 2019 in Shenzhen City, China. Our study was aimed at determining the epidemiology of the outbreak, investigating its origins, and making a clear identification of the SaVs genetic diversity. METHODS: Epidemiological investigation was conducted for this AGE outbreak. Stool samples were collected for laboratory tests of causative agents. Real-time reverse-transcription polymerase chain reaction (rRT-PCR) and conventional RT-PCR were used for detecting and genotyping of SaVs. The nearly complete genome of GII.8 SaV strains were amplified and sequenced by using several primer sets designed in this study. Phylogenetic analysis was performed to characterize the genome of GII.8 SaV strains. RESULTS: The single factor analysis showed that the students who were less than 1.5 m away from the vomitus in classroom or playgroundwere susceptible (P < 0.05). Seven of 11 fecal samples from patients were positive for GII.8 SaV genotype. In this study, we obtained the genome sequence of a SaV GII.8 strain Hu/SaV/2019008Shenzhen/2019 /CHN (SZ08) and comprehensively analyzed the genetic diversity. The phylogenetic analysis showed that the GII.8 strain SZ08 formed an independent branch and became a novel variant of GII.8 genotype. Strain SZ08 harbored 11 specific amino acid variations compared with cluster A-D in full-length VP1. CONCLUSIONS: This study identified SaVs as the causative agents for the AGE outbreak. Strain Hu SZ08 was clustered as independent branch and there was no recombination occurred in this strain SZ08. Further, it might become the predominant strain in diarrhea cases in the near future. Constant surveillance is required to monitor the emerging variants which will improve our knowledge of the evolution of SaVs among humans.

Case Study-Spiking Neural Network Hardware System for Structural Health Monitoring.

This case study provides feasibility analysis of adapting Spiking Neural Networks (SNN) based Structural Health Monitoring (SHM) system to explore low-cost solution for inspection of structural health of damaged buildings which survived after natural disaster that is, earthquakes or similar activities. Various techniques are used to detect the structural health status of a building for performance benchmarking, including different feature extraction methods and classification techniques (e.g., SNN, K-means and artificial neural network etc.). The SNN is utilized to process the sensory data generated from full-scale seven-story reinforced concrete building to verify the classification performances. Results show that the proposed SNN hardware has high classification accuracy, reliability, longevity and low hardware area overhead.

Glycan Binding Specificity and Mechanism of Human and Porcine P[6]/P[19] Rotavirus VP8*s.

Rotaviruses (RVs), which cause severe gastroenteritis in infants and children, recognize glycan ligands in a genotype-dependent manner via the distal VP8* head of the spike protein VP4. However, the glycan binding mechanisms remain elusive for the P[II] genogroup RVs, including the widely prevalent human RVs (P[8], P[4], and P[6]) and a rare P[19] RV. In this study, we characterized the glycan binding specificities of human and porcine P[6]/P[19] RV VP8*s and found that the P[II] genogroup RV VP8*s could commonly interact with mucin core 2, which may play an important role in RV evolution and cross-species transmission. We determined the first P[6] VP8* structure, as well as the complex structures of human P[19] VP8*, with core 2 and lacto-N-tetraose (LNT). A glycan binding site was identified in human P[19] VP8*. Structural superimposition and sequence alignment revealed the conservation of the glycan binding site in the P[II] genogroup RV VP8*s. Our data provide significant insight into the glycan binding specificity and glycan binding mechanism of the P[II] genogroup RV VP8*s, which could help in understanding RV evolution, transmission, and epidemiology and in vaccine development.IMPORTANCE Rotaviruses (RVs), belonging to the family Reoviridae, are double-stranded RNA viruses that cause acute gastroenteritis in children and animals worldwide. Depending on the phylogeny of the VP8* sequences, P[6] and P[19] RVs are grouped into genogroup II, together with P[4] and P[8], which are widely prevalent in humans. In this study, we characterized the glycan binding specificities of human and porcine P[6]/P[19] RV VP8*s, determined the crystal structure of P[6] VP8*, and uncovered the glycan binding pattern in P[19] VP8*, revealing a conserved glycan binding site in the VP8*s of P[II] genogroup RVs by structural superimposition and sequence alignment. Our data suggested that mucin core 2 may play an important role in P[II] RV evolution and cross-species transmission. These data provide insight into the cell attachment, infection, epidemiology, and evolution of P[II] genogroup RVs, which could help in developing control and prevention strategies against RVs.

Glycan binding patterns of human rotavirus P[10] VP8* protein.

