发热伴血小板减少综合征病毒热稳定性和灭活条件初步研究

目的 阐明新发危害严重的发热伴血小板减少综合征病毒的稳定性和理化灭活条件.方法 评估细胞培养制备的SFTS病毒在不同温度下的热稳定性,对紫外线、酸性环境、常用消毒剂、有机溶剂敏感性.处理后病毒感染Vero细胞,用间接免疫荧光法确定病毒复制,并采用基于病毒核蛋白的双抗体夹心ELISA方法滴定病毒与无相应处理的对照组,比较分析各种理化条件对病毒感染性的影响.结果 SFTS病毒在37℃能够存活较短时间,感染性下降较快,在4℃能保持相对稳定,1周内感染性无明显下降.对热敏感,60℃30 min能够完全灭活病毒.对紫外线敏感,185 μW/cm2紫外线照射30 min可灭活病毒.对乙醚、氯仿等有机溶剂,β-丙内酯、甲醛和常用有机氯消毒剂敏感,在合适的浓度下可在较短时间内有效灭活病毒,400 mg/L的有效氯灭活病毒需室温放置10 min以上,在pH3.0条件对病毒活力有损害,但不能完全灭活病毒.结论 研究结果初步客观的评价了SFTSV热稳定性和灭活条件,为科学研究和疾病控制中样本采集、病毒灭活、安全防护等工作提供了科学依据.     

次氯酸钠溶液和钴-60辐照对脱细胞结膜基质中病毒灭活效果的研究

目的验证猪源脱细胞结膜基质中病毒灭活工艺的灭活效果。方法用不同浓度次氯酸钠溶液对猪睾丸(ST)细胞和牛肾(MDBK)细胞行细胞毒性实验;选择猪眼结膜、脱细胞结膜基质各3个批号,用ST细胞和MDBK细胞进行次氯酸钠灭活处理和钴-60(25 kGy)辐照处理后的样品/终止产物的细胞毒性实验;以猪细小病毒(PPV)和牛病毒性腹泻病毒(BVDV)为指示病毒,采用细胞病变法分别测定次氯酸钠工艺和钴-60(25 kGy)辐照工艺对猪脱细胞结膜基质(中间品)病毒灭活效果,并进行细胞盲传实验。结果次氯酸钠溶液稀释至质量浓度100.00 mg/L(1.0‰)时,ST细胞增殖率大于70%;质量浓度50.00 mg/L(0.5‰)次氯酸钠溶液的MDBK细胞增殖率大于70%。1.0‰次氯酸钠溶液处理的猪眼结膜样品在10倍体积稀释后,对ST细胞和MDBK细胞均无抑制作用。脱细胞结膜基质辐照处理,经100倍体积稀释后对MDBK细胞无毒性,经10倍体积稀释后对ST细胞无毒性影响。3个批号猪眼结膜样品经过次氯酸钠溶液处理30 min后,PPV和BVDV的灭活平均降低系数分别为≥4.542 logs和≥4.333 logs;负载PPV病毒样品的盲传3代为阳性,即仍能检测到PPV。3个批号脱细胞结膜基质样品经钴-60辐照后,PPV和BVDV的灭活平均降低系数分别为≥4.792 logs和≥3.667 logs,负载BVDV的3个批号样品盲传结果为阴性,负载PPV病毒的3个批号样品盲传结果为阳性。结论用次氯酸钠溶液和钴-60辐照两个工艺分别处理脱细胞结膜基质(中间品)后的病毒灭活降低系数累加为:BVDV总降低系数大于8 logs,PPV总降低系数大于9 logs。两个工艺联合可以有效灭活猪源脱细胞结膜基质中污染的PPV和BVDV。     

