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Background: COVID-19 epidemiology changed with the emergence of SARS-CoV-2 variants of concern (VOC). Pakistan administered mostly inactivated vaccines. We investigated the association between VOC and breakthrough infections in a mixed-vaccination-status population of Karachi. Methods: We investigated SARS-CoV-2 VOC tested in 392 respiratory specimens collected between May and December 2021. Data for age, sex, hospital admission, vaccinations, together with CT values of the diagnostic PCR test were analyzed. Results: The median age of COVID-19 cases tested was 40 (27−57) years and 43.4% were female. Delta variants were most common (56.4%) followed by Alpha (15.9%), Omicron (12.2%), Beta/Gamma (11.3%), and others (4.3%). Eighteen percent of cases were hospitalized whereby, predominant VOC were Beta/Gamma (40.8%), Alpha (35.2%) and Delta (22.5%). Overall, 55.4% of individuals were fully vaccinated, 7.4% were partially vaccinated and 37.2% were unvaccinated. Most (74.6%) inpatients were unvaccinated. Vaccines comprised inactivated (85.34%), single-shot vector (8.62%), two-shot vector (3.02%) and mRNA (3.02%) types. Omicron variants showed lower viral loads as compared to Alpha, Beta/Gamma, and Delta (p = 0.017). The risk of infection with Delta and Omicron variants was higher, 8 weeks after vaccination. The majority of those with breakthrough infections after receiving inactivated vaccines acquired COVID-19 within 4 months of vaccination. Conclusion: Our data highlights the shifting of VOC from Delta to Omicron during 2021 and that COVID-19 vaccinations reduced both hospitalizations and viral transmission. It informs on the increased risk of breakthrough infection within 8 weeks of vaccination, indicating the need for booster vaccinations.

Objective: To compare the effectiveness of Methotrexate (MTX) versus Cyclosporine (CyA) in the patients of moderate to severe atopic dermatitis (AD) Study Design: Systematic review and Meta-analysis. Methodology: After formulating our research team and PICO question rationally, a search strategy and literature search were carried out as per Cochrane guidelines. Full-text articles were retrieved after title and abstract screening per our inclusion criteria. The whole search process was documented in the form of a PRISMA flow chart. Team members made and filed data extraction sheets, and qualitative data synthesis was performed. Afterwards, a meta-analysis was carried out, involving a total of 03 randomized controlled trials. Revman was used to process the continuous and dichotomous data collected for our outcomes. RoB 2 was used to assess the risk of bias in the included studies. Grade pro-GDT was used to summarize the findings in the table. Results: Pooled results from 3 RCTs showed that CyA in a 2.5mg/kg dose is better than MTX @0.4 mg/kg/week at 12 weeks in AD. At follow-up, MTX maintains better disease control, i.e., until 24 weeks post-treatment. MTX has a better safety profile than CyA. Conclusion: MTX and CyA are both effective drugs for moderate to severe AD, but they have their pros and cons.

Abstract Chitinase-producing fungi have now engrossed attention as one of the potential agents for the control of insect pests. Entomopathogenic fungi are used in different regions of the world to control economically important insects. However, the role of fungal chitinases are not well studied in their infection mechanism to insects. In this study, Chitinase of entomopathogenic fungi Trichoderma longibrachiatum was evaluated to control Aphis gossypii. For this purpose, fungal chitinase (Chit1) gene from the genomic DNA of T. longibrachiatum were isolated, amplified and characterised. Genomic analysis of the amplified Chit1 showed that this gene has homology to family 18 of glycosyl hydrolyses. Further, Chit1 was expressed in the cotton plant for transient expression through the Geminivirus-mediated gene silencing vector derived from Cotton Leaf Crumple Virus (CLCrV). Transformed cotton plants showed greater chitinase activity than control, and they were resistant against nymphs and adults of A. gossypii. About 38.75% and 21.67% mortality of both nymphs and adults, respectively, were observed by using Chit1 of T. longibrachiatum. It is concluded that T. longibrachiatum showed promising results in controlling aphids by producing fungal chitinase in cotton plants and could be used as an effective method in the future.

