Prevalence of HIV, HBV and HCV and their co-infection during primary investigation and before ART in Eastern Region of Nepal

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1 IJAMBR 5 (2017) ISSN Prevlence of HIV, HBV nd HCV nd their co-infection during primry investigtion nd efore ART in Estern Region of Nepl Snjy Mhto 1,2 *, Asmit Mhto 1 nd Jyendr Ydv 3 1 Asr Reserch nd Eduction Acdemy Counsel, Birtngr 7, Zip code 56613, Nepl. 2 Deprtment of Microiology, Mhendr Morng Adrsh Multiple Cmpus, Trihuvn University, Birtngr 16, Zip code 56613, Nepl. 3 Deprtment of Pthology nd Microiology Unit, Koshi Zonl Hospitl, Birtngr, Zip code 56613, Nepl. Article History Received 03 Septemer, 2017 Received in revised form 01 Novemer, 2017 Accepted 07 Novemer, 2017 Keywords: Co-infection, Heptitis B viruses, Heptitis C viruses, HIV, Prevlence, Seroprevlence. Article Type: Full Length Reserch Article ABSTRACT The im of this study ws to determine the seroprevlence of heptitis B viruses (HBV), heptitis C viruses (HCV) nd humn immune deficiency viruses (HIV) nd their co-infection during primry screening efore ntiretrovirl therpy (ART) in the Estern region of Nepl. This ws cross-sectionl oservtionl study, in which 3,716 individuls, ttending Koshi Zonl Hospitl, Birtngr, Nepl, were investigted for HBV, HCV, nd HIV from June 2016 to My Among 3,716 ptients [femle 2880 (77.5%) nd mle 836 (22.5%)], HIV ws found in 101 (2.7%), HBV in 53 (1.4%) nd HCV in 15 (0.4%). HIV ws found in 65.3% of mles nd 34.7% of femles, HBV in 47.2% of mles nd 52.8% of femles, nd HCV in 93.3% of mles nd 6.7% of femles. The overll rte of co-infection with HBV nd/or HCV ws 5.9% (6 out of 101 HIV positives). Only 3.0% were positive for oth HIV nd HBV infection mrker nd 2.0% were positive for HIV nd HCV mrkers. Only isoltes (1.0%) ws positive for ll three mrkers of HIV, HBV nd HCV. A significnt ssocition ws oserved etween gender nd the prevlence of HIV, HBV or HCV (p<0.001). Mritl sttus nd the prevlence were nonsignificnt. Most of the HIV positive ws found in yers of ge group (32.2%), while HBV in yers group (45.3%) nd HCV in yers group (40.0%). The co-infections were restricted to the ge group of nd yers. Regulr screening for HIV, HBV nd HCV mong ptients with doutful history cn help in detecting mny new cses t the pproprite time BluePen Journls Ltd. All rights reserved INTRODUCTION Heptitis B viruses (HBV), heptitis C viruses (HCV) nd humn immune deficiency viruses (HIV) re the three most common chronic virl pthogen known. Despite their iologicl differences nd nturl history of chronic infection, the viruses hve common routes of trnsmission (such s lood nd lood products, shring needles to inject drugs nd sexul ctivities) nd similr *Corresponding uthor. E-mil: mhto.snjy@gmil.com. risk fctors (Kellermn et l., 2003; Chen et l., 2016). These viruses re ssocited with incresed moridity nd mortlity. An estimted 240 million people hve chronic heptitis B virus infection, million people hve chronic heptitis C virus infection (WHO, 2016) nd million people hve HIV (WHO, 2017). Moreover, heptitis B is estimted to result in 887,000 deths nnully versus 399,000 deths from heptitis C worldwide (WHO, 2017; WHO, 2017c). Prevlence of heptitis B nd C in HIV-infected individuls hs een reported to e higher thn tht of the generl popultion.

