HIV, hepatitis B virus, and hepatitis C virus co-infection among HIV positives in antiretroviral treatment program in selected hospitals in Addis Ababa: A retrospective cross-sectional study.

INTRODUCTION: HIV co-infection with hepatitis B (HIV-HBV) and hepatitis C (HIV-HCV) is known to affect treatment outcomes of antiretroviral therapy (ART); however, its magnitude is not well documented. We aimed to determine the magnitude of HIV-HBV and HIV-HCV co-infections simultaneously in people living with HIV (PLHIV) enrolled in ART care in Addis Ababa.
METHODS: We reviewed the medical records of adults ≥15 years who were receiving ART care in three high burden hospitals in Addis Ababa. Baseline clinical and laboratory test results were extracted from medical records. Co-infection was determined based on hepatitis B surface antigen (HBsAg) and hepatitis C virus antibody (anti-HCV) tests obtained from the medical records. A multivariable logistic regression model was used to identify the risk factors for hepatitis B and C co-infections.
RESULTS: A total of 873 HIV-positive participants were included in this study. The median age of the participants was 37.5 years, and 55.7% were women. Overall, HIV-HBV co-infection was 5.96% (95% CI: 4.56-7.74), and HIV-HCV co-infection was 1.72% (95% CI: 1.03-2.83). The multivariable logistic regression showed that the male sex was the most independent predictor for viral hepatitis B co-infection with an odds ratio of 2.42(95% CI:1.27-4.63). However, HIV-HCV co-infection did not show a significant association in any of the sociodemographic data of the participants.
CONCLUSION: HIV co-infection with hepatitis B was moderately high in individuals enrolled in ART care in Addis Ababa. Men had significantly higher HIV-HBV co-infection. HIV co-infection with hepatitis C was relatively low. Strengthening integrated viral hepatitis services with HIV care and treatment services should be emphasized to improve patient care in health facilities.

Introduction

HIV-associated morbidity and mortality have declined in resource-limited countries owing to the rapid scale-up of antiretroviral therapy [1]. However, co-infection with hepatitis B virus (HBV) and hepatitis C virus (HCV) has emerged as a clinical and public health challenge [2]. Studies have shown an increase in the number of liver-related deaths among antiretroviral users [2-4].The guidelines of the World Health Organization (WHO) in 2016 recommended the early detection and screening of viral hepatitis in people living with HIV (PLHIV) at the initiation of antiretroviral therapy (ART) [5]. Globally, less than 5% of PLHIV know their HBV or HCV status [6].Although Africa is known to harbor most of the HIV, HBV, and HCV infected people, little is known about viral hepatitis co-infection status in HIV-positives enrolled in ART care.

In Ethiopia, more than 669,000 people were living with HIV; 11,500 people died from an AIDS-related illness, and 71% of PLHIV were treated by the end of 2019 [7]. Viral hepatitis is endemic, with an estimated national hepatitis B prevalence of 9.4% in 2017 [8]. There is no recent national population-based HCV study; however, population-based studies from the northwest and southwest of the country showed hepatitis C prevalence of 1% in 2017 [9] and 1.9% in 2018 [10].

As in most parts of the world, there is a general insufficiency in the diagnosis of HBV and HCV as in Ethiopia, which remains a challenge in explaining the magnitude and pattern of co-infections and providing the necessary treatment to HIV positives. Studies in some hospitals in Addis Ababa have estimated co-infections [11, 12]. Understanding the magnitude of HIV-HBV and HIV-HCV co-infection is necessary to prioritize care and treatment services for co-infected individuals.

The lack of continuous studies to estimate HBV and HCV in Ethiopia remains a serious challenge in determining the pattern of co-infection. Therefore, we aimed to determine the magnitude of HIV-HBV and HIV-HCV co-infection in PLHIV who were on ART in Addis Ababa.

Methods

Study design and setting

A retrospective cross- sectional medical record review was carried out in three big hospitals in Addis Ababa; Zewditu Hospital, Alert Hospital, and Black Lion Hospital. The three hospitals together serve 16,400 ART clients. Addis Ababa, the capital city of Ethiopia, has a population of more than five million people [13]. Addis Ababa is the second highest (3.5%) HIV prevalent city, with an estimated 125,000 PLHIV in 2020. In addition, one-fourth of the total number of PLHIV in the country receive ART in health facilities in Addis Ababa [7].

The antiretroviral treatment program in Ethiopia is provided based on the national guidelines, which were adopted from the WHO guidelines issued in 2016. The guideline recommends screening all HIV-positives for both hepatitis B and hepatitis C viruses at ART initiation [14, 15].