BACKGROUND: Rotaviruses (RVs) are a major cause of acute children gastroenteritis. The rotavirus P [10] belongs to P[I] genogroup of group A rotaviruses that mainly infect animals, while the rotavirus P [10] was mainly identified from human infection. The rotavirus P [10] is an unusual genotype and the recognition pattern of cellular receptors remains unclear. METHODS: We expressed and purified the RV P [10] VP8* protein and investigated the saliva and oligosaccharide binding profiles of the protein. A homology model of the P [10] VP8* core protein was built and the superimposition structural analysis of P [10] VP8* protein on P [19] VP8* in complex with mucin core 2 was performed to explore the possible docking structural basis of P [10] VP8* and mucin cores. RESULTS: Our data showed that rotavirus P [10] VP8* protein bound to all ABO secretor and non-secretor saliva. The rotavirus P [10] could bind strongly to mucin core 2 and weakly to mucin core 4. The homology modeling indicated that RV P [10] VP8* binds to mucin core 2 using a potential glycan binding site that is the same to P [19] VP8* belonging to P[II] genogroup. CONCLUSION: Our results suggested an interaction of rotavirus P [10] VP8* protein with mucin core 2 and mucin core 4. These findings offer potential for elucidating the mechanism of RV A host specificity, evolution and epidemiology.

The suppression of innate immune response by human rhinovirus C.

Rhinovirus C (RV-C), a newly identified group of human rhinoviruses (RVs), is associated with exacerbation of severe asthma. The type I interferon (IFN) response induced by this virus and the mechanisms of evasion of IFN-mediated innate immunity for RV-C remain unclear. In this study, we constructed a full-length cDNA clone of RV-C (LZ651) from a clinical sample. IFN-ß mRNA and protein levels were not elevated in differentiated Human bronchial epithelial (HBE) cells at the air-liquid interface infected with RV-C, except in the early stage of infection. The ability to attenuate IFN-ß activation was ascribed to 3Cpro of RV-C, and the 40-His site of 3Cpro played an important role. Furthermore, RIG-I was degraded by 3Cpro in a caspase-dependent manner and 3Cpro cleaved MAVS at 148 Q/A, which inhibited IFN signaling. Taken together, our results demonstrate the mechanism by which RV-C circumvents the production of type I IFN in infected cells.

Functional and Structural Characterization of P[19] Rotavirus VP8* Interaction with Histo-blood Group Antigens.

Rotaviruses (RVs) of species A (RVA) are a major causative agent of acute gastroenteritis. Recently, histo-blood group antigens (HBGAs) have been reported to interact with human RVA VP8* proteins. Human P[19] is a rare P genotype of porcine origin that infects humans sporadically. The functional and structural characteristics of P[19] VP8* interaction with HBGAs are unknown. In this study, we expressed and purified the VP8* proteins of human and porcine P[19] RVs. In oligosaccharide and saliva binding assays, P[19] VP8* proteins showed obvious binding to A-, B-, and O-type saliva samples irrespective of the secretor status, implying broad binding patterns. However, they did not display specific binding to any of the oligosaccharides tested. In addition, we solved the structure of human P[19] VP8* at 2.4 Å, which revealed a typical galectin-like fold. The structural alignment demonstrated that P[19] VP8* was most similar to that of P[8], which was consistent with the phylogenetic analysis. Structure superimposition revealed the basis for the lack of binding to the oligosaccharides. Our study indicates that P[19] RVs may bind to other oligosaccharides or ligands and may have the potential to spread widely among humans. Thus, it is necessary to place the prevalence and evolution of P[19] RVs under surveillance. IMPORTANCE: Human P[19] is a rare P genotype of porcine origin. Based on phylogenetic analysis of VP8* sequences, P[19] was classified in the P[II] genogroup, together with P[4], P[6], and P[8], which have been reported to interact with HBGAs in a genotype-dependent manner. In this study, we explored the functional and structural characteristics of P[19] VP8* interaction with HBGAs. P[19] VP8* showed binding to A-, B-, and O-type saliva samples, as well as saliva of nonsecretors. This implies that P[19] has the potential to spread among humans with a broad binding range. Careful attention should be paid to the evolution and prevalence of P[19] RVs. Furthermore, we solved the structure of P[19] VP8*. Structure superimposition indicated that P[19] may bind to other oligosaccharides or ligands using potential binding sites, suggesting that further investigation of the specific cell attachment factors is warranted.

The Molecular Mechanisms of Ciprofloxacin Resistance in Clinical Campylobacter jejuni and Their Genotyping Characteristics in Beijing, China.

