Detection and Typing of Human Rotavirus in Reference to Repeated Acute Gastroenteritis in Infants

Stool specimens from infants who visited a clinic because of acute gastroenteritis were tested for the presence of human rotavirus. Among the samples obtained were specimens taken from seven patients who had visited the clinic at two different times. In six of these seven children, human rotavirus (HRV) was detected in only one of the specimens taken (i.e. during only one of the two visits). One patient was shown to have excreted HRV twice; in both cases the HRV was serotyped to be type 1. The present results indicate that the symptomatic reinfection of HRV was not a widely occurring phenomenon in the group of infants tested.     

Yeast基因下游二级结构与多聚腺苷作用信号

形成真核生物mRNA 3′末端的多聚腺苷（poly(A)）作用涉及前体mRNA下游的三个元件：效率元件（EE）、定位元件（PE）以及实际的剪切和poly(A)作用位点,实验研究提出了一些EE和PE的碱基序列组成.对180个Yeast基因下游（终止密码子后200个碱基）二级结构进行的详细分析显示,约86%的EE、89%的PE与二级结构中碱基非配对的环(发夹环、膨胀环、内环或多分支环)区或连接单链区有关.这个结果提示,反式因子对EE和PE的识别和作用在一定程度上有赖于EE和PE的二级结构特征.借助mRNA二级结构可以提高对EE和PE位点预测的准确性.     

信号肽去除的耐热碱性磷酸酯酶FD-TAP在大肠杆菌中的亚克隆和高表达

通过计算机辅助分析,发现在耐热碱性磷酸酯酶（简称ＦＤ－ＴＡＰ）的Ｎ端存在２６个氨基酸的信号肽序列。但一般信号肽切点并非是完全专一的,所以利用基因工程手段将ＦＤ－ＴＡＰ的Ｎ端分别缺失２４、２５、２６和２７个氨基酸,得到了Ｎ端分别缺失２４、２５、２６和２７个氨基酸的克隆子ｐＴＡＰＮＤ２４、ｐＴＡＰＮＤ２５、ｐＴＡＰＮＤ２６和ｐＴＡＰＮＤ２７。考虑到这样的无隆子其翻译起始区所形成的能量较低结构稳定的二组     

Rotaviruses code for two types of glycoprotein precursors

Rotaviruses are nonenveloped viruses that code for two glycoproteins: a structural glycoprotein (VP7) and a nonstructural glycoprotein (NS29). The precursor to VP7 (37K) was shown to contain a 1.5K cleavable signal sequence. The 37K precursor was authentically processed (signal sequence cleaved and the polypeptide "core" glycosylated) when synthesized in a cell-free system supplemented with dog pancreatic microsomes. Similar experiments were performed with the nonstructural glycoprotein precursor (20K); however, the 20K precursor contained an integral (noncleavable) signal sequence. Both precursors were inserted into membranes cotranslationally and both glycosylated products underwent posttranslational oligosaccharide processing. The results suggest a morphogenetic scheme for the simian rotavirus SA11.     

A组人轮状病毒全基因组克隆和基因型分析

运用基因克隆和重组技术,从一急性胃肠炎患儿样品中克隆到一株轮状病毒（RV）全基因组cDNA。核苷酸序列测定结果表明,该株RV基因组11条RNA共含有18613个核苷酸（3302bp—751bp）,编码5791个氨基酸。全基因组研究结果表明,该株RV（TB-Chen）为A组,II亚组,基因型为G2P[4]/NSP4[A]。这是首个A组人RV全基因组研究结果的报道,对研究和开发RV疫苗具有积极的意义。     

人B组轮状病毒vp7基因的克隆和序列分析

利用逆转录—聚合酶链反应(RT-PCR)技术,扩增了人B组轮状病毒WH—2vp7基因,连接到克隆载体pUCm—T,并对其基因序列进行了分析。WH—2与ADRV的同源性达98％,与印度加尔各达分离株CAL—1达92％,而与动物B组轮状病毒同源性差别较大,如与IDIR(鼠)同源性仅为58％,与WD653(牛)B组轮状病毒同源性为63％,与ATI(牛)同源性为61％。对vp7基因的二级结构分析发现其mRNA折叠形成多达18个发卡环状结构。VP7蛋白是249个氨基酸组成,分子：量为28．4kDa,含有3个潜在的N连接的糖基化位点和多个磷酰化位点,从氨基酸序列的同源性来看,WH—2与ADRV的同源性达99％,与CAL—1达95％,而IDIR仅为51％,说明了WH—2和ADRV的起源相同。此研究对了解B组轮状病毒基因的进化和变异规律具有重要意义,也将为B组轮状病毒预防性疫苗的研制提供科学的依据。     

Optimal cell density and multiplicity of infection for the propagation of human rotavirus in monkey kidney cells

The human rotavirus titer was optimal at an infection cell density of about 4–8 × 10⁴ cells cm^–2 in monkey kidney cell cultures. The highest viral titers (3.8 × 10⁷ TCID₅₀ ml^–1 and 3.7 × 10⁷ TCID₅₀ ml^–1) were obtained at an multiplicity of infection of 0.05 in well plate and T-flask, respectively.     