However, the guidelines are not fully practiced in most health facilities that provide ART services in the country. The recommended testing is a serological test for HB surface antigen (HBsAg) and anti-HCV antibodies. The guideline also recommends a first-line drug combination regimen of antiviral agents (tenofovir and entecavir) that are active against HBV. Direct acting drugs that treats HCV are very expensive and not easily accessible, even for those who can afford to pay for it until recently. In response to this, the Federal Ministry of Health has promoted and included HCV direct-acting drugs in the essential drug list. Following that private healthcare providers have started procuring direct-acting drugs to treat HCV. The government is also working to expand affordable access to direct-acting drugs through increasing procurement, private partnerships, and awareness. The second national five years (2021–2025) Viral Hepatitis Strategic Plan also included subsidized procurement of direct-acting drugs as one strategic direction to improve the health of the nation.

The outcome of this study was the magnitude of HIV-HBV co-infection and HIV-HCV co-infection. Adults ≥ 15 years who had documented viral hepatitis B and viral hepatitis C test results enrolled in ART care from September 2012 to December 2018 (started routine viral hepatitis screening) were eligible. Study participants without viral hepatitis test results were excluded from the analysis. A comparison was made between those who had test results and those without test results to check for selection biases using selected demographic and clinical characteristics of age, sex, education, and marital status.

Sample size determination and sampling procedures

The sample size for the study was determined using a single proportion formula, which required 256 and 240 HBV and HCV positives, respectively. The following assumptions were included in the calculation:Co-infection prevalence of HIV-HBV and HIV-HCV was 0.052 and 0.055, respectively [16], margin of error, d = 0.03, Z = 1.96, and a 95% CI. However, we included all eligible adults ≥ 15 years with documented viral hepatitis test results enrolled in ART care from September 2012 to December 2018 to increase the precision of the estimate and be able to conduct internal comparisons. A total of 873 (Zewditu hospitals, n = 523, ALERT, n = 186, and Black Lion hospital, n = 164) HIV positives with documented viral hepatitis B and viral hepatitis C were included in this study.

Data extraction procedures

The data extraction form was developed in English and pretested in two health facilities that were not part of the study sites. Four data abstractors (two BSc nurses and two data managers) were trained in the selected study hospitals. The data collectors were team-up in two groups with a data manager and a nurse in one group to abstract clinical and laboratory data from the patient folder directly into a tablet. An experienced supervisor with a master’s degree in Public Health joined the data abstractor team.

The folders of the HIV positives was the source of information for this study. The client folder included a follow-up card in which the HIV positive ‘ visits were recorded at ART initiation and during the follow-up visits. Laboratory results and history intake forms were attached to the patient folder. The patient folder is kept in the Health Information System (HMIS) archive, which is a dedicated card room where all patient cards are kept independent of the patient’s HIV status. The ART register and the electronic database were used to obtain a list of the unique ART numbers and eligible adults for the study.

The ART data manager sorted 20–30 eligible unique ART numbers from the database and sent them to a runner (HMIS card person) to remove the patient folder from the HMIS for data abstraction during the study period.

The data extracted from the medical records of HIV-positive included sociodemographic characteristics (age, sex, marital status, education, and religion), baseline laboratory and clinical characteristics, hepatitis B surface antigen (HBsAg) test, and anti-hepatitis C virus (anti-HCV) test. Extracted data were uploaded daily to the REDCAP server after the supervisor checked for completeness of the data. REDCAP is a secure web application for building and managing online surveys and databases. The data were collected from November 12, 2019, to March 15, 2020. Data were exported from REDCAP to STATA14 for further cleaning, data management, and analysis.

Statistical analysis

The background characteristics of the study participants were described with the median and interquartile range (IQR) for numeric variables and frequency and proportion for categorical variables. The normality of the distribution was tested first to determine the type of statistical test. For numeric variables, the Kruskal-Wallis equality-of-populations rank test or Wilcoxon test was used as appropriate. For categorical outcome variables, the chi-square test was used. Statistical significance was set at p <0.05. The co-infection of HIV-HBV and HIV-HCV was calculated using proportions and presented with a 95% confidence interval. Multivariable logistic regression was used to assess risk factors for HIV-HBV and HIV-HCV, and the odds ratio (OR) with 95% CI was presented.

Ethics approval and consent to participate

This study was approved by the Institutional Review Board of the University of Gondar in Ethiopia and the Armauer Hansen Research Institute, Addis Ababa, Ethiopia. No consent was required from participants. No external persons accessed the patient’s folder, and no patient identifier was included in our data set. The patient folder that was taken from the HMIS room for data abstraction was kept in a locked file cabinet until it was returned to the HMIS room to maintain participant’s confidentiality. The analysis used routinely collected patient data for program monitoring and evaluation purpose, and had no direct patient contact. As this was a retrospective study, the University of Gondar Institutional Review Board waived the need for informed consent. We confirm that all methods have been carried out in accordance with relevant guidelines and regulations.

Results

Characteristics of the study participants at ART enrollment

Of the 998 adults eligible for this study, 873 were included in this analysis, and 125 (15%) of the study participants were excluded from the analysis because they did not have documented viral hepatitis test results. Of the 873 participants included, their median age at enrollment was 37.5, with an inter-quartile range (IQR) of–31–45 years, and 55.7% were female. Married participants constituted the highest proportion 408(48.69%) of the study participants. Regarding educational status, 731(86.82%) of the study participants had attained basic education, 296(35.15%) attained secondary education, and 173(20.55%) attained tertiary education Table 1.

