Publications by authors named "Yasir Arshad"

11 Publications

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Rapid and Sensitive Direct Detection and Identification of Poliovirus from Stool and Environmental Surveillance Samples by Use of Nanopore Sequencing.

J Clin Microbiol 2020 Aug 24;58(9). Epub 2020 Aug 24.

Department of Infectious Disease Epidemiology, Imperial College London, London, United Kingdom.

Global poliovirus surveillance involves virus isolation from stool and environmental samples, intratypic differential (ITD) by PCR, and sequencing of the VP1 region to distinguish vaccine (Sabin), vaccine-derived, and wild-type polioviruses and to ensure an appropriate response. This cell culture algorithm takes 2 to 3 weeks on average between sample receipt and sequencing. Direct detection of viral RNA using PCR allows faster detection but has traditionally faced challenges related to poor sensitivity and difficulties in sequencing common samples containing poliovirus and enterovirus mixtures. We present a nested PCR and nanopore sequencing protocol that allows rapid (<3 days) and sensitive direct detection and sequencing of polioviruses in stool and environmental samples. We developed barcoded primers and a real-time analysis platform that generate accurate VP1 consensus sequences from multiplexed samples. The sensitivity and specificity of our protocol compared with those of cell culture were 90.9% (95% confidence interval, 75.7% to 98.1%) and 99.2% (95.5% to 100.0%) for wild-type 1 poliovirus, 92.5% (79.6% to 98.4%) and 98.7% (95.4% to 99.8%) for vaccine and vaccine-derived serotype 2 poliovirus, and 88.3% (81.2% to 93.5%) and 93.2% (88.6% to 96.3%) for Sabin 1 and 3 poliovirus alone or in mixtures when tested on 155 stool samples in Pakistan. Variant analysis of sequencing reads also allowed the identification of polioviruses and enteroviruses in artificial mixtures and was able to distinguish complex mixtures of polioviruses in environmental samples. The median identity of consensus nanopore sequences with Sanger or Illumina sequences from the same samples was >99.9%. This novel method shows promise as a faster and safer alternative to cell culture for the detection and real-time sequencing of polioviruses in stool and environmental samples.
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http://dx.doi.org/10.1128/JCM.00920-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7448630PMC
August 2020

Emergence of Chikungunya Virus, Pakistan, 2016-2017.

Emerg Infect Dis 2020 02;26(2):307-310

During December 2016-May 2017, an outbreak of chikungunya virus infection occurred across Pakistan. The East/Central/South African genotype was predominant. This study provides baseline data on the virus strain and emphasizes the need for active surveillance and implementation of preventive interventions to contain future outbreaks.
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http://dx.doi.org/10.3201/eid2602.171636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6986857PMC
February 2020

Genetic Epidemiology Reveals 3 Chronic Reservoir Areas With Recurrent Population Mobility Challenging Poliovirus Eradication in Pakistan.

Clin Infect Dis 2020 Oct;71(7):e58-e67

Department of Virology, National Institute of Health, Chak Shahzad, Islamabad, Pakistan.

Background: Pakistan is among 3 countries endemic for wild poliovirus type 1 (WPV1) circulation that are still struggling for eradication of poliomyelitis. Active clinical and environmental surveillance with meticulous laboratory investigations provide insights into poliovirus transmission patterns and genomic diversity to inform decisions for strategic operations required to achieve eradication.

Methods: We analyzed epidemiological and virological data to comprehend the current epidemiological status of WPV1 in Pakistan during 2015-2017. Stool specimens of patients with acute flaccid paralysis (AFP) and sewage samples collected from 60 environmental sites were tested. Viral culturing, intratypic differentiation by real-time polymerase chain reaction, and nucleic acid sequencing of the VP1 region of the poliovirus genome to determine genetic relatedness among WPV1 strains were applied.

Results: Poliovirus isolates were grouped into 11 distinct clusters, which had ≥95% nucleotide homology in the VP1 coding region. Most of the poliovirus burden was shared by 3 major reservoirs: Karachi, Peshawar, and Quetta block (64.2% in 2015, 75.4% in 2016, and 76.7% in 2017).

Conclusions: Environmental surveillance reveals importations and pockets of unimmunized children that dictate intensive target mop-up campaigns to contain poliovirus transmission. A decrease in the number of orphan isolates reflects effective combination of AFP and environmental surveillance in Pakistan. The genetic data reflect sustained transmission within reservoir areas, further expanded by periodic importations to areas of high immunity reflected by immediate termination of imported viruses. Improved immunization coverage with high-quality surveillance is vital for global certification of polio eradication.
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http://dx.doi.org/10.1093/cid/ciz1037DOI Listing
October 2020

High prevalence of G3 rotavirus in hospitalized children in Rawalpindi, Pakistan during 2014.

PLoS One 2018 30;13(4):e0195947. Epub 2018 Apr 30.

Department of Virology, National Institute of Health, Chak Shahzad, Islamabad, Pakistan.

