• Users Online: 102
  • Print this page
  • Email this page


 
 Table of Contents  
ORIGINAL ARTICLE
Year : 2020  |  Volume : 4  |  Issue : 2  |  Page : 67-72

Bacterial vaginosis and its association with human papilloma virus and increased risk of cervical intraepithelial lesions: An experience from Eastern India


1 Department of Gynaecological Oncology, Acharya Harihar Post Graduate Institute of Cancer, Cuttack, Odisha, India
2 Department of Zoology, Revenshaw University, Cuttack, Odisha, India
3 Department of Microbiology, S.C.B. Medical College, Cuttack, Odisha, India
4 Department of Pathology, S.C.B. Medical College, Cuttack, Odisha, India

Date of Submission25-Feb-2019
Date of Decision12-Jun-2020
Date of Acceptance23-Jul-2020
Date of Web Publication17-Aug-2020

Correspondence Address:
Manoranjan Mohapatra
Department of Gynaecological Oncology, Acharya Harihara Post Graduate Institute of Cancer, Cuttack - 753 007, Odisha
India
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/oji.oji_9_19

Rights and Permissions
  Abstract 


Background: Bacterial vaginosis (BV) is the most common vaginal disorder affecting women of reproductive age and has an influence in acquisition of certain genital infections. However, it is difficult to determine whether BV is actually a risk factor for human papilloma virus (HPV) acquisition or not and whether it may lead to cervical cancer. Aim of the Study: The aim of the present study is to determine the prevalence of BV and its association with HPV infection and cervical intraepithelial neoplasia (CIN). Materials and Methods: A multiinstitutional prospective study was conducted to analyze the vaginal samples collected from the women aged between 19 and 49 years during the period from December 2014 to January 2018. Results: A total of 333 women with vaginal samples were analyzed for BV. 103 (30.93%) samples were diagnosed with BV as per Amsel's criteria. The prevalence of HPV DNA was higher in BV-positive cases in comparison to that of BV-negative cases (44.7% vs. 9.6%; P = 0.000) showing a significant association between BV and HPV infection. There was a rising trend in the incidence of CIN for women diagnosed with BV when compared to BV negative women (62.1% vs. 43%; P = 0.506), although statistically insignificant. Moreover, the severity/high grading of CIN was not significantly associated with BV (P = 0.765). Conclusion: The result of our study hypothesized that BV was significantly associated with increased risk of HPV infection. There was a rising trend for the association of BV with CIN incidence although statistically insignificant.

Keywords: Association, bacterial vaginosis, cervical intraepithelial neoplasia, human papilloma virus


How to cite this article:
Nayak B, Patnaik P, Mohapatra M, Soren D, Patra P, Besra K, Giri SK. Bacterial vaginosis and its association with human papilloma virus and increased risk of cervical intraepithelial lesions: An experience from Eastern India. Oncol J India 2020;4:67-72

How to cite this URL:
Nayak B, Patnaik P, Mohapatra M, Soren D, Patra P, Besra K, Giri SK. Bacterial vaginosis and its association with human papilloma virus and increased risk of cervical intraepithelial lesions: An experience from Eastern India. Oncol J India [serial online] 2020 [cited 2020 Oct 27];4:67-72. Available from: https://www.ojionline.org/text.asp?2020/4/2/67/291906




  Introduction Top


Bacterial vaginosis (BV) is one of the most prevalent causes of abnormal vaginal discharge affecting women of reproductive age groups. It is characterized by the loss of indigenous Lactobacillus predominant vaginal flora and a concurrent massive overgrowth of anaerobic bacteria. Cervical cytological abnormalities are more commonly seen in women with altered vaginal flora which suggest an existence of possible link between BV and the cervical cancer development.[1],[2] Several studies have focused on the role of BV/vaginal microbiome for the acquisition and persistence of human papilloma virus (HPV) and cervical cancer development but with inconsistent findings.