We assessed the susceptibility of 182 Campylobacter jejuni isolates from patients with diarrhea to eight antibiotics and analyzed the molecular mechanisms of ciprofloxacin resistance as well as the genetic characteristics based on multilocus sequence typing (MLST). The C257T mutation was found on the quinolone resistance-determining region (QRDR) of the gyrA gene in all ciprofloxacin-resistant strains. Mutations on the QRDR of the gyrB gene were silent. A total of 74 strains had 7 inverted repeat (IR) (a 16-bp IR on the intergenic region between cmeR and cmeABC) mutation polymorphisms. Compared with strains without the IR mutations, strains with the IR mutations had higher resistance rates to ciprofloxacin (94.6% vs. 83.3%), nalidixic acid (94.6% vs. 83.3%), tetracycline (98.6% vs. 85.2%), doxycycline (91.9% vs. 71.3%), florfenicol (59.5% vs. 17.6%), chloramphenicol (25.7% vs. 4.6%), gentamicin (16.2% vs. 3.7%), and multidrug resistance than those without IR mutations (all p < 0.05). With C257T mutation alone, 89.9% strains with minimum inhibitory concentration (MIC) values focused on 16, 32, and 64 µg/mL, whereas strains with C257T mutation in combination with the IR mutations had a higher ciprofloxacin resistance level with 88.6% MIC values focused on 64, 128, and 512 µg/mL (p < 0.0001). The strains in this study showed a high genetic variability based on MLST with 117 sequence types (STs), 37 of which were novel. CC-21 was the most common clonal complex (CC) followed by CC-353 and CC-45. No association was found between STs and ciprofloxacin resistance. In conclusion, the C257T mutation on gyrA was the major mechanism for ciprofloxacin resistance, and the C257T mutation in combination with the IR mutations might result in more severe ciprofloxacin resistance to C. jejuni.

Roles of ERα during mouse trophectoderm lineage differentiation: revealed by antagonist and agonist of ERα.

During mouse early embryogenesis, blastomeres increase in number by the morula stage. Among them, the outer cells are polarized and differentiated into trophectoderm (TE), while the inner cells remain unpolarized and give rise to inner cell mass (ICM). TE provides an important liquid environment for ICM development. In spite of extensive research, the molecular mechanisms underlying TE formation are still obscure. In order to investigate the roles of estrogen receptor α (ERα) in this course, mouse 8-cell embryos were collected and cultured in media containing ERα specific antagonist MPP and/or agonist PPT. The results indicated that MPP treatment inhibits blastocyst formation in a dose-dependent manner, while PPT, at proper concentration, promotes the cavitation ratio of mouse embryos. Immunofluorescence staining results showed that MPP significantly decreased the nuclear expression of CDX2 in morula, but no significant changes of OCT4 were observed. Moreover, after MPP treatment, the expression levels of the genes related to TE specification, Tead4, Gata3 and Cdx2, were significantly reduced. Overall, these results indicated that ERα might affect mouse embryo cavitation by regulating TE lineage differentiation.

Inhibition of phosphorylated Ser473-Akt from translocating into the nucleus contributes to 2-cell arrest and defective zygotic genome activation in mouse preimplantation embryogenesis.

Phosphorylated Ser473-Akt (p-Ser473-Akt) is extensively studied as a correlate for the activity of Akt, which plays an important role in mouse oogenesis and preimplantation embryogenesis. However, little progress has been made about its effect on the mouse zygotic genome activation (ZGA) of 2-cell stage in mouse preimplantation embryos. In this study, we confirmed its localization in the pronuclei of 1-cell embryos and found that p-Ser473-Akt acquired prominent nucleus localization in 2-cell embryos physiologically. Akt specific inhibitors API-2 and MK2206 could inhibit the development of mouse preimplantation embryos in vitro, and induce 2-cell arrest at certain concentrations. 2-cell embryos exposed to 2.0 µmol/L API-2 or 30 µmol/L MK2206 displayed attenuated immunofluorescence intensity of p-Ser473-Akt in the nucleus. Simultaneously, qRT-PCR results revealed that 2.0 µmol/L API-2 treatment significantly downregulated the mRNA pattern of MuERV-L and eIF-1A, two marker genes of ZGA, suggesting a defect in ZGA compared with that of control group. Collectively, our work demonstrated the nuclear localization of p-Ser473-Akt during major ZGA, and Akt specific inhibitors API-2 and MK2206 which led to 2-cell arrest inhibited p-Ser473-Akt from translocating into the nucleus of 2-cell embryos with defective ZGA as well, implying p-Ser473-Akt may be a potential player in the major ZGA of 2-cell mouse embryos.

