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Abstract Background/Aim Efforts have been made to eliminate wild poliovirus transmission since 1988 when the World Health Organization began its global eradication campaign. Since then, the incidence of polio has decreased significantly. However, serotype 1 and serotype 3 still circulate endemically in Pakistan and Afghanistan. Both countries constitute a single epidemiologic block representing one of the three remaining major global reservoirs of poliovirus transmission. In this study we used genetic sequence data to investigate transmission links among viruses from diverse locations during 2005-2007. Methods In order to find the origins and routes of wild type 1 poliovirus circulation, polioviruses were isolated from faecal samples of Acute Flaccid Paralysis (AFP) patients. We used viral cultures, two intratypic differentiation methods PCR, ELISA to characterize as vaccine or wild type 1 and nucleic acid sequencing of entire VP1 region of poliovirus genome to determine the genetic relatedness. Results One hundred eleven wild type 1 poliovirus isolates were subjected to nucleotide sequencing for genetic variation study. Considering the 15% divergence of the sequences from Sabin 1, Phylogenetic analysis by MEGA software revealed that active inter and intra country transmission of many genetically distinct strains of wild poliovirus type 1 belonged to genotype SOAS which is indigenous in this region. By grouping wild type 1 polioviruses according to nucleotide sequence homology, three distinct clusters A, B and C were obtained with multiple chains of transmission together with some silent circulations represented by orphan lineages. Conclusion Our results emphasize that there was a persistent transmission of wild type1 polioviruses in Pakistan and Afghanistan during 2005-2007. The epidemiologic information provided by the sequence data can contribute to the formulation of better strategies for poliomyelitis control to those critical areas, associated with high risk population groups which include migrants, internally displaced people, and refugees. The implication of this study is to maintain high quality mass immunization with oral polio vaccine (OPV) in order to interrupt chains of virus transmission in both countries to endorse substantial progress in Eastern-Mediterranean region.

Abstract Background Eight genotypes of Hepatitis B virus designated A-H, have been known but in Pakistan, no such data is available on the prevalent HBV genotypes. Therefore, the subject study was conducted to determine HBV genotypes in the indigenous Pakistani population. Methods A total of 690 individuals were enrolled for HBV screening with EIA and nested PCR. Positive samples were further analyzed to determine HBV genotypes (A-F) by multiplex-PCR using type specific primers. Results 110 (15.94%) individuals were positive for HBV, including 64% males and 36% females. Out of these, 66 samples (65.34%) were classified into genotype D, 27 (26.73%) were of genotype B while 5(4.95%) had genotype A. In 3 (2.98%) samples, multiple genotypes were detected (genotype A+B; 2(1.99%) and genotypes B+D; 1(0.99%). Nine (8.18%) samples remained untyable. Conclusion In Asia, genotypes B and C are the most prevalent but our study reveals that genotype D is predominant and HBV infection constitutes a significant health problem in Pakistan.

Abstract Background The epidemiological significance of Hepatitis B virus genotypes has been well established and becoming an essential concern day by day however, much little is known about the mixed infection with more than one Hepatitis B virus genotypes and their clinical relevance. Methods Intravenous drug abusers are considered as a major risk group for the acquisition and transmission of blood borne infections like hepatitis B, however, in Pakistan, no such data has ever been reported about the epidemiology of HBV and its genotypes in Injecting Drug Users. 250 individuals were analyzed for hepatitis B virus genotypes after prior screening with serological assay for the detection of HBsAg. Results 56 (22.4%) individuals were found positive on ELSIA for HBsAg. The genotype distribution was found to be as: genotype D, 62.5%; genotype A, 8.92% while 28.57% individuals were found to be infected with a mixture of genotype A and D. Conclusion There is an urgent need of the time to develop public health care policies with special emphasis towards the control of HBV transmission through high risk groups especially Injecting Drug Users.

Abstract Background The infection rate of hepatitis B virus is continuously increasing in Pakistan. Therefore, a comprehensive study of epidemiological data is the need of time. Methods A total of 1300 individuals were screened for HBV infection markers including HBsAg, anti-HBsAg, HBeAg and anti-HBcAg. The association of these disease indicators was compared with patients' epidemiological characteristics like age, socio-economic status and residential area to analyze and find out the possible correlation among these variables and the patients disease status. Results 52 (4%) individuals were found positive for HBsAg with mean age 23.5 ± 3.7 years. 9.30%, 33.47% and 12% individuals had HBeAg, antibodies for HBsAg, and antibodies for HBcAg respectively. HBsAg seropositivity rate was significantly associated (p = 0.03) with the residing locality indicating high infection in rural areas. Antibodies titer against HBsAg decreased with the increasing age reflecting an inverse correlation. Conclusion Our results indicate high prevalence rate of Hepatitis B virus infection and nationwide vaccination campaigns along with public awareness and educational programs are needed to be practiced urgently.

Abstract Background Enteroviruses are among the most common viruses infecting humans worldwide and they are associated with diverse clinical syndromes. Acute flaccid paralysis (AFP) is a clinical manifestation of enteroviral neuropathy, transverse myelitis, Guillian-Barre Syndrome, Traumatic neuritis and many other nervous system disorders. The objective of this study was to understand the role of Non-Polio Enteroviruses (NPEV) towards this crippling disorder. Methods Stool specimens of 1775 children, aged less than 15 years, suffering from acute flaccid paralysis were collected after informed consent within 14 days of onset of symptoms during January 2003 to September 2003. The specimens were inoculated on RD and L20B cells using conventional tube cell culture while micro-neutralization test was used to identify the non-polio enterovirus (NPEV) serotypes. Detailed clinical information and 60-days follow-up reports were analyzed for NPEV-associated AFP cases. Results NPEV were isolated from 474 samples. The male to female ratio was 1.4:1. The isolation of NPEV decreased significantly with the increase in age. Cases associated with fever at the onset of NPEV-associated AFP were found to be 62%. The paralysis was found asymmetrical in 67% cases, the progression of paralysis to peak within 4 days was found in 72% cases and residual paralysis after 60 days of paralysis onset was observed in 39% cases associated with NPEV. A clinical diagnosis of Guillian-Barre syndrome was made in 32% cases. On Microneutralization assay, echo-6 (13%) and coxsackievirus B (13%) were the most commonly isolated serotypes of NPEV along with E-7, E-13, E-11, E-4 and E-30. The isolates (n = 181) found untypable by the antiserum pools were confirmed as NPEV by PCR using Pan-Enterovirus primers. Conclusion The present study suggests that NPEV are a dominant cause of AFP and different serotypes of NPEV are randomly distributed in Pakistan. The untypable isolates need further characterization and analysis in order to determine their association with clinical presentation of a case.

