抗生素废水批量处理的pH调控及其影响

在不同pH水平下进行了抗生素废水的批量处理,试验结束时利用扫描电镜(SEM)、荧光原位杂交-流式细胞术联用技术(FISH-FCM)分析了不同pH条件下获得的降解菌群中酵母和细菌的构成,基于Biolog方法比较了不同降解菌群的代谢多样性,并考察了降解菌群构成的变化对抗生素废水批量处理效果的影响.结果表明,批量处理过程的pH调控显著影响了降解菌群中酵母和细菌的比例,并进而影响了废水处理效果.当批量处理过程的pH值分别控制在4～5、5～6、6.5～7.5时,降解菌群中酵母构成比例分别达到88.20%、54.43%、1.75%,同时细菌的构成比例相应呈逆向变化;Biolog FF微平板分析表明,3种pH条件下的降解菌群具有类似的代谢多样性,但酵母占优势时降解菌群的代谢活性较低;随着酵母比例的下降或细菌比例的上升,批量处理的COD去除率分别为34.8%、44.8%和61.2%.     

Sanfilippo D syndrome: Correction of glucosamine-6-sulphatase deficiency following fibroblast culture in chang's media

The de-O-sulphation of α-linked glucosamine-6-sulphate residues in heparan sulphate requires a specific sulphatase, glucosamine-6-sulphatase, which has been shown to be deficient in tissues of Sanfilippo D, or mucopolysaccharidosis type IIID (MPS IIID), patients. MPS IIID fibroblasts cultured in Basal Eagle's medium supplemented with either fetal calf serum or heat-inactivated fetal calf serum, MDCB or Ultraserg media had residual glucosarnine-6-sulphatase activities towards a heparin-derived trisaccharide substrate, O-(α-N-acetylglucosamine-6-sulphate)-(1→4)-L -O-(α-iduronic acid-2-sulphate)-(1→4)-D -O-2,5-anhydro [1-³H]mannitol-6-sulphate, GlcNAc6S-IdoA2S-anM6S, which were less than 1 per cent of the normal range for fibroblasts cultured in Basal Eagle's medium supplemented with fetal calf serum. However, the glucosamine-6-sulphatase activities of MPS IIID fibroblasts grown in Chang's medium were similar to the activities in normal control fibroblasts which were cultured in Basal Eagle's medium. These results indicate that caution is required for prenatal diagnosis of MPS IIID patients using chorionic villi or amniotic cells cultured in Chang's medium.     

Lesch—Nyhan syndrome: Carrier and prenatal diagnosis

We report the results of carrier and prenatal diagnosis for hypoxanthine guanine phosphoribosyltransferase (HPRT) deficiency, Lesch—Nyhan syndrome, by carrier testing of 83 women and prenatal analysis of 26 pregnancies. Our diagnostic methodologies include mutation detection and linkage analysis for probands and their families and biochemical measurement of HPRT enzyme activity for at-risk pregnancies. Identification of the mutation in the index case of each family permits precise carrier diagnosis using polymerase chain reaction (PCR) amplification of HPRT gene sequences and automated DNA sequencing. We demonstrate 100 per cent sensitivity for the detection of mutations in the HPRT gene of affected males and highly efficient carrier testing of at-risk females. Two other molecular methods proven to have high utility include PCR-based dosage analysis and linkage analysis by PCR amplification of a short tandem repeat (STR) in intron 3 of the HPRT gene. As a result, 45 at-risk women, 56 per cent of those tested, were identified not to be carriers of their family's HPRT gene mutation. Seven of these women were the mothers of affected males and prenatal testing for future pregnancies was recommended because of the possibility of gonadal mosaicism. Thirty-eight of these women were more distant relatives of affected males, thereby eliminating the need for future prenatal procedures. These studies illustrate the utility and precision of molecular methodologies for carrier and prenatal diagnosis of Lesch—Nyhan syndrome. These studies also illustrate that molecular diagnostic studies of affected males and carrier testing prior to pregnancy can clarify genetic risk predictions and eliminate unnecessary prenatal procedures.     

