Genetic Diversity of Hepatitis B and C Among Patients in Jos, North Central Nigeria.



Bigwan Emmanuel Isa

Department of Medical Laboratory Science Faculty of Medical Sciences University of Jos, Nigeria.


Hepatitis B and C viruses (HBV and HCV) have been well known to be a menace globally causing chronic liver disease (CLD) and cancer. These viral agents have several genotypes and sub-genotypes due to their high replicative activity, which makes them prone to error resulting in high genetic diversity. The genotypes of these viruses show a wide geographical distribution globally. These viruses therefore exhibit high genetic diversity which is characterized by regional differences in genotype prevalence. This poses a great challenge to the development of vaccines and treatment options that require the consideration of global trends in HBV and HCV genotype prevalences. Most studies in the proposed study area-North Central Nigeria, were based on antibody detection for these viral agents. The need for molecular characterization is of great importance to guide treatment decisions, epidemiological studies, drug resistance studies as well as drug and vaccine development. This study aims at determining the genetic diversity of hepatitis B and C virus in the study area and the clinical/socio-demographic factors associated with each of the genotypes that will be determined. This study will be a retrospective study which will involve the use of de-identified samples of both HIV positive and HIV negative patients on which HBV and HCV viral load have been quantified at the AIDS Prevention Initiative in Nigeria (APIN) supported Laboratory at the Jos University Teaching Hospital (JUTH), Jos. Ethical clearance will be obtained from the Ethics Committee of JUTH. The patients’ confirmed HBV and HCV plasma samples stored at - 80 °C will be retrieved along with their socio-demographic and clinical information as well as their CD4+ counts and viral load results. These will be randomly selected; and de- identified by a person who is blinded (that is not part of the research project). Polymerase Chain Reaction (PCR) will be carried out at the APIN Laboratory on all samples using primers that will be synthesized at Inqaba Biotec, Pretoria, South Africa, and the amplicons generated will be detected using Agarose Gel Electrophoresis. Ten amplicon products of each viral agent will be taken to DNA Laboratory, Kaduna, Nigeria for competency training on HBV and HCV sequencing. The PCR products will be purified using DNA purification Kit, quantified and then sequenced using the ABI 3130xl Genetic Analyzer, Applied Biosythesis, USA (BigDye Terminator Version 3.1). The sequences generated will then be determined and phylogenetic analysis will be carried out on all the sequences generated by aligning them with reference strains from the NCBI GenBank using the CLC Work Bench 4.1, to determine the circulating genotypes/ subtypes in the study area. The Clinical/ socio-demographic variables/factors associated with of each viral genotype will be determined by multivariate analysis using Stata software and p values < 0.05 will be considered statistically significant.




OMB No. 0925-0001 and 0925-0002 (Rev. 10/15 Approved Through 10/31/2018)





NAME: Bigwan, Emmanuel Isa

eRA COMMONS USER NAME (credential, e.g., agency login):

POSITION TITLE: Assistant Lecturer



EDUCATION/TRAINING (Begin with baccalaureate or other initial professional education, such as nursing, include postdoctoral training and residency training if applicable. Add/delete rows as necessary.)






(if applicable)

Completio n Date





Ahmadu Bello University, Zaria



Federal College of Vet and Medical Lab.Tech., Vom














University of Jos






Medical Microbiology


Federal College of Vet and Medical Lab.Tech., Vom



Ahmadu Bello University, Zaria












Medical Microbiology



Microbiology (Virology)


  1. Personal Statement

I have the expertise, training that will be essential to carryout most part of the proposed research project. I will require a little training on sequencing analysis from one of the commercial laboratory in Nigeria, DNA Laboratory, Kaduna to add to the already acquired exposure I had on the subject. The technical expertise of my mentors on molecular studies will be an added advantage for me. I have been part of a study on genotyping of HCV in Kaduna State, Nigeria based on primers- specific PCR amplification and also the just concluded study on genotyping of the 5’ UTR region   of HCV in parts of North Central Nigeria, is also an added experience for me to carryout this study successfully. I have also carried out several studies either as a principal investigator or co-investigator in the field of Medical Microbiology.