Cervical cancer is the second most common malignancy among female worldwide.[1] Instead of the development in prevention strategies, the burden of disease remains a health problem, particularly in developing countries. Persistent genital HPV infection, particularly the high-risk groups are the basic concept for the development of cervical cancer. However, infection by oncogenic HPV is not a sufficient cause of cervical cancer. It has been assumed that additional cofactors in conjunction with HPV modulate the risk of transition from cervical HPV infection to cervical malignancy. The factors that may have a role in this progression include young age at first intercourse, multiple sexual partners, smoking, contraceptive use, low socioeconomic status, nutrition, and infection with sexually transmitted diseases (STDs), such as BV, chlamydia trachomatis, and trichomonas vaginalis.[3],[4]

Several hypotheses exist toward increased acquisition of HPV infection in women with BV and create a conductive environment for carcinogenesis. These are lactobacillus poor flora resulting changes in the vaginal ecosystem, increased mucin degrading sialidases in the vaginal fluid destroying the protective mucosal barrier, changes in the production of factors such as cytokines affect immunological balance within the cervical tissue, and production of carcinogenic nitrosamines increase the probability of DNA damage and altered cytokine profile reducing immune system's ability to eliminate HPV infection.

The magnitude of association of BV with HPV infection and cervical intraepithelial neoplasia (CIN) incidence has varied in epidemiological studies and remains controversial yielding conflicting results and ranging from the absence of any association to a clear positive relationship.[1],[2],[5] In this context, there is an acute need for specific clinical research and survey studies to determine whether BV is actually a risk factor for HPV acquisition or not and whether it may lead to cervical cancer. Therefore, the present prospective study was conducted to clarify the correlation of BV with HPV infection and CIN incidence.


  Materials and Methods Top


The present multiinstitutional, prospective study was conducted in the department of gynecological oncology of AHPGIC, Cuttack, in collaboration with the department of obstetrics and gynecology, microbiology, and pathology of SCB Medical College, Cuttack, from the Eastern zone of India. Women with the symptoms of presentation for the genital disease aged between 19 and 49 years attending out-patient department of both gynaecological oncology, and obstetrics and gynecology during the period from December 2014 to January 2018 were included in the study. The study was performed in accordance with the Helsinki Declaration (2000). The informed written consent was obtained from all participants before enrolment into the study. The study protocol was approved by the Institutional Ethics Committee (registration number 1045/E-74/2013).

Women who were unmarried, pregnant, having vaginal bleeding at the time of clinical examination, having urinary or fecal incontinence, who used antibiotics within 72 hour prior to presentation, and who have undergone hysterectomy were excluded from the study. A structured questionnaire was used to collect the information from the participating women regarding their socioeconomic status, douching, smoking, history of previous preterm deliveries, age of sexual debut, number of sexual partners during their life time, current contraception method, and history of STDs if any.

Four high vaginal swabs were collected for each patient by sterile cotton tip swabs from posterior vaginal fornix, the swabs were dipped in 0.5 ml normal saline, and the characteristics of vaginal discharge such as color, nature, consistency, and odor were recorded. Smear from the cervix was collected using Ayres spatula. Additional endocervical cells were collected using a Dracon swab and were transferred into tubes containing phosphate-buffered saline stored at −70°C until further processing. Extraction of total DNA was done using QI Amp DNA mini kit according to the manufacturer's instructions. The 4th swab was preserved for sialidase assay.

Diagnosis of BV was done by the Amsel criteria and Nugent scoring.[6],[7] In Amsel criteria, the clinicomicrobiological diagnosis of BV is considered when 3 out of 4 findings are present such as (a) vaginal PH >4.5, (b) a thin milky white homogeneous discharge independent of color and quantity, (c) positive whiff-amine test, i.e., accentuation of the fishy odor of the discharge with addition of 10% potassium hydroxide (KOH) and (d) >20% clue cells on microscopic examination of vaginal swabbing samples in saline. As per the Nugent scoring system, the Gram-stained vaginal contents are interpreted as Lactobacillus-predominant normal vaginal microbiota, intermediate, and BV-like conditions when score values are 0–3, 4–6, and 7–10, respectively.

The pH of the vaginal discharge was recorded using standard pH indicator paper with a range of 2–10.5. The amine test was performed by adding a few drops of 10% KOH solution directly over the soaked swab to find out if there was emission of ammine-like odor. The wet preparation was assessed for the presence of clue cells, motile trophozoites of trichomonas vaginalis, and budding yeast cells. One of the swabs was stained by Gram-staining technique. The film was examined for the presence of clue cells, Gram-negative or Gram variable bacilli, budding yeast cells, polymorphs, etc. A scoring of the vaginal flora pattern was done according to the scoring system of Nugent Scoring.