以腺病毒为模式分析影响病原微生物2种常用灭活方法可靠性的因素

目的以表达绿色荧光蛋白的重组腺病毒（rAdvGFP）为模式病毒,模拟实验室内利用次氯酸钠灭活含病毒废液和利用紫外线对生物安全柜台面进行消毒的实验室操作,并评价其可靠性及影响因素。 方法终浓度为0（阳性对照）、0．5％、0．2％、0．1％、0．05％的次氯酸钠溶液（0．5％、0．2％、0．1％、0．05％次氯酸钠组）分别与rAdvGFP溶液（1×10^6 TCID50）混合后,将混合液原液与混合液的1：10～1：10^5倍比稀释液分别接种人胚肾293细胞,在荧光显微镜下观察GFP蛋白的表达。将rAdvGFP溶液（1×10^3 TCID50）置于紫外灯下分别照射0（阳性对照）、15、30、60、120min（15、30、60、120min照射组）,各组再按垂直照射高度的不同分为5、10、20、30、67cm亚组,照射后收集各组病毒液接种293细胞,在荧光显微镜下观察GFP蛋白的表达。实验均重复3次。 结果各组次氯酸钠病毒混合液原液均造成细胞不同程度的损伤,其中0．5％次氯酸钠组混合液原液对细胞的毒性作用较强;稀释10倍后,除0．05％次氯酸钠组外,其余各浓度次氯酸钠组稀释液均导致细胞不同程度的死亡;稀释100倍后,仅0．5％次氯酸钠组稀释液仍对细胞具有毒性作用。荧光显微镜下观察显示,仅0．1％次氯酸钠组的1：10、1：10^2倍稀释液和0．05％次氯酸钠组的1：10、1：10^2、1：10^3倍稀释液接种细胞可见绿色荧光。紫外线照射后在荧光显微镜下观察显示,15min照射组的各亚组细胞均可见明显绿色荧光;30min照射组的20、30、67cm亚组与60、120min照射组的67cm亚组细胞仍可见绿色荧光。 结论有效氯浓度须在0．2％以上、安全柜内1．12mW／cm^2强度紫外线照射30min以上方能有效灭活重组腺病毒,建立实验室操作的可靠性评价技术体系并据此制定标准操作规程很有必要性。     

狂犬病病毒体外生存活力研究

目的了解狂犬病病毒在体外组织和体液样本中的生存能力。方法通过病毒滴度测定、直接免疫荧光、RT-PCR和病毒分离实验室技术手段对体外组织和悬液中狂犬病病毒生存活力进行验证。结果随着温度升高, 脑组织和悬液中狂犬病病毒活力逐渐下降, 在56℃, 30 min后病毒即失去感染力, 在37℃时, 组织中病毒活力可保持7 d;在pH9.6时, 1 h后无法检测病毒活力;终浓度30%以上乙醇, 500 mg/L以上次氯酸钠和100 mg/L以上苯扎溴铵对悬液中狂犬病病毒作用3 min后即可完全灭活;80%丙酮对组织中狂犬病病毒灭活作用最强, 在浸泡4 h后样本即无法进行病毒分离。结论狂犬病病毒不耐高温, 在pH6.8～7.4环境中较为稳定, 常用消毒剂即可对病毒发挥作用;本实验旨在为狂犬病病毒的体外生存活力提供详细数据, 为下一步狂犬病防控工作开展提供理论支持。     

过氧乙酸-乙醇对同种异体骨植入材料中病毒的灭活效果分析

目的 探讨过氧乙酸-乙醇对同种异体骨(同种骨)移植材料中的病毒的灭活效果.方法 本研究采用细胞感染方法对同种异体骨移植材料中的伪狂犬病毒、牛腹泻病毒、人免疫缺陷病毒采用过氧乙酸-乙醇(10g/1的过氧乙酸+体积分数为24％的乙醇)灭活的效果进行检测.结果 过氧乙酸-乙醇对同种异体骨(同种骨)移植材料中牛腹泻病毒、伪狂犬病毒在常温下浸泡10min即可达到灭活的效果,可以使同种异体骨(同种骨)移植材料中污染的牛腹泻病毒、伪狂犬病毒灭活对数值为4lg以上.对各组实验材料中未检出牛腹泻病毒、伪狂犬病毒的标本接种细胞盲传至第3代,无病毒检出.过氧乙酸-乙醇对同种异体骨(同种骨)移植材料中人免疫缺陷病毒在常温下浸泡30min即可达到灭活的效果,对各组实验材料中未检出人免疫缺陷病毒的标本接种细胞盲传至第3代,无病毒检出.结论 过氧乙酸-乙醇对同种异体骨(同种骨)移植材料中污染的伪狂犬病毒、牛腹泻病毒、人免疫缺陷病毒在常温下均具有较好的灭活效果.     

