四种结核分枝杆菌检测方法的临床应用评价

目的评估涂片、培养、PCR和增菌PCR检测结核分枝杆菌临床应用价值。方法对124例临床确诊的肺结核、可疑结核患者和非结核病人痰标本的涂片、培养、PCR和增菌PCR四种方法的检测结果进行比较。结果涂片、培养、PCR和4及7d的增菌PCR检测31例临床确诊的肺结核病人痰标本阳性率分别为22.5%、32.2%、54.8%、64.5%和87.1%;检测59例临床可疑肺结核病人痰标本阳性率分别为13.6%、18.6%、28.8%、37.3%和52.5%。比较四种方法的阳性检测率有显著性差异(P<0.05)。检测34例非结核病人痰标本,涂片、培养均为阴性,而PCR和增菌PCR均有1例假阳性,假阳性率2.9%。比较PCR与增菌PCR对菌阳和菌阴病人的痰标本阳性检测率,有显著性差异(P<0.05),而两种方法的假阳性率相同。结论增菌PCR检测结核分枝杆菌具有很高的敏感性和特异性,可作为结核病的有效辅助诊断方法之一。     

聚合酶链反应技术检测咽拭子标本中结核菌DNA的评价

为探讨聚合酶链反应（PCR）检测咽拭子标本中结核菌在诊断肺部疾病的价值。应用PCR法检测肺结核患者50例，非肺结核患者55例及100例正常健康人咽拭子标本中结核菌和咽拭子涂片找抗酸杆菌，并与痰涂片检查作比较。肺结核组、非肺结核组及正常健康人咽拭子PCR阳性率分别为60％、3O．9％及10％，涂片为6％、0％和0％。结核组及非结核组痰PCR阳，牲率为54％和10．9％，涂片为26％和0％。结果表明PCR检测咽拭子标本结核菌DNA是肺结核病早期诊断和鉴别诊断的一种有价值的检测手段。但应注意假阳性或假阴性出现的可能。     

建立增菌PCR方法对痰涂片阴性肺结核早期诊断

目的 建立增菌PCR方法，以达到对涂片阴性痰标本中结核菌早期检定和确认活菌的目的。方法 采用7H12半流体培养和PCR扩增相结合的增菌PCR方法，对46份初治肺结核患者痰标本进行检测，并对杂交结果进行凝胶呈像灰度分析。结果 应用增菌PCR检测涂片阴性痰标本，其中86.21%的标本在增菌7d内得到阳性结果，于4周时得到最大阳性率，较培养法的阳性检出时间明显缩短（P＜0.01)，较普通痰标本PCR阳性率明显提高（P＜0．05）；有63．04％的标本增菌7d时的杂交结果凝胶灰度值与0d时相比＞1：并有62.07%的增菌PCR7d内得到阳性的标本得到同一样本7H12半流培养阳性结果的支持和活菌判定。结论 增菌PCR不是单纯的PCR，增菌7d后PCR阳性率明显增加，且有杂交结果凝胶灰比值的增加和随后的培养结果对其进行支持和活菌判断，会有助于临床诊断的准确性。     

痰结核分枝杆菌DNA检测诊断肺结核的临床意义

目的评估痰结核分枝杆菌DNA检测诊断肺结核的可行性及临床意义。方法回顾性分析2016年5月—12月在我科住院的经痰结核分枝杆菌培养确诊的136例肺结核患者的临床资料,比较痰涂片找抗酸杆菌和痰结核分枝杆菌DNA检测2种方法的检出率。结果 136例确诊的肺结核患者中,痰涂片阳性的有64例(47.06%),痰结核分枝杆菌DNA检测阳性的88例(64.71%),痰结核分枝杆菌DNA检测阳性率明显高于痰涂片找抗酸杆菌(P0.05)。72例痰涂片阴性患者中痰结核分枝杆菌DNA检测阳性29例,占痰涂片阴性肺结核患者比例的40.28%。结论痰结核分枝杆菌DNA检测优于痰涂片找抗酸杆菌,能够辅助涂阳肺结核的诊断,并提高涂阴肺结核的诊断水平,是一种快速、敏感的病原学诊断方法。     