Prenatal diagnosis of pseudo arylsulphatase a deficiency

Prenatal diagnosis was performed on a pregnancy at risk for metachromatic leukodystrophy (MLD) in a family with the pseudo arylsulphatase A deficiency trait. Extracts of cultured amniotic fluid cells were deficient in arylsulphatase A indicating that the fetus was either affected with MLD or had the benign pseudodeficiency trait. In the cerebroside sulphate loading test, the at risk cells hydrolysed sulphatide like control cultured amniotic fluid cells implying that the fetus had pseudodeficiency. The pregnancy was carried to term and a male child was delivered. Placenta, urine and fibroblasts had very low activities of arysulphatase A. However, no sulphatide could be detected in urine and growing fibroblasts responded normally in the cerebroside sulphate loading test, suggesting pseudodeficiency. At 29 months, the infant is healthy and shows no stigmata of MLD. The prediction based on the results of the cerebroside sulphate loading test on cultured amniotic fluid cells appeared to be borne out.     

牡蛎鳃组织的培养及TBTCl对细胞活力的影响

采用无血清组织培养和台盼蓝排斥实验,从细胞形态、细胞成活率等方面分别研究了氯化三丁基锡(TributyltinChloride,TBTCl)对牡蛎鳃组织细胞的影响。实验结果表明,TBTCl对牡蛎鳃细胞的生长及活力均产生一定的影响,具有抑制牡蛎鳃细胞增生,造成牡蛎鳃细胞损伤以及致使鳃细胞死亡的作用。     

Effects of vegetation type on soil microbial community structure and catabolic diversity assessed by polyphasic methods in North China

Soil microbes play a major role in ecological processes and are closely associated with the aboveground plant community.In order to understand the effects of vegetation type on the characteristics of soil microbial communities,the soil microbial communities were assessed by plate counts,phospholipid fatty acid (PLFA) and Biolog microplate techniques in five plant communities,i.e.,soybean field (SF),artificial turf (AT),artificial shrub (AS),natural shrub (NS),and maize field (MF) in Jinan,Shandong Province,North China.The results showed that plant diversity had little discernible effect on microbial biomass but a positive impact on the evenness of utilized substrates in Biolog microplate.Legumes could significantly enhance the number of cultural microorganisms,microbial biomass,and community catabolic diversity.Except for SF dominated by legumes,the biomass of fungi and the catabolic diversity of microbial community were higher in less disturbed soil beneath NS than in frequently disturbed soils beneath the other vegetation types.These results confirmed that high number of plant species,legumes,and natural vegetation types tend to support soil microbial communities with higher function.The present study also found a significant correlation between the number of cultured bacteria and catabolic diversity of the bacterial community.Different research methods led to varied results in this study.The combination of several approaches is recommended for accurately describing the characteristics of microbial communities in many respects.     

Mucolipidosis type IV: Accumulation of phospholipids and gangliosides in cultured amniotic cells. A tool for prenatal diagnosis

Cultured amniotic fluid cells from two mucolipidosis type IV (MLIV)-affected fetuses demonstrated accumulation of phospholipids and gangliosides when compared with normal controls. Like cultured skin fibroblasts from MLIV patients, cultured amniotic cells from the affected fetuses accumulated primarily lyso phospholipids and this could be demonstrated by radioactive labelling with appropriate precursors, either inorganic phosphate or oleic acid. Furthermore, like cultured skin fibroblasts, there was significant retention of exogenously supplied GDIA ganglioside in the affected amniotic cells. This storage was previously demonstrated to be unique to MLIV and thus can be used at present as a specific procedure for prenatal diagnosis of MLIV.     

Separation of amniotic fluid cell types in primary culture by percoll density gradient centrifugation

Amniotic fluid cells obtained by transabdominal amniocentesis at 15–17 weeks of gestation and cultured for 15 and 21 days were separated into three fractions by density gradient centrifugation in Percoll. Each fraction (or peak) corresponded to the following densities: peak A, 1·02–1·03 g/ml; peak B, 1·04–1·05; peak C, 1·05–1·06. Peak A was composed of both non-viable and viable cells; the latter adopted the morphology of epithelial cells in culture. Peak B contained a mixture of fibroblasts and epithelioid cells and peak C had only epithelioid cells. The variability of N-acetyl-β-hexosaminidase and alkaline phosphatase activities was reduced in peaks B and C as compared to that of peak A and of unseparated cells suggesting that more defined and homogeneous cell types for enzymatic determinations can be obtained by centrifugation in Percoll density gradient. In prenatal diagnosis of biochemical defects, the separation of cells would permit a more precise diagnosis by eliminating enzyme variability due to the presence of different cell types or non-viable cells.     

Prenatal diagnosis of galactosialidosis

The second prenatal diagnosis of galactosialidosis is reported. Neuraminidase and β-galactosidase activities in cultured amniotic cells were deficient, this being confirmed by skin fibroblast enzyme assay on the affected fetus after interruption of the pregnancy. Cultured placental cells demonstrated the same enzyme deficiencies. Analysis of deproteinized amniotic fluid showed the presence of abnormal oligosaccharides specific for a-neuraminidase deficiency.     