  1. Positions and honours Positions and Employment

1992-1993       Industrial Training Fund-National Veterinary and Research Institute,

Vom, Plateau State

1994-1995       Teacher, National Youth Service Corps-Christian Commercial College Oro-Ago, Kwara State

1996-1998       Teacher, Mangu Secondary School, Mangu, Plateau State

2001                Medical Laboratory Scientist, Rainbow Clinics and Maternity Home, Garki Village, Abuja

2001-2002       Medical Laboratory Scientist, Arewa Poly Clinics Wuse Zone 4, Abuja 2002-2003                        Medical Laboratory Scientist 1, Winners Medical Diagnostic/Research

Institute, Abuja

2003-2006       Medical Laboratory Scientist 2, Plateau State Hospital Management Board Jos

2005-2009       Medical Laboratory Scientist 2, Jos University Teaching Hospital 2009- Date                        Assistant Lecturer, University of Jos.

Other Experience and Professional Memberships

    1. Association of Medical Laboratory Scientist of Nigeria (AMLSN)
    1. American Society of Microbiology (ASM)
    1. African Society for Laboratory Medicine (ASLM).
  1. Contribution to Science
    1. My earliest research works were in the area of HIV/AIDS in which we carried out a study on the assessment of the impact of HIV/AIDS awareness campaign strategies on HIV sero-incidence in Abuja metropolis and its environs. The study showed that the HIV/AIDS campaign strategies have substantially helped to reduce the HIV sero-incidence in this part of the country, but their contents should be restructured for maximum impact in the target population (those aged 21 – 40 years). I served as the Co-investigator in this study.
      1. Bonnie P.R., Bigwan E.I., David J.S. Njab J.E., Abdullahi J.J. ,Agbonlahor D.E., Nwobu G. (2003).                                Assessment of the Impact of HIV/AIDS Awareness Campaign Strategies on HIV Sero Incidence in Abuja Metropolis and its Environs. Book of Abstracts, National Scientific Conference of the Association of Medical Laboratory Scientists of Nigeria.
    1. We carried out a study on the comparison of two methods for the enumeration of T-Lymphocytes subsets (CD4 and CD8) in whole blood. The study showed that, the manual Dynabeads T4, T8 protocol compared and correlated well with the standard immunoflow-cytometer assay on its CD4 protocol, while CD8 protocol was significantly at variance and poorly correlated with the standard immunoflow-cytometric assay. The Dyenabeads protocol for CD8 protocol should therefore be reviewed, especially if CD4/CD8 ratio must be reliable and dependable. I served as the Co- investigator or co-investigator in all of this study.
      1. Ondobo C.,Njab J.E.,Bigwan E.I.,Bonnie P.R.I.,Okoye McPaul,Abdullahi J.J.,Nwobu G.,Agbonlahor D.E., David J.S.(2003). Comparison of Two Methods of Enumeration of T-Lymphocytes Subsets in Whole Blood. Book of Abstracts, National Scientific Conference of the Association of Medical Laboratory Scientists of Nigeria.
    1. In our study on a comparative study on bacterial growth from fresh and delayed urine samples. The showed that inoculation of samples within two hours of collection yielded the best results and thus should be the best practice in our laboratories and where it is not visible refrigeration should only be reconsidered. Delay in culturing urine samples can lead to issuance of misleading results as such it should be discouraged as much as possible. I served as the Primary investigator or co-investigator in all of this study.
      1. E.I. Bigwan; H.I. Okonkwo; I.S. Udosen; G.C. Markus; Z. Sheyin. A Comparative study on bacterial growth from Fresh and Delayed urine samples. Nigerian Biomedical Science Journal 2012; 8(2):22-25.
    1. We conducted a study on the prevalence of urinary tract infections among HIV patients attending a non-governmental health facility in Jos, Nigeria and the study showed a higher prevalence of bacterial uropathogens among HIV/AIDS patients than for non-HIV/AIDS patients. I served as the Primary investigator or co- investigator in all of this study.
      1. Bigwan E. I. and Wakjissa F. D. Prevalence of Urinary Tract Infections among HIV Patients Attending a Non- Governmental Health Facility in Jos, Plateau State, Nigeria. International Journal of Biomedical and Advance Research 04 (08):528 -533, 2013.
    1. In a study on the Socio-economic dependence on the incidence of Mycobacterium tuberculosis in Jos, the study revealed that pulmonary tuberculosis was recorded only among patients with low socio-economic status. I served as the Primary investigator or co-investigator in all of this study.
      1. Bigwan E. I., Ohaeri M. C., Vem T. S., Sheyin Z., Umar A., Olukose O. J., Wuyep P., Gyang B., Chollom S. C.. Socio-Economic Dependence on the Incidence of Tuberculosis in Jos, North Central Nigeria. Science Journal of Public Health. Vol. 1, No. 5, 2013, pp. 235-238.
    1. Recently, we carried out a study on the genotyping of HCV using the 5’ UTR amongst blood donors and pregnant women in parts of North Central Nigeria, the study revealed the circulating HCV genotypes 1, 2 and 4. The is need to consider genotyping the NS5B region or the full genome sequencing for epidemiological studies that is why I still added this virus in this study. I served as the Primary investigator or co-investigator in all of this study, findings yet to be published.