Rotavirus A species (RVA) is the leading cause of severe diarrhea among children in both developed and developing countries. Among different RVA G types, humans are most commonly infected with G1, G2, G3, G4 and G9. During 2003-2004, G3 rotavirus termed as "new variant G3" emerged in Japan that later disseminated to multiple countries across the world. Although G3 rotaviruses are now commonly detected globally, they have been rarely reported from Pakistan. We investigated the genetic diversity of G3 strains responsible RVA gastroenteritis in children hospitalized in Rawalpindi, Pakistan during 2014. G3P[8] (18.3%; n = 24) was detected as the most common genotype causing majority of infections in children less than 06 months. Phylogenetic analysis of Pakistani G3 strains showed high amino acid similarity to "new variant G3" and G3 strains reported from China, Russia, USA, Japan, Belgium and Hungary during 2007-2012. Pakistani G3 strains belonged to lineage 3 within sub-lineage 3d, containing an extra N-linked glycosylation site compared to the G3 strain of RotaTeqTM. To our knowledge, this is the first report on the molecular epidemiology of G3 rotavirus strains from Pakistan and calls for immediate response measures to introduce RV vaccine in the routine immunization program of the country on priority.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0195947PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5927433PMC
July 2018

Epidemiological and molecular investigation of a measles outbreak in Punjab, Pakistan, 2013-2015.

J Med Virol 2018 08 25;90(8):1297-1303. Epub 2018 May 25.

Department of Virology, National Institute of Health, Chak Shahzad, Pakistan.

Despite the availability of an effective vaccine, the measles virus continues to cause significant morbidity and mortality in children worldwide. Molecular characterization of wild-type measles strains is an invaluable component of epidemiological studies or surveillance systems that provides important information pertinent to outbreak linkages and transmission pathways. Serum samples and throat swabs were collected from suspected measles cases from the Punjab province of Pakistan (2013-2015) and further tested for measles immunoglobulin M (IgM) through enzyme-linked immunosorbent assay and reverse-transcriptase polymerase chain reaction for molecular characterization. Among the total of 5415 blood samples, 59% tested positive for measles IgM. Males had a higher infection rate (55%) than females (45%), and the highest frequency of positive cases (63%) was found in the age group of 0 to 5 years. Partial sequencing of the nucleoprotein gene showed that 27 strains belonged to the B3 genotype, whereas 2 viruses were identified as D4. On phylogenetic analysis, Pakistani B3 strains were found to be closely related to previously reported indigenous strains and those from neighboring countries of Iran and Qatar. This is the first report on the detection of the measles B3 genotype from Punjab, Pakistan. The current study shows a high burden of measles infections in Punjab province owing to poor routine immunization coverage in major cities. It is imperative that national health authorities adopt strategic steps on an urgent basis for improvement of routine immunization coverage. Molecular epidemiology of the measles viruses circulating in different parts of the country can provide useful data to manage future outbreaks.
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http://dx.doi.org/10.1002/jmv.25206DOI Listing
August 2018

Outbreak of dengue virus type-3 in Malakand, Pakistan 2015; A laboratory perspective.

Acta Trop 2017 May 17;169:202-206. Epub 2017 Feb 17.

Department of Virology, National Institute of Health,Park Road, Chak Shahzad, Islamabad, Pakistan. Electronic address:

An outbreak of dengue fever was reported in Malakand district, Khyber Pakhtunkhwa (KP) province of Pakistan during 2015. Detection of viral RNA by real-time PCR confirmed dengue virus serotype-3 (DENV-3) to be the causative agent causing the outbreak. Phylogenetic analysis based on partial E-NS1 gene sequences showed that the DENV-3 viruses belonged to genotype III with maximum homology with the dengue-3 strains previously reported from Pakistan and India. Our current report provides updated information on molecular epidemiology and phylogenetic analysis of dengue virus serotypes responsible for 2015 outbreak in KP.
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http://dx.doi.org/10.1016/j.actatropica.2017.02.011DOI Listing
May 2017

Dengue Virus Serotypes Circulating in Khyber Pakhtunkhwa Province, Pakistan, 2013-2015.

Ann Lab Med 2017 Mar;37(2):151-154

Department of Virology, National Institute of Health, Park Road, Chak Shahzad, Islamabad, Pakistan.

From 2013 to 2015, the National Institute of Health, Pakistan, received 1,270 blood samples of suspected dengue cases reported from inpatient and outpatient departments of various hospitals in Khyber Pakhtunkhwa (KPK) province. In this study, we determined the circulating dengue virus (DENV) serotypes using real-time reverse transcriptase (RT)-PCR to understand the serotype-based epidemiology of DENV. All four serotypes (DENV-1 [6%], DENV-2 [33%], DENV-3 [47%], and DENV-4 [0.1%]) were found circulating during the study period. Our findings suggest the need for an active surveillance system coupled with the laboratory diagnosis, especially in the chronic endemic areas of the country. Public awareness programs are needed for effective control and prevention of outbreaks in the future.
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http://dx.doi.org/10.3343/alm.2017.37.2.151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5203993PMC
March 2017

NS1 antigen: A new beam of light in the early diagnosis of dengue infection.