Background: Dialysis patients are at high risk for surgery, but their outcomes after splenectomy are unclear. We compared postoperative complications between dialysis and non-dialysis patients., Methods: Data were retrieved from the National Surgical Quality Improvement Program for this retrospective cohort. Adult patients undergoing elective splenectomy between 2005 and 2020 were included., Results: Among 10,339 included patients, 143(1.4%) were on chronic dialysis. Postoperative mortality was higher in dialysis vs. non-dialysis patients (9.1% vs. 1.8%). Dialysis patients were more likely to have 30-day major morbidity, infectious and non-infectious complications, reoperation, and prolonged hospital stay. On multivariable regression, dialysis dependence significantly increased odds of mortality, major morbidity, blood transfusion, prolonged length of stay, reoperation, and failure-to-rescue (FTR)., Conclusion: Dialysis patients were at higher risk of postoperative morbidity following splenectomy. Additionally, the risk of FTR in this patient population is also significantly more compared to non-dialysis patients., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Introduction  Despite the evidence against drain placement after thyroidectomy, there is a lack of consensus on drain use in patients with substernal goiter. Objective  To assess the factors that increase the likelihood of drain placement and its impact on postoperative hematoma and other 30-day complications among adult patients undergoing thyroidectomy for substernal goiter. Methods  A retrospective cohort study that used data from the American College of Surgeons National Surgical Quality Improvement Program (ACS-NSQIP). Adult patients (aged ≥ 18 years) who underwent elective thyroidectomy for substernal goiter from 2016 to 2020 were included. Cases with closed suction neck drains placed upon completion of surgery were included in the drain group, and the remaining cases formed the nondrain group. Results  A total of 1,229 patients were included (46.5% with drain placement). The factors that increased the likelihood of drain placement included body mass index (BMI) ≥ 30 kg/m 2 , score between 3 and 5 on the American Society of Anesthesiologists (ASA) physical status classification, sternal split/transthoracic surgical approach, operative time ≥ 90 minutes, and surgery conducted by otolaryngologists. Patients with clean-contaminated or contaminated wound classifications were less likely to be submitted to drain placement. In addition, drain use had no impact on postoperative hematoma formation but was found to independently increase the risk of prolonged length of hospital stay. Conclusion  Thyroidectomy without drain placement might be safe for substernal goiter. However, this decision should be individualized for each patient. Level Of Evidence: 3., Competing Interests: Conflict of Interests The authors have no conflict of interests to declare., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution 4.0 International License, permitting copying and reproduction so long as the original work is given appropriate credit ( https://creativecommons.org/licenses/by/4.0/ ).)

Purpose: We hypothesize that lower grade gliomas (LGG) can be identified and classified into two distinct subtypes: radiologically circumscribed Lower-Grade Gliomas (cLGG) and infiltrating Lower-Grade Gliomas (iLGG) based on radiological parameters and that these two different subtypes behave differently in terms of clinical outcomes., Methods: We conducted a retrospective cohort study on surgical patients diagnosed with lower grade glioma over five years. Patient records and MRIs were reviewed, and neurosurgeons classified tumors into cLGG and iLGG groups., Results: From the 165 patients in our cohort, 30 (18.2%) patients were classified as cLGG and 135 (81.8%) patients were classified as iLGG Mean age in cLGG was 31.4 years while mean age in iLGG was 37.9 years ( p  = 0.004). There was significant difference in mean blood loss between cLGG and iLGG groups (270 and 411 ml respectively, p  = 0.020). cLGG had a significantly higher proportion of grade II tumors ( p  < 0.001). The overall mean survival time for the iLGG group was 14.96 ± 1.23 months, and 18.77 ± 2.72 months for the cLGG group. In univariate cox regression, the survival difference between LGG groups was not significant (HR = 0.888, p  = 0.581), however on multivariate regression cLGG showed a significant (aHZ = 0.443, p  = 0.015) positive correlation with survival. Intense contrast enhancement (HZ = 41.468, p  = 0.018), blood loss (HZ = 1.002, p  = 0.049), and moderately high Ki-67 (HZ = 4.589, p  = 0.032) were also significant on univariate analyses.Conclusion: cLGG and iLGG are radiologically distinct groups with separate prognoses, surgical experience, and associations., Competing Interests: The authors have no relevant financial or non-financial interests to disclose., (© 2024 Published by Elsevier Inc.)

Objective: To assess associations between various clinic-demographic factors and clinical outcomes among patients treated for sepsis., Methods: The retrospective study was conducted at the Aga Khan University Hospital, Karachi, and comprised data of all patients aged >18 years diagnosed with sepsis from January to December 2019. Multivariable logistic regression was used to evaluate independent associations between predictors and outcomes. Data was analysed using R packages., Results: Of the 1,136 patients, 621(54.6%) were male and 515(45.3%) were female. The overall mean age was 59.05±16.91 years. Female gender (odds ratio: 1.029; 95% confidence interval: 1.03-1.64) was found to be an independent predictor of septic shock, while hypertension (odds ratio0.75; 95% confidence interval: 0.59-0.95) emerged as a protective factor. Chronic kidney disease (odds ratio: 1.539; 95% confidence interval: 1.14-2.07) was an independent predictor of prolonged length of stay, while older age appeared to be protective (odds ratio: 0.98; 95% confidence interval: 0.98-0.99). Mortality was associated with a significantly lower odds of Escherichia coli on culture (odds ratio: 0.26; 95% confidence interval: 0.12-0.54)., Conclusions: Independent associations were found between specific patient characteristics and adverse clinical outcomes.