Table 1

Socio-demographic characteristics of the participants at ART enrollment (September 2012-December 2018), Addis Ababa, n = 873.

Variables	n(%)
Age in (years), (n = 873)
15–29	166 (19.01)
30–44	469 (53.72)
45–59	198 (22.68)
60+	40 (4.58)
Median (IQR)	37.5 (31–45)
Sex, (n = 869)
Male	385(44.30)
Female	484(55.69)
Marital status, (n = 838)
Never married	210(25.05)
Married	408(48.68)
Separated/Divorced	140(16,70)
Widowed/er	80(9.54)
Education, (n = 842)
No education	111(13.18)
Primary	262(31.11)
Secondary	296(35.15)
Tertiary	173(20.54)
Religion, (n = 843)
Muslim	97(11.50)
Orthodox	646(76.63)
Protestant	89(10.55)
Catholic	4(0.47)
Other	7(0.83)

IQR:Inter-quartile range; ART: Antiretroviral Therapy.

Magnitude of HIV-HBV co-infection

The overall HIV-HBV co-infection was 5.96% (95% CI: 4.56–7.74, n = 52) in PLHIV enrolled for ART care. HBV co-infection was highest in men (8.83%;95% CI: 6.37–12.12, n = 34) than in female participants and 3.72% (95% CI: 2.35–5.83, n = 18, p = 0.0001). In terms of age group, HBV co-infection was the highest in the age group of 45–59 years (7.57% [95% CI: 4.60–12.23,]) and lowest in the age group 15–29 years (4.21% [95% CI: 2.01–8.64, p = 0.626]).

Married study participants had the highest HIV-HBV co-infection of 7.59%(95% CI: 5.39–10.62, p = 0.316), followed by separated/divorced (5.71%;95% CI: 2.86–11.10), and the lowest HBV was reported among those who had never been married (3.80%;95% CI: 1.91–7.47) Table 2.

Table 2

HIV-HBV and HIV-HCV co-infection in HIV positives enrolled in ART care by selected background characteristics (September 2012-December 2018), Addis Ababa, N = 873.

Variables	Overall	HIV-HBV Co-infection		HIV-HCV Co-infection
	N	N	Estimated Prevalence (%) (95% CI)	N	Estimated Prevalence (%) (95% CI)
Total	873	52	5.96(4.56–7.74)	15	1.72(1.03–2.83)
Age in (years)
15–29	166	7	4.21(2.01–8.64)	4	2.41(0.89–6.30)
30–44	469	28	5.97(4.15–8.52)	4	0.85(0.32–2.25)*
45–59	198	15	7.57(4.60–12.23)	4	2.02(0.75–5.30)*
60+	40	2	5.00(1.18–18.87)	3	7.50(2.32–21.71)
Sex
Male	385	34	8.83(6.37–12.12)*	7	1.82(0.87–3.77)
Female	484	18	3.72(2.35–5.83)	7	1.45(0.69–3.01)
Marital status
Never married	210	8	3.81(1.91–7.47)	5	2.38(0.99–5.63)
Married	408	31	7.59(5.39–10.62)	4	0.98(0.37–2.59
Separated/Divorced	140	8	5.71(2.86–11.10)	2	1.43(0.35–5.62)
Widowed/er	80	4	5.00(1.85–12.82)	2	2.50(0.61–9.72)
Education
No education	111	6	5.40(2.42–11.64)	1	0.90(0.12–6.29)
Primary	262	14	5.34(3.18–8.84)	4	1.52(0.57–4.02)
Secondary	296	24	8.11(5.48–11.83)	3	1.01(0.33–3.11)
Tertiary	173	8	4.62(2.31–9.03)	4	2.31(0.86–6.05)
Religion
Muslim	97	6	6.18(2.76–13.26)	1	1.03(0.13–7.18)
Orthodox	646	41	6.34(4.70–8.51)	11	1.70(0.94–3.05)
Protestant	89	3	3.37(1.06–10.13)	1	1.12(0.15–8.81)
Catholic	4	1	(**)	-	-
Other	7	1	(**)	-	-

HBV: hepatitis B virus; HCV: hepatitis C virus; HIV: human immunodeficiency virus; CI: confidence interval

*p-value < 0.05 is considered as statistically significant level (

** Observation smaller than 25)

Magnitude of HIV-HCV co-infection

HIV-HCV co-infection among HIV-positive enrolled for ART care was 1.72% (95% CI: 1.03–2.83, n = 15). HCV co-infection was highest, 7.50% (95% CI: 2.31–21.71, p = 0.014) in the age group of 60+, followed by the age group 15–29 years 2.41% (95% CI: 0.89–6.30). With regard to the marital status of HIV positiveswith HCV, widowed/er showed the highest HCV co-infection, 2.50% (95% CI: 0.61–9.72) followed by never married, 2.38% (95% CI: 0.99–5.63), and the lowest was recorded among the married study participants (0.98% [0.37–2.59]). Male’s sex had a higher 1.82(0.87–3.77) HCV co-infection compared with female 1.45(95% CI: 0.69–3.01, p = 0.665) study participants Table 2.

