Correlation between Asymptomatic Bacteriuria and HIV-1 Viral Load Level and CD4 Count in Pregnant Women on Antiretroviral Therapy in N’djamena (Chad)

A cross-sectional study was conducted at the “Centre de l’Appui Psycho-Médico-Social (APMS)” which is a centre for Psychological and Medical Support in N'Djamena (Chad) from January to March 2014. The aim of this study was to evaluate the correlation between asymptomatic bacteriuria (ASB) and viral load level and CD4 count in seventy-six (76) HIV-1 infected pregnant women on antiretroviral therapy (ART). Urine culture and bacteria identification were performed by using a chromogenic culture medium (UriselectR4). T CD4+ lymphocytes count and viral load measurement were done respectively on PIMATM test and Abbott m2000 RealTime HIV-1. In this study, 25 (32.9%) pregnant women were carrying ASB and major bacteria; Escherichia coli and Streptococcus agalactiae known to cause neonatal meningitis to newborns were identified. Bacteria were isolated mainly in women with CD4 lymphocytes < 200 cells/mm3 (60%) (15/25) and a viral load > 3log (70%) (19/25). Besides the prevention of mother to child transmission of HIV, which remains a goal, it is important to prevent also the transmission of other microorganisms causing neonatal Corresponding authors. A. F. Abderrazzack et al. 309 infections. Our findings support the needs to do bacteriological analysis of urine in every HIV-infected pregnant woman at least in late pregnancy.


Introduction
Successful antiretroviral therapy (ART) in people living with HIV (PLHIV) may leads to an undetectable viral load (VL) after 6 months treatment and be maintained undetectable or below 50 copies/ml later [1] [2].Besides this antiretroviral drugs' effectiveness, it turned out that the immunosuppression state induced by the Human Immunodeficiency Virus (HIV) infection seems to increase asymptomatic bacteriuria (ASB) in HIV infected pregnant women in about 6% to 20% of cases [3]- [7].ASB is a urinary tract infection without clinical signs [8] [9].In this case, urine culture is recommended at the first prenatal visit [10].
This study is conducted to evaluate the correlation between ASB and CD4 count and HIV-1 viral load in a cohort of HIV infected pregnant women followed-up for prevention of mother to child transmission (PMTCT) in N'djamena (Chad).

Study Site and Population
This was a cross sectional study conducted from January 1 st to March 31 st , 2014 among pregnant HIV-1 infected women on ART and followed-up at Centre de l'Appui Psycho-Médico-Social (APMS) of Ndjamena (Chad), a health center dedicated to people living with HIV.Pregnant women attending the site for antenatal care and not presenting any symptoms of a urinary tract infection (UTI) and not taking antibiotics for any current infection, who gave their oral consent, were included in the study.

Samples Collection
Clean-catch midstream urine samples were collected in sterile universal containers from pregnant women.They were instructed on how to collect samples and the need for prompt delivery to the laboratory the next day following their inclusion.About 10 ml venous blood were collected at the elbow on 2-Ethylene Diamine Tetra Acetate (EDTA) tubes.

Laboratory Analyses
Urine samples were seeded within 30 minutes to 1 hour after the collection onto Uriselect TM 4 (Bio-Rad, France)a chromogenic non-selective agar medium consisted of a rich nutrients base of four peptones ensuring the growth of all the germs of the urinary tract, two chromogenic substrates for the detection of bacterial enzymes (β-galactosidase and β-glucosidase), and tryptophan for the detection tryptophanase's activity (indole production) and tryptophan deaminase (TDA).Cultures were incubated at 37˚C for 24 hours.Bacterial colonies growing on the agar after the incubation period were identified as described by the Uriselect TM 4 user guide.
CD4 count was performed on whole blood using Alere Pima TM (Alere Technologies GmbH, Germany), a point-of-care device.Viral load samples were brought to the Virology Unit of the General Reference Hospital of N'Djamena (HGRN, Chad), using an insulated refrigerated container at 4˚C.On site, two plasma aliquots of about 1 ml were made and stored at −20˚C until the VL test was carried out.The VL was determined using Real-Time-Reverse Transcription Polymerase Chain Reaction (Real-Time RT-PCR) method using Abbott m2000 RealTime System (Abbott Laboratories, Abbott Park, Illinois, USA).

Demographic and Clinical Information
Socio-demographic information and antiretroviral treatment histories were collected using a standardized form.

Ethical Considerations
The study was approved by the ethical committee of the Centre de l'Appui Psycho-Médico-social (APMS) of Ndjamena (Chad) Coordination of the Psychological and Medical Support Unite (APMS) under the reference No. 196/PR/PM/MSPASSN/DG/PSLS-IST/PMSS/2013.

Statistical Analysis
Continuous variables were compared using the Wilcoxon rank-sum test, and comparison between two categorical variables was performed using the Chi-square test and Fisher's exact test when cell values were less than 5.For all analyses p-value of less than or equal to 0.05 was used to determine statistical significance.

Results and Discussions
During the 3-month study period, 76 HIV-infected pregnant women were included in the study.They aged 15 to 45 years with a median of 30 years (IQR) [22 -28 years].The gestational age ranged from 5 to 9 months.The women were treated by a combination ART (cART) represented by (Tenofovir (TDF) + Lamivudine (3TC) + Efavirenz (EFV)) (71.03%).The median duration on ART was 4 months.The socio-demographics characteristics of study population are summarized in Table 1.
We found 25 (32.9%)pregnant women of the study population carrying ASB and none of them carried more than one bacteria strain in the urinal tractus.They aged 24 to 35 years with a median of 34.5 years.All age groups were concerned by the ASB.The ASB prevalence was 32.9% CI95% [22.3% -43.5%] and it was higher than 18.6% reported from a similar study conducted in South Africa [11].

Table 1 .
Characteristics of study population.