次氯酸钠溶液对不同时期粪肠球菌生物膜的药物作用

背景:有研究指出粪肠球菌能在营养物质缺乏、抗菌药存在的恶劣环境中生存,形成再感染而影响根管再治疗效果。也有实验表明次氯酸钠溶液作为根管冲洗液对残留的粪肠球菌具有很强的清除作用。目的:建立不同时期粪肠球菌生物膜,研究不同浓度次氯酸钠溶液对其的作用效果。方法:建立对数期、稳定期、饥饿期粪肠球菌生物膜模型,以1%,2.5%,5.25%次氯酸钠溶液分别作用于各时期粪肠球菌生物膜表面30 s、5 min、10 min,同时用激光共聚焦显微镜观察用药后生物膜情况。结果与结论:在相同浓度次氯酸钠溶液作用下,饥饿期粪肠球菌生物膜中的活菌降低量低于稳定期与对数期粪肠球菌生物膜中的活菌降低量(P0.05);在1%次氯酸钠溶液作用下,稳定期与对数期粪肠球菌生物膜中的活菌降低量差异有显著性意义(P0.05);在2.5%,5.25%次氯酸钠溶液作用下,稳定期与对数期粪肠球菌生物膜中的活菌降低量差异无显著性意义(P0.05)。在同一时期生物膜模型下,5.25%次氯酸钠溶液作用下的活菌降低量高于2.5%、1%次氯酸钠溶液作用下的活菌降低量(P0.05);2.5%次氯酸钠溶液作用下的活菌降低量高于1%次氯酸钠溶液作用下的活菌降低量,但仅在药物作用30 s时差异有显著性意义(P0.05)。结果表明,在相同药物浓度、相同作用时间内,饥饿期粪肠球菌生物膜较对数期、稳定期粪肠球菌生物膜对次氯酸钠溶液更具耐药性;在相同作用时间内,5.25%次氯酸钠溶液对饥饿期粪肠球菌生物膜的杀菌效果最佳。     

热和紫外线照射对丙型肝炎病毒JFH-1株灭活效果的研究

目的 应用HCV JFH-1株细胞培养系统,研究热和紫外线照射对HCV的灭活效果.方法 感染性滴度为2.5×104 Ffu/ml的丙型肝炎病毒JFH-1株(HCV JFH-1 strain)原液,经56℃水浴或紫外线照射不同时间后,感染Huh7-25-CD81细胞系,应用间接免疫荧光法测定病毒感染性滴度的动态变化.若被感染的细胞经旨传3代后,用问接免疫荧光法检测为阴性,则判定病毒原液已被完全灭活.结果 感染性滴度为2.5×104 FFU/ml的HCV JFH-1株原液,经56℃水浴孵育10 min、20 min、30 min后,其细胞的感染性滴度分别降至1.6×103FFU/ml、3.1×102FFU/ml和3.3×10FFU/ml;该HCV JFH-1株原液暴露于紫外灯下(波长253.7 nm,辐照强度≥60 μW/cm2)30 cm处照射15 s、30 s、45 s后,其细胞的感染性滴度分别降至1.0×103FFU/ml、1.1×102FFU/ml和2.7×10FFU/ml.经56℃水浴孵育40 min或紫外灯下照射(波长253.7 nm,辐照强度≥60 μW/cm2)1 min后,应用间接免疫荧光法检测为阴性,被感染的细胞经盲传3代后,间接免疫荧光法检测仍为阴性,证明该病毒液已被完全灭活.结论 HCV JFH-1株对热和紫外线照射较为敏感,56℃下40 min或紫外线照射(波长253.7 nm,辐照强度≥60 μW/cm2,距离30 cm)1 min,可完全灭活HCV JFH-1株.     

消毒剂及紫外线照射对细菌L型的灭菌效果观察

目的探讨消毒剂及紫外线对L型菌的诱导及杀灭情况,为指导现场消毒杀菌工作提供实验依据。方法分别采用常用消毒剂与紫外线作用于金黄色葡萄球菌、大肠埃希氏菌或枯草杆菌,观察L型菌的诱导及杀灭情况。结果采用常用消毒剂消毒,发现仅75％乙醇作用1min内可诱导金黄色葡萄球菌形成L型,L型菌仍具有DNA酶活性、溶血作用,凝固酶仍为阳性;在紫外线消毒中,在不干燥的条件下,即使杀灭照射剂量足够,仍不能达到完全杀灭细菌的目的。结论采用乙醇消毒,应严格保证有效作用浓度和足够的消毒时间;采用紫外线消毒．应严格控制空气及物体表面的湿度,从而达到完全杀灭细菌的目的。     

新冠肺炎疫情下结直肠癌标本采集和处理的对策

正2019年12月以来,新型冠状病毒肺炎(新冠肺炎,COVID-19/2019-nCoV)在我国武汉暴发,并迅速向全世界蔓延~([1])。2020年1月20日,国家卫生健康委员会将该疾病纳入乙类传染病并按照甲类传染病进行防治~([2])。在疫情的影响下,我国的各行各业均受到很大影响,尤其是医疗卫生事业。相关卫生部门在疫情影响下制定了各种应对策略。COVID-19病毒对紫外线和热敏感(56℃30 min),乙醚、75%乙醇和过氧乙酸等条件下均可有效灭活病毒~([3])。COVID-19的主要传播途径包括呼吸道飞沫和密切接触,在相对封闭的环境中长时间暴露于高浓度气溶胶情况下还可能存在气溶胶传播~([4])。疫情期间,医院中患者来源结构复杂、     

