Quantitative detection of Streptococcus mutans and bacteria of dental caries and no caries groups in permanent teeth from a north China population / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Streptococcus mutans (S. mutans) is the prime pathogen of dental caries. There are few reports that studied the relationship between S. mutans, bacteria and dental caries in permanent teeth when compared to those in primary teeth. This study aimed to detect S. mutans and bacteria of dental caries and non-caries groups in permanent teeth from a north China population by real-time polymerase chain reaction (PCR) and compare the relationship between the number of these bacteria and the prevalence of dental caries in permanent teeth.</p><p><b>METHODS</b>Human saliva samples were collected from 142 subjects with permanent teeth. According to their dental tooth (DT), 142 subjects were divided into a dental caries group (DT ≥ 1) and a non-caries group (DT = 0). With specific primers for S. mutans and 16S rRNA, the total number of S. mutans and total bacteria of 142 saliva samples were detected by real-time PCR and statistically analyzed.</p><p><b>RESULTS</b>There was no significant difference between the detection rates of S. mutans (P = 0.118) and medians of S. mutans (P = 0.115). The ratio of S. mutans to total bacteria in people with dental caries was significantly higher than in those without caries (P < 0.001), but the total number of bacteria in people with dental caries was significantly lower than in those without caries (P < 0.001).</p><p><b>CONCLUSIONS</b>S. mutans had different effects on caries in the permanent teeth of several individuals from a north China population. The ratios of S. mutans to total bacteria in saliva detected by real-time PCR with Sm479F/R and 16S RNA primers were closely associated with the prevalence of dental caries in the same population. These assays may be useful for the assessment of an individual's risk of dental caries.</p>

A family associated outbreak of Mycoplasma pneumoniae infection in a hospital ward / 中华流行病学杂志

Objective To describe the epidemiological and serological features on a family associated outbreak caused by Mycoplasma pneumoniae (MP) infection occurred in Beijing in August 2007.Methods Mutual exposure of the family members was investigated and retrospective medical record was reviewed for the hospitalized patients.Serum antibodies to MP were measured and chest X-rays were taken for all the family members.Results This family consisted of 5 members,with fixed members as the boy (13 years old ),his father (43 years old) and mother (44 years old),grandmother (64 years old) and uncle (32 years old ) who was involved in taking care of the sick boy and his father.During 23 days of the event,four of all the five family members were ill.Three (boy,father and uncle) had radiographic pneumonia,whose paired sera all showed a ≥ fourfold increase in antibody titer,and two of them were confnrmed by chest X-ray on day 2 after onset of fever.The grandmother suffered from bronchitis,with positive(PA) serum antibody to MR Serum MP-IgG from the father and uncle was positive,3 days and 2 days after the onset of fever.The chances of contact between grandmother with the boy and uncle with the father were both only in the hospital wards.Only the mother remained asymptomatic,with her serum MP-IgM (-)and MP-IgG ( + )for which the blood sample was collected 37 days after close contact with the boy.The longest time of exposure to the patients was between mother and the boy but only the mother did not increase her total workload or feeling for fatigue.Conclusion Results of MP-IgG from post-infection did not completely defend against the repeated MP infection.Combined risk factors as index patients with severe cough,prolonged close contact,poorly ventilation of the environment,and family members with excessive fatigue might work as the causes of this family MP outbreak.

Quantitative detection of Streptococcus mutans in different people with dental caries by real-time polymerase chain reaction / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To establish a quantity detection method of Streptococcus mutans (Sm) and bacteria and compare the relationship between the number of these bacteria and the prevalence of dental caries in different people.</p><p><b>METHODS</b>With specific primers for a unique sequence in a 14 kb HaeIII restriction fragment consistently presenting during detecting Sm by chromosomal DNA fingerprints, the total number of Sm and bacteria of 99 saliva samples were detected by real-time polymerase chain reaction (PCR) and statistically analyzed.</p><p><b>RESULTS</b>The primers were specific for Sm and the minimum detectable level by real-time PCR was 0.1 microg/L. The total number of bacteria in the dental caries and people without caries was 51.4 x 10(8) cell copies/L and 221.6 x 10(8) cell copies/L respectively, in which the ratio of Sm to bacteria was 0.0193 and 0.0059 respectively. The differences were significantly different between the people with dental caries and those without caries in the total number of bacteria and the ratio of Sm to bacteria.</p><p><b>CONCLUSIONS</b>The primers can be used to detect the Sm by real-time PCR. The ratio of Sm to bacteria was closely associated with the prevalence of dental caries.</p>

A study on the (CA)n in FVIII gene in Han ethnic group in Guangxi Zhuang Autonomous Region by amplification polymorphisms combined with silver staining / 中华儿科杂志

