Effect of silicate on Gram staining and viability of pneumococci and other bacteria

Application of silicate solutions to living or heat-killed pneumococci and to certain "viridans" streptococci causes their conversion from a Gram-positive to a Gram-negative state. The original staining properties can be restored by suspending the silicate-treated bacteria in alkaline solutions of various salts but not by simple washing in water. Living pneumococci and the strains of streptococci whose staining properties are similarly affected are killed when suspended in silicate solutions. In other Gram-positive species silicate causes conversion to Gram negativity but restoration to positivity occurs upon washing in water. In a third group of Gram-positive organisms silicate has no effect on the Gram reaction. The viability of organisms in these two groups is unaffected by silicate under the conditions employed. No effect on staining or viability of Gram-negative bacteria has been observed. The effects of silicate on staining and viability are inhibited by nutrient broth or whole serum but not by purified serum albumin. Lecithin, choline, and other substituted ammonium compounds also inhibit the effects of silicate on pneumococci.     

中国麻风分枝杆菌种型分布与特征

目的了解中国目前传播的麻风分枝杆菌(M.leprae)菌种及单核苷酸多态性(SNPs)型分布与特征。方法对来自中国22个省市171例麻风患者皮损组织样本,采用巢式PCR扩增麻风分枝杆菌特异16S rRNA保守片段,并对PCR阳性产物直接测序,经BLAST进行序列比对;采用PCR产物限制性酶切基因多态性方法对麻风分枝杆菌进行SNP分型。结果 171例标本扩增片段与来自巴西的麻风分枝杆菌Br4923相似性达99%,均为M.leprae,未检出M.lepromatosis。在85例SNP分型标本中,SNP3型、SNP1型和SNP2型分别占78.8%(67/85)、20%(17/85)和1.2%(1/85),未检出SNP4型。130例16S rRNA序列存在C251T位碱基变异,不同临床型别标本中16S rRNA序列突变及SNP型别分布差异无统计学意义;16S rRNA基因C251T突变与麻风分枝杆菌菌株的SNP分型有一定的关联,存在突变的菌株多为SNP3型,少见SNP1型,未见SNP2型;不同地区的SNP型别分布之间差异有统计学意义,内陆地区的SNP3型菌株分布率97.1%(34/35)显著高于沿海66%(33/50)(χ~2=11.96,P0.01)。不同地区的16S rRNA序列突变率差异也有统计学意义,内陆地区的16S rRNA突变率94.8%(92/97)显著高于沿海51.4%(38/74)(χ~2=43.56,P0.01)。结论麻风分枝杆菌16S rRNA基因序列突变C251T与SNP分型有关,可提示不同基因型麻风分枝杆菌的地理分布。未发现麻风分枝杆菌M.lepromatosis菌种。     

荧光染色法与萋-尼染色法检测结核分枝杆菌的效果评价

目的比较荧光染色与抗酸染色(萋-尼染色)检测结核分枝杆菌的结果差异,比较亚甲蓝液、Haris苏木素液、高锰酸钾液作为荧光染色法复染试剂的效果。方法采集疑似结核病症状的患者痰液标本198份,分别进行萋-尼染色和荧光染色,比较分析两种方法结核分枝杆菌检出率的差异。荧光染色分别用0.3%亚甲蓝液、0.5%Haris苏木素液、0.5%高锰酸钾液作为复染剂,比较不同复染剂在荧光显微镜下镜检的效果。结果萋-尼染色分枝杆菌检出率为66.67%(132/198)。荧光染色分枝杆菌检出率为94.9%(188/198),其中,亚甲蓝复染检出率为94.95%(188/198)、苏木素复染检出率94.44%(187/198)、高锰酸钾复染检出率为94.44(187/198),与萋-尼染色比较,差异均有统计学意义(P0.05)。结论荧光染色法阳性检出率明显高于萋-尼染色法,其中0.3%亚甲蓝液是一种良好的背景淬灭复染液。     

