Development of multiplex real-time quantitative PCR for simultaneous detection of Chlamydia trachomatis,Mycoplasma hominis,Ureaplasma urealyticum,and Mycoplasma genitalium in infertile women

PurposeSexually Transmitted Diseases (STDs) can cause sterility and many other problems for women planning pregnancy. Currently, almost 340 million people worldwide suffer from Sexually Transmitted Infections (STIs). This study made attempts to quickly identify STDs' most critical infectious agents using dedicated primers and probes.MethodsThe present study was done on the cervical samples of 200 infertile women. After extracting the total DNA of Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, and Mycoplasma genitalium, quantitative methods were employed to determine the rate of target bacteria using multiplex real-time PCR.ResultsThe multiplex qPCR showed the rates of 47%, 16%, 46%, and 16.5% for Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, and Mycoplasma genitalium in infertile women, respectively. In some patients, there were co-infections with two or three bacteria. The diagnostic approach used in our research could be employed as an alternative detection tool to identify the four most common STD-associated bacterial agents while detecting mixed infections.ConclusionsInfertile women with no biological problems could have their genital tract checked using this newly designed identification technique and get proper treatment for their infections as quickly as possible.     

Detection of Chlamydia trachomatis,Ureaplasma urealyticum and Mycoplasma hominis in infertile Bulgarian men with multiplex real‐time polymerase chain reaction

The effect of Chlamydia trachomatis, Ureaplasma urealyticum and Mycoplasma hominis over the sperm quality is still unclear. The aim of this study was to determine their prevalence in infertile Bulgarian men. A total of 281 men were examined by applying mRT‐PCR. The registered prevalence was as follows: C. trachomatis – 13.9%, U. urealyticum – 19.2%, M. hominis – 9.9%. Co‐infection was established in eight swabs. This first in Bulgaria to study for detection of chlamydia and mycoplasmas in infertile men by mRT‐PCR demonstrates higher prevalence of the tested microorganisms in the infertile group toward the control one.     

Evaluation of the Mycoplasma Duo kit for the detection of Mycoplasma hominis and Ureaplasma urealyticum from urogenital and placental specimens

This study compares the Mycoplasma Duo kit for the detection of genital mycoplasmas with conventional culture using A7 differential agar for the detection of Mycoplasma hominis and Ureaplasma urealyticum in clinical samples. Detection of the mycoplasmas is based on the specific metabolic properties of each organism to hydrolyse either arginine or urea. The Mycoplasma Duo test showed a significantly higher detection rate than did culture, although many of the culture-negative results may have been due to the presence of bacterial overgrowth.     

Activity of moxifloxacin against the urogenital mycoplasmas Ureaplasma spp., Mycoplasma hominis and Mycoplasma genitalium and Chlamydia trachomatis

The activity of moxifloxacin was compared with that of other antimicrobial agents against 54 strains of Ureaplasma spp., 54 strains of Mycoplasma hominis , 14 strains of Mycoplasma genitalium , and 44 strains of Chlamydia trachomatis . Moxifloxacin inhibited 90% of all isolates at a concentration ≤1 mg/L, being the most active compound against C. trachomatis and sharing the highest activity with garenoxacin and gemifloxacin against mycoplasmas. Moxifloxacin killed the 30 mycoplasma isolates tested at a concentration ≤1 mg/L, except those resistant to fluoroquinolone. Thus, moxifloxacin has attracted interest as a potential therapy for mycoplasmal or chlamydial urogenital infections.     

Mycoplasma hominis and Ureaplasma urealyticum in different gynecologic diseases

The incidence of Mycoplasma hominis (M. hominis) y Ureaplasma urealyticum (U. urealyticum) was investigated in 113 endocervical samples obtained from women who were seen for different gynecological pathologies. Forty-seven (42%) patients were positive to these microorganisms; 26 cases (23%) were positive for M. hominis and 21 (19%; p = NS) for U. urealyticum. Average age was 32.1 +/- 7.7 years; the average number of sexual partners was 1.7 +/- 1.1. Eleven of 17 patients with 3 o more sexual partners were positive for Genital Mycoplasma (GM), and U. urealyticum was found more often in this group. A higher incidence of GM was found in women between 26 and 30 years (34%); 57.5% of the patients with positive cultures for GM had begun sexual activity before 20 years of age. M. Hominis was found in 61% of women with no parity and U. urealyticum in 71% of parous women. The cultures were positive in 10 of 14 patients with pelvic inflammatory diseases (PID). A cervical biopsy was taken from 52 cases and the diagnosis of cervical intraepithelial neoplasia (CIN) was made in 49 (94%) but only 24 of them were positive for GM (50%). Thirty-five patients suffered sterility, and 12 (34%) were positive for GM, however all positive cases consulted because of primary sterility. The conclusions obtained from this study are: 1) Near half of the patients was positive for GM and none of the species was predominant over the other. 2) The more sexual partners the higher was the incidence of GM, especially U. Urealyticum. 3) The lower the age of the first sexual intercourse the higher the probability of contamination with these microorganisms. 4) M. hominis was more common in nulliparous women and U. urealyticum was found more often in parous patients; the number of deliveries did not have influence in these findings. 5) A statistical significance between GM and PID was found (p = 0.03). 6) GM have no influence on spontaneous abortion. 7) No statistical significance was found between GM and the beginning and evolution of CIN. 8) No relation statistically significative was found between GM and sterility.     