The results of PAP smears were reported according to the Bethesda III system. DNA extraction done from cervico-vaginal smear by manual extraction using QIAamp DNA Kits. Polymerase chain reaction amplification of the extracted DNA was performed using the consensus degenerate primers (MY09 and MY11) derived from the highly conserved L1 open reading frame of the HPV genome with an amplicon size of 450 bp. Sialidase assay was conducted using BV Blue Test Kit. The results are interpreted on the basis of color change. A positive result is indicated by the appearance of blue/green color in the test vessel or on the head of the swab, while the appearance of yellow color indicates a negative result.

Descriptive statistics were expressed as percentages. The association of the presence of BV with sociodemographic factors was calculated using the Chi-square test. The measures of diagnostic accuracy such as sensitivity, specificity, positive predictive value (PPV), and negative predictive value were evaluated for clue cell and amine test separately. Furthermore, the association between BV with CIN, ASCUS, and severity of CIN and HPV infection was evaluated using the Chi-square test.


  Results Top


The present prospective study comprises the evaluation of vaginal/cervical samples/swabs collected from 368 women. Among these, 26 (7.06%) were positive for Candida species and 9 (2.44%) revealed trichomonas vaginalis. Hence, these 35 cases were excluded from further study.

A total of 333 cases were analyzed for the diagnosis of BV. According to Amsel's criteria, 103 out of 333 women (30.93%) were diagnosed with BV (≥3 criteria), and the rest 69.07% women were BV negative. According to Nugent's criteria, Gram stains were positive for BV in 26.12% of cases [Table 1]. In our study, BV diagnosed cases were common among women aged 35–39 years (33.01%), and majority of cases above 34 years age (69.9%).
Table 1: Bacterial vaginosis diagnosed by Amsel's criteria and Nugent's scoring (n=333)

Click here to view


Out of the total 333 women included in the study, majority of cases belonged to Hindu, literate, coming from urban slums, and were not working/homemaker group constituting 73.27%, 72.97%, 59.76%, and 74.17% of cases, respectively. While comparing the demographic characteristics of BV diagnosed women with BV undiagnosed cases, we found significantly higher illiterate rate (58.25% vs. 13.04%; P = 0.000), a smaller number of urban slum location (38.83% vs. 69.13%; P = 0.0004), relatively higher number of women with first sexual contact before 18 years of age (72.82% vs. 54.35%; P = 0.001), and relatively more women of unskilled laborers (31.07% vs. 15.22%; P = 0.003) among BV diagnosed women. There was no significant difference in other base line demographic characteristics such as religion and number of sexual partners between the BV positive and negative cases [Table 2].
Table 2: Association of bacterial vaginosis with socio demographic and risk factors

Click here to view


Abnormal vaginal discharge and lower abdominal pain were the common symptoms encountered in the study, and these symptoms were more commonly seen in BV-positive cases. Abnormal vaginal discharge was the most common presentation among BV diagnosed cases by Amsel's criteria consisting of 84.47% (87/103) cases, followed by lower abdominal pain (63.11%; 65/103), both and Pruritus. Whereas, lower abdominal pain was the most common presentation among women with BV-negative cases followed by others.

Clue cells were positive in 40/103 BV diagnosed cases, whereas amine test found positive in 85/103 BV cases. The sensitivity and specificity of clue cells were found to be 38.83% and 98.7%, respectively, whereas Amine test had higher sensitivity (82.52%) but lower specificity (84.78%). However, both the above criteria were significantly associated with BV with P = 0.000 for each [Table 3].
Table 3: Statistical evaluation of clue cell and Amine test in the diagnosis of bacterial vaginosis

Click here to view


Sixty eight of 333 vaginal samples (20.42%) examined were positive for HPV DNA. However, when compared, HPV DNA was positive more in BV-positive cases than that of BV-negative cases (44.66% vs. 9.57%; P = 0.000). We suggested the presence of significant association between HPV infection and BV. The incidence rate of CIN in our study was 48.95%. There was a relatively rising trend of CIN incidence among BV diagnosed cases (62.14%; 64/103) than that of BV-negative cases (43.04%; 99/230) with the P = 0.506. When CIN patients were subdivided into low grade CIN 1 group and high grade CIN II/III groups, both low grade and high grade CIN had no significant difference between both the groups (P = 0.765) showing the presence of BV was not associated with the severity of CIN [Table 4].
Table 4: Association between bacterial vaginosis with cervical intraepithelial neoplasia, atypical squamous cells of undetermined significance, and severity of cervical intraepithelial neoplasia and human papilloma virus infection