在中国卢龙县发现G5型人A组轮状病毒

轮状病毒是引起我国儿童重症腹泻的主要病原。按照WHO轮状病毒监测方案,对2003年间河北省卢龙县开展了以医院和社区为基础的小于5岁儿童轮状病毒腹泻的监测,发现一株新型轮状病毒。该病毒用传统分型引物(G1、G2、G3、G4)扩增不出条带,对其VP7基因全序列测定和分析后确定该毒株为G5型。这是我们在亚洲首次发现的人类G5型轮状病毒。该毒株与猪的G5型C134毒株核苷酸和氨基酸序列同源性分别为88·6%和95·4%,与非洲发现的人的G5型毒株MRC3105核苷酸和氨基酸的同源性为89·9%和94%,与巴西发现的IAL-28毒株核苷酸和氨基酸的同源性为87·2%和93·3%。系统发生树分析表明:卢龙毒株LL36755与其他已经报告的两种猪和一种人类的G5型毒株可能具有相同的起源。这是人轮状病毒G5型首次在亚洲国家发现,而且该毒株可能是由人类轮状病毒与动物轮状病毒毒株自然重组产生。     

Secondary structure of β-hydroxydecanoyl thiol ester dehydrase, a 39-kDa protein, derived from Hα, Cα, Cβ and CO signal assignments and the Chemical Shift Index: Comparison with the crystal structure

Summary Nearly complete backbone ¹H, ¹⁵N and ¹³C signal assignments are reported for -hydroxydecanoyl thiol ester dehydrase, a 39-kDa homodimer containing 342 amino acids. Although ¹⁵N relaxation data show that the protein has a rotational correlation time of 18 ns, assignments were derived from triple-resonance experiments recorded at 500 MHz and pH 6.8, without deuteration. The Chemical Shift Index, CSI, identified two long helices and numerous -strands in dehydrase. The CSI predictions are in close agreement with the secondary structure identified in the recently derived crystal structure, particularly when one takes account of the numerous bulges in the -strands. The assignment of dehydrase and a large deuterated protein [Yamazaki et al. (1994) J. Am. Chem. Soc., 116, 11655–11666] suggest that assignment of 40–60 kDa proteins is feasible. Hence, further progress in understanding the chemical shift/structure relationship could open the way to determine the structures of such large proteins.Supplementary Material is available on request, comprising Table S1 listing the spectral parameters; Table S2 listing the assignments; Fig. S1 showing the 2D ¹H–¹⁵N HSQC spectrum; Fig. S2 showing sequential NOEs, secondary shifts, H-exchange and ³J_HN data; and Fig. S3 showing plots of the H, C, CO and C Chemical Shift Indexes.To whom correspondence should be addressed.     

Detection of human rotavirus in hospitalized diarrheic children in central India

During the present study, group A human rotaviruses were detected among diarrheic children using polyacrylamide gel electrophoresis (PAGE) technique, with a typical RNA migration pattern of 4:2:3:2, suggestive of group A rotavirus. During the study, a total of 46 fecal samples collected from hospitalized children with acute diarrhea as well as children inhabiting nearby animal farms with history of presence of animal rotaviruses on the farms were processed for detection of human rotavirus. Out of 33 diarrheic children, 12 showed presence of rotavirus infection (36.36%), however, none of the children from animal farm areas showed presence of rotavirus. Female children were more susceptible to rotavirus infection (46.15%) than males (30%). Majority of the cases of rotavirus gastroenteritis belonged up to one year of the age, with an incidence of 40.91%. RNA profile of rotaviruses suggested circulation of 5 different electropherotypes in this geographical locale of the country, indicating existence of genomic diversity among human rotaviruses. Majority of the isolates were of long pattern (66.67%), whereas short pattern was detected only in one third of the viruses. This preliminary study emphasizes for further detailed studies on the molecular characterization of rotaviruses circulating in this part of country and their relationship with other human rotavirus strains and animal strains in the country.     

