Characteristic Description of Multiparous Women

with

Mycoplasma hominis

and

Ureaplasma urealyticum

Infection

at Outpatient Clinic in Medan

L. Feriyawati

, D. R. Anggraini

and T. A. Nasution

Anatomic Department, Medical Faculty, Universitas Sumatera Utara, Medan,

Indonesia

Microbiology Department,Medical Faculty, Universitas Sumatera Utara, Medan,Indonesia

Keywords: Mycoplasma hominis, Ureaplasma urealyticum, Multiparous Women.

Abstract:

Among the organisms that cause genital infection are

Mycoplasma hominis

and

Ureaplasma urealyticum

which

are commensally in women

urogenital

but may become pathogens and were associated as a cause of

complications of genital tract infection in pregnant women, such as ascending chorioamnionitis, premature

rupture of membranes, preterm birth, miscarriage, weight and neonatal birth low and newborn deaths..The

objective of this study was to describe the characteristic of multiparous women with

Mycoplasma hominis

and

Ureaplasma urealyticum

infection. Design of the study

was

a descriptive study with cross-sectional approach

collecting cervical swab of 50 multiparous women

from

outpatient clinic in Medan. Duplex PCR assay was

perfomed using two primers: RNAH1 and RNAH2 that amplify the 16sRNA

M. Hominis

gene at 334bp;

UMS125 and UMA226 amplifying serovar3 genes multiple banded antigen which can amplify biovar1 that

appeared at 403bp and biovar2 appeared at 448bp. The result of this study showed

Mh-Uu

duplex PCR results

revealed that 3(6.0%) respondents were positive Mycoplasma infection and 9(18%) respondents were positive

Ureaplasma infection and 5(10.0%) from infected respondents had abortion history.

1 INTRODUCTION

Mycoplasma hominis and Ureaplasma spp. is a

commensal organism that found in 30-80% women’s

urogenital tract, thus causing urogenital tract

infection. In pregnant woman, these organisms could

stay in utero and then transmission by placenta to the

fetus, causing several infections and trigger

premature labor (Otgonjargala, 2017). In rare cases

these microorganisms may infect the central nervous

system in healthy neonates and present a risk of

severe complications and poor prognosis

(Wildenbeest, 2016). The role of these pathogens in

women with chronic urinary tract symptoms remains

a problem due to difficult to detect and its

intracellular nature makes conventional antibiotics

ineffective (Nasution, 2007).

Polymerase Chain Protein (PCR) analysis of

these bacteria should be performed if symptomatic

sterile leukocytosis is present, chronic urethritis

and bladder hyperactivity or interstitial cystitis/

painful bladder syndrome, recurrent infections or if

microbiological culture is negative (Combaz-

Söhnchen, 2017). The difficulty of detecting M.

hominis on persistent neonatal CNS infections with

unknown causes requires diagnostic protocols using

a specific real-time PCR. Physicians should be

aware of the pathogens as possible causes of

neonatal meningoencephalitis if corrective failure is

found in empirical antibiotic treatment

(Wildenbeest, 2016).

A study by Manhart et al, 2003, found that from

719 young women Mycoplasma genitalium was

detected as much as 50 (7%) in a sample previously

negative for bacterial vaginosis. Nasution et al, 2007,

in 40 Malaysia women, used duplex PCR Mh / Uu

method (Mycoplasma hominis/ Ureaplasma

urealyticum) and showed that Ureaplasma is the most

commonly discovered pathogen (positive in 90.5% of

women and 47.5% of newborns), followed by

Mycoplasma (32.5% and 7.5%), and the rest are

chlamydia, trichomonas and gonococcus.

The purpose of this study is to describe the

characteristic of multiparous women with

Feriyawati, L., Anggraini, D. and Nasution, T.

Characteristic Description of Multiparous Women with Mycoplasma hominis and Ureaplasma urealyticum Infection at Outpatient Clinic in Medan.

DOI: 10.5220/0010093108370840

In Proceedings of the International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches (ICOSTEERR 2018) - Research in Industry 4.0, pages

837-840

ISBN: 978-989-758-449-7

837

Mycoplasma hominis and Ureaplasma urealyticum

infection detected by using duplex PCR method.

2 METHOD

2.1 Study Design

The study design is cross sectional with observational

approach.

2.2 Study Area

This study was conducted at an outpatient clinic in

Medan North Sumatera in 2018.

2.3 Sampling

Protocol of this study has been approved by Medical

Ethics Committee Universitas Sumatera Utara

(No.375/TGL/KEPK FK USU-RSUP HAM/2018)

2.2.1 Respondents Characteristics

Respondents were 50 female adolescents,

multiparity, age above 18 years and signed the

informed consent.

3 DATA COLLECTION

3.1 Socio-demographics Background

A questionnaire consist of social demographics, using

contraception, first age of sexual intercourse and

history of abortion was administered by self-reports

of the participants,

3.2 Sample Collection and Laboratory

Processing

3.2.1 DNA Extraction

Cervical swabs from 50 respondents on the micro

tube contained 0.9% NaCl firstly centrifuged at a

speed of 14,000rpm for 4 minutes. The precipitated

portion is added to 1.5 ml of the PBS (phosphate

buffer salin). After that tube were repeating

centrifuged at a speed of 14.000 rpm for 4 minutes.

Furthermore, a DNA isolation procedure is performed

based on the protocol of the Invitrogen® kit. There

are 200 µL sample inserted into another micro tube.

Moreover, inside the tube was added 20 µL of

proteinase K and 200 µL of lysis buffer, then vortexed

for a few minutes. After that, the tube was incubated

at 55°C for 10 minutes. Followed step is the tube were

added 250 µL ethanol 96%, then vortexes for 15

seconds and move the fluid into the column spin.