Click here to view



  Discussion Top


The prevalence of BV in different clinical settings varies widely from 10% to 64%.[8],[9],[10] Such wide variation in BV prevalence may be due to the presence of different patient populations, demographical variation, and variation in diagnostic criteria among different studies. Studies carried out for the prevalence of BV between developed and developing countries did not differ significantly. However, this insignificant difference cannot be extrapolated easily due to the heterogeneity in study, and the less studies from developing countries included in the analysis.[1] In the present study, the overall presence of BV was 30.93% supported by the published data. A Nigerian study by Abdullateef et al. revealed BV prevalence of 40.1% among asymptomatic women of reproductive age.[11] An earlier study by Sodhani et al. reported 41.5% of the prevalence of BV in urban slum of Delhi.[8] Nugent's scoring of a Gram-stained smear is the gold standard method to diagnose BV. However, Gram stain is rarely available or utilized in clinical settings, and Nugent's scoring may not be performed on all women. Amsel's criteria is most commonly used for the diagnosis of BV.[12] Following Amsels microbiological criteria and Nugent's score, we found 30.93% and 26.12% women were diagnosed with BV, respectively.

In our study, BV was common among women aged 35–39 years (33.01%). We found majority of BV cases above 34-years age (69.9%). However, majority of research revealed increase in BV detection rate among younger women. According to Abdullateef et al., BV was common among women aged 25–34 years (58.8%).[11] Furthermore, a study by Lu et al. found that most of the women with BV positive was younger than 30 years in comparison to the BV-negative group.[13] The possible explanation of our late age presentation was due to the exclusion of unmarried and pregnant women from the study. 27.03% of the women were illiterate in our study population and did not receive any schooling. However, when compared illiteracy was more in BV diagnosed cases (58.25%) than BV-negative cases (13.04%). This is due to the low level of education associated with high-risk sexual behaviors resulting in increased acquisition of BV as supported by Ness et al. and Ashraf-Ganjoei.[14],[15] However, the Nigerian study by Abdullateef et al. reported that majority of women had tertiary education explained by the fact that the study was hospital-based and women of higher education level more likely able to afford the cost of care attended the hospital. Therefore, the reports from different studies varies.[11] Majority of our BV diagnosed cases were either homemakers or domestic maids (P = 0.003) and had made their sexual debut before 18 years of age (P = 0.001). In our study, women living with >1 sexual partner had no significant association with BV prevalence (P = 0.528). Sexual habits and characteristics of the male partner may play a vital role in the development of BV. Studies reported that women with multiple sexual partners have a three-fold increase in probability for BV.[16],[17] Our study does not have adequate information on partner's sexual behavior and the possible explanation is that they are unaware of the partners' high-risk sexual behavior or unwilling to reveal them. There is, however, some limitation in our study. Since risk factors were self-reported, and it is possible that there may be under-reporting and misclassification of risk behaviors. This study involved the collection of data pertaining to sensitive sexual behaviors as well as information on women's sex partners so there is a possibility of measurement error that may lead to residual confounding effect obscuring the relationship between BV and risk factors.

We found vaginal discharge (84.4%) as the most common symptom of presentation followed by lower abdominal pain (63.10%) and pruritus (27.18%) among women with BV. A number of studies explored the association of vaginal discharge with vaginal infections. Gutman et al. reported that women with BV were more likely to be symptomatic in comparison to women without BV with individual symptoms of vaginal discharge and foul odor being more common (P < 0.01).[18] In our study, clue cells in the wet mount and positive amine test were found to be statistically significantly associated with BV (P = 0.000). We found high specificity (98.7%) and PPV (93.02%) for clue cells toward the diagnosis of BV. The findings on positive clue cells also support previous studies that found it to have the higher specificity of the individual clinical criteria for diagnosing BV. Gutman et al. found sensitivity of 67% and high specificity of 93% for amine test. They also stated that sensitivity and specificity of clue cells at a cutoff of >20% clue cells on wet mount appears to be maximized, i.e., 74% and 86%, respectively.[18]