荧光定量PCR检测结核杆菌DNA与涂片抗酸染色结果的比较

目的:探讨荧光定量PCR检测结核杆菌DNA的拷贝数与涂片抗酸染色镜检阳性程度的相关性.方法:对可疑结核病患者的124份痰样本同时进行荧光定量PCR检测和涂片抗酸染色镜检,按涂片抗酸染色镜检结果阴性(-)、可疑(±)、1+、2+、3+、4+分类,对荧光定量PCR检测结核杆菌DNA的拷贝数进行统计分析.结果:98例涂片阴性痰样本中,24例样本荧光定量PCR检测阳性,结核杆菌DNA拷贝数多为102～103数量级;26例痰涂片阳性样本中,24例样本荧光定量PCR检测阳性,2例检测阴性,结核杆菌DNA拷贝数多为103～104数量级,和涂片阳性程度基本呈正相关.结论:荧光定量PCR检测结核杆菌DNA与涂片抗酸染色镜检阳性程度有较好的相关性,可以作为临床诊疗较为可靠的实验方法.     

液基夹层集菌技术检测痰标本中抗酸杆菌对诊断肺结核的应用价值

目的 评价液基夹层集菌技术检测抗酸杆菌对肺结核诊断的应用价值.方法 对134例肺结核病人晨痰标本同时进行液基夹层集菌技术与直接痰涂片进行镜检检测抗酸杆菌,比较两种方法的阳性检出率.选取10份抗酸杆菌阳性痰标本,将处理后的消化痰液接种到罗氏培养基,37℃培养,来评价液基夹层集菌技术的安全性.结果 液基夹层集菌技术和直接痰涂片检测抗酸杆菌的阳性率分别为28.4％（38/134）和16.4％（22/134）,检测结果差异有统计学意义.安全性试验发现经消化液处理的痰标本,培养8周后未见分枝杆菌生长.结论 液基夹层集菌技术提高了肺结核病人痰标本的阳性检出率,且操作简单、安全性高,是一种很有推广价值的检测方法.     

支气管镜肺泡灌洗液改良抗酸染色法诊断菌阴性肺结核的价值探讨

目的：建立改良抗酸染色法,探讨纤维支气管镜肺泡灌洗液改良抗酸染色法对痰菌阴性肺结核患者的诊断价值。方法对50例痰菌阴性肺结核患者进行纤维支气管镜肺泡灌洗,应用传统、改良两种抗酸染色法对灌洗液进行检查,了解两种方法对肺结核诊断的阳性率。结果传统、改良两种抗酸染色法的阳性率分别为38％和82％（P ＜0．05）。结论改良抗酸染色法大大提高痰菌阴性肺结核患者的阳性诊断率,值得临床推广应用。     

标本固定时间对免疫组化染色结果的影响

目前最常用的组织标本固定剂是4%甲醛液,其甲醛醛基交联封闭抗原作用对免疫组化染色的影响是众所周知的。在适宜的固定时间内可使抗原保存完好,固定时间过短或过长则免疫组化染色减弱以至消失。本文选择23个固定时相点的人体正常胃组织标本,用7种常用抗体进行标记,试图进一步证明固定时间对免疫组化染色结果的影响,寻找适用于多数抗原表达的组织固定时间段,为提高免疫组化染色方法的准确性和稳定性提供参考依据。1材料与方法11材料外科手术切除的新鲜正常胃壁组织5例。已固定1~10年的陈旧胃壁组织9例。7种常用抗体,分别为CK(AE1/AE3)(Dak…     

Effect of alkali and alkaline earth metal species on the combustion characteristics of cattle manures

This study investigates the effects of alkali and alkaline earth metal (AAEM) species on the combustion characteristics of cattle manures (CM). Different AAEM species (K, Na, Ca, and Mg) were mixed with CM and deashing CM (D-CM) samples. The combustion characteristics of raw and char samples were compared. The effects of AAEM species on CM char were analyzed based on the structural characteristics of the char sample. Results show that K and Na exert a positive effect, and this effect varies depending on the addition amount. Ca and Mg also exhibit a positive effect, but this effect does not change with the addition amount. The positive effect of K, Na, and Ca is related to the decrease in graphitization degree and increase in specific surface area. However, the positive effect of Mg is negligible. In conclusion, CM can be mixed with fuels containing K or Na in an appropriate ratio. The amount of Ca to be mixed with fuels has no specific requirement, whereas that of Mg to be mixed with fuels should be controlled.