创伤弧菌紫外线抗性观察

目的:观察创伤弧菌对紫外线(UV)的抵抗力,确定紫外线有效灭杀创伤弧菌的时间,为创伤弧菌消毒提供实验依据.方法:创伤弧菌ATCC27562、大肠杆菌ATCC25922悬液及平板在强度值达90μW/cm2的紫外线照射不同时间后,进行培养并定量测照射前后的活菌浓度,统计杀菌率,绘制照射前后灭活曲线(t-△lg).结果:创伤弧菌在强度值达90 μW/cm2的紫外线照射下,15 min后定性培养为阴性,定量检测1 min平均灭活对数值LIV悬液法为4.37±0.13、平板法为5.10±0.19,平均杀菌率悬液法为(99.99±0.01)%、平板法为(99.99±0.05)%,照射15min后无菌存活.大肠杆菌在照射1 min后达到消毒水平,10min后达到无菌状态.结论:创伤弧菌经强度值达90 μW/cm2的紫外线照射1 min达到消毒水平(LIV≥4.00),照射15 min创伤弧菌能被迅速有效灭杀.     

Effect of chemicals on the infectivity of chicken anaemia virus

None of five commercial disinfectants, invert soap, amphoteric soap, orthodichlorobenzene, iodine disinfectant and sodium hypochlorite, was completely effective in destroying the infectivity of chicken anaemia virus (CAV) in liver material at 5% concentration. However, the iodine disinfectant and sodium hypochlorite completely inactivated the virus in tissue culture (TC) material when used at 1% concentration. CAV was resistant to organic solvents such as methyl alcohol, ethyl alcohol, acetone and chloroform. Beta-propiolactone and glutaraldehyde inactivated CAV. Fumigation with formaldehyde for 24 h only partly inactivated both liver and TC materials. It is presumed very hard to disinfect CAV in poultry facilities.     

Morphological response of human rotavirus to ultra-violet radiation, heat and disinfectants

The morphological damage induced in human rotavirus particles by exposure to ultraviolet (UV) radiation at a wavelength of 254 nm increased progressively with length of treatment. Exposure of the virus in suspension to 9000 ergs/cm2/s was sufficient to remove the smooth capsid layer from 50% of particles after 1 min and from all the virions within 10 min. By this time, the number of stain-penetrated or empty particles increased markedly, along with the appearance of virus-derived debris in the form of disrupted and isolated capsomeres. After treatment for 120 min no intact virus particles were observed. The action of wet (100 degrees C) or dry (60 degrees C) heat resulted in changes similar to those effected by UV radiation, with a rapid loss of viral outer capsid shell from the virions followed by stain penetration and disintegration of particles. Sodium hypochlorite, cetrimide and 70% ethanol induced a rapid loss of the outer capsid layer, but, compared with UV radiation or heat, a slower increase in the number of stain-penetrated particles was noted. This was particularly evident with cetrimide. Chlorhexidine and phenol had effects on virus structure only after extended periods of exposure, whilst glutaraldehyde treatment had little influence on virus morphology. Glutaraldehyde 2% v/v would appear to be most suitable for the disinfection of rotavirus-containing electronmicroscope grids before their examination.     

Inactivation of the infectivity of two highly pathogenic avian influenza viruses and a virulent Newcastle disease virus by ultraviolet radiation

Exposure of a virulent isolate of Newcastle disease virus (NDV) and two highly pathogenic avian influenza (HPAI) viruses, one of H7N1 subtype and the other H5N1 subtype, to a continuous ultraviolet B flux of approximately 90µW/cm², which models solar ultraviolet radiation, resulted in an exponential decline in infectivity with time. The time taken for a reduction in titre of 1 log₁₀ median tissue culture infectious dose for each virus was: NDV, 69 min; H7N1 HPAI virus, 158 min; and H5N1 HPAI, virus 167 min.     

Comparative study of inactivation of herpes simplex virus types 1 and 2 by commonly used antiseptic agents.