Complete List of Published Work in My Bibliography:

  1. Damen J.G., Luka, J., Biwan, E.I, Lugos, M. Prevalence of Intestinal Parasites among Pupils in Rural North Eastern Nigeria. Nigerian Medical Journal 2011; 52(1)4-6.
  2. Emmanuel I. Bigwan, Gabriel C. Okezie and Sheyin, Z. Prevalence of Gastrointestinal Parasitic Infection in a Secondary Health Facility in Dadin Kowa Community, Jos Metropolis, Central Nigeria. International Journal of Current Research 2012; 4(07): 092- 095.
  3. E.I. Bigwan; H.I. Okonkwo; I.S. Udosen; G.C. Markus; Z. Sheyin. A Comparative study on bacterial growth from Fresh and Delayed urine samples. Nigerian Biomedical Science Journal 2012; 8(2):22-25.
  4. Bigwan E.I., Tinja B., Damen J.G. Prevalence of Schistosomiasis Among Secondary School Boarding Students in Potiskum Metropolis,Yobe State, North-Eastern Nigeria. Bayero Journal of Pure and Applied Sciences (Bajopas).2012; 5(1):155-158.
  1. Chollom S.C., Agada G.O.A., Bot D.Y., Okolo M.O, Dantong D.D., Choji T. P., Echeonwu B. C., Bigwan E.I., Lokason S And Banwat E.Phytochemical Analysis and Antiviral Potential of Aqueous Leaf Extract of   Psidium   Guajava against Newcastle Disease Virus in Ovo. Journal of Applied Pharmaceutical Science 2 (10); 2012: 045- 049.
  2. Sheyin Z, Jatau E.D, Mamman A.I, Randawa A.J, Bigwan I.E. Detection of Hepatitis C Virus amongst Pregnant Women, in Kaduna State, Nigeria. Wudpecker Journal of Medical Sciences; 2012, 1(2): 012 – 015.
  3. Vem T. S., Pondei J., Nimzing L., Bigwan E., and Abdullahi, A. Prevalence of Hepatitis B Surface Antigen Amongst Blood Donors in Jos, North Central Nigeria

.International Journal of Current Research; 2012: 4(11): 059-062.

  1. Bigwan E.I. and Elijah David. Prevalence of Escherichia Coli among Uropathogens in Asymptomatic Bacteriuria in a Nigerian Tertiary School in Jos, Nigeria. International Journal of Biomedical and Advance Research; 2013, 04 (03):198 -202.
  2. Bigwan E.I., Kunihya R.Z., John T.J. Epidemiological Survey of Urinary Schistosomiasis among Primary School Children in Michika, Adamawa State, North-Eastern Nigeria. International Journal of Current Research and Review. 2013; 5(5): 111-116.
  3. Sheyin Z., Bigwan EI., Galadima M. Comparison of Formol-Ether, Direct Smear and Nigrosine Methylene Blue for the Diagnosis of Human Intestinal Parasites. Journal of Microbiology Research and Reviews; 2013, 1 (3):30-34.
  4. Bigwan E. I. and Wakjissa F. D. Prevalence of Urinary Tract Infections among HIV Patients Attending a Non- Governmental Health Facility in Jos, Plateau State, Nigeria. International Journal of Biomedical and Advance Research; 2013, 04 (08):528 -533.
  5. Sheyin Z., Bigwan IE. Comparism of CARE START HRP2 rapid malaria test with light microscopy for guiding patient’s treatment of fever in Nigerian endemic areas. Journal of Medicine and Medical Sciences; 2013, 4 (9):353-356.
  6. Bigwan Emmanuel Isa, Egah Daniel Zanyu, Badung Bitrus Pam, Danung Monday. Seroepidemiology of Rubella IgG among Unvaccinated Pregnant Women Attending Antenatal Clinics from Two Rural Communities in Plateau State, Nigeria. European Journal of Preventive Medicine. 2013, 1(3), 2013: 58- 62.
  7. Bigwan E. I., Ohaeri M. C., Vem T. S., Sheyin Z., Umar A., Olukose O. J., Wuyep P., Gyang B., Chollom S. C.. Socio-Economic Dependence on the Incidence of Tuberculosis in Jos, North Central Nigeria. Science Journal of Public Health. 2013; 1(5): 235-238.
  8. Bigwan E.I., Ohaeri M.C., Elijah David, Florence D. Wakjissa, Sheyin Z. Some risk factors associated with Acid-Alcohol-Fast Bacilli in Patients with suspected Pulmonary Tuberculosis in Jos, Central Nigeria. African Journal of Infectious Diseases. 2014; 8(2):22-26.
  9. Bigwan E.I., Ohaeri M.C., Okonkwo H.I., Udosen I.S., Markus G.C., Sheyin Z. Prevalence of Acid-Alcohol-Fast Bacilli among Patients with suspected cases of