2 Int. J. Appl. Microiol. Biotechnol. Res. 124 Aout 2.9 million people (2-15%) living with HIV re coinfected with heptitis C virus nd 2.6 million (5-20%) with heptitis B virus (WHO, 2016). These co-infections hve een linked with reduced survivl, drug-relted heptotoxicity, drug resistnce, cross-resistnce nd suoptiml response (Blogun et l., 2012; Pennp et l., 2016). Severl studies highlight tht the rte of HBV is higher thn tht of HCV in HIV-infected ptients (Nouip et l., 2015; Brndão et l., 2015; Muriuki et l., 2013). On the contrry, few results re conflicting (Muriuki et l., 2013; Suprm et l., 2015). Not only su-shrn Afric is the most ffected region with 25.6 million HIV infection, ut lso ccounts two-third of the glol totl of new HIV infections (WHO, 2017). Prevlence of heptitis B virus is highest in su-shrn Afric nd Est Asi with chronic infection in 5-10% dult popultion. Centrl nd Est Asi (3.8%) nd north nd West Afric (2.3%) re the most ffected regions with heptitis C. These infections re ecuse of unsfe medicl injections nd other medicl procedures (WHO, 2016; Khyriyyh et l., 2013; Mutgom et l., 2017). Interestingly 18-34% of cute heptitis infections re spontneously clered (Westrook nd Dusheiko, 2014). MATERIALS AND METHODS The informed consent ws tken from the ptients. The study ws pproved y AASRA Reserch nd Eduction Acdemy Counsel Ethics Committee (Approvl no. AREC 1606 A002). This cross- sectionl oservtionl study ws conducted in the estern region of Nepl t Koshi Zonl Hospitl, Birtngr, Morng for durtion of 12 months (June 2016 to My 2017). The study ws crried out in the Pthology nd Microiology Lortory unit of Koshi Zonl Hospitl nd the dt nlysis ws done t Asr Reserch nd Eduction Acdemic Counsel, Birtngr, Nepl. A totl of 3716 ptients were tested during the study period. The ptients investigted for HIV, HBV, nd HCV were not wre of their infection sttus s no test were done erlier. So the ptients were not hving ny sort of ART. If ny ptient ws wre HIV positive nd ws hving ART, they were excluded from the study. This ws lso confirmed y questioning the ptients. The study ws imed t studying the prevlence of HIV, HBV, nd HCV s well s co-infection of HBV nd/or HCV mong HIV-positive ptients. Demogrphic profile like ge, gender nd mritl sttus of the ptients ws tken. The ge ws ctegorized into different groups with regulr clss intervl. The prevlence of HIV with or without HBV nd/or HCV in mle ws compred with the femle. The HBV, HCV, nd HIV were tested y kits HEPACARD, HCV TRI-DOT nd HIV TRI-DOT (J. Mitr nd Co. Pvt. Ltd, New Delhi, Indi.), respectively. For HIV, nother kit test with Uni- Gold TM HIV (Trinity Biotech, Irelnd) ws lso performed to verify the result. HBV test Bsed on the ntigen cpture, or sndwich principle; HEPACARD (J. Mitr nd Co., Indi) is visul, rpid, sensitive nd ccurte one-step immunossy for qulittive detection of Heptitis B Surfce Antigen (HBsAg) in humn serum or plsm. The ssy is intended to e used s n id in the recognition nd dignosis of cute infections nd chronic infectious crriers of HBV ( dtestkits/hvrnge/hepcrd.spx). HCV test The fourth genertion HCV TRI-DOT is visul, rpid, sensitive nd qulittive in vitro dignostic test for the detection of ntiodies to heptitis C virus in humn serum or plsm. The test is designed with incresed sensitivity for core nd NS3 ntiodies using unique comintion of modified HCV ntigens. They re highly purified HCV ntigens like the puttive core (structurl), protese/helicse NS3 (non-structurl), NS4 (nonstructurl) nd replicse NS5 (non-structurl) regions of the virus tht is immoilized on the device ( dtestkits/hcvrnge/hcv_tri_dot.spx). These ntigens re in the form of two test dots T1 nd T2 to provide highly sensitive nd specific dignostic test. Testing ws for ntiodies ginst HCV, which does not differentite ctive (HCV RNA+) versus previously clered (HCV RNA-) HCV infection. The prevlence estimtes should, therefore, e