Objective: To determine the association between internet use and anxiety among people during the coronavirus disease- 2019 pandemic., Methods: The cross-sectional study was conducted across Pakistan from January 14 to February 21, 2021, which was the active phase of the coronavirus disease-2019 pandemic in Pakistan. The participants were aged at least 13 years having internet access regardless of gender or their location across Pakistan. The anonymous web-based survey was conducted using a questionnaire generated on Google Forms and disseminated through various social media platforms and WhatsApp groups. Anxiety symptoms were screened using the Depression, Anxiety and Stress Scale-21, while the Young Internet Addiction Test was used to evaluate symptoms of internet addiction. Data was analysed using STATA 16., Results: Of the 1,145 subjects, 686(60%) were females and 459(40%) were males. A total of 257(22.5%) participants were found to have extremely severe anxiety and internet usage pattern was significantly associated with the level of anxiety (p<0.05). Age, gender, social class and marital status were not significantly different (p>0.05), while family income and area of living were significantly different (p<0.05) in terms of anxiety levels. The odd of addictive internet use was 10.2 (95% confidence interval: 5.7-18.5) times greater in extreme anxiety individuals compared to individuals having no anxiety after controlling for other sociodemographic, health-related, behavioural and environmental factors during the pandemic., Conclusion: A significant association of anxiety was found with internet addiction during the coronavirus disease-2019 pandemic.

The objective of this study is to report clinical, radiological, and histopathological characteristics of three paediatric patients diagnosed as Choroid plexus carcinoma seen at our hospital, between 2015 and 2020. Three patients were diagnosed with choroid plexus carcinomas between 2015 and 2018. The mean age at diagnosis was 1.3 years (range 8 months to 1.5 years). All the three patients had subtotal resection and received adjuvant chemotherapy. One patient also received adjuvant radiotherapy. Despite these treatment measures, residual disease was noted in all three patients and two patients were subsequently treated on palliative care grounds. The average duration of follow-up after the first surgery for all three patients was approximately 33 months. Attaining satisfactory outcome in patients with CPC is challenging. Our case series reflects the difficulty in achieving gross total resection and ensuring that the disease does not recur.

Acute respiratory distress syndrome (ARDS) is the most severe and devastating form of acute lung injury. Neutrophil to lymphocyte ratio (NLR) and C-reactive protein have been used to predict severity and prognosis of certain viral illnesses., Methods: A retrospective cohort study was conducted in hospitalized COVID-19 patients. Serial measurements of NLR and C-reactive protein were recorded and patients were followed for development of ARDS., Results: Out of 388 patients admitted with COVID-19, 43 patients developed ARDS compared with 345 patients who did not develop ARDS. The median NLR at presentation was significantly higher in patients who developed ARDS (8.89 vs. 4.25) compared with those who did not develop ARDS ( P <0.001). Moreover, serial measurement of NLR at presentation, day 3, day 5 and day 7 was significantly associated with development of ARDS ( P <0.001). In multivariable analysis, age of greater than or equal to 50 years (adsjusted odds ratio=3.28; 95% CI=1.40-7.69) and unit increase in NLR at presentation (adsjusted odds ratio=1.07; 95% CI=1.03-1.11) were independently associated with development of ARDS., Conclusion: Serial measurement of NLR can predict patients who are at a greater risk for developing ARDS in COVID-19., Competing Interests: The authors have no conflicts of interest., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc.)

Epilepsy is the third most common neurological disease in the world associated with a high frequency in the paediatric age group. This study aims to evaluate the prevalence, types and aetiologies of epilepsy within the Pakistani population. A retrospective review of the charts of all patients, below the age of 18 years, presenting with epilepsy to the Department of Neurology at The Children's Hospital and Institute of Child Health, Lahore, from January 2016 to December 2020, was carried out. Analysis was performed using SPSS Version 26. A p value of <0.05 was considered statistically significant. A total of 1,097 patients were studied, of which 644 (58.8%) were males and 451 (41.2%) females. a vast majority, i.e. 1,021 (96.1%), of the study participants, belonged to the province of Punjab. Afebrile seizures [n=798 (72.7%)] were more commonly reported than febrile seizures [n=299 (27.3%)]. Among seizure types, generalised seizures were the most common type of seizure reported in 520 (49.8%) patients. Refractory seizures were the least common type reported in 3 (0.3%) patients. Aetiology was mostly idiopathic [n=540 (49.2)], followed by congenital [n=228 (20.8%)]. The most frequently reported duration of seizure was between one and three minutes [n=116 (42.3%)]. The most common ictal features seen were a combination of up-rolling of eyes and frothing from the mouth [n=206 (34.9%)]. Results from this study can be used by health care providers to better formulate therapeutic interventions for a timely diagnosis and effective treatment of epilepsy.

Introduction: The effect of chronic steroid therapy on postoperative outcomes after craniotomy for tumor resection remains understudied., Research Question: This study aimed to fill this gap and to identify risk factors of postoperative morbidity and mortality among patients on chronic steroid use undergoing craniotomy for tumor resection., Materials and Methods: Data from the American College of Surgeons National Surgical Quality Improvement Program were used. Patients who underwent craniotomy for tumor resection between 2011 and 2019 were included. Perioperative characteristics and complications were compared among patients with and without chronic steroid therapy, defined as steroid use for at least 10 days. Multivariable regression analyses were conducted to assess the impact of steroid therapy on postoperative outcomes. Subgroup analyses involving patients on steroid therapy were conducted to explore risk factors of postoperative morbidity and mortality., Results: Of 27,037 patients, 16.2% were on steroid therapy. On regression analyses, steroid use was significantly associated with any postoperative complication, infectious complication, urinary tract infection, septic shock, wound dehiscence, pneumonia, non-infectious, pulmonary, and thromboembolic complications, cardiac arrest, blood transfusion, unplanned reoperation, readmission, and mortality. On subgroup analysis, risk factors for postoperative morbidity and mortality among patients on steroid therapy included older age, higher American Society of Anesthesiology physical status, functional dependence, pulmonary and cardiovascular comorbidities, anemia, dirty/infected wounds, prolonged operative time, disseminated cancer, and a diagnosis of meningioma., Discussion and Conclusion: Preoperative brain tumor patients on steroids for 10 or more days are at a relatively high risk of postoperative complications. We recommend a judicious use of steroids in brain tumor patients, both in terms of dosage and duration of treatment., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier B.V. on behalf of EUROSPINE, the Spine Society of Europe, EANS, the European Association of Neurosurgical Societies.)