Emergence of human caliciviruses among diarrhea cases in southwest China.

BACKGROUND: Acute diarrhea is one of the most serious problems in global public health that causes considerable morbidity and mortality worldwide. Human caliciviruses (HuCV) including norovirus (NoV, genogroup GI and GII) and sapovirus (SaV), is a leading cause of acute sporadic diarrhea in individuals across all age groups. However, few studies had been conducted clarifying the characteristics of HuCV in diarrhea cases across all age groups in China. Our study was aimed at assessing the HuCV-related diarrhea burden and NoV genotypes distribution in southwest China. METHODS: The study was conducted in four hospitals in Kunming city, Yunnan province, from June 2014 to July 2015. Stool specimens were collected from 1,121 diarrhea cases and 319 healthy controls in outpatient departments. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect NoV (GI, GII) and SaV. Sequencing was applied to confirm the three viral infections and phylogenetic analysis was performed to determine their genotypes. A structured questionnaire was used to record the demographic information and clinical symptoms of subjects. RESULTS: HuCV was detected at an 11.0 % infection rate in 1,121 diarrhea cases and at 3.4 % rate in 319 non-diarrhea subjects (p < 0.0001, OR = 3.5, 95 % CI 1.8-6.5). The prevalence of the NoV genogroup GII and genotype GII.4 in diarrhea cases was significantly higher than that found in healthy controls (p < 0.0001, p = 0.018, respectively). NoV GII (n = 118, 10.5 %) was the most common HuCV subtype in diarrhea cases, followed by SaV (n = 3, 0.3 %) and NoV GI (n = 2, 0.2 %). Of 118 NoV GII strains isolated from diarrhea patients. GII.4 (n = 55, 46.6 %) was the predominant strain, followed by GII.3 (n = 28, 23.7 %), GII.12 (n = 25, 21.2 %), GII.17 (n = 8, 6.8 %), and GII.5 (n = 2, 1.7 %). Of the 55 GII.4 strains, the GII.4 Sydney 2012 variant had absolutely predominant prevalence (n = 52, 94.5 %), followed by the NoV GII.4-2006b variant (n = 3, 5.5 %). The GII.4 Orleans 2009 variant was not found in diarrhea cases of the study. CONCLUSIONS: NoV GII was the major genogroup and GII.4 was the most predominant strain detected in diarrhea patients. The GII.17 is an emergent variant in sporadic diarrhea and might become the predominant strain in diarrhea cases in the near future. Rapid, accurate detection kits need to be developed to help us find and treat NoV-associated diarrhea in clinical settings in a timely manner.

Cytokine responses and correlations thereof with clinical profiles in children with enterovirus 71 infections.

BACKGROUND: Severe complications associated with EV71 infections caused many infants death. However, the pathogenesis of EV71 infection in the severe cases remained poorly understood. METHODS: In this study we collected plasma and cerebrospinal fluid (CSF) specimens drawn in the acute and/or recovery phases from EV71-infected individuals, and plasma specimens from healthy children served as normal controls. We compared the levels of cytokines and chemokines determined by a Luminex-based cytokine bead array. RESULTS: The plasma levels of IL-1ß and IL-6 were significantly higher in severe and critical cases than in mild patients and normal controls. Higher plasma levels of IL-6, IL-10, and IL-8 were evident in critical than severe cases. The CSF levels of IL-6, IL-8, and IP-10 were higher, and that of RANTES lower (compared to plasma), in severe and critical patients. Significantly lower CSF levels of cytokines and chemokines were recorded in the recovery than the acute phase in severe and critical cases treated with intravenous immunoglobulin (IVIG) and glucocorticoids. Only the CSF levels of IL-6, IP-10, and IL-8 were significantly correlated with white blood cell counts, and absolute neutrophil and monocyte counts, in severe cases. Furthermore, the CSF levels of IL-6 were correlated with temperature in both cases. CONCLUSIONS: These data indicate that a major cytokine response and inflammation, in both plasma and the CNS, are features of disease caused by EV71 infection. Systemic inflammation caused by EV71 infection exacerbated the deterioration of the disease, and resulted in the disease progression to the critical illness stage.

Characterization and cancer cell targeted imaging properties of human antivascular endothelial growth factor monoclonal antibody conjugated CdTe/ZnS quantum dots.