Fetal presentation of morquio disease type A

A fetus with mucopolysaccharidosis type IV A (Morquio type A) is described. The family had one affected child exhibiting symptoms of classical Morquio A disease, and late in the subsequent pregnancy prenatal diagnosis was requested. At 23 weeks' gestation, moderate ascites was detected by detailed ultrasound scan and keratan sulphate was found in the amniotic fluid. The pregnancy was terminated by prostaglandin induction and the diagnosis of mucopolysaccharidosis type IV A was confirmed by demonstration of a deficiency of N-acetylgalactosamine-6-sulphate (GalNac-6-S) sulphatase in cultured amniotic cells and in post-mortem fibroblast cultures. The activities of β-galactosidase and arylsulphatase A were normal, ruling out Morquio disease type B and multiple sulphatase deficiency. These results indicate that mucopolysaccharidosis IV A (a disease that predominantly affects the skeletal system) may produce ascites in the fetus to such an extent that it can be detected by ultrasound.     

Correct prenatal diagnosis of a hurler fetus where amniotic fluid cell cultures were of maternal origin

Contamination of amniotic fluid cell cultures by maternal cells can be expected to lead to misdiagnosis of fetal genotype in 0·1 to 0·5/100 cultures, when assays are carried out directly on cultured cells. Chemical analysis of the cell-free amniotic fluid supernatant may overcome this source of error and has the added advantages of speed and independence from amniotic cell culture failure. We describe a pregnancy at risk for Hurler's disease where amniotic cells cultured at amniocentesis had a female karyotype and an α-iduronidase activity towards both phenyl and 4-methylumbelliferyl substrates at the lower end of the normal range, suggesting a heterozygous fetus. An affected fetus was predicted, however, because of a high concentration of dermatan sulphate in the amniotic fluid. The discrepancy between these findings was shown to be due to maternal cell contamination of amniotic fluid cell cultures by the birth of a male infant with Hurler's disease.     

The distinction between arylsulphatases in chorionic villi

The relatively high activity of arylsulphatase C (ASC) in the placenta is a potential risk for the misdiagnosis of arylsulphatase A (ASA) or arylsulphatase B (ASB) deficiency in chorionic villus sampling when assayed by synthetic substrates. A clear distinction between these enzymes can be achieved in either the direct villi or the cultured villi cells. Interestingly, the activity of ASC differed significantly in cultured villi cells when prepared by two different methods, namely, minced villi versus treatment with trypsin and collagenase, while ASA and ASB were not affected by these treatments. Whether ASC was directly affected by one of these treatments or whether a selection of cells with different ASC levels was achieved is not yet clear, but this phenomenon clearly indicates the importance of precise definition of CVS preparations to correlate with the enzyme activity data.     

Trisomy 20 mosaicism in amniotic fluid cells

The significance of trisomy 20 mosaicism in cultured amniotic fluid cells is still confusing. We report a case of amniotic cell normal/trisomy 20 mosaicism diagnosed prenatally. The pregnancy was carried to term and a normal baby girl was delivered. The authors consider that in cases of amniotic fluid cell normal/trisomy 20 mosaicism the termination of pregnancy may not be advised, however, the parents should be fully informed.     

Prenatal diagnosis of Chediak-Higashi syndrome

We report the first prenatal diagnosis of an affected fetus with Chediak-Higashi syndrome (CHS). Diagnosis was accomplished via fetal blood sampling at 17 menstrual weeks and was confirmed after birth. Retrospective measurement of the largest acid phosphatase-positive lysosomes in cultured amniotic fluid cells and chorionic villus cells showed that in CHS these lysosomes are significantly larger than those in normal cells. This method may be used for prenatal diagnosis of CHS by amniocentesis and chorionic villus sampling (CVS).     

Prenatal diagnosis of inborn errors in peroxisomal β-oxidation

In recent years, an increasing number of inherited diseases in man have been recognized in which there is an impairment in the peroxisomal β-oxidation of very-long-chain fatty acids. In general, these disorders are associated with severe neurological and physical abnormalities and death within the first years of life. In this paper we describe our experience with regard to the prenatal diagnosis of a number of different inborn errors of peroxisomal β-oxidation. Eleven pregnancies at risk were monitored by measuring very-long-chain fatty acid levels as well as very-long-chain fatty acid β-oxidation in cultured chorionic villous fibroblasts and/or amniotic fluid cells. Five affected fetuses were identified. It is concluded that prenatal diagnosis in this group of diseases can be done reliably using cultured chorionic villous fibroblasts or amniotic fluid cells.     