<p><b>OBJECTIVE</b>Hemophilia A is an inherited bleeding disorder caused by defects in factor VIII (FVIII) gene. In the present study, the frequencies of the microsatellite alleles at introns 13 and 22 in the factor VIII gene were analyzed in the group of Han nationality in Guangxi Zhuang Autonomous Region to explore their diagnostic value for hemophilia A. These two sites were also used to detect the carriers in 13 hemophilia A families.</p><p><b>METHODS</b>Ninty-one individuals of Han ethnic group in Guangxi Zhuang Autonomous Region (135 X chromosomes) and 13 HA families were subjected to molecular studies. First, these two fragments were PCR amplified simultaneously. Then, silver staining was used later to show their polymorphisms. The investigators selected one sample at random to obtain its lengths of the PCR products at these two sites by ABI310 PCR amplifier. After counting its repeated numbers of (CA) according to the documents concerned, the repeated numbers of the other samples could be counted easily.</p><p><b>RESULTS</b>In the 91 individuals, 6 and 4 alleles were detected at these two sites, respectively. At intron 13 the allele frequencies ranged from 0.0002 to 0.5408 and polymorphism information content (PIC) was 0.5899. At intron 22 the allele frequencies ranged from 0.0444 to 0.4963 and its PIC was 0.5359. The actual heterozygosity for intron 13 and intron 22 were 0.6364 (28/44) and 0.5227 (23/44), respectively. In 13 hemophilia A families with positive history, 9 of them were diagnosed by this method and the diagnosis rate was 69%.</p><p><b>CONCLUSION</b>With high PICs, (CA)n at intron 13 and intron 22 were two valuable sites in the diagnosis of hemophilia A in the population of Han ethnic group in Guangxi Zhuang Autonomous Region. Compared with some other HA restrictive fragment length polymorphisms (RFLP), intron 22 (GT)n (AG)n was more informative.</p>

Detection of alpha thalassemia using real-time PCR and dissociation curve analysis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish an automatic, high throughput, quick detection method of alpha thalassemia.</p><p><b>METHODS</b>The genotypes of -alpha(4.2) and -alpha(3.7) were detected by two real-time fluorescence PCRs using SYBR-Green1 (SYBR-PCR) with the analysis of dissociation curve (DC) and melting temperature (Tm). The PCR products were recombined into T-vector and the correct cloning was selected as positive control. Sensitivities were gained from a serious dilution of the recombinant as SYBR-PCR template, from which detection deadlines for two kinds of alleles could be determined. Totally 110 samples were detected by this technique.</p><p><b>RESULTS</b>The length of product of -alpha(4.2) and -alpha(3.7) were 1.65 kb and 1.9 kb respectively, and the Tm were (81.5+/-0.5) degrees C and (82.5+/-0.5) degrees C respectively. The detection deadline were 9 x 10(2 ) copies and 4.3 x 10(2) copies respectively. The sensitivity of the technique was much higher than that of regular PCR plus gel electrophoresis method, and the detection results of the technique were the same as that of multiplex PCR.</p><p><b>CONCLUSION</b>The genotypes of -alpha(4.2), -alpha(3.7), non-deletion alpha alpha/and --(SEA) can be sensitively, exactly diagnosed by the real-time PCR with SYBR-Green1 combined with the dissociation curve analysis. The assay is automatic, low-cost, high throughput, and easy for quality control without fluorescence probe.</p>

Improvement of gene analysis method in hemophilia A and its application of prenatal diagnosis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish a simple and rapid system for carrier detection and prenatal diagnosis in hemophilia A (CHA) family.</p><p><b>METHODS</b>Long distance-polymerase chain reaction (LD-PCR) was selected for detection factor VIII intron 22 inversion. Polymorphism of factor VIII intragenic restriction fragment length polymorphism (RFLP) of Xba I and Hin d III, short tandem repeat (STR) within intron 13 and 22, as well as extragenic DXS52 (ST 14) variable number of tandem repeat (VNTR) were assayed by PCR and linkage analysis.</p><p><b>RESULTS</b>Seventy-one females were diagnosed as carriers within 52 HA families. Twenty-one families were diagnosed to be factor VIII intron 22 inversion and 28 families were diagnosed by linkage analysis, whereas 3 families could not been diagnosed. Seventeen of 18 fetuses at risk were male. Ten of 17 male fetuses were shown to be affected and were subsequently aborted. Seven male fetuses were diagnosed to be not affected. One female fetus was identified to be HA carrier. One-year follow-up study demonstrated that these babies were normal and living well.</p><p><b>CONCLUSION</b>LD-PCR combined with multiple locus linkage analysis enables the direct and indirect detection of HA for carrier testing and prenatal diagnosis.</p>

High-level expression of phenylalanine ammonia-lyase in Lactococcus Lactis via synthesized sequence based on bias codons / 生物工程学报

To construct a safer and more efficient gene engineering Lactococcus Lactis for expressing phenylalaine ammonia lyase (PAL) which will be benefit for PKU therapy, pal cDNA of Parsly and synthesized sequence based on Lactococcus Lactis bias codons were recombined into two Lactococcus Lactis NICE systems. The activities of the expressed PAL were detected, and the effect of Lactococcus Lactis bias codons on the expression of exterior protein was analyzed. The results showed that the expression level of PAL was increased by using Lactococcus Lactis bias codons in both Lactococcus Lactis NICE systems. Through which several safer andmore efficient strains of the gene engineering Lactococcus Lactis were obtained.

