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Bacterial Vaginosis in preterm labour at a tertiary care centre.

INTRODUCTION: Bacterial Vaginosis is of special public health concern in India because of the high burden of reproductive and pregnancy related morbidity. (1) BV is a very important cause of preterm labour. (2)

Since a long time, it has been conceived that genital tract infection during pregnancy, especially the cervix, may cause premature rupture of the membranes (PROM) and initiate preterm labor (PL) with subsequent maternal and perinatal morbidities and mortalities. Bacterial Vaginosis (BV) has been highlighted by its association with PL and PROM and other adverse pregnancy outcomes which represent a real challenge to obstetricians as they are the leading causes of high neonatal morbidity and mortality associated with prematurity.

Vaginal flora of a healthy woman is a dynamic ecosystem that can be altered easily. BV is defined as an imbalance in the normal vaginal flora with diminution in the normally-predominant lactobacilli and the proliferation of other anaerobic bacteria. A cause and effect relationship between BV and adverse pregnancy outcome can be better determined in cohort studies, in which BV is diagnosed before the onset of a number of pregnancy complications. In large percentage of cases of preterm labour (PL) the etiological factor is obscure and leads to an increased incidence of idiopathic PL.

BV is a very important cause of preterm labour (PL), hence diagnosis of BV plays a critical role in the outcome of the pregnancy.

Global literature suggests that asymptomatic or minimally symptomatic infections are related to premature births. BV is believed to be a risk factor for preterm delivery, as well as being associated with peripartum complications such as chorioamnionitis, and postpartum endometritis. (3)

BV has also been linked to several puerperal morbidities including fever, and endometritis. Many studies revealed the association of BV with established preterm labor.

Bacterial Vaginosis [BV] is the most common variety of vaginal infection in which the normal predominant vaginal flora Lactobacillus is replaced with Gardnerella vaginalis, anaerobic bacteria and Mycoplasma hominis. The prevalence of BV is relatively high in women with poor hygiene, malnutrition, improper sanitation.

BV is polymicrobial condition with a decrease in vaginal acidity (towards alkalinity 4.5-6.0) and decrease in Lactobacilli, accompanied by an increase in the number of other micro-organisms. Bacterial Vaginosis is currently thought to be a polymicrobial clinical syndrome distinguished by characteristic abnormalities of vaginal secretion and disturbances in vaginal ecology with displacement of normal lactobacillary flora by anaerobic microrganisms. (4) The term Vaginosis is used instead of vaginitis for this condition because there is no inflammatory response in vagina.

Routine screening for BV in higher risk group can reduce neonatal morbidity and mortality to significance extent in the developing countries.

This prospective study aims to screen for BV in patients with threatened PL and at high risk of PROM and to define a simplified screening plan applicable for the developing countries.

MATERIALS AND METHODS: The study was carried out in the Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences (RIMS), Kadapa from March 2014 to Mar 2015, involving 100 women admitted in the Department of Obstetrics and Gynaecology, RIMS, Kadapa with idiopathic preterm labour as per selection criteria mentioned below.

Inclusion Criteria:

1. Singleton pregnancy.

2. Gestational age between 28-36 wks.

3. Intact membranes.

4. Painful uterine contractions > 2 in 10 minutes each lasting > 45 seconds.

5. Cervical dilatation between 1 to 4 cm.

6. Cervical effacement > 25%.

Exclusion Criteria:

1. Gestational age <28 weeks.

2. History of Antepartum hemorrhage, urinary tract infection, respiratory tract.

3. Infections, Diarrhea or any other obvious cause for preterm labour.

4. Medical complications of pregnancy such as severe Anaemia, PIH, DM.

5. History of leaking membranes or absent membranes.

6. Multiple pregnancy.

7. Intrauterine growth restriction.

8. Intrauterine foetal death.

9. Antibiotic therapy in the last 30 days.

The study group included 100 women with preterm labour without any obvious cause for the same. Control group included 100 women carrying singleton pregnancy at term gestation. Detailed history taken, a thorough general and systemic examination was done to exclude exclusion criteria.

Patients were examined in lithotomy position. Local examination was done to exclude any local genital infections. Unlubricated cusco's speculum was introduced and per speculum examination was done after excluding leaking of the membranes.