镉致背角无齿蚌消化腺组织损伤及调节消化腺和斧足的抗氧化酶活性

采用酶学和组织学方法,研究了镉(Cd)对背角无齿蚌(Anodonta woodiana)消化腺和斧足组织中超氧化物歧化酶(SOD)和过氧化氢酶(CAT)活性和组织形态结构的影响.Cd浓度设置为4.22、8.43、16.86、33.72和67.45 mg·L-1,处理时间为24 h、48 h、72 h和96 h.结果显示,与对照组相比,同一时间随着Cd浓度的增加,消化腺中SOD和CAT活性,在24 h呈现升高的趋势;96 h消化腺中SOD活性逐渐升高,差异极显著(p0.01);48 h和72 h呈现先升后降的趋势.斧足中SOD和CAT在72 h和96 h时活性升高明显,差异显著或极显著(p0.01,p0.05).Cd浓度67.45 mg·L-1处理24 h,消化腺组织细胞结构变化不明显;在48 h以后,消化管间结缔组织松散,腺体结构损伤严重,上皮细胞形态结构异常,细胞界限模糊,甚至消失;细胞膜肿胀、变形;有的细胞核碎裂、明显肿胀甚至消失,上皮细胞部分与基膜分离、脱落;斧足组织未出现显著变化和损伤.研究表明,消化腺在酶活性和组织结构上对镉的反应较快且更具规律性,而且具有明显的时间和剂量效应关系.但是,斧足对镉的胁迫反应与消化腺相比较为迟缓,说明组织之间对重金属的敏感性具有差异性.     

Prenatal diagnosis of hunter syndrome

Sixteen pregnancies at risk for Hunter syndrome have been monitored by amniocentesis. Iduronate 2-sulphate sulphatase levels were measured in amniotic fluid, cultured amniotic fluid cells and cord blood. Thirteen of the pregnancies resulted in normal livebirths, two are continuing and one affected pregnancy was terminated. Reduced enzyme levels were observed in either amniotic fluid, cells or cord blood for four female fetuses. Such fetuses are likely to be carriers expressing reduced enzyme levels. The affected male fetus had reduced enzyme activity in amniotic fluid; insufficient cells were cultured for enzyme estimation, however no enzyme activity was detected in fetal liver after termination. Eight cord blood enzyme estimations have been performed, five confirming normal male infants.     

Prenatal diagnosis of mucopolysaccharidosis by two-dimensional electrophoresis of amniotic fluid glycosaminoglycans

Amniotic fluid from 72 pregnancies at risk for mucopolysaccharidosis have been analysed for glycosaminoglycans (GAG) by means of two-dimensional electrophoresis. Definite results were obtained in all cases and required only 3 days to report. Of the 66 cases for which GAG analysis was accompanied by enzyme assays on cultured amniotic fluid cells, there was agreement of results in 65. In the one case of disagreement the result of GAG analysis proved to be correct. Fourteen pregnancies were predicted to be affected and the predictions were confirmed by studies on fetal tissues and/or cultured cells, or in one case the birth of an affected infant. Of the 58 pregnancies predicted to be unaffected, 48 have so far gone to term and produced healthy infants. There have been no false negative predictions. The analytical method is simple, rapid, and, in this study, has been found completely reliable for prenatal diagnosis.     

Resorbed co-twin as an explanation for discrepant chorionic villus results: Non-mosaic 47,XX, +16 in villi (direct and culture) with normal (46,XX) amniotic fluid and neonatal blood

Non-mosaic trisomy 16 was observed in chorionic villus cytotrophoblasts (direct) as well as cultured mesenchymal core cells derived from the pregnancy of a 38-year-old woman. Chromosome preparations from amniotic fluid and neonatal cultures (cord blood) were 46,XX. Normal fetal growth as determined by serial ultrasound examinations occurred throughout the pregnancy, which resulted in a healthy 2724 g female. Multiple biopsies taken from the umbilical cord, placental cotyledons, and fetal membranes were 46,XX. However, a placental nodule and three of six cultures initiated from membranes (amnion and chorion) showed 46,XX/47,XX, + 16 mosaicism. We propose that the trisomy 16 cells arose from residual villi derived from a trisomic cotwin that never developed. This case further demonstrates that normal fetal growth may presage normal outcome irrespective of cytogenetic findings in cytotrophoblasts (direct) and cultured mesenchymal core cells.