Objective: We aim to document the frequency of HAAA cases among AA patients presenting at a tertiary care hospital, and to determine the most common agents (viral/drug induced) and Clinico-haematological features among HAAA patients at a tertiary care hospital., Methods: This study was a retrospective review, conducted at a tertiary care hospital in Karachi, Pakistan., Results: A total of 21 patients were included in the study. Hepatitis among the HAAA patients was viral in 17 cases, while 4 were idiopathic. All the patients acquired aplastic anaemia within 3-12 months of the Hepatitis episode and most presented with bleeding, bruises and petechiae., Conclusions: This study indicates and proves that presence and prevalence of this disease in the Pakistani population is quite significant. Unlike the rest of the world, HAAA in Pakistan is not entirely of unknown aetiology, most of the cases can be associated with one of the Hepatitis viruses.

Background: Due to the differences in size and invasiveness when compared to non-giant macroadenomas (nGPAs), giant pituitary adenomas (GPAs) are considerably harder to resect. This study aimed to differentiate GPAs from nGPAs, based on the presenting complaints, surgical approaches, peri- and postoperative outcomes., Methods: We retrospectively analyzed cases of pituitary macroadenomas that underwent surgical resection at a tertiary care hospital. GPAs were tumors greater than 4 cm in the largest dimension, while nGPAs were tumors smaller than 4 cm. 55 GPA patients and 70 nGPA patients from 2006 to 2017 were included. Demographic, perioperative, and post-operative outcomes were evaluated. Group comparisons for continuous variables were made using an independent t-test/Mann Whitney U test and categorical data was analyzed on Chi-square/Fisher exact test; a p-value of < 0.05 was considered significant., Results: Visual deterioration was the most common complaint, reported by 61.4% of nGPA patients and 81.8% of GPA patients. The mean extent of gross total resection was 47.1% in nGPA patients and 18.2% in GPA patients (p = 0.001). After surgery, tumor recurrence was seen in 1.4% of nGPA patients and 18.2% of GPA patients (p = 0.001). First re-do surgery was required in 5.7% of nGPA patients and 25.5% of GPA patients (p = 0.004)., Conclusion: Compared to nGPAs, GPAs are more likely to present with a higher number of preoperative symptoms, and lesser chances of gross total tumor resection. GPAs are also associated with a higher rate of recurrence, which results in more follow-up procedures. Larger, multi-center longitudinal studies need to be done to validate these findings., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: The endoscopic endonasal approach (EEA) has been gaining popularity for resection of adult craniopharyngiomas. However, the safety and effectiveness of the procedure in comparison to the traditional transcranial approach (TCA) remains unestablished as previous reviews are outdated., Methods: A literature search without language restriction was conducted in PubMed, Cochrane database, and Web of Science from conception to July 9, 2021. Cohort studies and case series that compared EEA with TCA and assessed postoperative complications, recurrence, and 30-day mortality were included. Articles, where data for adult populations could not be extracted or calculated, were excluded. Article selection and data extraction in a predesigned data extraction form were conducted in duplicate. Pooled participant data were included in a random-effects model., Results: The search yielded 227 articles, from which eight cohort studies containing 11,395 patients were included (EEA: 6,614 patients, TCA: 4,781 patients). Six studies were good quality and two were fair quality according to the Newcastle Ottawa Scale. There were significantly higher rates of cerebrospinal fluid leak (risk ratio [RR]=0.23, 95% confidence interval [CI] 0.17-0.32, p <0.00001, I²=0%) and lower rates of postoperative hypopituitarism (RR=1.40, 95% CI 1.30-1.51, p <0.00001, I²=0%), hydrocephalus (RR=6.95, 95% CI 5.78-8.36, p <0.00001, I²=0%), visual impairment (RR=1.52, 95% CI 1.34-1.73, p <0.00001, I²=0%), and 30-day mortality (RR=5.63, 95% CI 3.87-8.19, p <0.00001, I²=0%) after EEA. Non-significant lower rates of postoperative diabetes insipidus (RR=1.12, 95% CI 0.78-1.61, p =0.53, I²=85%) and recurrence of tumor (RR=2.69, 95% CI 0.35-20.81, p =0.34, I²=47%) were seen after EEA., Conclusion: EEA may be associated with reduced postoperative hypopituitarism, hydrocephalus, visual impairment, and 30-day mortality and higher rates of cerebrospinal fluid leak. These findings do not account for differences in tumor size and extension between the EEA and TCA cohorts. Further research on patients with comparable tumor characteristics is required to fully assess outcomes., Competing Interests: The authors have no potential conflicts of interest to disclose., (Copyright © 2022 The Korean Brain Tumor Society, The Korean Society for Neuro-Oncology, and The Korean Society for Pediatric Neuro-Oncology.)

Background No scoring system is available to predict the extent of resection of giant pituitary adenomas (GPAs) based on magnetic resonance imaging (MRI) parameters. We developed a novel AKU Giant Pituitary Adenoma (AGPA) score and assessed the predictive ability of the scoring system concerning the extent of resection of GPAs. Methodology We retrospectively collected data of patients presenting with GPAs and used our scoring system to assess the surgical resection of these tumors. The Lundin-Pederson (ABC/2) method was used to calculate the pre- and post-resection tumor volume. The relationship between the extent of resection and the AGPA score was assessed using linear regression. The AGPA score considered the tumor's extension into various planes. The maximum total score was 9. Results The scoring system was applied to 45 patients with GPA who underwent surgical resection. The mean resected tumor volume (%) was 82.0 ± 16.7, and the overall mean AGPA score was 4.2 ± 0.8. The pairwise correlation between the resected tumor volume and the overall AGPA scores showed a strong inverse association (r = -0.633, p < 0.001). A significant difference was detected between the estimated scores of 3 and 5 and 4 and 5 (p < 0.001). Conclusions AGPA score is inversely related to the extent of the tumor to be resected, which would help surgeons predict the amount of tumor resection possible as well as predict the difficulty of surgery and plan optimal preoperative patient counseling. In addition, it can predict if staging and a transcranial approach are required., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2022, Enam et al.)

