1. INTRODUCTION

Mycoplasma hominis and Ureaplasma urealyticum are microorganisms involved in many pathologies and intrauterine infections. They are generally associated with pyelonephritis, chorioamnionitis, pelvic inflammatory disease, endometritis, and other important complications such as prematurity, low-birth weight, spontaneous abortion, and infertility issues [1-5]. Pathologies due to such bacteria are often undiagnosed because of the non-specificity and poverty of their clinical presentations.

Such non-specificity problem has been increased ever since HIV pandemic emergence, which has complicated the germs diagnosis [6]. Mycoplasmas are also bacterial agents, of which several species are found in commensal state in humans they are responsible for respiratory, genital, and systemic infections in immunosuppressed individuals [7,8]. Urogenital colonization varies in the world between 20 and 30% for M. hominis and 60–80% for U. urealyticum [9]. Treatment of urogenital infections due to mycoplasmas is mainly based on cyclins (tetracyclines), macrolides, and quinolones [10].

In the number of quinolones, fluoroquinolones are attractive choices for treating genitourinary tract infections [11]. Biological data obtained in CIRMF Public Health Laboratory on ambulatory patients show a spread of Mycoplasma and Ureaplasma spp. infections in recent years, what is, furthermore, associated with antibiotic resistance. As a result, a better knowledge of the epidemiology of resistance to such antibiotics will, on the one hand, improve the therapeutic management of patients while reducing broad-spectrum antibiotics prescription and reduce the impact of such infection in the population, on the other hand. No study to date has considered looking into a comprehensive epidemiology and evaluation of mycoplasma resistance to antimicrobials in Gabon. The purpose of our study was on determining M. hominis and U. urealyticum strains prevalence in asymptomatic and assessing the resistance of such strains to some antibiotics.

2. MATERIALS AND METHODS

The study was conducted in Franceville at the International Center for Medical Research, (CIRMF). It was a cross-cutting and descriptive study, which was conducted over a period of 4 months on women living in Franceville and vicinities, which have been checked either during gynecological consultation or voluntarily come for mycoplasma bacteriological screening at the medical analysis laboratory of CIRMF. To be included in the study, women had to be at least 13 years old and sexually active. Their consent was to be sought before enrolling.

On inclusion in the study, a detailed medical history and clinical examination were performed for each woman. Such information was recorded on a patient card aimed at collecting first-hand socio-demographic, gestational, and behavioral data. Sampling was performed with a swab after a highlighting of cervix using speculum. In that process, leucorrhea losses were picked at endocervix level for the research of mycoplasma. Since such bacteria have high affinity for cells membranes and mucous, it was deemed important to vaginally scratch the mucosa to collect as many cells as possible for diagnosis and antimicrobial susceptibility testing.

The diagnosis was performed using Mycoplasma IST2 Kit (bioMérieux) in consonance with manufacturer’s instructions, specifically for urogenital mycoplasmas. This kit allows the culture, identification, indicative blood count, and determination the susceptibility of U. urealyticum and M. hominis to antibiotics. Mycoplasma IST2 principle combines a selective culture broth with a 22-well gallery. The broth is suitable for optimal growth of urogenital mycoplasmas.

The result has been interpreted according to the manufacturer’s recommendations (positive for identification), yellow to red color (positive for germ number ≥ 104 UFC/sample). The infection threshold for such microorganisms is set to a specific value greater than or equal to 104 ufc/ml, what enables to determine a colonization for a turn of <104 ufc/ml and an infection for a value greater than or equal to 104 ufc/ml. Wells were read in 24 and 40 h at 36 ± 2°C and the reading was valid if only the control was positive. As far as antibiotic susceptibility study was concerned, nine antibiotics molecules from Mycoplasma IST2 Kit identification gallery were tested. These molecules are Tetracycline, Doxycycline, Josamycin, Erythromycin, Azithromycin, Clarithromycin, Ofloxacin, Ciprofloxacin, and Pristinamycin. Such molecules belong, respectively, to the antibiotic families of Cyclins, Macrolides, Fluoroquinolones, and Streptogramines.

The data collected on information sheets were recorded in a database (MS Excel® spreadsheet). Data analysis was conducted using SPSS software, version 20. The value of P ? 0.05 was considered statistically significant.

3. RESULTS AND DISCUSSION

4. DISCUSSION

The study is the first of its kind to assess in a comprehensive manner the prevalence and antibiotic resistance profile of Mycoplasma strains (U. urealyticum and M. hominis) in Franceville, Gabon. The exclusion of women under childbearing age and the free availability of mycoplasma research examinations at the CIRMF Medical Analysis Laboratory allowed us to eliminate certain selection biases. We were able to document in 278 women enrolled in the study, 64.7% of infection caused by U. urealyticum, 22.7% by M. hominis, and 19.4% coinfection involving both germs (U. urealyticum and M. hominis). Infections alike were observed in age groups between 18 and 42 years. Carriage rate was more significant in such age groups. Carriage and infections caused of mycoplasma this kind may to be explained by this population’s sexual activity. In China, authors reported that 31.2% infections caused by U. urealyticum, 0.7% by M. hominis, and 1.9% coinfections due to the two bacterial germs were believed to be in a population of 6051 women over 4-year period [12].