Asian Pac J Trop Med 2016 12 9;9(12):1212-1214. Epub 2016 Nov 9.

Department of Virology, National Institute of Health, Park Road, Chak Shahzad, Islamabad, Pakistan. Electronic address:

Objective: To evaluate NS1 antigen detection ELISA for the early laboratory diagnosis of dengue virus infection.

Methods: The present study was conducted to evaluate the overall positivity of NS1 antigen detection ELISA and its comparison with viral RNA detection via real time PCR and IgM antibodies detection by ELISA.

Results: A total of 1270 serum samples were tested 86% (1097/1270) were detected positive by one or more than one diagnostic test. Out of 1 270, 64% (807/1270) were positive by NS1 ELISA and 52% (662/1270), 51% (646/1270) were positive by real-time RT-PCR and IgM ELISA respectively.

Conclusions: NS1 antigen detection ELISA is highly suitable diagnostic tools and it also has great value for use in outbreak and epidemic situation.
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http://dx.doi.org/10.1016/j.apjtm.2016.10.007DOI Listing
December 2016

Respiratory syncytial virus and influenza are the key viral pathogens in children <2 years hospitalized with bronchiolitis and pneumonia in Islamabad Pakistan.

Arch Virol 2017 Mar 24;162(3):763-773. Epub 2016 Nov 24.

Department of Virology, National Institute of Health, Chak Shahzad, Park Road, Islamabad, 44000, Pakistan.

Pneumonia remains a leading cause of morbidity and mortality in developing countries. Comprehensive surveillance data are needed to review the prevention and control strategies. We conducted active surveillance of acute lower respiratory infections among children aged <2 years hospitalized at two hospitals of Islamabad, Pakistan. Viral etiology was determined using real-time PCR on respiratory specimens collected during March 2011-April 2012. The overall mean age was 7.83 ± 5.25 months while no statistical difference between age or sex distribution of patients with positive and negative viral etiology (p > 0.05). The average weight of the study group was 6.1 ± 2.25 kg. ≥1 viral pathogens were detected in 75% cases. Major respiratory viruses included RSV-A: 44%, RSV-B: 23%, Influenza-A: 24.5%, Influenza-B: 7%, Adenovirus: 8.4% and HmPV: 5.2%. A single, dual or multiple viral pathogens were detected in 43%, 27% and 5.2% patients respectively. Common symptoms were cough (95%), apnoea (84%), fever (78%), wheeze (64.5%), nasal congestion (55%) and rhinorrhea (48%). Among the RSV positive cases, 2-6 months age group had highest detection rate for RSV-A (30%, n = 21/69) and RSV-B (20%, n = 14/69) while patients infected with Influenza-A were in 2.1-6 months age group (61%, 23/38). Statistically significant difference was observed between RSV-positive and negative cases for nutrition status (p = 0.001), cigarette/wood smoke exposure (p = 0.001) and concomitant clinical findings. Most patients had successful outcome on combination therapy with bronchodilators, inhaled steroids and antibiotics. Our findings underscore high burden of ALRI in Pakistan. Interventions targeting viral pathogens coupled with improved diagnostic approaches are critical for better prevention and control.
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http://dx.doi.org/10.1007/s00705-016-3146-7DOI Listing
March 2017

Dengue outbreak in Swat and Mansehra, Pakistan 2013: An epidemiological and diagnostic perspective.

Asian Pac J Trop Med 2016 Apr 9;9(4):380-384. Epub 2016 Mar 9.

Department of Virology, National Institute of Health, Park Road, Chak Shahzad, Islamabad, Pakistan. Electronic address:

Objective: To high light some epidemiological, clinical and diagnostic features of dengue fever during an outbreak and the role of different diagnostic techniques to achieve the highest level of accuracy in results.

Methods: Blood samples (n = 323) were collected along with epidemiological and clinical data from suspected dengue patients who visited different hospitals in Swat and Mansehra district of Pakistan between May-November 2013 during a dengue outbreak. Samples were tested for the detection of viral nucleic acid by real-time PCR, non structural protein-1 (NS1) antigen and IgM antibodies by ELISA.

Results: Out of 323 cases with clinical dengue infection, 304 were positive by one or more diagnostic parameter; 201 samples were positive by real-time PCR, 209 were positive by NS1 ELISA and 190 were positive by IgM antibodies. Sensitivities of real-time PCR and NS1 ELISA were comparable for early diagnosis of dengue virus infection, IgM antibody detection assay was found useful for the diagnosis in the samples collected later than day 5 of onset.

Conclusions: The use of real-time PCR or detection of non structural protein NS1 by ELISA followed by IgM antibodies detection can be recommended for early diagnosis of dengue virus infection with a high level of accuracy.
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http://dx.doi.org/10.1016/j.apjtm.2016.03.010DOI Listing
April 2016