Background: Elimination of poliovirus in Pakistan and Afghanistan is challenged by notions against the role of oral poliovirus vaccine (OPV) in eradicating contemporary wild poliovirus (WPV) strains., Methods: A total of 1055 WPV type 1 (WPV1) strains isolated between 2013 and 2018 were categorized into 68 antigenic groups and tested for neutralization by OPV-derived antibodies. Molecular docking was conducted to determine neutralization efficiency of antibodies against WPV. The clinical significance of WPV1 variants was assessed to ascertain their role in patient outcomes., Results: We found that 88% of WPV1 strains isolated from paralytic children belonged to a single antigenic lineage identical to the WPV1 strain detected in 1993. WPV1 antigenic variants were effectively neutralized by OPV-derived antibodies, with geometric mean titers comparable to the neutralization titers found for 3 strains in OPV (OPV1-3, 7.96-9.149 [95% confidence interval, 6.864-10.171]; WPV1 strains, 7.542-8.786 [6.493-9.869]). Docking examination underscored a strong antigen-antibody interaction despite variations within the viral protein 1 epitopes. There was no significant association (P = .78) with clinical prognosis among patients infected with antigenically diverse WPV1 strains and patient outcomes, including death., Conclusions: Our findings substantiate the robustness of OPV for neutralizing the contemporary WPV1 strains endemic in Pakistan and Afghanistan. Vaccination coverage must be augmented to achieve early eradication., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Awake-craniotomy (AC) allows maximum tumour resection while reducing the risk of permanent postoperative neurological deficits without compromising the outcomes of the treatment. AC is a potentially stressful procedure for the patients, which may lead to long-term effects on patients' psychology, and may lead to post-traumatic stress. However, there is a scarcity of data suggesting actual association of any neuropsychiatric conditions after AC. Therefore, we sought to review the neuropsychiatric conditions post awake craniotomy.

Background: This study aimed to document the evolution of perceptions of frontline healthcare workers (FHCW) regarding their well-being and the quality of health systems' response to the COVID-19 pandemic over four months in Pakistan., Methods: We conducted this prospective longitudinal qualitative study during the four months (June-September 2020) coinciding with the peak and trough of the first wave of Pakistan's COVID-19 pandemic. We approached frontline healthcare workers (physicians and nurses) working in emergency departments (ED) in two hospitals using the WhatsApp group of the Pakistan Society of Emergency Physicians (PSEM). Participants were asked to self-record their perception of their wellness and their level of satisfaction with the quality of their hospitals' response to the pandemic. We transcribed, translated, and analysed manually using MAXQDA 2020 software and conducted the thematic analysis to identify themes and sub-themes., Results: We invited approximately 200 FHCWs associated with PSEM to participate in the study. Of the 61 who agreed to participate, 27 completed the study. A total of 149 audio recordings were received and transcribed. Three themes and eight sub-themes have emerged from the data. The themes were individual-level challenges, health system-level challenges, and hope for the future. Sub-themes for individual-level challenges were: fear of getting or transmitting infection, feeling demotivated and unappreciated, disappointment due to people's lack of compliance with COVID-19 protocols, physical exhaustion, and fatigue. For the healthcare system, sub-themes were: Infrastructure, logistics, management, and communications response of the hospital/healthcare system and financial stressors. For sub-themes under hope for the future were the improved disease knowledge and vaccine development. The overall perceptions and experiences of FHCWs evolved from fear, grief, and negativity to hope and positivity as the curve of COVID-19 went down., Conclusion: This study shows that the individuals and systems were not prepared to deal with the challenges of the COVID-19 pandemic. The findings highlight the challenges faced by individuals and health systems during the wake of the Covid-19 pandemic. The healthcare workers were emotionally and physically taxed, while the health systems were overwhelmed by COVID-19. The overall perceptions of FHCWs evolved with time and became negative to positive as the curve of COVID-19 went down during the first wave of COVID-19 in Pakistan., (© 2022. The Author(s).)

This study aims to determine the burden of COVID19-induced internet addiction and related psychosocial factors among the Pakistani sample. A web-based cross-sectional survey was conducted from January to March 2021. An analytical cross-sectional survey was broadcast on the internet via a Google form completed by 1145 Pakistanis. Individuals aged 13 years and above with competency to comprehend English or Urdu language, currently residing in any province of Pakistan, having access to the questionnaire, and willing to participate were eligible to participate. Overseas Pakistanis were excluded from the study. The outcome is COVID19-induced internet addiction was measured using the validated tool Young's Internet Addiction Test (IAT). In addition, symptoms of depression, anxiety, stress, and other psychosocial factors were assessed using the validated tool Depression, Anxiety, and Stress Scale-21 (DASS-21). Adjusted odds ratios with a 95% confidence interval were reported using multinomial logistic regression. Most participants were females and youth (between 20 and 24 years). The prevalence of problematic internet users (PIU) and addictive internet users (AIU) was 27.3% and 11.3%, respectively. The odds of extremely severe anxiety among AIU were approximately three times (Adj OR: 2.6 (1.1-7.1), followed by the odds of having extremely severe stress being about five times higher among AIU (Adj OR: 5.4 (1.6-17.6)) as compared to normal internet users (NIU). Amid COVID-19, the burden of internet addiction has surged among the Pakistani populace. This study identified that gender, marital status, depression, stress, anxiety, work situation, and mood changes during the COVID-19 pandemic are significantly correlated with problematic and addictive internet use. Preventative measures against the addictive use of the internet are needed to avoid or mitigate any serious mental health problems.

Surveillance and detection of polioviruses (PV) remain crucial to monitoring eradication progress. Intratypic differentiation (ITD) using the real-time RT-PCR kit is key to the surveillance workflow, where viruses are screened after cell culture isolation before a subset are verified by sequencing. The ITD kit is a series of real-time RT-PCR assays that screens cytopathic effect (CPE)-positive cell cultures using the standard WHO method for virus isolation. Because ITD screening is a critical procedure in the poliovirus identification workflow, validation of performance of real-time PCR platforms is a core requirement for the detection of poliovirus using the ITD kit. In addition, the continual update and improvement of the ITD assays to simplify interpretation in all platforms is necessary to ensure that all real-time machines are capable of detecting positive real-time signals. Four platforms (ABI7500 real-time systems, Bio-Rad CFX96, Stratagene MX3000P, and the Qiagen Rotor-Gene Q) were validated with the ITD kit and a redesigned poliovirus probe. The poliovirus probe in the real-time RT-PCR pan-poliovirus (PanPV) assay was re-designed with a double-quencher (Zen™) to reduce background fluorescence and potential false negatives. The updated PanPV probe was evaluated with a panel consisting of 184 polioviruses and non-polio enteroviruses. To further validate the updated PanPV probe, the new assay was pilot tested in five Global Polio Laboratory Network (GPLN) laboratories (Madagascar, India, Philippines, Pakistan, and Democratic Republic of Congo). The updated PanPV probe performance was shown to reduce background fluorescence and decrease the number of false positives compared to the standard PanPV probe., Competing Interests: The authors have declared that no competing interests exist.

