Epidemiological investigation and antimicrobial susceptibility analysis of mycoplasma in patients with genital manifestations

Abstract Background The aim of this study was to investigate the infection and antimicrobial resistance of Ureaplasma urealyticum and Mycoplasma hominis in patients with genitourinary symptoms among Hakka population in Meizhou, China. Methods A total of 12 633 females and 3315 males who presented urogenital symptoms and were subjected to mycoplasma tests from 2014 to 2018 were enrolled in this study. The mycoplasma detection and antimicrobial susceptibility were tested using the Mycoplasma ID/AST kit. Results The total incidence of mycoplasma infection, as well as the incidence of U urealyticum in Hakka population was annually increasing from 2014 to 2018. The total incidences and U urealyticum infection were more prevalent in females than males. Higher positive rate of mycoplasmas infection was observed in women aged 16‐20 (50.9%) and men aged 26‐30 (25.4%). The occurrence of antimicrobial resistance of mycoplasma to antibacterial agents remained relatively similar in the past five years. Ureaplasma urealyticum infection, M hominis infection, and co‐infection of resistance to levofloxacin, erythromycin, ciprofloxacin, ofloxacin, roxithromycin, azithromycin, clarithromycin, and sparfloxacin were dramatically higher in females than in males. Conclusion Our findings indicate a high burden of mycoplasmas infection and antimicrobial resistance of mycoplasmas infection among females, and josamycin and minocycline may be recommended as the primary choice in clinical treatment of anti‐mycoplasmas.

Previous study reported that genital mycoplasmas were related to urogenital infections, including urethritis, vaginitis, cervicitis, and pelvic inflammatory disease. 5 However, it is difficult to prove their pathogenic effect because of their existence in genital tract of healthy human. The prevalence of these organisms is significantly associated with age, socioeconomic status, physiological cycle, pregnancy, and multiple sex partners. 6,7 As Uu and Mh lack of peptidoglycan, β-lactams are completely inactive against them. Generally, quinolones, tetracyclines, and macrolides are used for the treatment of mycoplasma infection. 8 However, drug resistance of mycoplasma increased due to the improper use of the antibiotics. Since antibiotic resistance of many pathogens is continually changing, surveillance studies are required for assisting in the optimization of antimicrobial treatment.
The city of Meizhou, located in southern China, is known as the world's Hakka population capital, with unique populationbased culture and food, as well as different physiological characteristics. 9 However, little was known regarding the prevalence and antimicrobial susceptibility of mycoplasma infection in Hakka population.
The purpose of this study was to determine the prevalence and antimicrobial resistance of Uu and Mh in patients of Hakka population and presented genital manifestations. Our study would provide useful information for local epidemiology of mycoplasma, and thus help make the prevention and treatment strategy.

| Specimen collection
Cervical and urethral swabs (Kangjian Medical, Jiangsu, China) were used to collected samples from urogenital tracts. In female patients, cervical samples were obtained by cervical swabs from the cervix area after cleaning the exocervical mucus. In male patients, urethral samples were slowly taken from urethra inside 2 cm after external meatus had been cleaned; semen and prostatic fluid were collected and placed in a sterile cup (Kangjian Medical, Jiangsu, China). All samples were sent at room temperature to the clinical laboratory for examination within 2 hour.

| Culture and antimicrobial susceptibility test of Uu and Mh
The culture and susceptibility testing of Uu and Mh were performed using Mycoplasma ID/AST kit (DL medical), following the manufacture's protocol. Briefly, the specimen swab was inserted into medium flask and mixed intensively to make sample completely dissolve.
Then transfer 100 μL of mixture medium to the mycoplasma ID/AST strip. The negative control was added with 100 μL medium. Then the wells were added two drops of the sterile mineral oil and inocu-

| Statistical analysis
Data are presented as n (%) prevalence or mean ± SD and were statistically analyzed using SPSS 20.0 software (IBM Corp., Armonk, NY, USA). Quantitative data were analyzed using ANOVA test, and categorical data were analyzed using chi-squared test. P < .05 was considered statistically significant.

| Age distribution of mycoplasma infection
The Figure 2 presents an age-specific mycoplasma infection in Hakka population. As for female participants, the positive rates remained high in the age group of 16-50 and began to go down gradually more than 50 year. To be noted, the infection rates were relatively high in the age groups of 16-20 and 36-50. Of the 3315 males enrolled, the positive rate was stable in the age group of 16-60, and dramatically dropped after 60 years. Comparatively, the occurrence rate of mycoplasma infection was obviously higher in female than male in all of the age groups.

| Antimicrobial susceptibility patterns from 2014 to 2018
Our study found that Uu displayed low resistance rates to TET, LEV, ERY, JOS, DOX, OFX, MIN, ROX, AZM, CLR, and SPA, while relatively high resistance rates to CIP (>60%), as shown in Table 1 As shown in Table 4, the drug resistance was also different between female and male patients. The mycoplasma, either Uu or Mh, that infected females presented higher resistance to TET, LEV, ERY, DOX, OFX, ROX, AZM, CLR, and SPA, as compared with those infected males.

| D ISCUSS I ON
To our knowledge, this is the first large-scale study to investigate the prevalence of mycoplasmas infection and antimicrobial resistance in patients with genital symptoms among Hakka population in Meizhou.
The present study suggested that the incidence of mycoplasmas was 40.0% in females, which is lower than the previous findings in other areas of China, 10,11 but higher than those in Italy. 12 However, as previous studies suggested, mycoplasmas infection may not cause diseases, but remained as a normal symbiotic colonization.
In the present study, we investigated the prevalence of mycoplasmas in patients who showed urogenital symptoms and were excluded from other bacterial or fungal infections. The high positive rate in these samples highlighted the necessity of mycoplasmas examination, thus to facilitate the diagnosis and develop treatment strategy.

Previous research suggested that the infection level of Uu and
Mh was associated with gender and age. 15 In the current study, positive rates of three patterns of infection in females were much higher than in males. For one thing, the structure and environment of female genital system is more susceptible for the colonization of mycoplasmas. For another, the cervical specimen had the highest positive rate as reported in other studies. 13 Our study found that in Hakka population, the mycoplasmas infection rates peaked in individuals at the age groups of 16-20 and 36-50, and dramatically dropped in patients older than 55. The agespecific infection were similar to other studies. 7