A comparative study of the different reactions of herpes simplex virus types 1 and 2 to Lysol, Listerine, bleach, rubbing alcohol, Alcide disinfectant (Alcide Corp., Westport, Conn.), and various pHs, temperatures, and UV light exposures was performed. Both types of stock virus (titers of approximately 10(6) and 10(5.5) for types 1 and 2, respectively) were inactivated by 0.5% Lysol in 5 min; by Listerine (1:1 mixtures) in 5 min; by 2,000 ppm (2,000 microliters/liter) of bleach in 10 min; by rubbing alcohol (1:1 mixtures) at zero time; by Alcide disinfectant (0.2 ml of virus plus 2.0 ml of Alcide) at zero time; by pHs 3, 5, and 11 in 10 min; and by a temperature of 56 degrees C in 30 min. A germicidal lamp (model G30TB; General Electric Co., Schenectady, N.Y.) (30 W) at a distance of 48 cm failed to completely inactivate the two types in 15 min. Type 1 showed slightly more resistance to Listerine and bleach and significantly more resistance to heat; moreover, pH 9 did not affect the infectivity of either type after 10 min.     

Effect of glutaraldehyde on the antigenicity and infectivity of hepatitis A virus

The effect of glutaraldehyde on the antigenicity and infectivity of hepatitis A virus (HAV) was examined. The CF 53 strain, adapted to human hepatoma PLC/PRF/5 cells, was treated with glutaraldehyde using three different concentrations, 0.02, 0.10, and 0.50%, for various periods of time, 3, 10, and 30 min, respectively. After the virucidal assays, glutaraldehyde and HAV were separated by gel filtration, then the antigen (radioimmunoassay) titer and the infectivity titer were determined. The greatest antigen titer reduction was about 80% after 30 min using 0.10% glutaraldehyde and within only 3 min using 0.50% glutaraldehyde. Glutaraldehyde is an effective disinfectant against HAV: the infectious virus titer decreased by more than 3 log10 after 30 min using 0.10% glutaraldehyde and within only 3 min using 0.50% glutaraldehyde. Statistical studies showed that the decrease of antigen or infectious virus titer was affected by both glutaraldehyde concentration and exposure time.     

Inactivation of hepatitis B virus by intermediate-to-high-level disinfectant chemicals

In five separate tests, hepatitis B virus in dried human plasma was exposed for 10 min at 20 degrees C to disinfectant chemicals having activity levels ranging from intermediate (e.g., 70% isopropyl alcohol) to high (e.g., 2% aqueous glutaraldehyde at pH 8.6). Five chimpanzees (one animal per disinfectant chemical) received treated material intravenously, and none showed signs of infection after post-inoculation periods of 9 months. Two animals were rechallenged with inoculum treated in the same manner, except that saline was used instead of a disinfectant chemical; both were infected within 4 weeks. Our results showed that hepatitis B virus was not as resistant to disinfectant chemicals as once thought and suggested that chemicals with similar activity levels (intermediate to high) might possibly be used on hepatitis B virus contamination with a margin of safety.     

Survival of HIV-1 activity after disinfection, temperature and pH changes, or drying.

A recently developed assay for measuring infectious HIV-1 particles was used to determine the stability of the virus under various storage conditions as well as the effect of commonly used disinfectants. At the optimum pH of 7.1 the half life of the virus ranged from approx. twenty-four hours at 37 degrees C to no significant loss over 6 months at -75 degrees C. Drying the virus on a glass surface or freezing caused a 5-12 fold and 4-5 fold decrease of activity, respectively. The dried preparations, however, were about as stable as when stored in a buffered solution. A solution of iodine and detergent (2% Jodopax) was the only disinfectant examined which removed all detectable HIV-1 activity. Isopropanol and ethanol were more potent than acetone; however, all three solvents left some viable particles after a 30 min treatment with 70% solutions.     

Protective effects of a water-soluble extract from cultured medium of Ganoderma lucidum (Rei-shi) mycelia and Agaricus blazei murill against X-irradiation in B6C3F1 mice: Increased small intestinal crypt survival and prolongation of average time to animal death

Radioprotective effects of a water-soluble extracts from cultured medium of Ganoderma lucidum (Rei-shi) mycelia (designed as MAK) and Agaricus blazei (Agaricus) against the shortening of survival time or the injury of crypt by X-irradiation were investigated in male B6C3F1 mice. MAK and Agaricus at three different doses were mixed into basal diet into biscuits at 5, 2.5 and 1.25% and administered from 1 week before irradiation. MAK (5% group) significantly prolonged animal survival as compared with basal diet group (control group) after 7 Gy of X-ray irradiation at a dose rate of 2 Gy min(-1). At doses of 8, 10 and 12 Gy X-irradiation at a dose rate of 4 Gy min(-1) MAK (5% group) significantly increased crypt survival as compared to other groups. These results suggest that MAK can act as a radioprotective agent.