The ongoing COVID-19 pandemic is caused by SARs-CoV-2. The virus is transmitted from person to person through droplet infections i.e. when infected person is in close contact with another person. In January 2020, first report of detection of SARS-CoV-2 in faeces, has made it clear that human wastewater might contain this virus. This may illustrate the probability of environmentally facilitated transmission, mainly the sewage, however, environmental conditions that could facilitate faecal oral transmission is not yet clear. We used existing Pakistan polio environment surveillance network to investigate presence of SARs-CoV-2 using three commercially available kits and E-Gene detection published assay for surety and confirmatory of positivity. A Two-phase separation method is used for sample clarification and concentration. An additional high-speed centrifugation (14000Xg for 30 min) step was introduced, prior RNA extraction, to increase viral RNA yield resulting a decrease in Cq value. A total of 78 wastewater samples collected from 38 districts across Pakistan, 74 wastewater samples from existing polio environment surveillance sites, 3 from drains of COVID-19 infected areas and 1 from COVID 19 quarantine center drainage, were tested for presence of SARs-CoV-2. 21 wastewater samples (27%) from 13 districts turned to be positive on RT-qPCR. SARs-COV-2 RNA positive samples from areas with COVID 19 patients and quarantine center strengthen the findings and use of wastewater surveillance in future. Furthermore, sequence data of partial ORF 1a generated from COVID 19 patient quarantine center drainage sample also reinforce our findings that SARs-CoV-2 can be detected in wastewater. This study finding indicates that SARs-CoV-2 detection through wastewater surveillance has an epidemiologic potential that can be used as supplementary system to monitor viral tracking and circulation in cities with lower COVID-19 testing capacity or heavily populated areas where door-to-door tracing may not be possible. However, attention is needed on virus concentration and detection assay to increase the sensitivity. Development of highly sensitive assay will be an indicator for virus monitoring and to provide early warning signs., Competing Interests: The authors have declared that no competing interests exist.

Ocular tissues can serve as a reservoir for the SARS-CoV-2 virus which can not only cause conjunctivitis but also serve as a source of infection transmission to others. Additionally, the eye and its tear drainage apparatus can track the SARS-CoV-2 from the eye into the respiratory tract of the patient. The potential ocular presence of the SARS-CoV-2 in the eye of a patient can target ACE2 receptors in the endothelium of the conjunctival vessels and use the lacrimal sac a potential space to evade immune detection and clinical isolation. The recently reported case of COVID-19 after the acquisition of SARS-CoV-2 from a COVID-19 patient should alert the healthcare professionals dealing with COVID-19 patients that wearing masks alone cannot guarantee protection against infection transmission. Further studies, like isolation of SARS-CoV-2 from the eyes of patients with COVID-19, are needed to identify the eyes as a potential source of SARS-CoV-2 infection transmission., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

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., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

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., (Copyright © 2020 Shaw et al.)

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., (© 2018 Wiley Periodicals, Inc.)

Background: Measles virus infection remains a significant cause of childhood mortality and morbidity despite continued global efforts and the availability of a safe and effective vaccine. Molecular analysis of indigenous measles viruses could provide critical information on outbreak linkages and transmission pathways that can aid the implementation of appropriate control programs in Pakistan., Methods: Blood samples and throat swabs were collected from subjects suspected with measles in Islamabad, Pakistan from 2013 to 2015. Serum samples were tested for the presence of measles immunoglobulin M (IgM) antibodies using enzyme-linked immunosorbent assay (ELISA) while throat swabs were used for the isolation (Vero/SLAM cell line) and subsequent characterization and phylogenetic analysis of measles strains., Results: Of 373 blood samples, 66% tested positive for measles IgM. Male subjects were more often infected (58%) than female (42%) with the highest frequency of positive cases (63%) in the 0-5-years age group. Among the positive cases, only 13% had received one or two doses of the measles vaccine, while 87% were unvaccinated. Of 80 throat swabs, 29 (36%) showed a measles virus-specific cytopathic effect (CPE) and were characterized as genotype B3 through partial sequencing of the nucleoprotein (N) gene. Phylogenetic analysis revealed the Pakistani B3 strains to be closely related to strains from neighboring countries (Iran and Afghanistan) as well as with B3 viruses from the USA, Germany, and the UK., Conclusions: The study results showed that despite the availability of an effective vaccine, the burden of measles infections is very high in Pakistan due to poor routine immunization coverage even in major cities, including the capital city of Islamabad. It is imperative that national health authorities take urgent strategic steps to improve routine immunization and implement adequate molecular identification methods to tackle future measles outbreaks., (Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Measles continues to be a major public health issue causing substantial outbreaks worldwide, mostly affecting young children. Molecular analysis of measles viruses provides important information on outbreak linkages and transmission pathways that can be helpful towards implementation of appropriate control programs. In Pakistan, the control of measles is still tenuous, and progress towards elimination has been irregular and challenging. In the 2013 measles outbreak we received 4,682 sera collected from suspected patients in 23 districts across Sindh. A total of 3,283 samples were confirmed measles positive using IgM ELISA with the highest infection rate in children aged 1-12 months. Males were more affected than females and a visible peak was observed from January to April. Among the 3,283 cases, 59.1% were unvaccinated, 29.6% had received 1 dose and 10.3% had received 2 doses of measles vaccine while 0.85% had an unknown vaccination status. For genotype detection and phylogenetic analysis, 60 throat swab samples were collected from suspected patients below 15 years of age in eight districts of Sindh province. Forty four (73%; 44/60) throat swab samples were successfully genotyped using RT-PCR. Phylogenetic analyses based on partial sequences of the nucleocapsid protein gene revealed that all Pakistani measles virus strains belonged to genotype B3 and were closely related to those isolated from neighboring countries such as Iran, Afghanistan (99.1-100%) and India with 98.6 - 99.6% nucleotide homology. This is the first report on the phylogenetic analysis of measles B3 genotype strains from Pakistan and highlights the need for strengthening the surveillance systems and improving immunization coverage across the country.

