Conceived and designed the experiments: NKV MS JSH MAS AR VV. Performed the experiments: NKV RN SA. Analyzed the data: NKV MS JSH MAS CD RN. Contributed reagents/materials/analysis tools: AR SA VV CD. Wrote the paper: NKV MS JSH MAS RN AR. Study supervision: MS AR VV.
The authors have declared that no competing interests exist.
To determine the prevalence and risk factors for
Pregnant women (≥28 weeks gestation) in Vellore, South India were approached for enrollment from April 2009 to January 2010. After informed consent was obtained, women completed a socio-demographic, prenatal, and sexual history questionnaire. Endocervical samples collected at delivery were examined for CT by a rapid enzyme test and nucleic acid amplification test (NAAT). Neonatal nasopharyngeal and conjunctival swabs were collected for NAAT testing.
Overall, 1198 women were enrolled and 799 (67%) endocervical samples were collected at birth. Analyses were completed on 784 participants with available rapid and NAAT results. The mean age of women was 25.8 years (range 18–39 yrs) and 22% (95% CI: 19.7–24.4%) were primigravida. All women enrolled were married; one reported >one sexual partner; and six reported prior STI. We found 71 positive rapid CT tests and 1/784 (0.1%; 95% CI: 0–0.38%) true positive CT infection using NAAT.
To our knowledge, this is the largest study on CT prevalence amongst healthy pregnant mothers in southern India, and it documents a very low prevalence with NAAT. Many false positive results were noted using the rapid test. These data suggest that universal CT screening is not indicated in this population.
Most epidemiologic data on CT is from industrialized nations; however, it is important to characterize CT disease from resource limited regions where most infants are born. Scarce information is available on laboratory-confirmed incidence and prevalence of chlamydia infection in otherwise healthy males and females in India. Furthermore, the available Indian data show a wide variation in CT prevalence and methods of laboratory confirmation
Area | Year | Age group | Study population | N | Sample | Test | Prevalence | Ref |
Vellore, India | 1993 | Not available | Pregnant women | 273 | Endocervical swab | Chlamydiazyme kit (Abbott, USA) | 3.3% (95% CI: 1.2%–5.4%) | 5 |
New Delhi, India | 1995 | Mean Age: 34.9 yrs | Generally healthy women, Gynecological clinic(outpatient department) | 257 | Endocervical swab | Chlamydiazyme kit (Abbott, USA) | 23.3% | 14 |
Mumbai, Inner City | 1998 | ≤35 yrs | Suspected PID, Infertility | 446 | Endocervical swab | ELISA | 0.5% | 7 |
Mumbai | 2001 | 18–42 yrs | Gynecological clinic, complications associatedwith reproductive health | 123 | Endocervical swab | Chlamydiazyme kit (Abbott, USA) | 1.7 to 20% among different risk categories | 11 |
New Delhi | 2003 | 18–40 yrs | Symptomatic women Gynecological clinic | 280 | Endocervical swab | NAAT | 28% (18–25 yrs) | 13 |
New Delhi | 2003 | 19–36 yrs | Pregnant women | 350 | Endocervical swab | DFA & PCR | 18.8% (95% CI 14.76–22.96%) | 12 |
Tamil Nadu, South India | 2004 | 15–45 yrs | Healthy adult population, clinic | 1066 (serum)841 (urine)Female samples | Serum, urine | IgM-ELISA,urine NAAT | 3.3% ELISA, 1.1% PCR (95% CI: 0.4–1.8%) | 8 |
Chennai | 2005 | N/A | STI clinic(high risk) | 143 men & women | Serum, genital swab endocervical/urethral | Culture/nested PCR (MOMP): CT Serum: IgG | 30.8% by nested PCR(MOMP) | 9 |
Aligarh, North India | 2009 | 18–40 yrs | Obstetric clinic, secondary infertility, pregnant women (control subjects) | 70 | Endocervical swab | Cell culture, ELISA | 55%–2°infertility; 5.5% pregnant women | 10 |
Karnataka State,South India | 2010 | Mean age: 30.7 yrs | Symptomatic women Gynecological clinic | 412 | Endocervical swab | NAAT | 2.6%, vaginal discharge; 2.7% vaginal discharge with clinical cervicitis | 6 |
This table shows a review on Indian data which show a wide variation in CT prevalence and methods of laboratory confirmation.
The prevalence of CT in pregnant women in India has been shown to vary by geographic region. In a study conducted in Vellore, TN India in 1993, a prevalence of CT in pregnant women was found to be 3.3% (95% CI: 1.2%–5.4%) using an enzyme immunoassay (EIA) which detects the presence of chlamydia antigen (Chlamydiazyme® kit, Abbott®,USA)
Current obstetric practice in southern India does not include universal prenatal screening for CT. This practice is based on evidence from older local data
Furthermore, although there have been only a few studies documenting prevalence of
In order to provide data for development of evidence-based guidelines for resource-limited regions, the objectives of our study were to determine the prevalence of CT genital infection in delivering women using highly sensitive NAAT testing and to study the maternal to child transmission of CT in Vellore, India.