We found a significant association of HPV infection with BV diagnosed women (P = 0.000). Most of the studies showed an increased association of HPV infection with BV-positive group supporting our data. A study by Lu et al. reported that the HPV infection rate was more in BV positive women than that of BV negative group (P = 0.0000). They also found the rate of BV infection in HPV positive group was more than that in HPV negative group (P = 0.0000). They suggested the presence of consistency or synergies between HPV infection and BV infection.[13] The association between BV infection and cervical cancer/CIN is still confusing from no relation to significant relation. We found a rising incidence of CIN among women having BV diagnosed cases than that of BV-negative cases though statistically not significant (P = 0.506). Our data revealed no significant association of BV with CIN incidence. Furthermore, the presence of BV was not associated with the severity of CIN (P = 0.765). Denslow et al. in a study reported no association of BV with an increased risk of high-grade squamous intraepithelial lesion or cervical lesion progression.[19] Boyle et al. also stated no association of BV with CIN after testing 379 women.[20] Both the literature supported our data. Whereas, Gillet et al. in a meta-analysis confirms a positive association between BV and cervical precancerous lesions suggesting the potential role of a disturbed vaginal microflora in gynaecologic complications.[1] According to Lu et al., CIN and cervical cancer were significantly increased among women with BV and HPV infection and suggested that BV promoting the CIN and cervical cancer from the epidemiological point of view.[13]

However, the question still remains whether BV and genital HPV infection are simply related because there is a biological interaction between them or because both occur frequently in sexually active women. A positive corelationship between BV and HPV might be explained by the fact that sexual risk behavior and promiscuity are found more often in women with BV than in comparison groups. A number of variables are contributing to the observed heterogeneity in previous studies. Various social habits and ethnogeographical risk factors may explain the wide BV prevalence range observed. Technical biases such as collection of specimen, subjectivity, sensitivity, and specificity of diagnostic methods also attribute to detected heterogeneity. Complete STD screening was not performed for the present study which may have confounding effects to a certain degree on the results of the present study as well.


  Conclusion Top


The present study suggests a significant association between BV and acquisition of HPV infection. There is a trend of increased CIN incidence among BV-diagnosed women although statistically insignificant. However, the presence of BV was not associated with the severity of CIN. Therefore, screening guidelines must adapt and implement a sensitive tool like HPV DNA testing in primary screening of BV positive women instead of cytological testing. Women with BV should be considered as a priority group for prophylactic vaccination against HPV.

Women with a history of recurrent or persistent BV should be kept under close follow-up. In that case restoring the vaginal micro flora should be a promising and feasible answer to the high prevalence of HPV infections. In the meantime, there is a need to evaluate the potential of BV treatment to prevent HPV acquisition and transmission. Taking into consideration that these conditions are common among women worldwide, further research in this field is imperative.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Gillet E, Meys JF, Verstraelen H, Verhelst R, De Sutter P, Temmerman M, et al. Association between bacterial vaginosis and cervical intraepithelial neoplasia: Systematic review and meta-analysis. PLoS One 2012;7:e45201.  Back to cited text no. 1
    
2.
Platz-Christensen JJ, Sundström E, Larsson PG. Bacterial vaginosis and cervical intraepithelial neoplasia. Acta Obstet Gynecol Scand 1994;73:586-8.  Back to cited text no. 2
    
3.
Nam KH, Kim YT, Kim SR, Kim SW, Kim JW, Lee MK, et al. Association between bacterial vaginosis and cervical intraepithelial neoplasia. J Gynecol Oncol 2009;20:39-43.  Back to cited text no. 3
    
4.
Boyle DC, Smith JR. Infection and cervical intraepithelial neoplasia. Int J Gynecol Cancer 1999;9:177-86  Back to cited text no. 4
    
5.
Peters N, Van Leeuwen AM, Pieters WJ, Hollema H, Quint WG, Burger MP. Bacterial vaginosis is not important in the etiology of cervical neoplasia: A survey on women with dyskaryotic smears. Sex Transm Dis 1995;22:296-302.  Back to cited text no. 5
    