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., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: Dengue virus is the causative agent of dengue fever, a vector borne infection which causes self-limiting to life threatening disease in humans. A sero-epidemiological study was conducted to understand the current epidemiology of dengue virus in Pakistan which is now known as a dengue endemic country after its first reported outbreak in 1994., Methods: To investigate the prevalence of dengue virus in Pakistan during 2009-2014, a total of 9,493 blood samples were screened for the detection of anti-dengue IgM antibodies using ELISA. Clinical and demographic features available with hospital records were reviewed to ascertain mortalities related to dengue hemorrhagic shock syndrome., Results: Out of 9,493 samples tested, 37% (3,504) were found positive for anti-dengue IgM antibodies. Of the seropositive cases, 73.6% (2,578/3,504) were male and 26.4% (926/3,504) were female. The highest number (382/929; 41.1%) of sero-positive cases was observed among the individuals of age group 31-40 years. The highest number of symptomatic cases was reported in October (46%; 4,400/9,493), and the highest number of sero-positive cases among symptomatic cases was observed in November (45.7%; 806/1,764). Mean annual patient incidence (MAPI) during 2009-2014 in Pakistan remained 0.30 with the highest annual patient incidence (11.03) found in Islamabad. According to the available medical case record, 472 dengue related deaths were reported during 2009-2014., Conclusion: The data from earlier reports in Pakistan described the dengue virus incidence from limited areas of the country. Our findings are important considering the testing of clinical samples at a larger scale covering patients of vast geographical regions and warrants timely implementation of dengue vector surveillance and control programs., Trial Registration Number: It is an epidemiological research study, so trial registration is not required.

Echovirus 13 (E-13) is reported worldwide and is mostly related to aseptic meningitis but it is also isolated from cases of acute flaccid paralysis (AFP). Unfortunately, all studies conducted on non polio enterovirus in Pakistan only confirm E-13 isolation based on microneutralization assay but there is lack of molecular epidemiological data on this serotype. In this study, 113 stool samples were collected from AFP patients during 2008-2010. An enterovirus primer mediated real-time reverse transcriptase polymerase chain reaction, a standard microneutralization assay and sequencing of viral protein 1 gene (VP1) identified the predominant serotype E-13. For molecular characterization, genetic relationship between 12 clinical isolates of echovirus 13 was investigated by partial sequencing of viral protein 1 gene. These strains, combined with related sequences from GenBank were divided phylogenetically into two different genogroups A and B (>30% divergence) and were found genetically distinct from the circulating strains in the world. Additionally, phylogenic grouping pattern revealed that the study strains clustered into three distinct subgroups (A3, A7 and B3) having >23% nucleotide divergence representing three new genotypes. The genotype A7 seems to be restricted geographically. In conclusion, the current study provides an overview of the molecular epidemiology and evolution of E-13 in the country. This study strongly suggests that enterovirus surveillance system should be established in the country to determine the temporal and geographical trends and disease pattern of different enterovirus serotypes in the community., (Copyright © 2016 Elsevier B.V. All rights reserved.)

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., Competing Interests: Authors' Disclosures of Potential Conflicts of Interest: No potential conflicts of interest relevant to this article were reported.

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., (Copyright © 2016 Hainan Medical University. Production and hosting by Elsevier B.V. All rights reserved.)

Objective: To determine the prevalence of cytomegalovirus in pregnant women and types of overt congenital infection in neonates., Methods: This cross-sectional study was conducted at the Pakistan Institute of Medical Sciences and Federal Government Services Hospital in Islamabad, Pakistan, from March 2010 to June 2011, and comprised blood samples of pregnant women. Seroprevalence of human cytomegalovirus, immunoglobulin G and immunoglobulin M was determined by enzyme-linked immunosorbent assay while its deoxyribonucleic acid was detected by nested polymerase chain reaction.The congenital human cytomegalovirus infection was also identified in newborn babies from actively infected pregnant women. SPSS 18 was used for data analysis., Results: Of the 409 pregnant women enrolled, 399(97.55%) were seropositive for cytomegalovirus immunoglobulinG and 52(12.71%) for immunoglobulinM, while cytomegalovirus deoxyribonucleic acid was detected in 82(20%). Of the cytomegalovirus immunoglobulinM-positive women, sera of 40(80%) had immunoglobulinG avidity >50%. The remaining 12(23%) sera had avidity assay value <50%. Among the 82(20%) infected pregnant women, 70(85.4%) were successfully followed up. Among them, the virus was isolated from 41(58.5%) newborns babies, of which 15(21%) were symptomatic while 26(47.2%) were asymptomatic. Of the former, 4(26.6%) had hepatosplenomegaly., Conclusions: Human cytomegalovirus infection in pregnant women was the main reason of congenital defects among neonates.

Background: Congenital cytomegalovirus (cCMV) infection contributes to considerable long-term sequelae in neonates and children all over the world. The association between viral genotypes and severity of clinical cytomegalovirus (CMV) infection is yet to be defined. The objective of this study was to find the impact of active CMV infection during pregnancy and the clinical significance of genotypes in neonates with congenital cytomegalovirus infections in Pakistan., Methods: A total of 409 blood samples from pregnant women seeking health care services at the two antenatal hospitals of Islamabad during January to December 2012 were tested by ELISA and nested-PCR. Pregnant women with active infection (detected as IgM positive, PCR positive or positive on both assays) were followed until delivery, to detect the outcome of overt cCMV infection in neonates. Genetic characterization of CMV strains was performed by sequence analysis of envelope glycoproteins: gB, gN and gH to detect the contributing CMV genotypes., Results: The seroprevalence of anti-CMV IgG and IgM was 97.5% (399 out of 409) and 12.7% (52 out of 409), respectively, while 20% (82/409) pregnant women were found positive for CMV DNA by PCR. Logistic regression analysis showed a significant association of active infection with parity [OR = 2.56, 95% CI = 1.82-2.62, p = 0.04], febrile illness [OR = 1.84, 95% CI = 1.76-3.65, p = 0.01] and jaundice [OR = 22.5, 95% CI = 4.53-85.02, p = 0.002]. We were able to isolate virus in 41 out of 70 neonates; 36.6% (15 out of 41) of them were symptomatic at birth while 63.4% (26 out of 41) were asymptomatic. The most prominent clinical feature observed in symptomatic neonates was hepatosplenomegaly (26.6%; 4 out of 15). All three genotypes gB, gN and gH were found with the highest frequency of gB1 genotype, found in 75% infants with hepatic damage. Phylogenetic analysis of Pakistani strains showed 96%-100% homology to their prototype strains., Conclusions: Active CMV infection during pregnancy is a major cause of congenital CMV infection with comparable distribution of all three genotypes: gB, gN and gH in symptomatic and asymptomatic neonates. Our findings emphasize to conduct a comprehensive large scale survey and introduction of country wide routine screening at maternity clinics for early diagnosis of CMV to reduce its associated devastating outcomes.