Factor associated with co-infection

The multivariant logistic regression showed that the male sex was the most independent predictor for viral hepatitis B co-infection with an OR of 2.42(95% CI: 1.27–4.63) Table 3. However, the multivariate logistic regression analysis of HIV-HCV co-infection did not show a significant association with HCV co-infection in all the socio-demographic data of the participants (age groups, sex, educational status, marital status, and religion) Table 4.

Table 3

Factors associated with viral hepatitis B co-infections in HIV positives enrolled in ART care (September 2012-December 2018) Addis Ababa, n = 873.

Variables	Viral Hepatitis B Co-infection
	Bivariate		Multivariable
	Odds ratio	p-value	Odds ratio	p-value
Sex
Male	2.50(1.39–4.51)	0.002	2.42(1.27–4.63)	0.007
Female	1		1
Age group
15–29	0.83(0.16–4.18)	0.828	1.30(0.23–7.13)	0.75
30–44	1.20(0.27–5.25)	0.803	1.34(0.29–6.13)	0.69
45–59	1.55(0.34–7.09)	0.567	1.37(0.28–6.51)	0.69
60+	1		1
Marital Status
Single	1		1
Married	2.07(0.93–4.60)	0.072	2.13(0.92–4.92)	0.074
Divorced/Separated	1.53(0.56–4.17)	0.406	1.87(0.65–5.35)	0.242
Widowed/er	1.32(0.38–4.54)	0.650	1.51(0.40–5.62)	0.535
Education
No Education	1		1
Primary	0.98(0.36–2.64)	0.981	0.78(0.28–2.16)	0.640
Secondary	1.54(0.61–3.88)	0.356	1.24(0.47–3.25)	0.662
Tertiary	0.84(0.28–2.51)	0.767	0.63(0.20–2.02)	0.441
Religion
Orthodox	1		1
Muslim	0.97(0.40–2.35)	0.952	0.99(0.40–2.47)	0.997
Protestant	0.51(0.15–1.69)	0.276	0.53(0.15–1.81)	0.317
Catholic	4.91(0.50–48.33)	0.172	7.01(0.596–82)	0.121
Other	2.45(0.28–20.91)	0.410	2.2(0.24–20.62)	0.479

Table 4

Factors associated with viral hepatitis C co-infections in HIV positives (September 2012-December 2018) Addis Ababa, n = 873.

Variables	Viral Hepatitis Co-infection
	Bivariate		Multivariable
	Odds ratio	p-value	Odds ratio	p-value
Sex
Male	1.26(0.43–3.62)	0.666	1.27(0.32–4.96)	0.728
Female	1		1
Age group
15–29	0.30(0.06–1.41)	0.130	0.54(0.04–6.38)	0.630
30–44	0.10(0.02–0.49)	0.004	0.12(0.01–1.51)	0.101
45–59	0.25(0.05–1.18)	0.008	0.49(0.04–5.16)	0.558
60+	1		1
Marital Status
Never married	1		1
Married	0.40(0.10–1.52)	0.182	0.29(0.06–1.43)	0.131
Divorced/Separated	0.59(0.11–3.10)	0.537	0.29(0.03–2.92)	0.298
Widowed/er	1.05(0.19–5.53)	0.953	0.48(0.04–5.24)	0.554
Education
No Education	1		1
Primary	1.70(0.18–15.43)	0.635	1.32(0.13–12.80)	0.808
Secondary	1.12(0.11–10.94)	0.918	0.88(0.08–9.10)	0.916
Tertiary	2.60(0.28–23.60)	0.395	0.90(0.07–11,12)	0.937
Religion
Orthodox	1		1
Muslim	0.60(0.07–4.71)	0.628	1.02(0.12–8.67)	0.979
Protestant	0.65(0.08–5.14)	0.688	0.92(0.11–7.72)	0.940
Catholic	-		-
Other	-		-

Discussion

We found that the magnitude of HIV-HBV co-infection was 5.96% and HIV-HCV co-infection was 1.72% in adults ≥15 years in Addis Ababa. The factor that was statistically and strongly associated with HBV co-infection was the male sex, which had a higher likelihood of being co-infected.

We demonstrated findings similar to those of other studies conducted in HIV-positive enrolled for ART care in hospital settings. For instance, our finding of HIV-HBV is similar to those of studies from the northern part of the country (5.6%,5.9%, and 5.7%) [17-19]. Overall, our finding of HIV-HBV is also consistent with the findings of other similar studies in Ethiopia, ranging from 4.7% to 7.7% [20-23].