In Serbia in 2010, it has been reported following a study comparing mycoplasma infections that the prevalence in a symptomatic and asymptomatic infections and coinfections rates in pregnant women population was variable [13]. Such difference observed in all the alike studies could be caused not only to the methodology but also to the epidemiological facies of different epidemiological zones.

Our outcomes show that the prevalence of Mycoplasma in Franceville is higher than that in Ouagadougou [14] and Bangui among a women population [15]. Such variation between Northern and Southern countries in terms of mycoplasma infection rate could be explained by the fact that the studies performed in northern countries focused on symptomatic populations, whereas the ones we performed focused on all women broadly speaking a broader comprehensive way focused on achieved in rates.

In all such studies infections and coinfections, distribution profile is the same decreasing order, dominated by U. urealyticum followed by M. hominis and finally coinfection caused to the two germs (U. urealyticum and M. hominis) [12,14-15]. The study of behavioral parameters allowed us to document that single women were the majority population. In response to our questionnaire, 131 single women enrolled in the study did not protect themselves during sexual intercourse that had no real influence on mycoplasma infection rates. Few authors having performed that mycoplasma studies have addressed behavioral parameters. In pregnant women, urogenital mycoplasmas often only bear witness to oestroprogestative impregnation in this particular population [16].

Mycoplasmas naturally resist to antibiotics that act on bacterial cell wall. This is because they do not have a wall. However, they are generally susceptible to antibiotics that inhibit bacterial proteins synthesis [11]. Of all the mycoplasma isolates tested, the overall resistance to fluoroquinolones ranged from 95.2% to 100% for Ciprofloxacin and Ofloxacin. For isolates of U. urealyticum and M. hominis, overall resistance rates for fluoroquinolones were, respectively, 96.8% and 100% for Ciprofloxacin and 95.2% and 97.2% for Ofloxacin. Such results are higher that, those reported in 2011 in a study which looked at determining the susceptibility of U. urealyticum and M. hominis strains to antibiotics in northeastern Romania. In the course of said study, a resistance was documented to Ciprofloxacin 75% and 53.7%, respectively, for M. hominis and U. urealyticum, against 30% and 16.13% for Ofloxacin in M. hominis and U. urealyticum strains [17].

The explanation for the high percentage of mycoplasma resistance to fluoroquinolones from strains isolated from human patients could not only cause to the frequency of such family of molecules prescription by prescribers in infection treatments but also caused to their mild side effects that could cause such molecules. Macrolide mycoplasmas resistance rates ranged from 69.9% to 100% for all test molecules (Erythromycin, Azithromycin, and Clarithromycin).

Studies achieved in New Guinea and Romania reported resistance rates of mycoplasma ranging from 33.33% to 97.8% for Erythromycin, confirming a high level of resistance to macrolides [17,18]. Such variable rates observed between the results of different studies for the same therapeutic molecules against mycoplasmas, could be due not only to the variability of isolates but also to the attitude of public as regard the therapeutic regimens given by the prescribers. However, mycoplasmas expressed a resistance to Josamycin ranging between 18.3% and 25.4%. Such levels are lower than those of the other molecules of the same family of macrolides. Cyclins resistant (Tetracycline and Doxycycline) was variable in the range of 26.1–44.4%. Such resistance rates are higher than those reported by some authors in New Guinea, Senegal, and Serbia [18-20].

Observed differences could be due to the use of molecules of this family since birth in some African populations. Some mycoplasmas acquire genetic resistance to Tetracycline through a Tet (M) mutation [7,21]. It would be interesting to study the prevalence of Tet (M) mutations, but also mutation genes to fluoroquinolones. In contrast to fluoroquinolones, macrolides, and cyclins, our isolates had a very low resistance to Pristinamycin in the range of 2.8–7.4%. This result shows the good activity of such a molecule on mycoplasma and is a significant a therapeutic agent for infections mainly caused by M. hominis and U. urealyticum. Studies on the identification of antibiotic resistance genes should be considered in Franceville, where the resistance of mycoplasma to certain antibiotic molecules reaches 100%. Such wide variability in antibiotic resistance profile tested has been reported by several authors throughout the world [22-24].

5. CONCLUSION

The results obtained in this study show that carriage of mycoplasma infection or colonization is high in the female population in Franceville. The treatment based on mycoplasmas probabilistic antibiotic therapy should focus on first-line use of Pristinamycin, followed by Cyclins, although somewhere, their effectiveness is probable on such mycoplasmas.