viewed s potentil overestimtes of the ctul rte of ctive HCV infection in the study group. HIV test Bsed on flow through technology, HIV TRI-DOT test is visul, rpid, sensitive nd ccurte immunossy for the differentil detection of HIV-1 nd HIV-2 ntiodies (IgG) in humn serum or plsm using HIV-1 nd HIV-2 ntigens immoilized on n immunofiltrtion memrne. It uses envelop ntigens gp41 nd C-terminus of gp120 for HIV-1 nd gp36 for HIV-2 ( dtestkits/hivrnge/hiv_tri_dot.spx). The test is screening test for nti-hiv-1 nd nti-hiv- 2 nd is for in vitro dignostic use only. The Trinity Biotech Uni-Gold HIV test is single regent ssy for the detection of ntiodies to humn immunodeficiency

3 Mhto et l. 125 Tle 1. Gender wise ge group distriution of ptients sujected for tests of HBV, HCV nd HIV. Age group (yers) Femle (%) Mle (%) Totl (%) <10 24(0.8) 36(4.3) 60(1.6) (14.5) 77(9.2) 496(13.3) (60.8) 211(25.2) 1962(52.8) (12.7) 188(22.5) 555(14.9) (4.8) 122(14.6) 261(7.0) (2.6) 106(12.7) 180(4.8) (2.3) 49(5.9) 115(3.1) (0.8) 36(4.3) 59(1.6) (0.6) 11(1.3) 28(0.8) Totl 2880(100.0) 836(100.0) 3716(100.0) Tle 2. Gender wise distriution of ptients with positive results of HBV, HCV, HIV nd their co-infection. Femle Mle Totl HBV only HCV only HIV only HIV + HBV HIV + HCV HIV + HBV + HCV HBV + HCV virus types 1 nd 2 in serum, plsm or whole lood. Dt nlysis The distriution of HIV, HBV, nd HCV ws recorded ccording to the gender nd ge. The sttisticl nlysis ws done with Sttisticl Pckge for Socil Sciences (SPSS 21.0) softwre pckge (SPSS Inc., Chicgo, USA). Fisher s exct test ws done. The p-vlue ws derived using the chi-squre test nd p-vlue of <0.05 ws considered sttisticlly significnt. RESULTS A totl of 3716 individuls ws involved in this study nd exmined for the prevlence of HIV, HBV, nd HCV. Out of 3716 individuls, 2880 (77.5%) were femle nd 836 (22.5%) were mle (Tle 1). They were ged etween 2-89 yers with men ge of yers (±0.223). The results showed tht the prevlence of HIV, heptitis B nd heptitis C virus mong different groups of popultion ws 2.7% (femle 0.9% nd mle 1.8%), 1.4% (femle 0.8% nd mle 0.7%) nd 0.4% (femle 0.0% nd mle 0.4%) respectively. The overll rte of coinfection with HBV nd/or HCV ws 5.9% (6 out of 101 ptients) s listed in Tle 2. The femle to the mle proportion who prticipted in the study ws 3:1. Interestingly, the positive results were in proportion of 1:2 (femle: mle) in HIV while nerly equl s 1:1 in HBV. Regrding the gender, the prevlence of HIV ws higher in mles (65.3%) compred to femles (34.7%). Interestingly, the prevlence of HCV ws exceptionlly higher in mles (93.3%) compred to femles (6.7%). Unlike HCV, the prevlence of HBV ws mrginlly higher in femles (52.8%) compred to mles (47.2%). The co-infection of HBV in HIV-infected ptients ws recorded s 4.0% while co-infection of HCV in HIV ptients ws 3.0%. However, 1.0% of HIV-infected were co-infected with oth HBV nd HCV. There ws highly significnt ssocition etween gender nd prevlence of HIV (χ 2 =109.33, =1, p<0.001) (Tle 3). Such significnt ssocition ws lso found etween gender nd prevlence of HBV (χ 2 =18.77, =1, p<0.001) (Tle 4). Considering Fisher s exct test, the ssocition etween gender nd prevlence of HCV (χ 2 =43.34, =1, p<0.001) ws found to e

4 Int. J. Appl. Microiol. Biotechnol. Res. 126 Tle 3. Chi-squre test for gender nd HIV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (0.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. Tle 4. Chi-squre test for gender nd HBV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (0.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. Tle 5. Chi-squre test for gender nd HCV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (25.