This study investigates the effects of alkali and alkaline earth metal (AAEM) species on the combustion characteristics of cattle manures (CM). Different AAEM species (K, Na, Ca, and Mg) were mixed with CM and deashing CM (D-CM) samples.     

基因芯片快速分枝杆菌鉴定技术与涂片抗酸染色技术的临床应用比较

目的比较基因芯片快速鉴定分枝杆菌与经典涂片抗酸染色镜检技术的临床应用效果,评估两种方法的优劣性。方法2011~2014年间以基因芯片与涂片抗酸染色两种技术对深圳市所有综合性医院临床怀疑有分枝杆菌感染的标本进行检验。χ2检验两种方法阳性率的差异性。结果共有标本2 481例,涂片抗酸染色技术检出阳性标本193例,阳性率7.8%。基因芯片技术检出阳性标本317例,阳性率12.8%。基因芯片鉴定技术比涂片抗酸染色技术阳性率高,两者间比较差异有统计学意义(P0.05)。317例阳性标本经基因芯片技术分类为263例结核分枝杆菌(MTB),27例脓肿分枝杆菌,18例胞内分枝杆菌,3例胃溃疡分枝杆菌,3例鸟分枝杆菌,1例戈登分枝杆菌,1例海分枝杆菌,1例堪萨斯分枝杆菌。结论基因芯片技术快速、准确,阳性率比涂片抗酸染色技术高。菌株的分类鉴定,有利于临床明确病源,对个体化抗分枝杆菌感染治疗有指导意义。     

Studies of in vitro activities of voriconazole and itraconazole against Aspergillus hyphae using viability staining

The minimal fungicidal concentrations (MFCs) of voriconazole and itraconazole for five clinical isolates each of Aspergillus terreus, Aspergillus fumigatus, Aspergillus flavus, and Aspergillus niger were determined by a broth macrodilution method. Conidial suspensions as inocula were compared to hyphae as inocula since the invasive form of aspergillosis is manifested by the appearance of hyphal structures. In addition, cell viability staining with the dye FUN-1 was performed to assess time-dependent damage of hyphae exposed to various concentrations of the antifungal agents. With conidial inocula the MFC ranges of voriconazole were 0.5 to 4 microg/ml and those of itraconazole were 0.25 to 2 microg/ml, whereas the MFCs (2 to >16 microg/ml) with hyphal inocula were substantially higher (P < 0.01) for both itraconazole and voriconazole. Only minor differences between the tested antifungals were observed since 16 of 20 and 17 of 20 of the isolates of Aspergillus spp. tested appeared to be killed by voriconazole and itraconazole, respectively. The results of FUN-1 viability staining correlated closely to colony counts, but various time- and dose-dependent levels of viability of hyphae were also observed. In conclusion, our study demonstrates the importance of the type of inoculum used to test antifungals and the applicability of FUN-1 staining as a rapid and sensitive method for assaying the viability of hyphae.     

Effect of Isoniazid on the Protoplasmic Viscosity in Mycobacterium tuberculosis

The effect of isoniazid on the protoplasmic viscosity in the H37Ra strain of Mycobacterium tuberculosis was determined by using electron spin resonance spectroscopy and a small spin label tempone (2,2,6,6-tetramethylpiperidone-N-oxyl radical). Isoniazid (0.5 μg/ml) caused the internal cellular viscosity to increase gradually over the first 15 h of exposure from a rotational correlation time value (T_c) of 2.4 × 10⁻¹⁰ to 3.4 × 10¹⁰ s and then decrease linearly to the control level after 27 h. These results could be interpreted to mean that isoniazid allows a continued and normal synthesis of the protoplasmic components while the rate of increase in the cell volume is reduced. A degradative process may begin after the initial 15-h exposure time, which would cause the reduction in the internal viscosity.     