Then the column was centrifuged at 10.000 rpm for 2

minutes. After that, replace the collection tube, then

washed with 500 µL wash buffer 1. Centrifuged at

10.000rpm for 2 minutes. Next step was replacing the

collection tube then washed again with 500 µL wash

buffer 2, centrifuge with maximum speed

(14.000rpm) for 3 minutes. Finally, replaced the

collection tube again and added 50 µL delution

buffer, then tube was centrifuged at 14.000rpm for 1.5

minutes.

3.2.2 Mycoplasma hominis and Ureaplasma

urealyticum Detection

M. hominis and U.urealyticum duplex PCR (Mh-Uu

duplex PCR), using 2 primers: RNAH1 and RNAH2

that amplify the 16sRNA M. hominisgene at 334 bp;

UMS125 and UMA226 amplifying serovar 3

genes multiple banded antigen which can amplify

biovar1 that appeared at 403 bp and biovar 2 appeared

at 448 bp. The amplification mixture was carried out

in 12,5l master mix PCR which consists of Taq

polymerase enzyme, MgSO

, and dNTP (Go Taq®

PCR Core System, Promega); 7,5 l nuclease-free

water and 4l DNA template. PCR was performed in

a thermocycler (Verity 96-well Thermal Cycler,

AppliedBiosystems) with an initial denaturation

C for 1 minute 30 seconds, annealing in 55

C for

2 minutes, extension for 1 minute 30 seconds and

ending with a final extension step at 72

C (Nasution

2007).

4 RESULTS AND DISCUSSION

4.1 Socio-demographics Background

In this study, respondents mostly women between 35-

40 years old (66%), followed by women above 45

years old (22%) and the least under 35 years old

(12%). Most respondents were housewife (58%)

Respondents whose using contraception accounted

for 26%. The first age of sexual intercourse was found

mostly women between 26-30 years old (56%),

following women between 20-25 years old (40%) and

the least were under 20 years old (4%). The

respondents whose having abortion history accounted

for 23%.

ICOSTEERR 2018 - International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches

838

Table 1: Socio-demographics characteristic

No Socio-demographics

characteristic

Number of

Respondent

(n=50)

1 Age

< 35 years

35-40 years

> 45 years

12.0

66.0

22.0

2 Occupation

Working woman

Housewife

58.0

42.0

3 Use of contraception

Yes

26.0

74.0

First age of sexual

intercourse

< 20 years old

20 – 25 years old

26 – 30 years old

4.0

40.0

56.0

History of abortion

Yes

46.0

54.0

4.2 Polymerase Chain Protein

Mh-Uu duplex PCR results revealed that 3(6.0%)

respondents were M. hominis positive and 9(18.0%)

respondents were U. urealyticum positive (Figure. 1)

Table 2: Distribution of detection Mycoplasma hominis

(Mh) and Ureaplasma urealyticum (Uu) using Duplex PCR

n No (%) Detection

Mh Uu

Age

< 35 years

35-40 years

> 45 years

1(16.0)

2(6.06)

2(33.0)

4(12.1)

3(27.2)

Occupation

Working woman

Housewife

3(10.3)

4(19.0)

5(17.2)

Use of contraception

Yes

3(8.1)

2(15.4)

7(0.2)

First age of sexual

intercourse

< 20 years old

20 – 25 years old

26 – 30 years old

1(5.0)

2(7.1)

6(30.0)

3(10.7)

History of abortion

Yes

3(13.0)

2(8.7)

7(25.9)

Figure 1: Mh-Uu duplex PCR results of 50 respondents

This study found that the respondents who had

Mycoplasma infection were all housewives with the

most 35-40 years old range. It is interesting that

almost all respondents have had abortion; even one of

the respondents had experienced abortion twice.

While respondents who have Ureaplasma infection

were most are housewives with the most age range

35-40 years old. Abortion history is also found in this

group who was 2 respondents ever aborts 1 time.

This study was consistent with another previous

study conducted in a cohort of females with bacterial

vaginosi (BV) or asymptomatic, in which U.

urealyticum infection was detected significantly more

often than M. Hominis (Verteramo, 2013 and Padang,

2015)

The high prevalence of Ureaplasma spp. infection

was also found according to previous study in Brazil

with 6,810 patients with the age range of 11 to 80

years who in gynecological routine examination,

found that M. hominis (n = 79), Ureaplasma spp. (n =

2,026) and the co-colonization of both (n = 199)

which extract from cytological sample and detect by

PCR (Milanezi, 2016). Another study in Iran using

multiplex PCR to urine and genital samples from

symptomatic females (20-54 years old), found that

the highest incidence of M. hominis and U.

urealyticum and were highly associated with habitual

abortion in symptomatic females (Maleki, 2013). The

inconsistent result found in case control study in Iran

which conclude that no association between

mycoplasma infection and spontaneous abortion

(Ramazanzadeh, 2016).

Characteristic Description of Multiparous Women with Mycoplasma hominis and Ureaplasma urealyticum Infection at Outpatient Clinic in

Medan

839

5 CONCLUSION

This study has revealed that from 50 multiparous

women, 3(6.0%) were positive Mycoplasma infection

and 9(18.0%) were positive Ureaplasma infection and

had abortion history. Further research is needed to

explore whether the cause of their previous abortion

was associated with this bacterial infection. Early

screening is needed to detect bacterial infection of M.

hominis and U. urealyticum in pregnant women with

a history of preterm and premature rupture of

membranes, thus decreasing the complication of this

infection to newborns’ morbidity and mortality

ACKNOWLEDGMENTS

The authors would like to thank the funding support

from the Universitas Sumatera Utara.

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