Evaluation of pH: The vaginal secretion which collected on the posterior blade of Cusco's speculum was placed on a sterile glass slide for wet mount preparation. Vaginal pH measured by dipping pH paper into discharge collected on the speculum. The pH was tested by the pHydrion vivid refill papers with a pH range of 1 to 12 procured from the Himedia laboratory supplies. Color change was noted and the results were recorded.

Whiff Test (Amine Test): Whiff test was done by placing a drop of vaginal secretions on a sterile glass slide and few drops of 10% KOH (wt/vol) to the vaginal discharge and noting the emission of an offensive amine like odor (fishy odor) which indicated a positive amine test.

Wet Mount Examination: Vaginal secretions which were placed on sterile glass slide, diluted with a drop of normal saline and cover slip was placed over it and examined under Microscope (x40 magnification) for identification of clue cells, bacterial morphology and Trichomonas with special flagellated motility.

Evaluation of Vaginal Smear: With a sterile cotton swab the posterior vaginal wall was swept and was smeared on a clean and sterile glass slide. Smear is heat fixed and gram staining done and observed under Oil immersion objective (x1000).

A score of 0 to 10 was assigned based on Nugent's scoring system. In vaginal smear, three bacterial morphotypes are recognized. Lactobacillus morphotype (gram positive rods) Gardnerella vaginalis and bacteriodes species morphotypes (small gram negative to variable rods), Mobiluncus species morphotype (curved gram negative rods).

The amount of each morphotype detected on the smear is graded and then allocated a score as below.

Lactobacillus:     4+ = 0,   3+ = 1,    2+ =2,    1+ = 3,   0 = 4
Gardnerella and    0 = 0,    1+ = 1,    2+ =2,    3+= 3,    4-= 4
Bacteroides
Mobiluncus:        0 = 0,    + or       3+ or     4+= 2
                             2+ = 1,


The individual score are then summed. The criterion for Bacterial Vaginosis is a total score of 7 or higher, a score of 4 to 6 considered and a score of 0 to 3 is considered normal.

Findings of Gram Staining are classified as follows:

Normal Flora: Lactobacilllus morphotypes predominate with few other morphotypes. Intermediate flora--Lactobacillus morphotypes reduced with other Normal flora--Lactobacillus morphotypes dominate with other morphotypes increased.

Bacterial Vaginosis: Absent Lactobacillus with a great increase in other morphotypes.

Aerobic Vaginitis: A condition in which destructed vaginal flora has been replaced by Esch coli. Group B streptococci or enterococci.

STATISTICS AND RESULTS:

Table 1: Study Group (n = 100)

AGE           No. of       BV         BOH        Gram
(in years)     Cases    positive    History    Staining
                          Cases

< 19            02         --          --         --
20-24           80         39          10         38
25-29           14         03          03         04
30-34           02         --          01         --
>35             02         --          --         --
TOTAL           100        42          14         42

AGE           Whiff Test   Clue Cells   pH >4.5
(in years)     Positive     Positive

< 19              --           --          --
20-24             38           37          45
25-29             05           02          04
30-34             --           --          --
>35               --           --          --
TOTAL             43           39          49

Age           No. of       BV         BOH        Gram
(in years)     Cases    positive    History    Staining
                          Cases

< 19            08         02          --         02
20-24           55         13          03         11
25-29           27         03          06         05
30-34           07         --          01         --
>35             03         --          --         --
TOTAL           100        18          10         18

Age           Whiff Test   Clue Cells   pH >4.5
(in years)     Positive     Positive

< 19              02           02          02
20-24             15           08          17
25-29             01           05          08
30-34             --           --          --
>35               --           --          --
TOTAL             18           15          27

Control group (n = 100)

Table 2: Age distribution in study and control group (n = 100)

Age            Study     Control       BV
(in years)     Group      Group     Positive

<19              02         08         0
20-24            80         55         39
25-29            14         27         03
30-34            02         07         0
>35              02         03         0


In the study group of 100 cases, most (80) were in the age group of 20-24 yrs. In this group BV was seen in 39 cases. In under the 19 yrs age, there were 2 cases in the study group and 8 cases in the control group. None of these were positive for BV. In 25 to 29 yrs age group there were 14 cases in the study group and 27 in the control group. 3 in the study group were positive for BV.

In the 30-34 yrs age group, there were 2 cases in the study group and 7 in the control group. In the age group of above 35 yrs, 2 were in the study group and 3 were in control group. There were no cases of BV in these age groups.

In the study group, 42 women had BV and in the control group 18 had BV.