A rapid silver staining method for identification of Mycobacterium leprae in histologic sections

A few methods have been reported for the purpose of staining Mycobacterium leprae in paraffin sections, including Fite oil fuchsin method, auramine-rhodamine method, and Blanco-Fite silver method. Among these staining techniques, Fite oil fuchsin method and auramine-rhodamine method are popular. However, the Blanco-Fite silver method takes approximately 20 days. Therefore, we developed a new procedure for rapid identification of M. leprae in paraffin sections using another silver solution and found that the procedure gave stable and satisfactory results. This new method has proved superior to others in demonstrating a reliable staining for M. leprae.     

15082例抗酸染色镜检结果分析

目的：通过对15082例抗酸染色病例的分析,研究结核病在男女性别之间以及不同的标本种类之间的发病情况。方法统计2010年1月至2013年9月做抗酸染色检查的全部病例,比较不同时间、性别因素及不同年龄段结核病的好发情况。结果随着时间的增加,结核病的发病率越来越高;不同标本种类之间穿刺液的阳性率是最高的;男女性别之间的差异对于结核病也是不同的;结核病的好发年龄段均处于青年人群。结论结核病的发病率逐年增加,且穿刺液的阳性率比痰液、脑脊液的都高;女性比男性容易患肺结核;肺结核的发病趋于年轻化。     

pH值对硼砂甲苯胺蓝染色的影响

目的探讨不同pH值的缓冲液对硼砂甲苯胺蓝染色的影响。方法将血涂片分为两组,第1组用硼砂甲苯胺蓝染色10min,第2组采用瑞氏染色10min,为对照组,每组由9张血涂片组成,每张血涂片染色时采用不同pH值的缓冲液,观察染色效果。结果第1组与单纯用硼砂甲苯胺蓝染色相比,血涂片加用缓冲液后再用硼砂甲苯胺蓝染色效果更好,细胞结构清晰,随缓冲液pH值增高,细胞胞质与细胞核着色逐渐加深,当pH值大于9时,核浆对比不再清晰。pH值为4～11之间,淋巴细胞核仁均清晰可见,pH越低核仁越清晰。第1组成熟淋巴细胞核仁立体感强,第2组成熟淋巴细胞核仁不清晰,无法与异染色质相鉴别。结论硼砂甲苯胺蓝染色效果与缓冲液的pH值密切相关,硼砂甲苯胺蓝染色可使部分成熟淋巴细胞核仁显色。     

Effect of Ethambutol on the Viable Cell Count in Mycobacterium smegmatis

Soon after a strain of Mycobacterium smegmatis was exposed to ethambutol (EMB), the number of viable cells increased dramatically above the number in a drug-free control. This rapid rise did not occur when the culture was maintained at 4°C instead of 37°C, when an EMB-resistant mutant was used, when auxotrophs were exposed in medium lacking nutrients essential for growth, nor when the levo form of EMB was used. EMB caused no increase in deoxyribonucleic acid synthesis, nor in septum formation of dividing cells. Treated cells changed morphologically, resulting in a lower surface area-to-volume ratio. Whereas EMB did not eliminate cell clusters, the cluster size decreased markedly as detected by filtration and Coulter counter measurements. We concluded that EMB causes a reduced surface-to-volume ratio, leading to reduced cell cohesion and a consequent reduction in cluster size, reflected in an increase in colony-forming units.     

Killing activity of micafungin against Aspergillus fumigatus hyphae assessed by specific fluorescent staining for cell viability

We studied the anti-Aspergillus activity of micafungin by using two fluorescent dyes to detect cell viability. Micafungin induced flattened hyphae, caused by the bursting of cells, which had lost their viability. Micafungin has killing activity against actively growing hyphae, even though it is not fungicidal against the whole burden of Aspergillus fumigatus.