6.
Amsel R, Totten PA, Spiegel CA, Chen KC, Eschenbach D, Holmes KK. Nonspecific vaginitis. Diagnostic criteria and microbial and epidemiologic associations. Am J Med 1983;74:14-22.  Back to cited text no. 6
    
7.
Nugent RP, Krohn MA, Hillier SL. Reliability of diagnosing bacterial vaginosis is improved by a standardized method of gram stain interpretation. J Clin Microbiol 1991;29:297-301.  Back to cited text no. 7
    
8.
Sodhani P, Garg S, Bhalla P, Singh MM, Sharma S, Gupta S. Prevalence of bacterial vaginosis in a community setting and role of the pap smear in its detection. Acta Cytol 2005;49:634-8.  Back to cited text no. 8
    
9.
Kenyon C, Colebunders R, Crucitti T. The global epidemiology of bacterial vaginosis: A systematic review. Am J Obstet Gynecol 2013;209:505-23.  Back to cited text no. 9
    
10.
Sodhani P, Gupta S, Gupta R, Mehrotra R. Bacterial vaginosis and cervical intraepithelial neoplasia: Is there an association or is co-existence incidental? Asian Pac J Cancer Prev 2017;18:1289-92.  Back to cited text no. 10
    
11.
Abdullateef RM, Ijaiya MA, Abayomi F, Adeniran AS, Idris H. Bacterial vaginosis: Prevalence and associated risk factors among non-pregnant women of reproductive age attending a Nigerian tertiary hospital. Malawi Med J 2017;29:290-3.  Back to cited text no. 11
    
12.
Huppert JS, Hesse EA, Bernard MC, Bates JR, Gaydos CA, Kahn JA. Accuracy and trust of self-testing for bacterial vaginosis. J Adolesc Health 2012;51:400-5.  Back to cited text no. 12
    
13.
Lu H, Jiang PC, Zhang XD, Hou WJ, Wei ZH, Lu JQ, et al. Characteristics of bacterial vaginosis infection in cervical lesions with high risk human papillomavirus infection. Int J Clin Exp Med 2015;8:21080-8.  Back to cited text no. 13
    
14.
Ness RB, Hillier SL, Kip KE, Soper DE, Stamm CA, McGregor JA, et al. Bacterial vaginosis and risk of pelvic inflammatory disease. Obstet Gynecol 2004;104:761-9.  Back to cited text no. 14
    
15.
Ashraf-Ganjoei T. Risk factors for bacterial vaginosis in women attending a hospital in Kerman, Islamic Republic of Iran. East Mediterr Health J 2005;11:410-5.  Back to cited text no. 15
    
16.
Smart S, Singal A, Mindel A. Social and sexual risk factors for bacterial vaginosis. Sex Transm Infect 2004;80:58-62.  Back to cited text no. 16
    
17.
Nagot N, Ouedraogo A, Defer MC, Vallo R, Mayaud P, Van de Perre P. Association between bacterial vaginosis and herpes simplex virus type-2 infection: Implications for HIV acquisition studies. Sex Transm Infect 2007;83:365-8.  Back to cited text no. 17
    
18.
Gutman RE, Peipert JF, Weitzen S, Blume J. Evaluation of clinical methods for diagnosing bacterial vaginosis. Obstet Gynecol 2005;105:551-6.  Back to cited text no. 18
    
19.
Denslow SA, Westreich DJ, Firnhaber C, Michelow P, Williams S, Smith JS. Bacterial vaginosis as a risk factor for high-grade cervical lesions and cancer in HIV-seropositive women. Int J Gynaecol Obstet 2011;114:273-7.  Back to cited text no. 19
    
20.
Boyle DC, Barton SE, Uthayakumar S, Hay PE, Pollock JW, Steer PJ, et al. Is bacterial vaginosis associated with cervical intraepithelial neoplasia? Int J Gynecol Cancer 2003;13:159-63.  Back to cited text no. 20
    



 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

Top
 
 
  Search
 
Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

 
  In this article
Abstract
Introduction
Materials and Me...
Results
Discussion
Conclusion
References
Article Tables

 Article Access Statistics
    Viewed205    
    Printed13    
    Emailed0    
    PDF Downloaded34    
    Comments [Add]    

Recommend this journal


[TAG2]
[TAG3]
[TAG4]