Examination of Ureaplasma urealyticum and Mycoplasma hominis in 4082 Chinese patients

The objective of this study was to analyze the infection rate and drug resistance of Ureaplasma urealyticum (UU) and Mycoplasma hominis (MH) in the genitourinary tract of Chinese patients. From December 2018 to June 2019, vaginal secretion or urinary secretion of outpatients in our hospital were selected for culture and drug sensitivity analysis of Ureaplasma urealyticum and Mycoplasma hominis. In 4082 Chinese samples, 1567 Mycoplasma were detected, a detection rate of 38.39%, among which 1366 cases were UU single positive, accounting for 33.47%, 15 cases were MH single positive, accounting for 0.36%, 186 cases were UU and MH mixed positive, accounting for 4.56%. The most affected age groups were 21-30 years and 31-40 years, accounting for 19.09 and 15.05%, respectively. The results of drug sensitivity showed that doxycycline, minocycline, josamycin, clarithromycin, and roxithromycin were more sensitive to mycoplasma infection. The distribution of Ureaplasma urealyticum and Mycoplasma hominis in the human genitourinary system and their sensitivity to antibiotics is different for sex and age groups.     

Experimental animal infections with Mycoplasma hominis and Ureaplasma urealyticum.

Subcutaneous tissue cavities in mice and guinea pigs were infected with human isolates of Ureaplasma urealyticum and Mycoplasma hominis. The minimal infective dose for M. hominis was as low as less than 10 color-changing units (CCU) for mice and 10(2) CCU for guinea pigs. The minimal infective dose for U. urealyticum was as low as less than 10 CCU for mice and 10(4) CCU for guinea pigs. Mouse infections with either U. urealyticum or M. hominis persisted for 1 day to greater than 4 months. Guinea pigs remained infected for up to 4 weeks. Two M. hominis isolates were similar in their ability to infect subcutaneous tissue cavities but two U. urealyticum isolates varied in their ability to infect the cavities. The histopathology of the M. hominis and U. urealyticum infections was similar: an initial intense polymorphonuclear response with giant cells, followed in 4 weeks by histiocytes and giant cells with some plasma cells and lymphocytes.     

In vitro activity of the two new 4-quinolones A56619 and A56620 againstNeisseria gonorrhoeae,Chlamydia trachomatis,Mycoplasma hominis,Ureaplasma urealyticum andGardnerella vaginalis

The in vitro activity of tetracycline, ciprofloxacin and two recently developed 1-aryl-fluoro-quinolones, A56610 and A56620, was tested against 65 beta-lactamase-negative and 35 betalactamase-positive Neisseria gonorrhoeae strains, 12 Chlamydia trachomatis,50 Mycoplasma hominis,28 Ureaplasma urealyticum and 50 Gardnerella vaginalis strains. In the case of Chlamydia trachomatis and Mycoplasma hominis both the MIC and the MBC were determined. The MIC90 of ciprofloxacin for Neisseria gonorrhoeae was 0.008 g/mland of A56619 and A56620 0.03 g/ml.No difference was observed between the activity against beta-lactamase-negative and beta-lactamase-positive strains. The MIC90 values of ciprofloxacin and A56620 for Chlamydia trachomatis, Mycoplasma hominis and Ureaplasma urealyticum were identical, the values being 2 g/ml, 1g/mland 4 g/mlrespectively. The MIC90 of A56619 for Chlamydia trachomatis and Ureaplasma urealyticum was 0.5 g/mland 1 g/mlrespectively. The MBC90 values of the three quinolones for Chlamydia trachomatis and Mycoplasma hominis were 2 g/ml.The activity of the quinolones against Gardnerella vaginalis was rather low, the MIC90 being 4 g/ml.It is concluded that A56619 and A56620 might be useful for single-dose therapy of gonococcal infections.     