On the other hand, our finding of HIV-HBV is much lower than the study conducted in the eastern parts of the country (11%) in three regional hospitals of Diredawa, Somali, Harari, and in the southern part of the country at Wolita Sodo Hospital [24, 25]. This difference seems to be related to traditional practices, such as female genital mutilation between regions, which may facilitate transmission. For instance, female genital mutilation is most common in the eastern part of the country, with 99% in Somali, 91% in Afar, 82% in Harari, and 75% in Dire Dawa compared with 54% in Addis Ababa or other parts of the country [26].

The HIV-HBV prevalence in this study was moderately high. More than 90% of HBV transmission occurs during childbirth in Africa and Asia [27]. The vaccination against the HBV was not yet in place in Ethiopia until 2006, when the country introduced pentavalent vaccination, which includes a vaccine against HBV at birth 14 years prior. Currently, adults ≥ 15 years were not part of the vaccination cohort. This may have contributed to the moderately high HIV-HBV co-infection findings in this study. Therefore, the moderately high HIV-HBV found in this study may remain the same for some time until the new vaccinated cohort joins the adult population.

In this study, we also found that HIV-HBV co-infection was twice as high in men than in women in the same age group. A similar significant difference between the sexes in HIV-HBV co-infection was reported in Ethiopia and other African countries [17, 19, 28]. The reason why men have higher HBV levels than women is beyond the scope of this study. However, previous large cohort and 12 years follow-up studies showed that biological factors are more important than environmental or behavioral factors. This explains that the higher HBV in men indicates that females develop HBV antibodies faster than males, which helps them to clear HBsAg early, so most may not progress to chronic HBV. Second, the female sex hormone, estrogen, may be a protective factor for females and androgen as a risk factor for men contributing to higher HBV [29, 30]. This may call for emphasis to encourage men to target HBV screening without compromising efforts to women.

We found a low (1.72%) HIV-HCV co-infection among HIV-positive enrolled in ART care in three Addis Ababa hospitals. Others also found extremely low HIV-HCV co-infections, as low as 1.0%in Addis Ababa, 1.3% in Debretabor Hospital, and 1.1% in Gondar Hospital [11, 31, 32].

In contrast, studies from Hawassa, Bahirdar, and Adawa hospitals showed a high HIV-HCV co-infection ranging from 5% to 11% [33-35]. The Hawassa study indicated that only 9.1% of the HIV-HCV cases were confirmed by polymerase chain reaction and were viremic. Studies have shown that 88% of HCV persists with chronic HCV, as opposed to what is reported in the Hawassa study [36]. Therefore, a direct comparison with this study in the absence of HCV viremic results from our study may not be reasonable.

The higher HIV-HCV prevalence from Adawa and Bahirdar hospitals may be due to the associated risk factors identified, including hospitalization, blood transfusion, and tooth extraction, which may contribute to high HIV-HCV co-infection in the area. Our finding of HIV-HCV co-infection is consistent with the Sub-Saharan Africa estimates of the general population, but much lower than the estimate in the HIV population [37].

The high HIV-HCV co-infection in sub-Saharan Africa can be explained by the prevalent behavioral factors such as injection of drugs in the HIV population in some parts of Africa. For instance, in Western and Central Africa, injecting drug use is common in the HIV population [38], while in Ethiopia, the HIV prevalence in people with injecting drug users is not different from the general population in urban setting. In 2018, co-infection of HIV-HBV and HIV-HCV among people with injecting drug users in Addis Ababa was 5.1% and 2.9% respectively [39].

The limitations of this study are as follows: We extracted hepatitis B and C test results from those who had documented viral hepatitis test, and we excluded those who did not have documented viral hepatitis test results (15%) of the eligible population, which may introduce selection bias. However, we compared HIV-positive who had documented viral hepatitis test versus those without based on their biological and background characteristics and did not find any difference. Second, we could not include some potential confounding variables and possible predictors related to behavior and route of transmission because they were not routinely collected at ART initiation. The absence of data on the route of transmission and behavior may affect the predictors of co-infection but does not alter the magnitude of co-infections. Test kits variation at health facilities with possible variation in sensitivity and specificity could be another limitation of the study.

Conclusions

HIV co-infection with hepatitis B was moderately high in individuals enrolled in ART care in Addis Ababa. Men had significantly higher HIV-HBV co-infection. HIV co-infection with hepatitis C was relatively low. Strengthening integrated viral hepatitis services in HIV care and treatment services should be emphasized to improve patient care in health facilities, and a more detailed study is needed to understand the factors associated with viral hepatitis B and C.

Raw data used for analysis.

(DTA)

Click here for additional data file.

15 Feb 2022

A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Desalegn Admassu Ayana, Ph.D

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. Please provide additional details regarding participant consent. Specifically, if the need for consent was waived by the ethics committee, please include this information. Thank you.

3. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match.

When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section.