We aimed 1) to estimate the prevalence of genital CT infection among women presenting for delivery at Christian Medical College Hospital (CMCH) Vellore, India using a NAAT technique 2) to identify risk factors associated with chlamydia infections in pregnant and delivering women in order to guide development of screening policies if needed; and 3) to estimate the prevalence of CT infection at birth and to identify factors associated with maternal-infant transmission.
The institutional review boards at both Cincinnati Children’s Hospital Medical Center (CCHMC) and the Christian Medical College Hospital approved the study protocol. Written informed consent was obtained during enrollment for the subject and her neonate. All clinical investigation was conducted according to the principles expressed in the Declaration of Helsinki.
This was a prospective hospital-based observational study.
CMCH, located in Vellore, India served as the surveillance site for this study. Vellore is a city in the Indian state of Tamil Nadu located in tropical southern India midway between the cities of Chennai (formerly Madras) and Bengaluru (formerly Bangalore). Vellore town has a population of 177,230 (2001 Indian census) of which 50.4% are female. CMCH is a private, non-profit, 2000+ bed tertiary referral hospital with primary care clinics (including pediatrics) and serves a diverse population of lower to middle class patients. The CMCH obstetrics department reports over 8,000 deliveries per year.
From April 2009 to January 2010, delivering women were approached and recruited daily by a research staff member during an antenatal outpatient visit, a scheduled labor/induction, or in the postpartum ward. During the antenatal outpatient visit, a physician would send his/her patient, based on the study’s inclusion criteria, to the designated research office for the research assistant to discuss the study with the delivering woman. On the inpatient setting, the research assistant approached subjects daily on the obstetric wards that met inclusion criteria. After the research assistant obtained the subject’s consent to the study, a questionnaire on socio-demographic factors, prenatal history, and sexual history was administered by the research assistant on initial encounter during a private interview using a pen and paper format.
The inclusion criteria required that study participants be 18 to 45 years of age; greater or equal to 28 weeks gestation; and planning to deliver at CMCH and use the CMCH Vaccine or Child Health clinic for infant care. Pregnant mothers who had a history of chronic disease including diabetes, congestive heart failure, and renal failure were excluded. Stillborn infants and infants with congenital disorders or malformations were excluded.
Endocervical swabs were collected by physicians or a research assistant during a pelvic exam performed either during an antenatal outpatient visit; prior to a scheduled labor/induction; or in the postpartum ward. The first swab was used to perform the rapid diagnostic test (RDT) and the second for NAAT testing. A third swab was stored frozen at −70°C for future testing. Swabs were collected before use of lubricant or insertion of a prostaglandin tablet for induction to prevent the possibility of inhibition of the test.
Within 48 hours after delivery, neonatal participant data was recorded by the research assistant and a conjunctival swab from the eye and a nasopharyngeal (NP) swab were collected. Swabs were collected after 24 hours in order to decrease the chance that a positive test would represent contamination by maternal secretions. After collection, the swabs were stored in a freezer at −70°C for NAAT testing.
The RDT used in the study was the Clearview Chlamydia Test Kit® (Inverness, Houston, TX 77038). This test is a one step immunochromatographic assay for direct antigen detection with an internal positive control indicator. The test was performed in the onsite research lab by study staff according to the package insert instructions.
Endocervical specimens were stored in a freezer at CMCH at −70°C. The NAAT samples were tested at Y.R. Gaitonade Centre for AIDS Research and Education Laboratory in Chennai, India using the Roche Amplicor® CT/NG test for
In addition, 102 samples from vials with remaining endocervical swab specimen had DNA extracted using the QIAamp DNA Minikit® (Qiagen, Hilden, Germany). These extracted samples were shipped, on dry ice, and retested independently in an established US laboratory using the Roche Amplicor PCR® (Roche Diagnostic Systems, Inc., Branchburg, N.J).
Subjects’ positive rapid test results were reported to the attending physicians in both the obstetric and neonatology departments following birth. Treatment decisions were made by the primary physicians.
For sample size estimations, we used estimated prevalence of CT from previous studies (
Data were analyzed using Statistical Analysis Systems software (SAS Institute, Cary, NC, Version 9.2). Means and standard deviations were used to characterize normally distributed data. T-tests were used to compare means of normally distributed data. Chi-square and Fisher’s exact tests were used to assess differences in proportions. All reported P values were 2-sided and p values <0.05 were considered statistically significant. Concordance between the rapid test and NAAT was determined by the Kappa statistic and Kendall’s Tau B.
