Drift of tetM determinant in urogenital microbiocenosis containing mycoplasmas during treatment with a tetracycline antibiotic

We studied the correlation between genetic transfer of tetM determinant in Tn916 conjugative transposon by urogenital mycoplasmas (Mycoplasma hominis and Ureaplasma urealyticum) and changes in the bacterial repertoire during treatment with a tetracycline antibiotic. Basic conditions favoring the nonspecific transfer of tetM determinant into mollicute cells are determined and the allele polymorphism of tetM determinant in clinical strains of M. hominis and U. urealyticum is evaluated. The structure of tetM gene in clinical mycoplasma and ureaplasma strains is characterized by a peculiar mosaic pattern and differs from all previously described alleles of this gene. The results suggest that tetracycline resistance in mollicutes is determined by mechanisms alternative to genetic transfer of tetM determinant.     

Subtyping of Mycoplasma pneumoniae isolates based on extended genome sequencing and on expression profiles

Mycoplasma pneumoniae isolates from patients, collected over a period of 12 years in Germany, were characterized by various methods (parameters) including multilocus sequence typing, restriction fragment length polymorphisms, Western blotting with mono-specific antibodies directed against selected proteins or with polyspecific antibodies directed against the Triton X-114-soluble protein fraction, and two-dimensional gel electrophoresis. The results for 91 isolates from Germany, which were complemented with 14 isolates from the USA and 10 isolates from France, clearly showed that M. pneumoniae is a highly uniform species and that most of the isolates could be assigned to one of the two subtypes 1 and 2. The members of one subtype differ from the other with respect to the sequence of the P1 gene, the ORF6 gene, the P65 gene, and by a typical DNA restriction fragment pattern. We observed four isolates (variants), which seemed identical by the above mentioned criteria, but did not belong to either one of the two subtypes. They showed most of the subtype 2-specific features, but differed in the sequence of the P1 gene and showed a variation in the restriction fragment pattern. The appearance of subtype 1 or 2 over the last 12 years in Germany showed a dominance of subtype 1 between 1989 and 1996 and a dominance of subtype 2 between 1997 and 1998. The variant (neither subtype 1 nor subtype 2) was only detected in 1991 and 1995 but it had no epidemiological consequences.     

Detection of Mycoplasma hominis antigen in clinical specimens by using a four-layer modification of enzyme immunoassay (EIA)

A 4-layer modification of enzyme immunoassay (EIA) was developed for the detection of Mycoplasma hominis antigen in clinical specimens. Microtiter plates were sensitized with rabbit anti-mycoplasma immunoglobulin, guinea pig anti-mycoplasma immunoglobulin was used as the secondary antibody, and horseradish peroxidase-conjugated anti-guinea pig immunoglobulin was used as the indicator antibody. The specificity of the assay was confirmed by using guinea pig immunoglobulins from preimmunization sera. The sensitivity of the assay is down to 10 ng/ml of antigen protein. Marked cross-reactivity was demonstrated for different strains within the species M. hominis, whereas the other genital mycoplasma species tested showed no reactivity in the assay. A comparison was made of EIA and conventional culture of vaginal specimens from 24 women. All 6 specimens positive by culture were also positive for M. hominis antigen by EIA. Antigen detection by EIA is a sensitive, rapid and simple method for the detection of M. hominis in clinical specimens.     

Immunohistochemical labelling of cytokines in lung lesions of pigs naturally infected with Mycoplasma hyopneumoniae

Mycoplasma hyopneumoniae (Mh) is the primary agent of porcine enzootic pneumonia (PEN), a chronic respiratory disease endemic to pig farms, and characterized histologically by infiltration of mononuclear cells in airways and prominent hyperplasia of the bronchus-associated lymphoid tissue (BALT). To gain further insight into the pathogenesis of PEN, cytokine expression in the lung, with particular attention to the BALT, was examined immunohistochemically in pigs naturally infected with Mh. An increase (P < 0.05) in proinflammatory and immunoregulatory cytokines (especially interleukin [IL]-2, IL-4 and tumour necrosis factor [TNF]-alpha, and to a lesser extent IL-1 [alpha and beta] and IL-6) was detected in the BALT, which showed intense lymphoid hyperplasia. IL-1beta and TNF-alpha were also detected in the bronchoalveolar exudate of infected pigs, and IL-6 and IL-8 were demonstrated in mononuclear cells of the alveolar septa. The results showed that in Mh infection, macrophage and lymphocyte activation results in the expression of a number of cytokines capable of inducing lung lesions and lymphoreticular hyperplasia of the BALT.     