4. Thank you for stating the following in the Acknowledgments Section of your manuscript:

We declar that there was no direct fund that the authors received. However,  the data collection fee at the three hospitals was covered thrugh AHRI by Federal HIV/AIDS Prevention and Contorl Office. The source of fund had no role in the design or conduct of the study; the collection, management, analysis, and interpretation of the data; or the preparation, review, or approval of the manuscript. The authors had full access to all the data in the study and takes the responsibility for the integrity of the data and the accuracy of the data analysis. The authors declars that there was no fund received from any organization.

We note that you have provided additional information within the Acknowledgements Section that is not currently declared in your Funding Statement. Please note that funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form.

Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows:

The author(s) received no specific funding for this work.

Please include your amended statements within your cover letter; we will change the online submission form on your behalf.

5. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions.

In your revised cover letter, please address the following prompts:

a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent.

b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories.

We will update your Data Availability statement on your behalf to reflect the information you provide.

6. We note that you have stated that you will provide repository information for your data at acceptance. Should your manuscript be accepted for publication, we will hold it until you provide the relevant accession numbers or DOIs necessary to access your data. If you wish to make changes to your Data Availability statement, please describe these changes in your cover letter and we will update your Data Availability statement to reflect the information you provide.

7. We note that you have included the phrase “data not shown” in your manuscript. Unfortunately, this does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. We require that authors provide all relevant data within the paper, Supporting Information files, or in an acceptable, public repository. Please add a citation to support this phrase or upload the data that corresponds with these findings to a stable repository (such as Figshare or Dryad) and provide and URLs, DOIs, or accession numbers that may be used to access these data. Or, if the data are not a core part of the research being presented in your study, we ask that you remove the phrase that refers to these data.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: No

Reviewer #2: Yes

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: No

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Line 38: the presented percentage might exaggerate the result as the observed outcome is only 1. Hence it is better if the author avoid calculating proportion for very small numbers.

The investigators included only sociodemographic factors in to the final multivariate model. However the exposure and clinical risk factors are more important for HBV and HCV coinfection. so it is better if the analysis is repeated with missed more relevant factors for decision making and policy purposes.

finally I think the conclusion is biased as the analysis and presentation lack the most relevant clinical and behavioral factors

Reviewer #2: Comments on the manuscript “HIV, hepatitis B virus, and hepatitis C virus Co-infection patients in Addis Ababa Antiretroviral Treatment Program, 2012-2018: a retrospective cohort study” PONE-D-21-34939

Line 1. HIV, hepatitis B virus, and hepatitis C virus Co-infection patients in Addis Ababa : Add “among “ between Co-infection and patients

Line 1-2 : HIV, hepatitis B virus, and hepatitis C virus Co-infection patients in Addis Ababa Antiretroviral Treatment Program, 2012-2018: a retrospective cohort study: How this study become a retrospective study as far as the authors took data in a one point time ? I felt it is a retrospective cross sectional study !.

The authors also considered the study was done in Addis Ababa, rather it was done among selected hospitals and it is good to revise the title in that sense.

Line 76-77, Study design and setting: the study design is not described well !! it mention the word retrospective only which is not adequate.

Line 90- 92: Direct acting drugs that threaten HCV are very expensive and not easily accessible, even for those who can 92 afford to pay for it. : There are national initiative in line with this and better to underscore that and your study could support that initiative.

Line 96. Study participants without viral hepatitis test results were excluded from the analysis: About 125 HIV patients had no HBSAg and HCV tests ? how can we assure bias happened from these group of HIV patients on Co-infection of HIV/HBV and HIV /HCV co-infection rate? You need to state this as a in the limitation part in more description.

Similarly, the authors did not know what types of rapid tests for HBSAg and HCV tests were implemented ? any effect of accuracy / sensitivity/ specificity issues on proportions of co-infections among the three hospitals ? Changes of test kits between time and years??

The authors did not mentioned the three hospitals in Addis Ababa and how many HIV patients each hospitals could have ? how many participants were enrolled from each hospital ? It is better to describe these under methods part.

It seems that the authors described the magnitude of HBSAg and HCV in the study site is moderately high, HIV-HBV co-infection was 5.96% (95% CI: 4.56-7.74) , HIV-HCV co-infection was 1.72% (95% CI: 1.03-2.83), and triple co-infection of HIV-HBV-HCV was 1(0.11%) . In reality this magnitude is determined for 7 years’ time (between 2012-2018). I think the authors should consider this while they describe the co-infection rate. Rather If they anticipate for yearly prevalence one can see trend of co-infections as far as they have adequate number of participants per year !.

Line 199-200 : We found that the magnitude of HIV-HBV co-infection was 5.96% and HIV-HCV co200 infection was 1.72% in adults ≥ 15 years in Addis Ababa.: how can we generalize your findings for Addis Ababa? Be specific to HIV patients and specific hospitals

Line 275 References: References

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

11 Mar 2022

Point-by-point response to reviewer’s

Dear Editor and Reviewers

Thank you very much for taking time in reviewing our manuscript and providing valuable comments. We took your suggestions and comments into consideration and revised our manuscript accordingly. The point-by-point response presents as below.