From April 2009 to January 2010, 7955 women delivered during the recruitment period. Of these 7955 women, a convenience sample of 1367 (17%) pregnant and delivering women was approached for recruitment into the study. Of those, 1198 (88%) women were enrolled; 45% during an antenatal outpatient visit, 55% before a scheduled labor/induction, and <1% in the postpartum ward.
April 2009 to January 2010, 7955 women delivered during the recruitment period. 1198 (88%) women were enrolled; 799 endocervical samples from the 1198 enrolled subjects were collected and data on 784 participants with both RDT and NAAT results are reported.
From the initial 1367 participants approached, 169 delivering women were not enrolled. Of these, 131 women were not enrolled in the study after refusing consent or not planning on follow up care at CMC. Another 38 women did not participate in the study for other reasons. Inquiries were made to determine the reasons for declining; 85% indicated they declined because their family did not want them to participate. Overall, women who declined were similar to the women who did participate in the study in regards to urban/rural residence (p = 0.77), religion (p = 0.47), and gravida (p = 0.1).
Enrolled Mothers | Enrolled Tested | Enrolled Not Tested | P-Value of Tested mothers vs. Untested mothers | ||||
N | Percent | N | Percent | N | Percent | ||
|
1198 | 783 | 415 |
|
|||
18–23 years | 384 | 32.05 | 231 | 29.50 | 153 | 36.87 | |
24–29 years | 625 | 52.17 | 425 | 54.28 | 200 | 48.19 | |
≥30 years | 189 | 15.78 | 127 | 16.22 | 62 | 14.94 | |
|
1198 | 783 | 415 |
|
|||
Other | 935 | 78.05 | 649 | 82.89 | 286 | 68.91 | |
Primigravida | 263 | 21.90 | 134 | 17.11 | 129 | 31.08 | |
|
1195 | 780 | 415 |
|
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Illiterate | 31 | 2.59 | 20 | 2.56 | 11 | 2.65 | |
Primary Education | 443 | 37.07 | 277 | 35.51 | 166 | 40 | |
Secondary/Tertiary | 293 | 24.52 | 183 | 23.46 | 110 | 26.51 | |
University Diploma | 428 | 35.82 | 300 | 38.46 | 128 | 30.84 | |
|
1121 | 731 | 390 |
|
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(<5000 Rs/mo) | 795 | 70.92 | 499 | 68.26 | 296 | 75.9 | |
(5000–10,000 Rs/mo) | 229 | 20.43 | 162 | 22.16 | 67 | 17.18 | |
(>10,000 Rs/mo) | 97 | 8.65 | 70 | 9.58 | 27 | 6.92 | |
|
1196 | 781 | 415 |
|
|||
Village | 604 | 50.50 | 393 | 50.32 | 211 | 50.84 | |
Urban | 592 | 49.50 | 388 | 49.68 | 204 | 49.16 | |
|
1193 | 779 | 414 |
|
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Hindu | 964 | 80.80 | 635 | 81.51 | 329 | 79.47 | |
Muslim | 165 | 13.83 | 96 | 12.32 | 69 | 16.67 | |
Christian | 62 | 5.20 | 47 | 6.03 | 15 | 3.62 | |
Jain | 2 | 0.17 | 1 | 0.13 | 1 | .24 | |
|
1183 | 775 | 408 |
|
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No | 1147 | 96.96 | 751 | 96.90 | 396 | 97.06 | |
Yes | 6 | 0.51 | 3 | 0.39 | 3 | .74 | |
Don’t Know | 30 | 2.54 | 21 | 2.71 | 9 | 2.21 | |
|
1190 | 775 | 415 |
|
|||
Only Husband | 1189 | 99.92 | 774 | 99.87 | 415 | 100 | |
Others | 1 | 0.08 | 1 | 0.13 |
(50Rs = 1 USD).
This table shows that tested mothers were significantly older, multiparous, and higher socio-economic group compared to untested mothers (p = 0.03, p = <0.0001, and p = 0.03; respectively).
We collected 799 endocervical samples from the 1198 enrolled subjects and data on 784 participants with both RDT and NAAT results are reported. 399 enrolled participants (33%) did not have endocervical samples collected after enrollment; see
The prevalence detected using the NAAT (considered the gold standard for this study) was 0.1% (95% CI: 0–0.38%). The mother with the positive NAAT specimen had no significant characteristics or risk factors. All 71 RDT samples that were positive were considered to be false positives. Thus, compared to NAAT this RDT had a sensitivity of 0%, a specificity of 90%. As anticipated, the statistical analysis by Kappa and Kendall Tau B revealed no agreement between the RDT and NAAT (
RDT | |||
NAAT | Positive | Negative | Total |
Positive | 0 | 1 | 1 |
Negative | 71 | 712 | 783 |
Total | 71 | 713 | 784 |
Cohen’s Kappa = −0.0025 | |||
Kendall’s Tau B = −0.0113 |
Cohen’s Kappa, which tests the agreement between two tests is negative, showing no agreement. Kendall’s Tau B is also negative, showing very slight negative association (inversion) between the two tests. The association here is very limited.