Relative contribution of vertical,within-farm and between-farm transmission of Mycoplasma synoviae in layer pullet flocks

ABSTRACT

In this study, the relative contribution of vertical transmission, within-farm transmission and between-farm transmission of Mycoplasma synoviae in layer pullet flocks was quantified using logistic regression analysis. Data from 311 Dutch pullet flocks, of which 172 (55%) were positive for M. synoviae, were included in the study. Also the M. synoviae status of the parent stock of these flocks was included. The M. synoviae status was determined with the M. synoviae rapid plate agglutination test. Data analysis showed that vertical transmission was the most important transmission route for M. synoviae in layers as is demonstrated by an odds ratio of 5.8 (P?=?0.000). A positive association with M. synoviae infections was found for layer pullet flocks on a multi-house farm where at least one other flock was M. synoviae-positive compared to single-house farms (odds ratio 3.1, P?=?0.022), while a negative association was found when no other M. synoviae-positive flocks were present (odds ratio?=?0.2, P?=?0.003). No association was found between M. synoviae status of pullet flocks and poultry farm density. Odds ratios were 0.54 (P?=?0.288) and 0.34 (P?=?0.073), respectively, for medium and highest poultry farm density compared to lowest poultry farm density. This is the first time that the relative contribution of horizontal and vertical transmission of M. synoviae has been quantified. These results can be extrapolated to M. synoviae control in general, and emphasize the importance of M. synoviae control in parent stock and practical channelling.     

建立不同民族永生细胞株质量控制方法的探讨

目的以建立普米族、独龙族和怒族永生细胞为基础,探讨EB病毒转化细胞、细胞培养、冻存、复苏以及支原体检测等建立永生细胞株的技术要点及质量检测方法.方法采用EB病毒转化技术建立3个民族永生细胞株;培养法和PCR法对细胞株进行支原体污染检测;染色体G显带及核型分析细胞株的遗传稳定性.结果成功建立了3个民族B淋巴细胞永生细胞株,转化率分别为98%、86%和76%.对已建株保存的3个民族的永生细胞进行复苏培养,复苏成活率为100%.支原体污染检测均为阴性.染色体计数和G带分析显示,细胞株经早期传代培养后,仍然保持二倍体特征,未发现染色体结构畸变.结论本研究中传代培养、细胞冻存、细胞复苏和支原体污染防范等一整套技术过程是满足建库要求的.同时为大规模永生细胞库的建立和进行相应的质量监测提供了科学的依据.另外,本研究还对一些影响转化的因素和可能的机理进行了探讨.     

Immunologic reactivity of (C57BL/6 × A/Sn)F1 mice in mixed Mycoplasma-virus infection

Mixed infection of hybrid mice, highly resistant to Rauscher virus, with this virus andMycoplasma arthritidis was accompanied by progressive inhibition of populations of splenic rosetteforming (REC) and plaque-forming (PFC) cells and led to induction of malignant erythroblastosis, cytologically identical with Rauscher's leukemia. During mixed infection of the hybrid mice withAcholeplasma laidlawii and Rauscher virus the immune response was almost completely suppressed on the 21st day and considerable splenomegaly was observed, but by the 62nd day of infection the RFC and PFC populations and also the weight of the spleens had regained the control level. The possible role of mycoplasmas in the induction and development of Rauscher's leukemia is discussed.N. F. Gamaleya Institute of Epidemiology and Microbiology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician O. V. Baroyan.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 88, No. 9, pp. 327–329, September, 1979.     