Identification ofMycoplasma hominis,Ureaplasma urealyticum,andChlamydia trachomatis by the polymerase chain reaction

A procedure for identifying chlamydia, mycoplasmas, and ureaplasmas by the polymerase chain reaction is described which uses a system of four primers complementary to fragments of the 16S RNA gene, one of which detects all three microorganisms and the other three are each specific for a particular organism. With this primer system, chlamydia, mycoplasmas, and ureaplasmas are detectable in a single reaction. The procedure may be used for routine diagnoses in diagnostic laboratories. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 120, N ^o 12, pp. 606–609, December, 1995 Presented by R. V. Petrov, Member of the Russian Academy of Medical Sciences     

Effect of carbohydrates on growth of Ureaplasma urealyticum and Mycoplasma hominis.

We examined the effect of 31 carbohydrates on the growth of Ureaplasma urealyticum and Mycoplasma hominis. Arbutin and its breakdown product, hydroquinone, inhibited growth of both species; the other substrates did not alter the extent of growth. Volatile and nonvolatile end products of carbohydrate metabolism were not detected by gas chromatography.     

Co-infections with Ureaplasma parvum,Mycoplasma hominis and Chlamydia trachomatis in a human immunodeficiency virus positive woman with vaginal discharge

A 30-year-old human immunodeficiency virus (HIV)-1 infected woman presented with vaginal discharge and associated vulval irritation. The vaginal swabs tested positive for Ureaplasma parvum and Mycoplasma hominis by both culture and polymerase chain reaction (PCR). The specimen also tested positive for Chlamydia trachomatis deoxyribonucleic acid (DNA) by cryptic plasmid and omp1 gene PCR assays. The patient was successfully treated with azithromycin based on the antibiotic susceptibility testing results of U. parvum and M. hominis by microbroth dilution. Since sexually transmitted infections enhance the transmission of HIV, HIV-positive patients should be screened routinely for these pathogens.     

孕妇解脲支原体、沙眼衣原体感染的诊断与药物干预

目的探讨妊娠期孕妇解脲支原体（UU）、沙眼衣原体（CT）感染的发病率及其对感染者的治疗效果。方法对108例妊娠妇女采用核酸杂交的方法检测宫颈分泌物,对标本UU、CT-DNA进行DNA扩增、核酸杂交检测。结果 108例妊娠早中期孕妇宫颈分泌物UU、CT的阳性率分别是33.33%,7.41%,口服阿奇霉素治疗后检测UU、CT阳性率分别是17.59%,1.85%。两组间有显著性差异（P=0.001或P=0.002）。结论解脲支原体、沙眼衣原体是妊娠期妇女感染的常见病原体,临床治疗效果显著值得推广应用。     

418例不孕妇女宫颈解脲脲原体和沙眼衣原体检测分析

目的了解解脲脲原体(Uu)与沙眼衣原体(CT)在不孕妇女中的感染率,探讨Uu、CT感染与不孕症的关系.方法对418例不孕妇女及52例正常早期妊娠妇女的宫颈分泌物进行Uu、CT的检测.结果不孕组Uu、CT阳性率为46.4%、17 7%,均显著高于对照组(P＜0.01).继发不孕组Uu阳性率为59.2%,显著高于原发不孕组(P＜0.01);继发不孕组CT阳性率为19.7%,高于原发不孕组,但无显著性意义(P＞0.05).225例Uu或/和CT感染患者经抗感染治疗后3个月内自然妊娠42例(18.7%).结论Uu、CT感染是导致不孕症的重要因素,把Uu、CT列为不孕妇女的常规检测项目,有利于不孕症的诊治.     

解脲支原体、沙眼衣原体和弓形虫感染与自然流产的关系

目的探讨解脲支原体(UU)、沙眼衣原体(CT)和弓形虫(TOX)感染与自然流产的关系。方法对310例自然流产患者,UU采用培养法检测,CT、TOX采用酶联免疫吸附试验(ELISA)法检测。同样对75例自愿人工流产者进行检测。结果310例自然流产患者中UU、CT和TOX阳性检出率分别为36.13%、29.25%、11.12%,对照组其UU、CT和TOX感染率分别为6.67%、5.13%、2.67%。两组间有显著性差异(P<0.01或P<0.05)。结论UU、CT和TOX感染与自然流产有相关性。     