The prevalence detected using the NAAT (considered the gold standard for this study) was 0.1% (95% CI: 0–0.38%).
There were 811 neonates enrolled which included 12 twin infants. There were 768 newborn specimens (NP and conjunctival) obtained from the neonates of the 784 enrolled mothers who had both NAAT and RDT results reported. During neonatal swab collection, there were 10 neonates who were lost to follow-up due to early discharge, 1 mother refused swab collection from her infant after delivery, and 5 neonates had incomplete swab collections. The neonatal characteristics included 52% born by normal vaginal delivery; mean gestation was 39 weeks, and mean birth weight was 2.99 kilograms (not including twins). Please see
N | Percent | ||
|
|||
Forceps | 164 | 21.35 | |
C Section | 201 | 26.17 | |
Normal Vaginal | 403 | 52.47 | |
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No | 681 | 88.67 | |
Yes (RDS, Sepsis, IUGR, NICU) | 87 | 11.33 | |
|
|
|
|
|
768 | 39.0 | 1.359 |
|
762 | 2.99 | 0.455 |
There were 768 newborn specimens (NP and conjunctival) obtained from the neonates of the 784 enrolled mothers who had both NAAT and RDT results reported. This table describes the neonatal characteristics.
This is the largest report from India on
In our study, we found that 10% of specimens were positive when evaluated by RDT. We chose the Clearview Chlamydia Test Kit® because it is one of the few FDA approved RDT for CT, and it had reasonable reported sensitivity (53–73%) and specificity (68–99%) on endocervical specimens when compared to culture or DNA probe assay/NAAT
If the true specificity of the RDT is between 68 and 99%
In newborns, the CT prevalence was found to be zero. Therefore, the transmission rate from Chlamydia positive mothers to infants could not be characterized because we did not have sufficient power to address this question. Further studies need to be done on pregnant mothers with a higher prevalence of CT infection to assess maternal to neonate transmission in this setting.
There are some limitations to our study. First, due to limited staff, we were only able enroll 17% of delivering women during our study period. However, 88% of women approached were enrolled. Second, the project sample population may not represent the local delivering female population. CMCH is a tertiary, private hospital and some patients may go to a nearby government hospital, or deliver at home. In addition, not all endocervical samples were obtained from recruited participants due to logistical reasons. Few endocervical specimens (n = 2) were also obtained postpartum which may have changed the RDT sensitivity and/or specificity. Tested mothers were significantly older, multiparous, and in the higher socio-economic group versus untested mothers (p = 0.03, p = <0.0001, p = 0.03; respectively). One reason for this finding may be because younger women in the lower socio-economic group delivered their neonate at home (especially if primigravida) or at a nearby government hospital.
In summary, we found that CT infection appears to be relatively rare in women delivering in this private tertiary care hospital in Vellore, India. We found a prevalence of 0.1% (95% CI: 0–0.38%) by NAAT which was much lower than that noted in other international studies including pregnant women
The local practice is to not perform prenatal screening in healthy pregnant women in this population based on evidence from twenty years ago which used older diagnostic methods. We conclude that our current study justifies that routine prenatal screening in this population would not be recommended given the low prevalence. However, future studies are important to reassess prevalence as sexual practices may change within this culture. Outpatient pre-natal testing may provide better information in all SES strata.
We gratefully acknowledge the Fogarty International Center and Vanderbilt University for allowing us this opportunity through the Fogarty International Research Clinical Fellows program. We thank the laboratory support of YRG Care (Dr. P. Balakrishnan and staff). We also thank CMC biostatistics department, Mrs. Alice Augustine, and Mrs. Geeta Marimuthu for recruitment and data management. We thank Stephanie Donnelly, Tasha Hughes, OG3 (CMCH) office for administrative support. This study would not have been possible without the help of the physicians (Dr. AK Jana, Dr. Ruby Jose, Dr. Gigi Matthews), OG residents, nurses, laboratory, and administrative staff affiliated with the Christian Medical College Hospital. We also thank Dr. Chuck Schubert,Dr. Connelly, Dr. David Bernstein, Gretchen Langdon, Dr. Joel Mortensen, and Amy Cassedy for their guidance with this project. We thank Dr. Charlotte Gaydos at Johns Hopkins University for her laboratory assistance. We deeply express our gratitude to the families who participated in the project.