Pulmonary Tuberculosis in Jos, Nigeria. African Journal of Experimental and Clinical Microbiology, 2014; 15(2): 103-108.

  1. Bigwan E.I., Ado S.A., Umoh V.J., Inabo H.I. Evaluation of two commonly used commercial immunochromatographic and ELISA screening kits for the detection of Anti-HCV antibodies among patients in North Central Nigeria. British Journal of Medicine and Medical Research 4(24) May,
  2. Sheyin, Z., Maimako, J., Shindang, J., Essien, C.U., Bigwan, E.I. and Ede, F.R. Antimicrobial Activity of Albizia lebbeck Leaf Extract on some Medically Important Bacteria. Int. J. Curr. Microbiol. App. Sci. 2015; 4(9): 473-477.
  3. Sheyin Z, Ede FR, Essien UC, Shindang J, Bigwan EI, Elisha P, Bako J, Banda JM. Detection of Measles Virus IgM Antibodies among Individuals Suspected of Measles in Kaduna State, Nigeria. International Journal of Applied Science and Technology Vol. 6, No. 1; February 2016; 87- 91.
  4. E. I. Bigwan, H. I. Inabo, S. A. Ado and E. D. Jatau. Seroprevalence of Hepatitis C Virus amongst Blood Donors in Parts of North Central Nigeria. British Microbiology Research Journal. 2016; 15(3): 1-6.
  5. E. I. Bigwan, E. D. Jatau, H. I. Inabo and S. A. Ado. Evidence of Hepatitis C Virus Antibodies amongst Pregnant Women in Parts of North Central Nigeria. International Journal of TROPICAL DISEASE & Health. 2016; 17(3): 1-6.
  6. Z. Sheyin, A.E. Edache1, U.C. Essien, J.M. Banda, J. Shindang and E. Bigwan. Etiology and Antibiotic Susceptibility Pattern of Community-acquired Urinary Tract Infection in Jos Metropolis. Science World Journal. 2016; 11 (1):22-25.


  1. Cost of procurement of reagents and consumables – ₦1, 279795.58 (See Appendix 1 for detailed list for cost of reagents and consumables
  2. Cost of sequencing 20 amplicons (10 amplicons for each viruses) and bench fee for proficiency training at DNA Laboratory, Kaduna - ₦ 160,000.00
  3. Honorarium for 1 Research Assistant – ₦ 93,642.64
  4. Article Processing Charges for Publication of two articles in International Journals – $500.00 (₦210,000.00).

Budget Justification:

Total cost of procurement of reagents and consumables will take ₦1, 279795.58, based on market prices at the time of the submission of the proposal and this will keep changing depending on the exchange rate in the market. The sequencing of selected amplicons that will be used for the proficiency testing and bench fee will cost ₦160,000. Honorarium for a Research Assistant will take ₦93,642.64 while publications in a high profile journal will cost ₦210,000.00, expenses for the one week proficiency training on sequencing at DNA Laboratory, Kaduna (accommodation, transport fares and feeding) will cost

₦50,000.00 all these are necessary for the work. The total sum for the work is ₦1, 793,438.22.