I. Editor comment

Comment 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming.

Response 1: As suggested we now organized our manuscript as per the PLOS ONE’s style and requirement and the naming formats.

Comment 2. Please provide additional details regarding participant consent. Specifically, if the need for consent was waived by the ethics committee, please include this information. Thank you.

Response 2: Thank you for this valuable suggestion, statement on consent wavier included in the manuscript, under method-Ethics section on page 8 line 169 to 173

Comment 3. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match.

When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section.

Response 3: We now removed statements regarding funding from the manuscript and no grant given.

Comment 4. Thank you for stating the following in the Acknowledgments Section of your manuscript:

We declare that there was no direct fund that the authors received. However, the data collection fee at the three hospitals was covered through AHRI by Federal HIV/AIDS Prevention and Control Office. The source of fund had no role in the design or conduct of the study; the collection, management, analysis, and interpretation of the data; or the preparation, review, or approval of the manuscript. The authors had full access to all the data in the study and takes the responsibility for the integrity of the data and the accuracy of the data analysis. The authors declares that there was no fund received from any organization.

We note that you have provided additional information within the Acknowledgements Section that is not currently declared in your Funding Statement. Please note that funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form.

Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows:

The author(s) received no specific funding for this work.

Please include your amended statements within your cover letter; we will change the online submission form on your behalf.

Response 4: We included amendment statements on the cover letter “The author(s) received no specific funding for this work”

Comment 5. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions.

In your revised cover letter, please address the following prompts:

a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent.

b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories.

We will update your Data Availability statement on your behalf to reflect the information you provide.

Response 5: We agree with data availability statement and amended on cover letter as

“All data are fully available without restriction”

Comment 6. We note that you have stated that you will provide repository information for your data at acceptance. Should your manuscript be accepted for publication, we will hold it until you provide the relevant accession numbers or DOIs necessary to access your data. If you wish to make changes to your Data Availability statement, please describe these changes in your cover letter and we will update your Data Availability statement to reflect the information you provide.

Response 6: Data Availability statement amended on the cover letter “Yes - all data are fully available without restriction “

Comment 7. We note that you have included the phrase “data not shown” in your manuscript. Unfortunately, this does not meet our data sharing requirements. PLOS does not permit references to inaccessible data. We require that authors provide all relevant data within the paper, Supporting Information files, or in an acceptable, public repository. Please add a citation to support this phrase or upload the data that corresponds with these findings to a stable repository (such as Figshare or Dryad) and provide and URLs, DOIs, or accession numbers that may be used to access these data. Or, if the data are not a core part of the research being presented in your study, we ask that you remove the phrase that refers to these data.

Response 7: The statement “data not shown” is removed from page 11 line 204 as it was also suggested by reviewer 1.

II. Review Comments to the Author

Reviewer # 1

Comment 1: Line 38: the presented percentage might exaggerate the result as the observed outcome is only 1. Hence it is better if the author avoids calculating proportion for very small numbers.

Response 1: Thank you very much for the comment and suggestions. It is true, if a percentage is presented out of a very small number it gives wrong impression. However, in our case, we have a total of 873 people tested for both (HBV and HCV) and the triple coinfection found is only 1/873(0.11%). We still believe that showing the percent does not exaggerate. However, our finding of triple coinfection is very small and may not be a big concern and we omitted from the analysis on page 3 line 47, page 11 line 205.

Comment 2: The investigators included only sociodemographic factors into the final multivariate model. However, the exposure and clinical risk factors are more important for HBV and HCV coinfection. so it is better if the analysis is repeated with missed more relevant factors for decision making and policy purposes.

Response 2. We highly appreciate the comment. We used secondary data; the analysis was limited to the available clinical and sociodemographic variables and we acknowledged this limitation on page 18 line 286 to 289.

Regarding the inclusion of clinical factors such as WHO staging, CD4 cell count, hemoglobin etc these could be an effect of disease (due to HIV, viral hepatitis, or TB) rather than a risk factor. However, as suggested, we included WHO staging, CD4 in the model and found still sex as an independent predictor. Hence, we would like to retain the risk factor analysis as it is.

Comment 3: I think the conclusion is biased as the analysis and presentation lack the most relevant clinical and behavioral factors.

Response 3. We understand the concern, however our conclusion emphasized only the overall magnitude of viral hepatitis coinfection and higher coinfection in men. We explained the limitation of the data and hence we believe our conclusion is reasonable.

Reviewer #2:

Comments 1: on the manuscript “HIV, hepatitis B virus, and hepatitis C virus Co-infection patients in Addis Ababa Antiretroviral Treatment Program, 2012-2018: a retrospective cohort study” PONE-D-21-34939

Line 1. HIV, hepatitis B virus, and hepatitis C virus Co-infection patients in Addis Ababa : Add “among “ between Co-infection and patients

Response 1: Thank you for your suggestion. We have now added the world “among” on the title on page 1 line 1 and the title has been rephrased accordingly.