In Vitro and in Vivo Evaluation of the Efficacy of Bovine Colostrum against Human Rotavirus Infection

We found that skimmed and concentrated bovine late colostrum (SCBLC) obtained from normal cows at 6–7 d after parturition exhibited high potency in inhibiting replication of human rotavirus (HRV) in vitro. Furthermore, prophylactic oral administration of SCBLC once before inoculation of HRV prevented the development of diarrhea in suckling mice in vivo. SCBLC from normal cows might be useful in the prevention of HRV-induced severe gastroenteritis in immunocompromised hosts.     

Human milk oligosaccharides shorten rotavirus-induced diarrhea and modulate piglet mucosal immunity and colonic microbiota

The impact of human milk oligosaccharides (HMO) on mucosal immunity, gut microbiota and response to rotavirus (RV) infection was investigated in the piglet model. Newborn piglets were fed with formula alone (FF) or formula supplemented with 4 g l⁻¹ HMO (HMO) or a prebiotic mixture of 9:1 short-chain galactooligosaccharides (3.6 g l⁻¹) and long-chain fructooligosaccharides (0.4 g l⁻¹) (PRE) (n=19–21 per group) for 15 days. Piglets (n=7–8) in each dietary group were orally infected with porcine rotavirus (RV) OSU strain on d10, and stool consistency was assessed daily. Blood, small intestine and colonic contents were collected at day 15. Serum RV-specific antibody concentrations, intestinal histomorphology, RV non-structural protein-4 (NSP4) and cytokine mRNA expression were assessed. Colonic content pH, dry matter (DM) and short-chain fatty acid concentrations were measured. Ascending colonic microbiota was analyzed by 16S rRNA gene v1-3 region pyrosequencing. HMO- and PRE-fed groups had shorter duration of diarrhea than FF piglets. Infection changed intestinal histomorphology, increased serum RV-specific antibody response and intestinal RV NSP4 expression, and modulated ileal cytokine expression. HMO enhanced T helper type 1 (interferon-gamma) and anti-inflammatory (interleukin-10) cytokines in the ileum, while prebiotics promoted RV-specific immunoglobulin M response to the infection. RV infection and HMO supplementation altered intraluminal environment and gut microbiota. HMO increased pH and lowered DM of colonic contents and enhanced the abundance of unclassified Lachnospiraceae, which contains numerous butyrate-producing bacteria. In conclusion, HMO and prebiotics did not prevent the onset of RV infection but reduced the duration of RV-induced diarrhea in piglets, in part, by modulating colonic microbiota and immune response to RV infection.     

p53蛋白质C端的三维结构及生物功能区

利用p53 C端118个氨基酸的mRNA二级结构和Chou-Fasman蛋白质二级结构预测原则,预测p53蛋白质C端289～325为卷曲肽段,368～393段包括两段螺旋结构: α₁ 368～373, α₂ 381～388.其中三段已知的蛋白质二级结构与此mRNA二级结构单元间有准确的对应关系.与四种以多重序列联配为基础的蛋白质二级结构预测方法（准确率均为73.20%左右）相对照,预测结果基本一致.结合单体聚合区31个氨基酸晶体结构,在SGI INDIGO²工作站上构建了p53 C端108个残基的三维结构.进一步揭示了p53 C端诸多生物功能区之间的空间构象关系.     

神经网络在蛋白质二级结构预测中的应用

介绍了蛋白质二级结构预测的研究意义,讨论了用在蛋白质二级结构预测方面的神经网络设计问题,并且较详尽地评述了近些年来用神经网络方法在蛋白质二级结构预测中的主要工作进展情况,展望了蛋白质结构预测的前景。     

Modeling evolution of the bacterial regulatory signals involving secondary structure

An algorithm for modeling the evolution of the regulatory signals involving the interaction with RNA secondary structure is proposed. The algorithm implies that the species phylogenetic tree is known and is based on the assumption that the considered signals have a conserved secondary structure. The input data are the extant primary structure of a signal for all leaves of the phylogenetic tree; the algorithm computes the signal primary and secondary structures at all the nodes. Concurrently, the algorithm constructs a multiple alignment of the extant (in leaves) sites of a regulatory signal taking into account its secondary structure. The results of successful testing of the algorithm for three main types of attenuation regulation in bacteria—classic attenuation (threonine and leucine biosyntheses in Gammaproteobacteria), T-box (in Actinobacteria), and RFN-mediated (in Eubacteria) regulations—are described.