Rapid detection of Mycoplasma genitalium,Mycoplasma hominis,Ureaplasma parvum,and Ureaplasma urealyticum organisms in genitourinary samples by PCR-microtiter plate hybridization assay

We present a method for detecting the presence of Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma parvum, and Ureaplasma urealyticum organisms, which are thought to be associated with nongonococcal urethritis (NGU) and other genitourinary infections, in clinical samples. This method consists of PCR amplification of a part of the 16S rRNA gene followed by 96-well microtiter plate hybridization assay using four species-specific capture probes to detect the targets. To test the efficacy of this method, we applied it to the detection of the four species in the urine of patients with NGU. There were no cross-reactions with other human mycoplasmas or ureaplasmas, and the PCR-microtiter plate hybridization assay detected as few as 10 copies of the 16S rRNA gene of each of the four species. Based on these results, this PCR-microtiter plate hybridization assay can be considered an effective tool for the diagnosis of genitourinary infections with mycoplasmas or ureaplasmas.     

100例不育症患者解脲支原体和沙眼衣原体定量分析

目的为了分析不育症和非淋茵性尿道炎患者感染解脲支原体和沙眼衣原体定量检测结果的差异.方法荧光定量多聚酶链式反应(FQ-PCR)技术连续对100例(有97例作了UU检测,84例作了CT检测)本院生殖中心不育症患者及同时时662例(有470例作了UU检测,540例作了CT检测)普通性病和妇科门诊非淋患者CTDNA和UUDNA进行了定量检测.结果不育组UU阳性率=57/97(58.76%),UU拷贝对数值6.016±1.044;CT阳性率=7/84(8.33%),CT拷贝对数值4.968±2.190.普通非淋组UU阳性率=209/470(46.60%),UU拷贝对数值6.131±1.385;CT阳性率=62/540(11.48%),CT拷贝对数值5.894±1.943.结论UU是引起非淋和不育症的主要病原体.UU相对于CT对人类生育的危害更为严重.不育组和普通非淋组UU的定量结果没有统计学差异,而普通非淋组的CT的拷贝数则高于不育组,P＜0.05.     

聚合酶链反应检测丝虫病患者泌尿道沙眼衣原体及解脲支原体的研究

本文采用聚合酶链反应方法,检测了５０ 例丝虫病患者尿中沙眼衣原体（ＣＴ） 及解脲支原体（ＵＵ）ＤＮＡ,同时选择３０ 例正常人作为对照。结果显示,丝虫病患者尿中ＣＴＤＮＡ、ＵＵ ＤＮＡ 和两者混合感染检出率分别为３０ ％ 、２５ ％ 和２０ ％ ,高于正常对３６３％ （Ｐ＜０－０１）,ＣＴ及ＵＵ 感染与病人的病情、病程有一定的关联。提示乳糜尿的发生、发展与泌尿道ＣＴ 和ＵＵ感染有关。     

Comparison of commercially available media for detection and isolation of Ureaplasma urealyticum and Mycoplasma hominis.

The Mycotrim Triphasic flask system (Irvine Scientific, Irvine, Calif.) was compared with a system composed of Mycotrim GU broth (Irvine Scientific) and A7 or A8 agar (Remel, Lenexa, Kans.) for the ability to detect Ureaplasma urealyticum and Mycoplasma hominis from 129 genital specimens. Of the 64 specimens positive for U. urealyticum, 25, 98, and 100% were detected on Mycotrim Triphasic agar and A7 and A8 agars, respectively. All 18 specimens that grew M. hominis were detected by A7 and A8 agars, and 94% grew on Mycotrim Triphasic agar. Mycotrim GU broth detected all of the positive specimens, and Mycotrim Triphasic broth detected all but one. Mycotrim GU broth inoculated simultaneously with either A7 or A8 agar was found to be more sensitive and cost-effective than the Mycotrim Triphasic flask system.     

148例男性前列腺炎患者病原体PCR定量分析

目的为了研究男性前列腺炎患者解脲支原体(UU)和沙眼衣原体(CT)感染特征.方法 FQ-PCR用于148例男性前列腺液的UUDNA和CTDNA定量检测.结果 UU和CT同时阳性10例(阳性率为6.76%),UU拷贝对数值为6.430±1.676,CT为5.904±0.729;UU单项阳性30例(阳性率20.27%),拷贝对数值为5.558±1.367;CT单项阳性8例(阳性率5.41%),拷贝对数值为5.046±2.220.结论 UU与CT相比较是引起男性前列腺炎的主要病原体,在合并和单项感染中两种病原体引起疾病急性发作的机会没有统计学差异.