基于ttr基因的mini-MPN-qLAMP法快速定量检测食品中沙门氏菌

该研究通过3管型微量MPN计数法(Mini Most Probable Number,mini-MPN)建立了一种快速定量检测食源性沙门氏菌的荧光定量环介导等温扩增(Quantitative Loop-Mediated Isothermal Amplification,qLAMP)方法。依据沙门菌属ttr基因设计了qLAMP和荧光定量PCR(Quantitative Polymerase Chain Reaction,qPCR)引物,结合5 h BPW增菌和MPN计数法建立了mini-MPN-qLAMP沙门氏菌快速定量检测方法;使用两种人工污染样品对mini-MPN-qLAMP法进行验证,使用Bland-Altman分析比较不同检测方法检测结果的一致性。结果表明,建立的qLAMP法与qPCR法反应特异性均良好,纯培养时qLAMP法检出限为500 CFU/mL。通过Bland-Altman分析表明所建立的mini-MPN-qLAMP法在速冻乌米饭中检测结果与mini-MPN-qPCR、mini-MPN计数法、平板计数法相比均具有较高的一致性,r^(2)≥0.994,检出限为-0.44 lg MPN/mL;而在速冻鸡胸肉中该法检测结果与mini-MPN-qPCR结果一致性最佳,r^(2)=0.990,检出限为-0.64 lg MPN/mL。肉制品中腐败杂菌会影响mini-MPN计数和平板计数结果,mini-MPN-qLAMP可排除肉制品中腐败杂菌对检测结果的影响。该研究所建立的mini-MPN-qLAMP法简单易行,准确度高,可用于食品中沙门氏菌的快速定量检测。

转甲状腺素蛋白淀粉样变性多发性神经病1例诊治体会

转甲状腺素蛋白淀粉样变性多发性神经病(transthyretin amyloid polyneuropathy,ATTR-PN)是由编码转甲状腺素蛋白的TTR基因致病变异导致的一种以周围神经损害为主,呈常染色体显性遗传的多系统疾病。本文报道1例ATTR-PN,希望提高临床医生对此病的认识,减少误诊率。

A New Arg54Gly Transthyretin Gene Mutation Associated with Vitreous Amyloidosis in Chinese

Purpose:To analyse the hereditary features of a Chinese pedigree with familial vitreous amyloidosis in Liaoning Province,China,and to investigate the correlation between the clinical appearance of the disease and transthyretin(TTR)gene mutation,including the locus and type of TTR gene mutation.Methods:Five patients (10 eyes) from one Chinese family were diagnosed with vitreous amyloidosis between July 1996 and April 2009.Family members were followed up subsequently,and peripheral venous blood was obtained from 13 subjects (including 2 patients,and 11 controls without clinical signs of disease).DNA samples were extracted and 4 exons of the TTR gene were amplified by polymerase chain reaction (PCR).The gene fragments were subjected to sequencing analysis.The results were analyzed with DNAMAN Windows 5.2.2.0 and Chromas sequence chart analysis software,TTR gene exons were compared between affected patients and normal controls.Results:Family pedigree analysis revealed that patients were distributed in three generations.Male and female subjects had equal prevalence,and only one parent of affected patients had signs of disease.TTR gene exon sequencing showed that the sequence of patients was identical to that of normal individuals.No TTR gene mutations were noted in 10 unaffected family members.However,a TTR Gly-54 point mutation in the 2nd exon was detected in two patients and 1 unaffected family member (one of the patients' daughters).Vitreous samples in 4 cases (7 eyes) showed positive Congo red staining,suggesting that these family members suffered from familial vitreous amyloidosis.Conclusion:This pedigree affected with familial vitreous amyloidosis was characterized by autosomal dominant inheritance;.a TTR Gly-54 point mutation in the 2nd exon is presumed to be the cause.This Gly-54 point mutation of the TTR gene is a novel mutation in vitreous amyloidosis.

甲状腺运载蛋白Gly103Arg突变致玻璃体淀粉样变的临床、病理和分子遗传学研究

目的探讨一玻璃体淀粉样变家系的临床、病理和分子遗传学特点。方法对该家系先证者进行详细的病史调查、全身及眼部检查,排除玻璃体炎、视网膜血管炎、玻璃体出血和全身其他系统淀粉样变。采用玻璃体切除进行治疗,术中通过玻切机采集法采集玻璃体液并进行病理检查。患者在术后第1天、第7天和1个月进行门诊随访复查。另外,采集该先证者的外周静脉血5 mL,提取基因组DNA,PCR扩增TTR和APOA1基因的外显子片段,用Sanger测序寻找致病突变,并在有限家系成员中进行家系共分离分析。结果先证者发病年龄为45岁,呈慢性进展性病程,表现为无痛性渐进性视力下降,无眼外其他系统受累的表现。患者术前的视力是指数/50 cm,术后1个月随访视力为1.0。玻璃体病理活检镜下见一些红染的、不定形的变性物质,刚果红染色阳性,病理诊断为右眼内淀粉样变性。对TTR和APOA1基因进行测序检测,发现先证者携带TTR基因的一个已知致病杂合错义突变,c.307G>C,p.Gly103Arg。另一个患病家系成员中也携带同一突变,证实该突变符合家系共分离。未发现APOA1基因的可疑致病突变。结论TTR基因Gly103Arg突变可导致以玻璃体淀粉样变为主要表现的家族性淀粉样多发性神经病,玻璃体病理活检有助于诊断,玻璃体切除可能是疗效较好的一种治疗方法。