套式PCR及测序方法用于痰标本中结核分枝杆菌rpoB耐药基因突变检测

目的应用套式PCR—DNA测序方法直接检测痰标本中结核分枝杆菌相关的rpoB基因突变，以期建立一种直接检测分枝杆菌耐利福平的快速方法，并评价其临床应用价值。方法采用套武PCR—DNA测序方法直接检测112例活动性肺结核患者和20例非结核性肺部疾病患者痰、标本中结核分枝杆菌rpoB基因突变情况。同份痰标本同时做涂片抗酸染色，罗氏培养及菌型鉴定。结果112例活动性肺结核患者痰标本套式PCR扩增87例呈阳性，产物DNA测序31例有rpoB基因突变。其中分离出耐利福平株的32例痰中29例发生了基因突变，耐药突变率90．6％（29／32），39例菌阴（涂阴培阴）痰中有2例发生突变。分离出对利福平敏感株的37例痰中未发生突变，20例非结核性肺部疾病患者痰标本套式PCR扩增均为阴性，特异性100％。结论套式PCR—DNA测序可望为直接检测临床痰标本中结核分枝杆菌耐利福平的准确、特异、快速的方法。     

纤支镜在菌阴肺结核中的应用评估

目的 观察纤支镜在菌阴肺结核诊断中的价值.方法 分析190例入院时痰涂片和PCR分析均为阴性疑诊为肺结核患者的病例资料,从经纤支镜抗酸杆菌涂片的阳性率、PCR检测结核分枝杆菌阳性率、上皮性肉芽肿支气管活检、结核分枝杆菌的培养阳性率等四个方面进行分析.结果 190例病例中,经纤支镜取样的阳性率:42.6％(痰涂片),63.6％(PCR分析),31.2％(肉芽肿支气管活检),54.2％(痰培养),将各种检查手段联合起来,诊断率可以达到85.2％.结论 经纤支镜取样有较高阳性率,可以提供快速明确的结核诊断.     

探针熔解曲线法联合交叉引物扩增技术快速检测耐药肺结核的研究

目的评价PCR熔解曲线法联合交叉引物扩增技术(CPA)快速检测耐药肺结核的临床价值。方法 2018年1月-2019年5月济宁市传染病医院结核门诊初诊疑似肺结核患者310例,采集痰标本,涂片,做萋苨氏染色、CPA检测、分枝杆菌BACTEC-MGIT 960快速分离培养及熔解曲线耐药检测,对培养阳性菌株进行菌型鉴定和比例法药敏试验。结果 310例患者痰标本涂片检查分枝杆菌阳性61例,阳性率19.7%(61/310),敏感度和特异度分别为62.2%和95.6%;痰培养分枝杆菌阳性141例,阳性率45.5%(141/310)。菌型鉴定为结核分枝杆菌(MTB)感染132例,非结核分枝杆菌(NTM)感染9例;CPA检测阳性148例,阳性率47.7%,方法的敏感度和特异度分别为94.2%和90.1%。以比例法药敏试验为金标准对132例患者的MTB分离菌株或CPA检测为阳性的痰标本进行探针熔解曲线耐药检测,RFP、INH、EMB、SM等4种抗结核药物检测的敏感度分别为96.82%、92.06%、86.79%和83.93%,特异度分别为95.65%、94.20%、78.48%和80.26%,诊断率分别为96.21%、93.18%、81.81%和81.81%。结论 PCR熔解曲线法和CPA检测技术操作简单、稳定、快速,特异度和敏感度较高,可提高MTB的检出率,两种方法联合使用可快速筛查患者MTB的耐药性。     

2月末结核分枝杆菌阳性初治肺结核患者菌型鉴定结果分析

目的分析深圳市南山区2月末结核分枝杆菌阳性肺结核患者的分枝杆菌分布情况,为诊断与治疗分枝杆菌感染提供参考依据。方法 2013年1月至2015年12月深圳市南山区痰结核分枝杆菌阳性的初治肺结核患者治疗2月末仍结核分枝杆菌阳性患者进行菌型鉴定并进行结果分析。结果 617例结核分枝杆菌阳性肺结核患者经2个月强化期治疗后,46例患者出现2月末结核菌阳性,其中涂片阳性为33例,涂片阴性而培养阳性为13例;涂片阳性患者中19例培养阴性,10例为结核药物敏感菌,4例为非结核分枝杆菌;涂阴培阳患者中1例为结核药物敏感菌,12例为非结核分枝杆菌;16例非结核分枝杆菌经分型鉴定后,7例脓肿分枝杆菌,9例戈登分枝杆菌。结论肺结核患者可能继发非结核分枝杆菌感染,抗结核治疗2月末肺结核病人应及时留取痰标本做结核菌涂片,培养和菌种鉴定,为进一步诊断和制定个体化疗方案提供依据。     