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., (Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.)

Background: Despite tremendous efforts in the fight against polio, Pakistan bears the highest proportion of poliomyelitis cases among the 3 endemic countries including Afghanistan and Nigeria. Apart from insecurity and inaccessibility challenges, the substantial shift of unimmunized children from North Waziristan due to recent military operations was presumed to favor the widespread poliovirus infection in Pakistan., Methods: To better understand the current epidemiological situation, we analyzed the virologic data of wild poliovirus type 1 (WPV1) strains detected in Pakistan during 2013-2015., Results: Five genetic clusters (A-E) were identified with at least 5% nucleotide divergence in the viral protein 1 (VP1) coding region. Peshawar, Quetta, and Karachi were found to be the major endemic foci where multiple discrete genetic lineages of WPV1 were detected. Phylogenetic analysis suggests that wild poliovirus strains from endemic regions were genetically distant (with 5%-15% VP1 nucleotide divergence) from those detected in North Waziristan cases, excluding the possibility of a recent progenitor of WPV1 instigating single-source transmission across the country. Orphan lineages detected in Rawalpindi, Lahore, Hyderabad, Sukkur, and Jacobabad revealed silent transmission and the need for vigilant surveillance. Sustenance of analogous genetic lineages over a period of 3 years highlights multiple unimmunized foci present to maintain viral genetic diversity., Conclusions: Our findings are inconsistent with the hypothesis that impoverished populations from North Waziristan serve as a possible determinant of widespread poliomyelitis infection in Pakistan and further emphasize the need to scale-up clinical and environmental surveillance as well as immunization activities., (© The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.)

Enteroviruses are known to cause childhood paralysis. The purpose of this study was to examine the genetic diversity and to determine the association of non-polio enteroviruses (NPEVs) with acute flaccid Paralysis (AFP). Stool samples (n = 1191) of children with AFP were collected from Khyber Pakhtunkhwa and Federally Administered Tribal Areas of Pakistan. Poliovirus was isolated in 205 (17.2%) samples and NPEVs were found in 215 (18.0%) samples. Out of 215 viruses, 124 (57.7%) were typed into 19 different types of enteroviruses while 91 (42.3%) remained untypeable on microneutralization assay that were reconfirmed as NPEVs by real time PCR. Echovirus 19 (20/35; 57.1%) was found the most prevalent type based on VP1 nucleotide sequencing with increased genetic diversity. Phylogenetic analysis revealed the circulation of a new genotype of E-19 in the country. The findings of this study are of great importance for future research and propose to establish the enterovirus surveillance system in the country to readily identify more enteroviruses and to monitor the emergence of new variants/genotypes especially at the moment when we are at the verge of polio eradication phase.

Background: Crimean Congo hemorrhagic fever (CCHF) has been reported from more than 30 countries in Africa, Asia, Eastern Europe and Middle East. The disease is considered endemic in Pakistan and neighboring countries like Iran and Afghanistan., Objectives: This study aimed to explore the genetic diversity of CCHF virus (CCHFV) detected in Pakistan and Afghanistan based on analysis of partial S-segment sequences., Study Design: During 2011, one hundred samples satisfying the CCHF case definition were tested by (ELISA) and RT-PCR for detection of IgM antibodies and viral RNA, respectively. Phylogenetic analysis was carried out on partial S-segment nucleotide sequences using MEGA 5.0., Results: Out of one hundred collected during 2011, 49 (49%) were positive for CCHF either by ELISA/RT-PCR or both. The mean age of the CCHFV positive cases was 30.32 years (range 18-56 years) and overall mortality rate was 20.4%. All CCHF virus isolates from this study clustered with strains previously reported from Pakistan, Iran and Afghanistan within the Asia-1 genogroup. Four distinct sub-clades were found circulating within Asia-1 genogroup. Six CCHFV strains found in Pakistan and Afghanistan grouped into a new sub-clade-D., Conclusions: Data from this study shows that endemic foci of CCHFV span the international border between Pakistan and Afghanistan with genetically diverse variants circulating in this region. Our findings emphasize to establish a laboratory based surveillance program and devise health policy measures to control CCHF infection especially in Baluchistan., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Human Bocaviruses (HBoV) have been detected in human respiratory and gastrointestinal infections worldwide. Four genotypes of HBoV (HBoV1-4) have been described; HBoV-1 is associated with respiratory tract infections while HBoV-2, -3, and -4 genotypes are considered as entero-pathogenic although the exact role largely remains unclear. The global prevalence of HBoV has been reported, but the epidemiological data from Pakistan is largely unavailable to date. This study was conducted to understand the genetic diversity and disease prevalence of HBoV in hospitalized Pakistani children with acute diarrhea. During 2009, a total of 365 stool samples were collected from children hospitalized with gastrointestinal symptoms (as per WHO case definitions) at Rawalpindi General Hospital, Pakistan. Demographic and clinical data were recorded using a standardized questionnaire. The samples were tested for HBoV and rotavirus using real-time RT-PCR and ELISA, respectively. There were 47 (13%) samples positive for HBoV with 98% (n = 46) showing co-infection with rotavirus. HBoV-1 was the most frequently detected and was found in 94% samples followed by HBoV-2 and HBoV-3 genotypes. The mean age of infected children was 7.57 ± 5.4 months while detection was more frequent in males (n = 32, 68%). All cases recovered after 2.43 ± 1.0 mean days of treatment. On phylogenetic analysis, HBoV strains from Pakistan clustered closely with viruses from neighboring Bangladesh and China. These findings represent the first known epidemiological study in Pakistan to investigate the role of HBoV in acute gastroenteritis. The clinical data demonstrates that HBoV is not significantly associated with gastroenteritis alone and predominantly co-infections with rotavirus are found., (© 2015 Wiley Periodicals, Inc.)