Comment 2 : Line 1-2 : How this study become a retrospective cohort study as far as the authors took data in a one point time? I felt it is a retrospective cross-sectional study.

Response 2: Thank you for this comment. As we used data only at enrollment the correct labelling is retrospective cross-sectional study. We have corrected that in the revised manuscript as the changes are indicated in the title and method sections.

Comment 3: The authors also considered the study was done in Addis Ababa, rather it was done among selected hospitals and it is good to revise the title in that sense.

Response 3: Thank you very much for the comment. We modified the title accordingly on page 1 line 1

As you suggested, we indicated that the study was done in selected three hospitals in the method section to be more specific: on page 5 line 88-89

Comment 4: Line 76-77, Study design and setting: the study design is not described well !! it mention the word retrospective only which is not adequate.

Response 4: We agree with the comment and included the word “cross-sectional” on page 5 line 87

Comment 5 : Line 90- 92: Direct acting drugs that threaten HCV are very expensive and not easily accessible, even for those who can afford to pay for it: There are national initiative in line with this and better to underscore that and your study could support that initiative.

Response 5: We agree with the comment completely. There are national initiatives regarding viral hepatitis C screening and treatment. Now statement added on national initiatives on page 5 line 103-109.

Comment 6: Line 96. Study participants without viral hepatitis test results were excluded from the analysis: About 125 HIV patients had no HBSAg and HCV tests? how can we assure bias happened from these group of HIV patients on Co-infection of HIV/HBV and HIV /HCV co-infection rate? You need to state this as a in the limitation part in more description.

Response 6: We agree with the comment. This was indicated in the limitation section on page 18 line 281 to 286

Comment 7: Similarly, the authors did not know what types of rapid tests for HBSAg and HCV tests were implemented? any effect of accuracy /sensitivity/ specificity issues on proportions of co-infections among the three hospitals? Changes of test kits between time and years??

Response 7: The type of test used during the data collection and their sensitivity and specificity was not available. This remains the limitation of the study as indicated on page 18 line 289 to 291.

Comment 8 : The authors did not mentioned the three hospitals in Addis Ababa and how many HIV patients each hospitals could have? how many participants were enrolled from each hospital? It is better to describe these under methods part,

Response 8: Thank you for the valuable comment. As suggested, we indicated the name of the hospitals and their respective sample on page 6 line 124 to 125

Comment 9. It seems that the authors described the magnitude of HBSAg and HCV in the study site is moderately high, HIV-HBV co-infection was 5.96% (95% CI: 4.56-7.74) , HIV-HCV co-infection was 1.72% (95% CI: 1.03-2.83), and triple co-infection of HIV-HBV-HCV was 1(0.11%) . In reality this magnitude is determined for 7 years’ time (between 2012-2018). I think the authors should consider this while they describe the co-infection rate. Rather If they anticipate for yearly prevalence one can see trend of co-infections as far as they have adequate number of participants per year !.

Response 9: As more than 80% of the sample was from year 2015-2018 the sample was not enough for trend analysis. The purpose of our study was not to see trend analysis rather to see the burden of viral hepatitis in ART clients among those who have got test access.

Comment 10: Line 199-200: We found that the magnitude of HIV-HBV co-infection was 5.96% and HIV-HCV co200 infection was 1.72% in adults ≥ 15 years in Addis Ababa.: how can we generalize your findings for Addis Ababa? Be specific to HIV patients and specific hospitals.

Response 10. Agree with the suggestion, we did not generalize for Addis Ababa, however, taking into consideration the homogeneity of ART users in Addis Ababa, and the likely hood of viral hepatitis transmission time (>90% at childhood) the finding can give a clue on the magnitude of viral hepatitis among ART users in Addis Ababa

Comment 11 : Line 275 References:

Response 11 : References: this was corrected accordingly on page 19 line 304

Submitted filename: Response to reveiwers.docx

Click here for additional data file.

5 Apr 2022

HIV, Hepatitis B Virus, and Hepatitis C Virus Co-infection Among HIV Positives in Antiretroviral Treatment Program in Selected Hospitals in Addis Ababa: a Retrospective Cross-sectional Study

PONE-D-21-34939R1

Dear Eleni Seyoum,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Desalegn Admassu Ayana, Ph.D

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: All comments are addressed

manuscript technically sound, and do the data support the conclusions

Limitations of this research is also indicated.

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Regassa, LD.

14 Apr 2022

PONE-D-21-34939R1

HIV, Hepatitis B Virus, and Hepatitis C Virus Co-infection Among HIV Positives in Antiretroviral Treatment Program in Selected Hospitals in Addis Ababa: a Retrospective Cross-sectional Study

Dear Dr. Seyoum:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Desalegn Admassu Ayana

Academic Editor

PLOS ONE