In the study group, raised pH (more than 4.5) was observed in 49 women where as in control group this was seen in 27 women.

Whiff test was positive in 43 women in the study group where as in the control group the test was positive in 18 women only.

Clue cells were identified and demonstrated in 39 women in the study group and in the control group only in 15 women.

Grams staining revealed--normal flora in 19 women, intermediate flora in 21 women, bacterial Vaginosis in 42 women and scanty flora in 16 women. Gram staining observations revealed Nugent score of 7-10 in all the cases of bacterial Vaginosis. (Table 7).

In the study group, 9 women showed the evidence of Trichomoniasis. Out of these, 6 women were diagnosed to be having BV. Candidiasis was seen in 21 women in the study group, out of which 3 were positive for BV. (Table 8).

Bad obstetric history suggestive of abortions and preterm labours was recorded in 14 cases of the study group. Out of these 14 cases, 6 women were diagnosed to be having BV. In 86 women there was no suggestive bad obstetric history. In this category 36 were diagnosed to be having BV. (Table 9).

In the study group, 32 cases and in the control group 14 cases were positive for BV in the gestational age of less than 34 weeks. In the gestational age of more than 34 weeks 10 cases in study group and 3 cases in control group were positive for BV (Table 10).

DISCUSSION: Study group of 100 women and a control group of 100 women admitted in the delivery room were tested for Bacterial Vaginosis. The diagnostic criteria in the present study were gram staining and Nugent criteria.

In the present study, 42 had BV (42%). This finding is almost similar to the work done by Vineeta Gupta et al (43.6%). (13) Similar studies by several authors yielded varying results which differ from our study. Kumar Aruna et al (12) reported 30.35%, Saharan SP et al (6) reported 37.5%, Atef M Darwish et al (8) reported 33.3%, Nelofar Saleem et al (11) reported 55.38% in their studies. The varying incidence of BV by different authors may be due to the factors like, different sample size, different geographic and socio-economic situations.

Almost similar incidence of BV by Vineeta Gupta et al (13) may be attributed to the similar sample size and the diagnostic criteria employed. Two diagnostic tests are commonly used for BV. Amsel criteria, (14) the test most commonly used in the clinical setting involves clinical conditions. There are inherent difficulties with each of the individual parameters in the Amsel criteria. The second most commonly used diagnostic test involves a Gram stain of vaginal discharge and applying Nugent criteria. The Nugent criterion (15) is the test most often used in epidemiologic studies in large scale, this method has several advantages that include 1) creating a permanent record that can be subsequently reviewed to confirm the diagnosis of BV and assess the reliability of the reading, 2) reporting intermediate stages of BV, which is particularly useful and 3) quantifying the amount of the three individual organisms, enabling assessment of the organism-specific risk of disease. These advantages are almost achieved in both the studies mentioned above. Thus, Nugent criteria is supposed to be ideal in diagnosing BV in the laboratory.

Gram staining in the present study revealed normal flora 19%, intermediate flora 21%, BV 42% and scanty flora 16%. A study by P Madhivanan et al (1) revealed normal flora 72.3%, intermediate flora 21% and BV in 68%.

In the present study, raised pH was seen in 49 cases accounting for 49%. All the BV diagnosed women showed raised pH. This is a very simple and rapid test which can be adapted to any health care setting. In a study by Atef M. Darwish et al (8) all the study group women with BV showed raised pH i.e., more than 4.5. Our study findings also reveal the same. Several studies in the developing countries for the screening of BV revealed similar findings. All these studies pointed out that for the diagnosis of BV the most sensitive test was pH of more than 4.5 due to its high negative predictive value coupled with the fact that it is an extremely simple and economical test to perform. Some other studies revealed a higher percentage of raised pH in BV.

Whiff test is most significant bed side test with high sensitivity, high specificity and high negative predictive value for diagnosis of BV. In our study Whiff test was positive for 43 cases which is less when compared to the study done by Atef M. Darwish et al (8) (75.6%).

In present study, 32 cases were positive for BV in the gestational age of less than 34 weeks which can be compared with study done by Desai Veena A et al (10) (31.73%).

In a study conducted by Awassanan et al, (3) 15% cases were positive for BV in gestational age group less than 34 weeks and 15% in age group more than 34 weeks.

