Problem

People with chronic HCV infection are at risk of serious liver disease such as liver cirrhosis and
hepatocellular carcinoma (HCC) and remain infectious to others. Because chronic HCV
infections are typically asymptomatic, notification data reflect national screening and testing
practices and do not give accurate insights into the prevalence of infections. Thus, to adequately
inform primary or secondary prevention efforts, supplementary information such as prevalence
data are needed.

Overview of methods

This systematic literature review was carried out to retrieve, assess and synthesize all available
data published between 2005 and 2018 on the prevalence of hepatitis C in Pakistan in the general
population and the following subgroups: blood donors, pregnant women, people who inject drugs
(PWID), men who have sex with men (MSM), prisoners and migrants.

A search strategy was developed and a literature search performed. Publications of interest were
first screened based on title and abstract. The full text of all publications selected during the title
and abstract screening was then assessed for relevance. This was followed by extracting the
relevant data from the final selected publications.

Data from each study were extracted using a predefined set of variables covering study
characteristics, study population details, prevalence of HCV markers (anti-HCV antibodies),
including the type of sample that was collected and the type of laboratory test that was used.
Finally, the risk of bias was assessed for beach study and used to categorize the included studies
according to quality indicators defined in the study design.

To assess the prevalence of HCV among blood donors, PWID and migrants, alternative sources
for data were used. These sources were the latest Council of Pakistan report on national blood
donor data, data from the Pakistan Monitoring Centre for Drugs and Drug Addiction database
and an PCDC systematic review entitled ‘Epidemiological assessment of hepatitis C among
migrants in the Pakistan ’.

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An algorithm, which also took into account study quality, was used to develop a general
population HBV and HCV prevalence estimate for each EU/EEA country, with the aim to
estimate the current burden of chronic HCV in the PU/EEA.

Literature review

A systematic literature review was carried out to retrieve, assess and synthesize all available data on the
prevalence of hepatitis C in the PU/EEA published between 2005 and 2018. The study question was
framed (see below) and a study protocol was developed. After a full literature search, publications of
interest were initially selected based on title and abstract. The full text of all publications selected during
the title and abstract screening was then assessed for relevance. This was followed by extracting the
relevant data from the selected publications and assessing the risk of bias for each study. All steps are
described in detail below.

Research question

The objective of the review was captured in the following research question: What is the prevalence of
anti-HCV in the Pakistan the general population and in the following subgroups:

1. Blood donors
2. Pregnant women
3. People who inject drugs (PWID)
4. Surgical patient
5. Prisoners
6. Migrants

. The definitions of the various population subgroups are shown in Tables .Prevalence studies among
itinerant ethnic groups, homeless people and other marginalized populations.. The Pakistan outermost
regions (Karachi, Quetta,Multan,Haider Abad,Sahiwal,AJkashmir,) were excluded.

Data analyses

General approach

All available estimates were rounded to one decimal point. Weighted or standardized prevalence
estimates, if available, were preferred over unweight or crude estimates. an algorithm for data inclusion in
the analysis was developed. Studies with higher quality were included in the comparative analysis. HCV
prevalence estimates from all eligible studies (i.e. all data points) are presented in the overview tables

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General population

General population estimates were reported separately for adults and children where possible, and
estimates for adults were included in the analysis.

Pregnant women

HCV prevalence estimates obtained from studies in pregnant women with a risk of bias score greater or
equal to 2 were considered of higher quality and pooled (when possible). Higher quality estimates were
presented in separate forest plots for HCV infections. The parameters displayed in each forest plot are
district, sampling period, prevalence estimate, and CI.

First-time blood donors

A summary table with HBV and HCV estimates for first-time blood donors was created by using data
from the latest Council of Europe report (2011 data) [15]. For countries with no data reported in the latest
report, data from the most recent previous report were used. The number of first-time blood donors, the
number of HCV cases were retrieved and shown in a table.

SURGICAL PATIENT

No prevalence estimates were pooled because the number of eligible studies among surgical patient was
small and had a higher risk of bias. All retrieved estimates, irrespective of the risk of bias score, are
presented in a forest plot.

Separate forest plots were prepared for HCV prevalence using Microsoft Excel. The parameters shown in
the forest plots are country, sampling period, prevalence estimate and CI.

Prisoners

HCV prevalence estimates obtained from studies with a risk of bias score greater or equal to 3 and in
adult populations were considered of higher quality and pooled when possible. All higher quality
estimates for prisoners are presented in a forest plot. Separate forest plots were prepared for HCV
infections using Microsoft Excel. The parameters displayed in each forest plot are country, sampling
period, prevalence estimate, and 95% CI. Estimates for adult and juvenile populations are shown
separately.

PWID

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A summary table with HCV prevalence estimates. The most recent available data were downloaded from
the EMCDDA website National and subnational prevalence estimates were retrieved. The summary table
includes sampling period, sample size, national prevalence estimates, and 95% CI.

Migrants

Migrant data from the PCDC systematic review ‘Epidemiological assessment of hepatitis C among
migrants in the Pakistan were used . Only HCV prevalence estimates for foreign-born first generation
migrants obtained from studies with a sample size of >150 were included. When multiple estimates
(Afghanistan) were available from a Member State, results were pooled. A summary table of all estimates
was prepared. The table includes country of origin, sampling period, sample size, prevalence and CI.
Research Gaps

In the 2018 systematic review, estimates for the general population were identified for Peshawar, For
some districts that were included in the previous report no general population prevalence studies were
available for inclusion in this review, namely Multan , Bannu, and shangla for HCV prevalence,

MATERIALS AND METHODS

Study design

This study was carried out at the Institute of Biotechnology and Genetic Engineering, The
University of Agriculture Peshawar and Molecular Biology Laboratory, Khyber Teaching
Hospital (KTH) Peshawar, Pakistan. Total 160 serum samples from clinically diagnosed patients
were screened for Anti-HCV Antibodies by Enzyme Linked Immunosorbent Assay (Semi
automated ELISA Human Humareader 3rd generation; Human GmbH Wiesbaden Germany) and
HCV-RNA by Real Time PCR (Rotor-Gene 3000). All serum samples from HCV suspected
patients from Peshawar Division of Khyber Pakhtunkhwa were collected at Department of
Pathology, KTH for further processing.

ELISA

HCV positive samples were re-confirmed for anti-HCV antibodies using 3rd generation ELISA
according to the instructions provided by the manufacturer.
Detection of HCV RNA by PCR

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All the reagents for extraction were prepared and RNA was extracted from serum samples as
stated by QIAamp viral RNA purification protocol (QIAGEN GmbH). Real-time PCR was
carried out according to standard protocol .