High luminescence quantum yield water-soluble CdTe/ZnS core/shell quantum dots (QDs) stabilized with thioglycolic acid were synthesized. QDs were chemically coupled to fully humanized antivascular endothelial growth factor165 monoclonal antibodies to produce fluorescent probes. These probes can be used to assay the biological affinity of the antibody. The properties of QDs conjugated to an antibody were characterized by ultraviolet and visible spectrophotometry, fluorescent spectrophotometry, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transmission electron microscopy and fluorescence microscopy. Cell-targeted imaging was performed in human breast cancer cell lines. The cytotoxicity of bare QDs and fluorescent probes was evaluated in the MCF-7 cells with an MTT viability assay. The results proved that CdTe/ZnS QD-monoclonal antibody nanoprobes had been successfully prepared with excellent spectral properties in target detections. Surface modification by ZnS shell could mitigate the cytotoxicity of cadmium-based QDs. The therapeutic effects of antivascular endothelial growth factor antibodies towards cultured human cancer cells were confirmed by MTT assay.

Astragaloside IV inhibits doxorubicin-induced cardiomyocyte apoptosis mediated by mitochondrial apoptotic pathway via activating the PI3K/Akt pathway.

Doxorubicin (DOX) is a widely used antitumor drug whose application is seriously limited by its cardiotoxicity. Mitochondria-mediated cardiomyocyte apoptosis plays a critical role in DOX-induced cardiotoxicity (DIC). The aim of the present study was to investigate the protective effect of astragaloside IV (3-O-beta-D-xylopyranosyl-6-O-beta-D-glucopyranosyl-cycloastragenol, AS-IV), a pure saponin isolated from Astragalus membranaceus, against DOX-induced cardiomyocyte apoptosis in primary cultured neonatal rat cardiomyocytes. Immunocytochemistry and Microculture Tetrazolium (MTT) assays showed that AS-IV significantly reduced DOX-induced cardiomyocyte loss. Additionally, AS-IV markedly ameliorated DOX-caused cardiomyocyte dysfunction via restoring the beating cell ratio and beating rate in cardiomyocytes. Furthermore, AS-IV substantially reduced the mitochondrial reactive oxygen species (ROS) production and lactate dehydrogenase (LDH), creatine kinase-MB isoenzyme (CK-MB) and cytochrome c (CytC) release, and restored the reduced ATP level, succinate dehydrogenase (SDH) and ATP synthase activities induced by DOX, suggesting that AS-IV significantly attenuated DOX-induced mitochondrial damage and dysfunction. It was further observed that DOX-induced cardiomyocyte apoptosis, as qualitatively evaluated by Hoechst 33258 staining and accurately quantified by flow cytometry, was markedly inhibited by AS-IV. Western blot analysis manifested that AS-IV significantly inhibited the activation of mitochondrial apoptotic pathway (MAP) via inducing the phosphorylation of Akt and Bad. Furthermore, phosphatidylinositol 3-kinase (PI3K) inhibitor 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002) remarkably inhibited the anti-apoptotic effect of AS-IV. Moreover, AS-IV didn't compromise the antitumor activity of DOX. Taken together, our findings indicate that AS-IV ameliorates DIC, and this beneficial effect appears to be dependent on the activation of the PI3K/Akt pathway. Thus, AS-IV may hold promise as an efficient cardioprotective agent against DIC.

Immunogenicity and protective efficacy of inactivated coxsackievirus B4 viral particles.

Coxsackievirus B4 (CVB4) is associated with a range of acute and chronic diseases such as hand, foot, and mouth disease, myocarditis, meningitis, pancreatitis, and type 1 diabetes, affecting millions of young children annually around the world. However, no vaccine is currently available for preventing CVB4 infection. Here, we report the development of inactivated viral particle vaccines for CVB4. Two types of inactivated CVB4 particles were prepared from CVB4-infected cell cultures as vaccine antigens, including F-particle (also called mature virion) consisting of VP1, VP3, VP2, and VP4 subunit proteins, and E-particle (also called empty capsid) which is made of VP1, VP3, and uncleaved VP0. Both the inactivated CVB4 F-particle and E-particle were able to potently elicit neutralizing antibodies in mice, despite slightly lower neutralizing antibody titres seen with the E-particle vaccine after the third immunization. Importantly, we demonstrated that passive transfer of either anti-F-particle or anti-E-particle sera could completely protect the recipient mice from lethal CVB4 challenge. Our study not only defines the immunogenicity and protective efficacy of inactivated CVB4 F-particle and E-particle but also reveals the central role of neutralizing antibodies in anti-CVB4 protective immunity, thus providing important information that may accelerate the development of inactivated CVB4 vaccines.