In our study group BOH was present in 14 cases out of which 6 were positive for BV; in control group BOH was present in 10 cases out of which 2 were positive for BV. This observation is in agreement with study conducted by Philip E Hay et al (16) who has shown that BV is associated with preterm delivery independent of recognized risk factors such as BOH.

Bacterial Vaginosis is one of the health problem accounting for the majority of cases of vaginitis and vaginal discharge in the developing countries. BV is an extremely prevalent condition in pregnancy, the true magnitude is not known because more than half of the BV cases are asymptomatic. This study focuses the importance of Nugent criteria and gram staining in the diagnosis of BV as a routine practice in the pregnant women. By this approach, the complications associated with BV in pregnancy can be prevented by diagnosing in early gestational age. This definitely reduces the adverse outcome of pregnancy associated with BV such as preterm labour, premature rupture of membranes, neonatal morbidity and mortality.

DOI: 10.14260/jemds/2015/1277

BIBLIOGRAPHY:

(1.) P. Madhivanan, K Krupp, C Karat, A Arun, CR Cohen et al. Prevalence and correlates of Bacterial Vaginosis among young women of reproductive age in Mysore, India. Indian J Med Microbiology. 2008; 26 (2): 132-7.

(2.) Bhalla P, Chawla R, Garg S et al. Prevalce of Bacterial vaginosis in Indian. J Dermatol Veneol Leprol 1994; 60: 8-14.

(3.) Awassanan Thanavuth, Amphan Chalemchockcharoenkit et al. prevalence of Bacterial Vaginosis in Thai pregnant women with preterm labour in Sirirag Hospital. J Med assoc Thai 2007; 90 (3): 437-41.

(4.) Azam Azargoon MD, Shohreh Darvishzadeh MD et al. Association of Bacterial Vaginosis, Trichomonas vaginalis and vaginal acidity without come of pregnancy. Rch Iranian Medicine 2006; 9 (3): 213-217.

(5.) Sharon L. Hillier, Robert P. Nugent, David A. Eschenbach, Marijane A. Krohn. Association between Bacterial Vaginosis and preterm delivery of a low- birth weight infant. New England J. 1995; Vol.333, No. 26.

(6.) Saharan SP, Surve C, Raut V, Bhattacharya M et al. Diagnosis and prevalence of Bacterial Vaginosis. J Postgrad Med. 1993; 39 (2): 72-73.

(7.) Carey JC, Kelbanoff MA, Hauth JC, Hillier SI, Thom EA, Ernest JM et al. Metronidazole to prevent preterm delivery in pregnant women with asymptomatic bacterial vaginosis. NICHD Network of Maternal and fetal medicine units. N Engl J med 2000; 342: 534-540.

(8.) Atef M Darwish, Mohammad H. Makaram, Ehab M Alnashar, Suha M. Hamadeh Screening for Bacterial Vaginosis in high risk pregnancy: The experience of a developing country.

(9.) C Karat, P Madhivanan, K Krupp, S Poornima et al. The clical and microbiological correlates of premature rupture of membranes. Ind J Med Microbiol 2006; 24 (4): 285-5.

(10.) Desai Veena A, Verma Ragini et al. Bacterial Vaginosis in patients with Idiopathic preterm labour. J Obstet Gynaecol India 2009; Vol.59, No. 1: Jan 53-57.

(11.) Nelofer Saleem, Habiba Sharaf Ali, Rubina Hussin et al. Prevalence of Bacterial Vaginosis in pregnant women and efficacy of rapid diagnostic tests in its diagnosis. Infectious Disease J Pakistan, 2006; Oct/Dec: 93-95.

(12.) Kumar Aruna, Khare Jyothi et al. Roleof Bacterial Vaginosis in preterm labour. J Obstet Gynaecol India, 2007; 57: 413-416.

(13.) Vineeta Gupta, Pratima Gupta et al, Clinicomicrobiological profile of women with vaginal discharge. J Ind Med Assoc, Vol 107, No 3, March, 2009.

(14.) Amsel R, Totan PA, Spiegel CA, Chen KCS, Eschenbach DA, Holmes KK. Non-specific vaginitis: Diagnostic Criteria and Microbiological and Epidemiologic association. Am J Med 1983; 74: 14-22.

(15.) Nugent RP, Krohn MA, Hillier SL, Reliability of diagnosing bacterial vaginosis is improved by a standardized method of gramstain interpretation. J Clin. Microbiology. 1991; 29: 297-31.