Budgetted timeline for the project



Time in month (s)













1.Ethical Clearance











2. Sample sorting











3. Reagents procurement











4. PCR amplification at APIN Lab.











5. Sequencing at 2 commercial Labs.











6.Sequencing at APIN Lab.











7.Data analysis






















9. Publication












Project Narrative: This work will provide a baseline genotyping data in the study area for HBV and HCV which will be useful for epidemiological studies. It will also provide useful information in treatment decision and future studies that may come up such as studies on pathogenesis, vaccine or drug development and drug resistance. The prospect on the In-house genotyping of these viral infections will help build local capacity for future research endeavours and will likely reduce cost of genotyping for patients in the area.

Research Plan:

Specific aims

  1. To characterize the circulating genotypes of HBV and HCV among HIV positive and HIV negative persons in the study area, Jos North Central Nigeria through sequencing and phylogenetic analysis.
  2. To determine the clinical/ socio-demographic correlates of the genotypes that will be determined.

Background and significance

Hepatitis B and C viruses (HBV and HCV) are the major causes of liver diseases globally. The relative importance of HBV and HCV infections differ greatly from one part of the world to another and changes over time 1. Worldwide, over two billion people are infected with HBV alone, of whom about one million die annually. Hepatitis C virus affects about 200 million people 2.

It has been reported that HBV/HIV co infection leads to increased morbidity and mortality as compared to HIV or HBV mono-infections3. With increased access to antiretroviral drugs for HIV patients, migrating populations and social networking by intravenous drug use, cases of HBV and HCV co infections have been on the rise 4.

Studies have confirmed that HIV co-infection can accelerate the clinical course of disease progression of chronic HCV or HBV infection and increase the risks of liver cirrhosis, hepatocellular carcinoma (HCC) and decompensated liver disease 5, 6, 7.

Worldwide, hepatitis C and B virus infections (HCV and HCV), are the two most common coinfections with human immunodeficiency virus and has become a major threat to the survival of persons infected with HIV8. Annually, approximately two million people die due to AIDS, more than 350 thousands people die from diseases associated with HCV and one million people die as a result of an HBV infection 9, 10, 11. Studies have shown that the prevalence of HBV, HCV, and HIV coinfection differs both by geographic region and behaviour of infected people 12,13,14,15.

The high error rate of HBV RT causes frequent nucleotide substitutions during viral replication, resulting in genetic diversity in the form of genotypes, sub-genotypes, quasispecies and a large number of mutations in different regions of the HBV genome 16. The high genetic diversity of HCV is due to high replicative activity and lack of proof- reading activity of RNA-dependent RNA polymerase 17.

Globally, HCV genotype 1 is the most prevalent with a prevalence of 46%; followed by genotype 3 with 30.1%; genotypes 2, 4, and 6 are responsible for a total of 22.8%; genotype 5 comprises the remaining <1%. While genotypes 1 and 3 dominate in most countries irrespective of economic status, the largest proportions of genotypes 4 and 5 are in lower-income countries 18.

The genotypes for both HBV and HCV display distinct geographical distribution which carries important implications such as treatment decisions, possible transmission route and vaccine development 19, 20, 21.

There is scarce information on HBV and HCV genotypes in Nigeria, amongst the few include the prevalence of HBV genotype E 22, HBV genotype E and subgenotype A3 23.

An earlier study on HCV genotypes in Lagos, Nigeria reported a prevalence of genotypes 1 and 4 24, a more recent study also in Lagos reported HCV genotypes 1, 3, 2, 4 and 6

with the following prevalence: 64.7%, 7.4%, 5.9%, 4.4%, and 2.9% respectively 25 while in a study in two remote communities in Nasarawa State, Nigeria reported HCV genotypes 1 and 2 with the prevalence of 85% and 15% respectively 26.

This study is aimed at answering these questions: what are the HBV and HCV genotypes in the area in relation to HIV/AIDS status? Is there diversity in the circulating genotypes? Are there cases of HIV/HBV/HCV coinfections? The Null hypothesis: There is no genetic diversity in HBV and HCV genotypes among registered patients at JUTH/APIN HIV clinic, while the alternate hypothesis is: there is genetic diversity in HBV and HCV genotypes among registered patients at JUTH/APIN HIV clinic.