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. significnt (Tle 5). No significnt ssocition ws found etween the mritl sttus nd the prevlence of HIV (χ 2 =3.369, =1, p=0.066), HBV (χ 2 =1.996, =1, p=0.158), nd HCV (χ 2 =2.999, =1, p=0.153) s shown in Tle 6, 7 nd 8. Aout 81.0% of the individuls were ged from 10 to 39 yers who visited the hospitl for screening of such diseses (Tle 1). Age group yers ccounted highest visit of 52.8% followed y yers (14.9%) nd yers (13.3%). The highest numers of femle (60.8%) were from ge group yers who were screened for HIV, HBV nd HCV. Age group yers ws followed y ge group yers (14.5%) nd yers (12.7%). The results show tht the femle of high sexul ctivity ge is frequent for screening. Comprtive to femle, mle ge group ws more distriuted. Among mle prticipnts, 25.2% were ged etween yers while 22.5% were ged etween yers. Age group yers ccounted 14.6% while yers were 12.7%. Even the ge group yers ws higher s 9.2%. These findings point out the risk of disese in wide rnge of ge group. Though the femle prticipnts were three times higher thn the mle, pproximtely two-third (65.3%) of HIV positive were mle comprtive to femle (34.7%). There were no cses of HIV ove 70 yers of ge in mle nd

5 Mhto et l. 127 Tle 6. Chi-squre test for mritl-sttus nd HIV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. Tle 7. Chi-squre test for Mritl-sttus nd HBV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. Tle 8. Chi-squre test for mritl-sttus nd HCV. Person Chi-squre Continuity correction Likelihood rtio Fisher's exct test cells (25.0%) hve expected count less thn 5. The minimum expected count is Computed only for 2 2 tle. ove 60 yers of the femle (Tle 9). In the femle, lrger shre of infection (40%) lied in the ge group yers followed y 31.4% of ge group yers, 20% of ge group yers, 17.1% of ge group yers, 8.6% of yers nd 2.9% of less thn 10 yers. Unlike femle, the mle of ge group yers topped the infection chrt with 33.3% followed y ge group yers with 25.8%, nd then, ge group yers with 19.7%. Age group less thn 10 yers showed 3.0% of infection mong mle while ge group yers showed the lest with vlue of 1.5%. Overll, the highly infected ge group were yers (32.7%) followed y the ge group yers (26.7%) nd then y yers (22.8%). The shre of ge group yers nd yers ws equl to 6.9%. Unlike HIV, the prevlence of HBV ws nerly 1:1 in proportion, tht is, mrginlly higher in femles (52.8%) compred to mles (47.2%) lthough the smple of femle to mle ws in 3:1 proportion. HBV ws not seen in the ge group of less thn 10 yers nd ove 70 yers group (Tle 9). Among femle, the highest portion of infection (60.7%) ws occupied y ge group yers followed y ge group yers (14.3%). Nerly 10.7% of ge group yers nd 7.1% of ge group yers were HBV positive. 3.6% of femle elonging to ge group yers nd yers

6 Int. J. Appl. Microiol. Biotechnol. Res. 128 Tle 9. Age group distriution of ptients with HBV, HCV, HIV, nd their co-infections. Age (yers) HBV only HCV only HIV only HIV+HBV HIV+HCV HIV+HBV+HCV F M F M F M F M F M F M < Totl ech were HBV positive. Mle of ge group yers showed the highest infection of HBV (32%) followed y ge group yers with 28%. Age group yers showed 24% of HBV infection while ge group yers ccounted 12% of HBV infection. Age group yers showed the lest HBV infection of 4%. Overll, the highly infected ge group ws yers (45.3%). Age group yers (22.6%) ws followed y ge group yers (13.2%). Age group yers hd HBV cses of 7.5% nd yers hd 9.4% of HBV. And the lest HBV positive cses were found in yers (1.9%). The prevlence of HCV ws pproximtely one-third of HBV. The prevlence of HCV ws higher in mles (93.3%) compred to femles (6.7%). Out of 15 HCV positive individuls, only one lied in femle. HCV ws not found in the ge group of less thn 20 yers nd ove 49 yers group. In the femle, the only infection ws found in the ge group yers. Unlike femle, ge group yers showed the highest infection of 42.9% followed y ge group yers with 35.7%, nd then, ge group yers with 21.4%. Overll, the infection of HCV ws 40.0% in the ge group yers, 33.3% in the ge group of yers nd 26.7% in the