(16.) Phillips E. Hay et al Abnormal colonization of genital tract and subsequent preterm delivery. BMJ Vol. 308.

Avula Shashikala [1], Bathala Nagasrilatha [2], Majeti Sasidhar [3], Barabari Manmohan [4], Thulamandi Lakshmi Suseela [5]

AUTHORS:

[1.] Avula Shashikala

[2.] Bathala Nagasrilatha

[3.] Majeti Sasidhar

[4.] Barabari Manmohan

[5.] Thulamandi Lakshmi Suseela

PARTICULARS OF CONTRIBUTORS:

[1.] Assistant Professor, Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences, Kadapa.

[2.] Associate Professor, Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences, Kadapa.

[3.] Professor, Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences, Kadapa.

FINANCIAL OR OTHER

COMPETING INTERESTS: None

[4.] Assistant Professor, Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences, Kadapa.

[5.] Associate Professor, Department of Gynaecology, Rajiv Gandhi Institute of Medical Sciences, Kadapa.

NAME ADDRESS EMAIL ID OF THE CORRESPONDING AUTHOR:

Dr. Avula Shashikala, Assistant Professor, Department of Microbiology, Rajiv Gandhi Institute of Medical Sciences, Kadapa-516002.

E-mail: dr.asasikala@gmail.com

Date of Submission: 01/06/2015.

Date of Peer Review: 02/06/2015.

Date of Acceptance: 17/06/2015.

Date of Publishing: 23/06/2015.

Table 3: Incidence of Bacterial Vaginosis

BV                  Study Group    Control Group

Women with BV            42              18
Women without BV         58              82

Table 4: Comparison of vaginal pH in both groups

PH          Study group        Control Group
         No. of Cases (%)    No. of Cases (%)

< 4.5           51                  73
> 4.5           49                  27

Table 5: Comparison of Whiff test among study and control groups

Whiff test      Study Group        Control Group
              No. of Cases (%)    No. of Cases (%)

Positive             43                  18
Negitive             57                  82

Table 6: Comparison of Clue cells in both groups

Clue cells       Study group        Control group
              No. of Cases (%)    No. of Cases (%)

Present              39                  15
Absent               61                  85

Table 7: Comparison of results of Gram staining

Gram Staining             Study Group        Control Group     Nugent
Results                    (n = 100)           (n = 100)        Score

                       No. of Cases (%)    No. of Cases (%)

Normal flora                  19                  45             0-3
Intermediate flora            21                  22             4-6
Bacterial Vaginosis           42                  18            7-10
Scanty flora                  16                  14
Aerobic Vaginitis             02                  01             --

Table 8: Association of Trichomoniasis and candida co-infection in the
study group

Name of           Positive    Negative    Total number    BV positive
confection                                  of cases

Trichomoniasis       09          91            100             6
Candidiasis          21          79            100             3

Table 9: BV distribution according to BOH

Bad Obstetrics    Study       BV       Control       BV
History (BOH)     group    positive     group     positive

PRESENT             14        06          10         02
ABSENT              86        36          90         16

Table 10: Incidence of BV according to gestational age

Gestational Age     BV positive cases    BV negative cases
(in weeks)
                   Study     Control    Study     Control
                    group     group      group     group

<34                  32         14        40         76
>34                  10         03        18         07

Table 11: Incidence of Bacterial Vaginosis

Sl. No.                  Author                  Incidence of BV

1               Madhivanan et al (1) 2008             19.0%
2          Awassanan Thanavath et al (3) 2007         19.0%
3             Azam Azargoon et al (4) 2006            16.0%
4           Saharan L Hillier et al (5) 1995          16.0%
5               Saharan SP et al (6) 1993             37.5 %
6                Carey JC et al (7) 2000              22.2%
7             Atef M Darwish et al (8) 2001           33.3%
8                Karat C et al (9) 2002               24.0 %
9              Desai veena et al (10) 2004            18.7 %
10           Nelofar Saleem et al (11) 2006           55.38%
11             Kumar Aruna et al (12) 2006            30.35%
12            Vineeta Gupta et al (13) 2009           43.6 %
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Title Annotation:ORIGINAL ARTICLE
Author:Shashikala, Avula; Nagasrilatha, Bathala; Sasidhar, Majeti; Manmohan, Barabari; Suseela, Thulamandi
Publication:Journal of Evolution of Medical and Dental Sciences
Date:Jun 25, 2015
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