Study design

This study will be a retrospective study using de-identified stored plasma samples of patients both HIV positive and HIV negative patients that have had HBV and HCV viral load assays done at APIN Laboratory, JUTH, Jos. The patients are registered for care at JUTH and APIN supported HIV clinic JUTH, Jos, Nigeria.

Some data will be extracted from patients’ case notes (hospital records) and other information not available in the case notes will be obtained through a telephone interview. The data to be extracted include age, sex, HIV status, CD4+ counts, viral loads, duration of infection, HIV/HBV/HCVcoinfection, educational status, history of antiretroviral therapy and liver function test/ biopsies.

Study subjects:

The study subject will be adults (males and females) aged 18- 60 years. Stored Plasma samples at -80oC of confirmed HBV and HCV positive patients with their socio- demographic and clinical information as well as their CD4+ counts and viral loads will be randomly selected; and de-identified by a person who is blinded (that is not part of the research project). Other clinical information that will be collected with includes patient’s HIV status, presence and classification of liver disease/ pathology and where present, liver function test results.

Exclusion criteria: samples with inadequate volume for the assays < 700μl, and those below the age of 18 and above 60 years.

Sample size

The sample size was calculated using OpenEpi 27 where p = x%, was the prevalence of 11% from a previous Nigerian study 2310. The calculated sample size is 151 for HBV and from available data on prevalences of HBV and HCV samples will be recruited in the ratio 3:1. One hundred (100) samples will be extracted for HBV while 50 for HCV.

Laboratory procedures

Nucleic acid extraction and molecular detection


Viral HBV DNA extraction will be carried out using QIAamp DNA Mini Kit (Qiagen, Germany) from which 200 μl plasma will be used according to the manufacturer’s instructions to obtain the HBV DNA.The precipitated DNA will be   resuspended in 200 μl of elution buffer and stored at -20°C until ready for use for PCR.


HBV genotype/subgenotype assignment will be based on phylogenetic analysis of the partial S-gene sequence (406 bp) as described previously but without the SP6 and T7 promoter sequences 23. The S-gene is considered suitable for genotype classification 28, 29. The samples will be amplified with primers previously described 30. The Dream Taq Green PCR Master Mix (Thermo Fisher) will be used for the Nested PCR amplification.


Nucleic acid (RNA) will be extracted from 200 μl of plasma by using a PureLink Viral RNA/ Kit (Invitrogen, Carlsbad, CA) according to the manufacturer’s instructions. The RNA will then be reverse-transcribed using the Superscript III First-Strand cDNA Synthesis Supermix Kit protocol using random hexamers (Invitrogen). Molecular detection will be performed by Nested PCR on the 5’ UTR and NS5B regions of the HCV genome. The amplifications of the two region will be carried out using primers previously described 31. Appropriate amplification controls will be included in the amplification to exclude the false negative results. PCR products will be detected using electrophoresis in 2% agarose gels, stained with Gel Red and visualized under ultraviolet illumination.

Determination of nucleotide sequences and phylogenetic analysis

PCR products will then be purified and quantified. Direct sequencing will be performed with 1.0 μM of the second-round primers using the automated DNA Genetic Sequencer ABI 3130xl (Applied Biosystems, Foster City, CA). Electrophoresis and data collection will be done with the instrument. The sequence chromatograph files that will be generated will be edited and analysed using CLC Main Workbench The edited sequences will be submitted to the Blast similarity software to check their similarity to related strains deposited in the GenBank. The reference strains with the highest similarity will be retrieved and aligned with the sequences that will be obtained in this study. Phylogenetic tress will be constructed on the edited sequences and the HBV/HCV prototypes that will be retrieved from the GenBank.

Statistical analyses

The independent variables include Socio-demographic and clinical variables (age, sex, marital status, education level, occupation, alcohol intake. Tobacco smoking, mode of HIV transmission, WHO clinical stage, TB co-infection, weight, height and BMI and laboratory variables (CD4+ and viral load). The outcome variable is each viral genotype identified. The initial univariate analysis will look at the association between each independent variable and a viral genotype, using Chi squared test or Fisher’s exact test for categorical variables and the Wilcoxon signed rank test for continuous variables. Then, those variables found to be associated with a viral genotype in the univariate analysis at p<0.05 will be included in a multivariate regression modelling, to determine the factors associated with the genetic diversity. All analysis will be carried out using Stata software and p values < 0.05 will be considered statistically significant.