Analysis of X ba I polymorphism of FVIII gene and its application on prenatal diagnosis for hemophilia A / 中华血液学杂志

<p><b>OBJECTIVE</b>To establish the linkage methods of X ba I polymorphisms specific for FVIII gene intron 22, and to find a rapid and simple system for haemophilia A (HA) carrier detection and prenatal diagnosis.</p><p><b>METHODS</b>A long PCR to amplify FVIII gene intron 22 followed by X ba I digestion was used to assay the gene rate and heterozygosity rate of 206 unrelated people. Detection of intron 22 inversion by long distance PCR (LD-PCR) and XbaI, BclI, Hind III, DXS52, STR polymorphism within intron 13 and 22 by hereditary linkage analysis were assays in 20 HA pedigrees.</p><p><b>RESULTS</b>The gene rate and polymorphism information contents of 206 people were 0.5475 and 0.4955 respectively, 7 of 20 HA families were diagnosed as intron 22 inversion, 6 of 13 non-inversion HA families were diagnosed by X ba I linkage analysis, 8 of 13 non-inversion HA families were diagnosed by two or more linkage analysis.</p><p><b>CONCLUSIONS</b>The improved X ba I linkage analysis is a specific and useful molecular diagnosis marker. LD-PCR and five-linkage analysis can be used in prenatal HA gene diagnosis.</p>

ELISA study on application of recombinant glycoprotein D of herpes simplex virus-1 in diagnosis of herpes simplex virus-1 infection with ELISA / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To establish a specific and sensitive serological ELISA, diagnostic method, for herpes simplex virus-1 (HSV-1) infection using yeast expressed glycoprotein D (gD) of HSV-1 as coating antigen.</p><p><b>METHODS</b>The yeast expressed products were collected at the most appropriate time and sonicated. After the optimum dilution titers of the coating antigens and sera were determined, 57 clinical sera were assayed using the recombinant gD and HSV-1-infected culture media as coating antigens respectively. The same sera were also assayed by homemade and Euroimmun ELISA kit, Germany. The results of German kit as a golden standard were compared with those of the other three methods in specificity, sensitivity and accordance rate.</p><p><b>RESULTS</b>Compared to the imported kit, the specificities of recombinant protein, HSV-I culture media and homemade kit were 57.1%, 57.1% and 100.0%, respectively, the sensitivities were 82.0%, 78.0%, 48.0% and the accordance rates were 78.9%, 75.4%, 54.4%, respectively. The results of repeated experiments of recombinant protein showed that there was no statistically significant difference between the two experiments (P > 0.05).</p><p><b>CONCLUSION</b>The ELISA with recombinant gD protein of HSV-1 as coating antigen is a specific, sensitive, quick and convenient method for diagnosis of HSV-1 infection.</p>

Inhibition effects of rhubarb ethanol extract on herpes simplex virus infection in vivo / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To know the anti-viral effects of rhubarb ethanol extract (REE) on herpes simplex virus(HSV) infection in vivo.</p><p><b>METHODS</b>BALB/c mice inoculated from tail vein with 0.15 ml of HSV (TCID50=10(3)) were injected hypodermically with REE next day. After divided into seven groups, three groups of mice were given different doses of REE respectively and the other groups as controls. Pathological sections from the liver, spleen, kidney were made at different times of postinfection, and their pathological changes were observed under microscope; the virus titers in viscera were assayed by using plaque formation technique and the rhubarb inhibitions to the infection of HSV in vivo?were observed.</p><p><b>RESULTS</b>No toxic response to mice were observed for REE injected hypodermically; no pathological changes were observed in different therapy groups of spleens. And those in livers and kidneys at medium- and high-dosed groups disappeared quickly. The effect of low-dosed group was equal to that of positive control group, acyclovir(ACV); the results of the titer tests showed that the virus decreased rapidly by using REE, especially in the medium- and high-dosed groups which were much more marked than the low-dosed group; Q test of the data showed that total mean value had statistical significance (F=49.1459, P<0.01); moreover there were statistical significance between therapy groups (ACV, DH1, DH2, DH3) and non-therapy groups (VC) (P<0.01 ) and between DH2, DH3 and DH1 (P<0.01); no statistical significance were found between DH1, DH2 or DH3 and ACV (P>0.05). Results show that as to the effect of decreasing the average of the total titer, rhubarb is as effective as ACV; furthermore, the medium- and high-dosed groups are superior to the low-dosed group.</p><p><b>CONCLUSIONS</b>REE has significant anti-viral effect on HSV in vivo; there will be a wide application foreground of it in clinical usage.</p>