ge group yers. HBV nd HCV co-infection The co-infection ws studied in individuls who even did not hve the ide of them suffering from HIV. The overll rte of co-infection with HBV nd/or HCV ws 5.9% (6 out of 101 ptients), mong which 4 [4.0% (Confidence Intervl 0.1, 7.8)] were positive for oth HIV nd HBV infection while 3 [3.0% (Confidence Intervl 0.0, 6.0)] were positive for HIV nd HCV. Among such co-infection, only one [1.0% Confidence Intervl (-1.0, 3)] ws positive for ll three (HIV, HBV, HCV) mrkers. Age The ge-specific seroprevlence ws not significntly different mong HBV nd HCV co-infection in HIVpositive ptients, lthough it ws mrginlly higher in the ge group yers. Gender Non-significnt ssocition of gender with co-infection of HBV or/nd HCV in HIV-positive ptients ws oserved, though the co-infection of HBV or/nd HCV in HIVpositive ptients were only found in the mle. Mritl sttus Though ll the cses of co-infection of HBV nd/or HCV in HIV-positive ptients were found in the mrried mle, the chi-squre test reveled no significnt ssocition mong them. Injectle drug users (IDUs) Less thn 2% of prticipnts were found to e IDUs. One IDU ws found to e suffering from HCV co-infection while 4 IDUs cses were positive for HCV infection. Two IDUs were suffering from HBV co-infection while 5 IDUs cses were positive for HBV infection. In totl 6 IDUs were suffering from HIV, of which 2 were co-infected with HBV nd 1 with HCV. DISCUSSION More thn three-fourth of the individuls in the study were

7 Mhto et l. 129 ged etween yers. Although the risk in mle ge group ws more widely distriuted over ge groups, the findings were true in oth mle nd femle. Among the ge group yers, pproximtely two-third were etween yers of ge. The results show tht the individuls of high sexul ctivity ge re frequent for screening. Mrrige, pregnncy, surgery, nd injection re high in femle ged etween yers. Fctors like ffirs, hving more thn one sexul prtner, unsfe sex re more frequent in this ge group. Our study lso reveled tht the prevlence of HCV (93.3%) nd HIV (65.3%) ws higher in mle, ut HBV ws higher in femle (52.8%). HCV ws evenly confined etween yers of ge in the mle. The distriution of HBV ws seen over wide rnge of ge group from 10 yers to 69 yers. An lrmingly high shre of infection of out 61% occupied the ge group yers in femle ws worrisome. Unlike femle, nerly 84% of HBV ws seen mong the ge group yers in mle. Overll, three-fourth of the HBV ws seen in the ge group yers. The study of the clinicl prevlence of HIV, HBV nd HCV nd their co-infection hve een very few nd decdes old s well (Ri et l., 1994; Shresth et l., 1998; Swym et l., 1999), yet the studies of HBV nd HCV in the lood donted to lood nk hve een in considerle numer (Chnder nd Phw, 2003; Krki et l., 2008; Shresth et l., 2009; Tiwri et l., 2010). A study y Kinkel et l. (2015) in Nepl points out the use of injection for drugs s mjor reson for the surge in the prevlence of such infection nd coinfections. The prevlence of HIV in our study (2.7%) ws higher thn the similr study (0.8%) conducted in western Nepl while the co-infection of HBV nd/or HCV (5.9%) in HIV positives in our study ws slightly lower thn the findings (7.3%) of western Nepl (Suprm et l., 2015). The prevlence of such infections nd co-infections worldwide displys gret vrition. It my e due to the regionl topogrphy, wreness, sexul prctice nd socil hesittion to visit hospitls, lck of surveillnce nd well-equipped hospitl nd lortory nd poverty in the different geogrphicl re. The overll co-infection (5.9%) of HBV nd/or HCV in HIV positives supports the ide of the negtive impcts of HIV infection with the progression of HBV nd HCV infection. High rtes of virl persistence, higher heptitis virl lod nd more rpid progression of liver firosis nd heptocellulr crcinom in co-infected ptients re the mjor negtive impcts (Westrook nd Dusheiko, 2014; Michielsen et l., 2005; Zmor