Future Directions:

This baseline study will provide useful data and impetus for future studies which may include comparative studies on partial and full genomic sequencing of the two viruses, epidemiological, drug resistance, pathogenesis, vaccine or drug development studies.

Plan for protection of Human Subjects:

The study will not involve direct contact with human subjects, it is their stored samples that will be used after they are de-identified by an assigned individual from the laboratory that will not be part of this research. Strict compliance on patient’s confidentiality will be observed.

Pending IRB Approval:

The proposal for the study will be presented to the Ethics Committee at the Jos University Teaching Hospital Jos, for review and approval before the commencement of the study.


  1. Te HS, Jensen DM: Epidemiology of hepatitis B and C viruses: a global overview. Clin Liver Dis 2010; 14(1):1–21.
  2. Parry J: At last a global response to viral hepatitis. Bull World Health Organ

2010; 88:801–802.

  1. Thio C: Hepatitis B and human immunodeficiency virus coinfection. Hepatology. 2009; 49:138–145
  2. Lacombe K, Boyd A, Gozlan J, Lavocat F, Girard PM, Zoulim F: Drug resistance and immune escape HBV mutants in HIV-infected hosts. Antivir Ther. 2010; 15:493–497.
  1. Thio CL, Seaberg EC, Skolasky R Jr, Phair J, Visscher B, et al. HIV-1, hepatitis B virus, and risk of liver-related mortality in the Multicenter Cohort Study (MACS). Lancet. 2002; 360: 1921–1926.
  2. Soriano V, Vispo E, Labarga P, Medrano J, Barreiro P. Viral hepatitis and HIV co-infection. Antiviral Res. 2005; 85: 303–315.
  3. Chen SL    and Morgan TR. The Natural History of Hepatitis C Virus (HCV) Infection. International Journal of Medical Sciences.2006; 3(2):47-52.
  4. Amiri FB, Mostafavi E, Mirzazadeh A. HIV, HBV and HCV Coinfection Prevalence in Iran - A Systematic Review and Meta-Analysis. PLOS ONE | DOI:10.1371/journal.pone.0151946 March 31, 2016 pp1-12.
  5. Lu F, Zhuang H. Management of hepatitis B in China. Chin Med J (Engl) .2009; 122: 3–4.
  6. Averhoff FM, Glass N, Holtzman D (2012) Global burden of hepatitis C: considerations for healthcare providers in the United States. Clinical Infectious Diseases. 2012; 55: S10–S15.
  7. Fauci AS, Folkers GK. Toward an AIDS-free generation. JAMA.2012; 308: 343– 344.
  8. Gupta S. and Singh S. “Hepatitis B and C virus co-infections in human immunodeficiency virus positive North Indian patients,” World Journal of Gastroenterology.2006;12(42): 6879–6883.
  9. Nelson P, Mathers B, Cowie B. et al., “The epidemiology of viral hepatitis among people who inject drugs: results of global systematic reviews,” The Lancet. 2011; 378( 9791): 571–583
  10. Tremeau-Bravard A, Ogbukagu IC, Ticao CJ, Abubakar JJ. “Seroprevalence of hepatitis B and C infection among the HIV-positive population in Abuja, Nigeria,” African Health Sciences. 2012; 12(3):312–317, 2012.
  11. WHO. Consolidated Guidelines on the Use of Antiretroviral Drugs for Treating and Preventing HIV Infection: Recommendations for a Public Health Approach, 2013.
  12. Zhang Z, Wu C, Chen X,   Li X, Lu J. Genetic variation of hepatitis B virus and its significance for pathogenesis. World J Gastroenterol 2016; 22(1): 126-144.
  13. Smith DB, Bukh J, Kuiken C, et al. Expanded classification of hepatitis C virus into 7 genotypes and 67 subtypes: Updated criteria and genotype assignment web resource. Hepatology 2014; 59: 318-27.
  14. Messina JP, Humphreys I, Flaxman A, Brown A, Cooke GS, Pybus OG, Barnes