应用蛋白芯片技术快速诊断肺结核病的研究

目的评价结核分枝杆菌蛋白芯片在检测临床标本中结核分枝杆菌抗体的应用价值。方法应用结核分枝杆菌蛋白芯片检测117例结核病患者的血清标本、103例肺部其他疾病患者和健康入的血清标本，并与痰涂片镜检法相比较。结果在菌阳肺结核、菌阴肺结核中，结核分枝杆菌抗体芯片检测的阳性率分别是100％（26／26）、49．6％（45／91），在肺部其他疾病患者和健康人中，结核分枝杆菌抗体芯片检测的阴性率是98．1％（101／103）。结论结核分枝杆菌抗体检测蛋白芯片是一种集基因工程技术、芯片技术和免疫学技术的检测一体化的新型结核病快速诊断方法，具有简便、快速、大量、敏感性高和特异性强、检测成本较低的特点，是结核病、尤其是菌阴结核病辅助诊断的有效方法。     

肺结核病的快速实验室诊断

检出痰中含有结核杆菌是临床诊断肺结核或鉴别肺结核与其他肺病的重要依据。痰中结核分枝杆菌（Mycobacterium tuberculosis，Mtb）的检测方法常规采用痰涂片及痰培养。涂片虽简单易行，但阳性率低，培养虽为金标准，但周期太长，均难以满足临床需要。近年来，作为直接检测分枝杆菌、种系鉴定和药敏试验的许多分子学方法得到长足发展，这些方法能将诊断时间从几周大大缩短到几天、几小时。其中，实时动态荧光定量PCR（FQ-PCR）因其技术成熟具有较高的灵敏度和特异性，已逐渐应用于临床。为明确使用FQ—PCR技术检测患者痰液、血液标本时在诊断肺结核病中的意义，我们对102例肺结核患者的痰和外周血标本进行MBDNA检测．其结果报道如下。     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

Microbiology of human immunodeficiency virus anorectal disease

PURPOSE: Individuals who are seropositive for the human immunodeficiency virus are at high risk for opportunistic infection and anorectal disorders. Little prospective information is available regarding anorectal pathogens in these patients. METHODS: One hundred sixty-three HIV-seropositive patients presented to the colorectal clinic between 1989 and 1992. Forty-seven (29 percent) patients were thought to have an infectious process and were prospectively studied using a standardized multiculture protocol. RESULTS: Mean age was 33 (range, 19–59) years. All were male; high-risk behavior accounted for 87 percent of HIV transmissions. Presenting complaints included anorectal pain (79 percent), pus per anum (28 percent), and blood per anum (26 percent). Examination revealed perianal tenderness (60 percent), condyloma (38 percent), perianal ulcers (38 percent), and anal fissures (34 percent). Sixty-six sets of cultures were performed; 28 patients had one set, 15 had two sets, and 4 had three sets. Thirty-two of these 47 patients (68 percent) had positive cultures including herpes (50 percent), cytomegalovirus (25 percent),Neisseria gonorrhoeae (16 percent), chlamydia (16 percent), acidfast bacilli (2 percent), and others (9 percent). Six of 32 patients with positive cultures had more than one organism cultured. Sixteen (50 percent) patients with positive cultures were treated medically, 8 (25 percent) were treated surgically and 8 (25 percent) were treated with both modalities. Sixty-one procedures were performed on 17 patients for condylomata. Eighteen patients had 20 procedures for abscesses, 50 percent of whom had positive cultures for other than common bowel flora; all improved. Fourteen patients underwent 33 procedures for perianal fistulas.Mycobacterium fortuitum was cultured from one patient who required 13 procedures for abscesses and fistulas. Forty-five (96 percent) patients were followed for an average of 12.5 months ±2.9 SEM (range, 1–94 months). Symptoms were improved or resolved in 22 of 32 (69 percent) patients with positive cultures and in 11 of 13 (84 percent) with negative cultures. CONCLUSIONS: Specific pathogens may often be identified in human immunodeficiency virus-seropositive patients with anorectal disorders if aggressively sought. Although patients without specific pathogens identified may be expected to improve with planned empiric treatment, positive identification allows more directed therapy.