et l., 2017). HIV-heptitis co-infection re t higher risk of developing liver enzyme elevtions on ntiretrovirl therpy (Hoffmnn et l., 2014). Co-infection with the three viruses my result erly deth due to increse in the risk of cute nd chronic liver insufficiency, cirrhosis, heptic filure in comprison to when person is mono-infected. Unlike other studies, our study reported tht only mle popultion of yers of ge ws co-infected with HBV nd/or HCV. Through questioning the infected ptients, it ws estlished tht most of them were drug ddicts nd few might hve cquired it from unsfe sex nd visit to prostitutes nd homosexulity. The ssocition etween older ge groups nd HBV is widely oserved fct resulting from the incresed risk of exposure with time nd greter vccintion coverge in younger popultions (Oliveir et l., 2014; Brndão et l., 2015). Unfortuntely, the prctice of screening for HCV nd HBV is not routine in Nepl t the initil ssessment of HIV-positive ptients. According to Chkrvrti (2011), the most common identifile risk fctor for HCV cquisition ws history of surgery followed y lood trnsfusion. Less thn 2% prticipnts reported history of injectle drug use. The prevlence of HCV nd HBV ws 0.4 nd 1.4% respectively while the IDUs ccounted 26.7% of totl HCV nd 9.4% of totl HBV which were lower thn the previous study in Nepl (Kinkel et l., 2015). In comprison to HCV, sexul trnsmission of HBV is higher followed y injection use y drug ddicts (Kinkel et l., 2015). IDUs ccounted 13.2% of totl HBV. The high rte of drug ddiction in Nepl is of gret concern. A totl of 5.9% HIV positives were identified s IDUs. The gender-wise HIV positive proportion reveled tht the mle popultion is t mximum risk. It my e ecuse of unsfe sex with sex workers, extr mritl ffirs hving more thn one sexul prtner, injection of the drug mong drug ddicts. As Nepl is remittncesed economy, severl individuls working rod nd their spouse residing loclly hve multiple sex prtners. In most of the cses, the person who returned home infected their spouse. Drug ddiction nd use of injection re lso contriuting to HIV mong the joless people. As per gender, nerly three-fourth mle re ccounting the HIV of injection group. Unemployment force femle to prostitution which is illegl y lw. This forces them to unsfe nd compromised sex. Government negligence nd mking prostitution illegl hve forced sex worker of not hving routine medicl check-ups for sexully trnsmitted diseses (STDs). Hence, the sexully ctive ge group re t mximum risk nd ccounts higher infections. Despite the methodologicl limittions, this study ws imed t prevlence estimtion nd outlining of the profile of HBV, HCV nd HIV nd HIV-infected ptients co-infected with HBV nd HCV coming for primry checkups efore ART. Conclusion The prevlence of HIV, HBV nd HCV is 2.7, 1.4 nd 0.4% respectively. 65.3% of mles nd 34.7% of femles

8 Int. J. Appl. Microiol. Biotechnol. Res. 130 suffer from HIV, 47.2% of mles nd 52.8% of femles suffer from HBV, nd 93.3% of mles nd 6.7% of femles infect from HCV. The overll rte of co-infection with HBV nd/or HCV is 5.9%. 3.0% re positive for oth HIV nd HBV infection mrker while 2.0% re positive for HIV nd HCV mrkers. 1.0% cse is positive for ll three mrkers of HIV, HBV nd HCV. Age group re t more risk for such n infection. Screening of HIV, HBV, nd HCV for ptients with suspected history should e done. Routine screening for HBV nd HCV mong ll HIV-positive individuls nd their sexul prtners re dvisle in order to stop liver-relted moridity nd mortlity due to co-infections. Higher prevlence of HIV nd HBV in this region highlights the need of wreness progrm nd government intervention to equip the hospitls for primry screening. For true picture of these infections, ntionwide study involving ll helth centers hs to e initited nd regulted. REFERENCES Blogun T. M., Emmnuel S. & Ojerinde E. 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