E. Global distribution and prevalence of hepatitis C virus genotypes. Hepatology. 2015; 61(1):77-87.

  1. McMahon BJ. The influence of hepatitis B virus genotype and subgenotype on the natural history of chronic hepatitis B. Hepatol Int 2009; 3:334–342.
  1. Mahfoud Z, Kassak K, Kreidieh K, Shamra S, Ramia S. (2010). Distribution of hepatitis C virus genotypes among injecting drug users in Lebanon. Virology Journal. 2010; 7:96.
  2. Pourkarim MR, Amini-Bavil-Olyaee S, Verbeeck J, Lemey P, Zeller M, Rahman M, Maes P, Nevens F, Van Ranst M. 2010. Molecular evolutionary analysis and mutational pattern of full-length genomes of hepatitis B virus isolated from Belgian patients with different clinical manifestations. J Med Virol 82:379–389.
  3. Odemuyiwa SO, Mulders MN, Oyedele OI, Ola SO, Odaibo GN, Olaleye DO, et al. Phylogenetic analysis of new hepatitis B virus isolates from Nigeria supports endemicity of genotype E in West Africa. J Med Virol. 2001; 65:463–9.
  4. Forbi JC, Vaughan G, Purdy MA, Campo DS, Xia GL, Ganova-Raeva LM, Ramachandran S, Thai H, Khudyakov YE . Epidemic History and Evolutionary Dynamics of Hepatitis B Virus Infection in Two Remote Communities in Rural Nigeria. PLoS One. 2010, 19; 5(7):e11615.
  5. Oni AO and Harrison TJ. Genotypes of hepatitis C virus in Nigeria. J Med Virol

1996; 49(3): 178-186.

  1. Okwuraiwe, AP Salu, AP Anomneze, E Audu, RA Ujah, IAO. Hepatitis C virus genotypes and viral ribonucleic acid titers in Nigeria. Nigerian Journal of Gastroenterology and Hepatology, 2012; 4(2).
  2. Forbi, JC, Purdy, MA, Campo, DS, Vaughan, G, Dimitrova, ZE, Ganova-Raeva, LM, Xia, G and Khudyakov, YE. Epidemic history of hepatitis C virus infection in two remote communities in Nigeria, West Africa. Journal of General Virology, 2012, 93: 1410–1421.
  3. OpenEpi[ /sspropor.htm]. Accessed on 05 October, 2016.
  4. Mahtab MA, Rahman S, Khan M, Karim F. Hepatitis B virus genotypes: an overview. Hepatobil Pan Dis Int: HBPD INT. 2008; 7:457–64.
  5. Ganova-Raeva L, Ramachandran S, Honisch C, Forbi JC, Zhai X, Khudyakov Y. Robust hepatitis B virus genotyping by mass spectrometry. J Clin Microbiol. 2010; 48:4161–8.
  6. Forbi JC, Ben-Ayed Y, Xia GL, Vaughan G, Drobeniuc J, Switzer WM, and Khudyakov YE. Disparate distribution of hepatitis B virus genotypes in four sub- Saharan African countries. J Clin Virol. 2013; 58(1): 59–66.
  1. Cantaloube JF, Gallian P, Attoui H, Biagini P, De Micco P, de Lamballerie X. Genotype distribution and molecular epidemiology of hepatitis C virus in blood donors from southeast France. J Clin Microbiol. 2005; 43: 3624-9.

Project Mentors:

  1. Prof. Godwin E. Imade

Research Professor /Laboratory Director

Department of Obstetrics and Gynaecology/APIN Centre Faculty of Medical Science, University of Jos, Nigeria.


  1. Dr. Edith N. Okeke

Professor of Medicine/ Consultant Gastroenterologist Department of Medicine

University of Jos/ Jos University Teaching Hospital, Jos, Nigeria.


  1. Dr. Jason Blackard

Director, Office of Global Health/ Associate Professor Department of Digestive Diseases

College of Medicine

University of Cincinnati, Ohio, USA.


Appendix 1: Cost of reagents and consumables Cost of Reagents, Consumables for Sequencing



COST (₦)


ZN Viral RNA extraction kit (100 Prep.)



DNA Extraction kit (200 Prep.)



cDNA synthesis kit (150 Prep.)



Master mix 500 Rxns



100bp DNA Ladder-125 gel lines



1 Agarose



DNA Clean-up kit (100 Prep.)



PCR Tubes (1000 Rxns)



Pipettes tips (2- 200ul)



Pipettes tips (50- 1000ul)



TAE, 25X Liquid concentrate






Cold Chain packaging






Sequencing 20 Amplicons and Bench fee at DNA Lab. Kaduna