柑桔黄龙病的直接荧光诊断法

试验证明,直接荧光诊断法是诊断柑桔黄龙病的一种快速、简便、准确的方法。 将健叶和黄龙病叶叶柄用保险刀片徒手切片,在透射式荧光显微镜下检查,可看到健叶叶柄的木质部导管细胞壁发黄色荧光,韧皮纤维细胞发绿色荧光,而病叶叶柄韧皮部中有1～多个鲜明的黄色或黄绿色荧光团块。这种特异性的荧光,在健叶柄韧皮部中是没有的,其它由病毒、类病毒、细菌、真菌等病原引起的病叶柄韧皮部中亦没有此种特异性荧光。     

探针ASPCR检测肺炎支原体 微量耐药突变A2063G方法的建立

[摘要]　目的　 建立能够特异性检测微量肺炎支原体（Mycoplasma pneumoniae, MP）A2063G耐药突变基因的特异性扩增等位基因的探针法实时定量PCR（probe-based allele-specific real-time PCR, 探针ASPCR）方法。方法?建立特异性检测A2063G耐药突变位点的探针ASPCR方法,并验证其灵敏度、特异度及准确度等性能。结果?特异性扩增2063G和非特异性扩增2063A/G的引物/探针组合分别扩增105拷贝野生基因型（2063A）模板的Ct值的差(△Ct)高达10.93,能够特异性检测A2063G突变。探针ASPCR方法检测2063G基因型占总MP的比例的准确度可低至1％;检测MP的灵敏度低至10拷贝,检测A2063G耐药突变比例的灵敏度低至0.01％。探针ASPCR方法与前期建立的染料ASPCR方法检测临床样本的MP感染结果一致,MP阳性检出率均为94.83%（55/58）,高于传统巣式PCR联合测序方法的检测结果（75.86%,44/58）;探针ASPCR和染料ASPCR 2种方法检测MP耐药率分别为63.64%（35/55）、70.91%（39/55）,高于传统巣式PCR联合测序方法检测结果59.09%（26/44）。结论?新建探针ASPCR方法是一种具有高特异度、准确度和灵敏度的快速检测MP微量A2063G耐药突变的方法;与染料ASPCR方法相比,探针ASPCR方法检测耐药MP的灵敏度略低,但其临床样本检测复查率也低于染料ASPCR方法,且其结果判读简单,更适合在临床中应用推广,能够为临床制定MP及耐药MP感染的治疗方案提供理论依据。     

儿童肺炎支原体感染的临床特征及混合感染相关因素研究

目的 了解儿童肺炎支原体（Mp）感染的临床特征,分析Mp与其他病原体混合感染的相关因素,为完善儿童社区获得性肺炎（CAP）防治提供证据支持。方法 基于在苏州大学附属儿童医院（SCH）开展的急性呼吸道感染病例监测,筛选2018-2021年在SCH住院的<16岁CAP病例,采用统一的调查表收集研究对象基本情况、基础疾病史、临床表现等信息,通过医院检验信息系统查询研究对象的病原检测结果,比较Mp、细菌、病毒等病原体感染者的临床特征,比较Mp混合其他病原感染对病情严重性的影响,采用logistic回归模型分析Mp混合其他病原感染的相关因素。结果 共收集8 274名CAP住院病例,其中Mp阳性2 184例（26.4%）;Mp检出率女童高于男童（P<0.001）,随月龄增加而升高（P<0.001）,夏秋季高于冬春季（P<0.001）。喘息、气促、喘鸣音及肺部呈片状阴影的发生率,以及发热和住院天数等指标在Mp、细菌和病毒感染病例中的差异均有统计学意义（均P<0.05）。<60月龄Mp混合感染病例出现喘息症状及痰鸣音和喘鸣音的比例高于单纯感染病例,≥60月龄混合感染病例较Mp单纯感染更易出现气促症状（均P<0.05）。多因素logistic回归模型分析显示,男童（aOR=1.38,95%CI：1.15~1.67）、<6月龄（aOR=3.30,95%CI：2.25~4.89）、6~月龄（aOR=3.44,95%CI：2.63~4.51）、24~月龄（aOR=2.50,95%CI：1.90~3.30）、48~71月龄（aOR=1.77,95%CI：1.32~2.37）和3个月内呼吸系统感染史（aOR=1.28,95%CI：1.06~1.55）为Mp混合其他病原感染的相关因素。结论 Mp是导致儿童CAP住院的主要病原体,单纯Mp感染病例较细菌、病毒感染病例发热天数更长;Mp常与细菌和病毒混合感染,男童、<72月龄和3个月内呼吸系统感染史是Mp混合感染的相关因素。