Abstract Background COVID-19 waves caused by specific SARS-CoV-2 variants have occurred globally at different times. We focused on Omicron variants to understand the genomic diversity and phylogenetic relatedness of SARS-CoV-2 strains in various regions of Pakistan. Methods We studied 276,525 COVID-19 cases and 1,031 genomes sequenced from December 2021 to August 2022. Sequences were analyzed and visualized using phylogenetic trees. Results The highest case numbers and deaths were recorded in Sindh and Punjab, the most populous provinces in Pakistan. Omicron variants comprised 93% of all genomes, with BA.2 (32.6%) and BA.5 (38.4%) predominating. The first Omicron wave was associated with the sequential identification of BA.1 in Sindh, then Islamabad Capital Territory, Punjab, Khyber Pakhtunkhwa (KP), Azad Jammu Kashmir (AJK), Gilgit-Baltistan (GB) and Balochistan. Phylogenetic analysis revealed Sindh to be the source of BA.1 and BA.2 introductions into Punjab and Balochistan during early 2022. BA.4 was first introduced in AJK and BA.5 in Punjab. Most recent common ancestor (MRCA) analysis revealed relatedness between the earliest BA.1 genome from Sindh with Balochistan, AJK, Punjab and ICT, and that of first BA.1 from Punjab with strains from KPK and GB. Conclusions Phylogenetic analysis provides insights into the introduction and transmission dynamics of the Omicron variant in Pakistan, identifying Sindh as a hotspot for viral dissemination. Such data linked with public health efforts can help limit surges of new infections.

Routable Digital Maps represent a recent advancement driven by the growth of Intelligent Transportation System applications. Efforts have been made to develop and utilize maps within computer systems that can autonomously identify optimal routes. Typically, online map visualizations lack essential information such as traffic patterns, construction zones, or lane counts, which are crucial for designing accurate and realistic routes. This research introduces a route planning method that aggregates and reuses various data points to establish a robust framework for developing new routing plans. The primary goal is to analyze the collected data and create a framework for estimating a geometrically smooth centerline of a road map route. This method involves generating a series of parallel curves, known as offset curves, to facilitate the creation of new routes and alternative lanes. The following examples illustrate some empirical results obtained from this approach.

Abstract Background and Aims COVID‐19 morbidity and mortality varied globally through the pandemic. We studied the relationship of SARS‐CoV‐2 variants of concern (VOC) with COVID‐19 severity and mortality among hospitalized patients in Pakistan. Methods A retrospective review of clinical, laboratory, and vaccination data of 197 COVID‐19 adult patients at the Aga Khan University Hospital, Karachi between April 2021, and February 2022 was performed. SARS‐CoV‐2 VOC identified in respiratory samples were analyzed. Univariate and multivariate analysis was conducted to identify factors associated with COVID‐19 outcomes. Results The median age of cases was 55 years and 51.8% were males. Twenty‐four percent of females were pregnant. Of COVID‐19 cases, 48.2% had nonsevere disease, while 52.8% had severe/critical disease. Hypertension (48%) and diabetes mellitus (41%) were common comorbids. SARS‐CoV‐2 VOC identified comprised; Omicron (55.3%), Beta (14.7%), Alpha (13.7%), Delta (12.7%), and Gamma (3.6%) variants. Most (59.7%) study subjects were unvaccinated. Of vaccines, 88% had received inactivated virus COVID‐19 vaccines. Increased risk of severe disease was associated with age ≥50 years (odds ratio [OR]: 5.73; 95% confidence interval [CI]: [2.45–13.7]), as well as with diabetes mellitus (OR: 4.24; 95% CI: [1.82–9.85]). Full vaccination (OR: 0.25; 95% CI: [0.11–0.58]) or infection with Omicron (OR: 0.42; 95% CI: [0.23–0.74]) was associated with reduced disease severity. The risk of mortality increased with age ≥50 years (OR: 5.07; 95% CI: [1.92–13.42]) and a history of myocardial infarction (OR: 5.11; 95% CI: [1.45–17.93]) whilst, infection with Omicron was found to reduce the risk (OR: 0.22; 95% CI: [0.10–0.53]). Conclusion Our study describes the relationship between the severity of COVID‐19, in‐hospital mortality in relation to SARS‐CoV‐2 variants, and the impact of COVID‐19 vaccination in Pakistan. Outcomes were more favorable in younger individuals, after vaccinations and with Omicron variant infections. Most cases received inactivated virus vaccines therefore these data highlight the protection provided against severe COVID‐19.

Objective: To look for migraine headaches and associated factors in patients being managed for psoriasis at a tertiary healthcare setup in Rawalpindi, Pakistan. Study Design: Comparative cross-sectional study. Place and Duration of Study: Department of Dermatology, Pak Emirates Military Hospital, Rawalpindi Pakistan, from Mar 2021 to Feb 2022. Methodology: One hundred and forty patients of psoriasis being managed at our Dermatology Unit were included in the analysis. All patients were screened with the Migraine Screen Questionnaire (MS-Q), and those having scores equal to or more than four were evaluated by medical specialists, and diagnosis of migraine was established based on the International Classification of Headache Disorders diagnostic criteria for migraine. An association of relevant risk factors was established with migraine among these patients. Results: Out of 140 patients with psoriasis, 75(53.4%) were male, while 65(46.6%) were female. Eighty-five (60.7%) patients had migraine confirmed by clinical criteria, while 55(39.3%) had no migraine. Statistical analysis revealed that more body surface area involvement and longer duration of psoriasis had a statistically significant relationship with migraine among the study participants (p-value

Abstract Lower respiratory illness is one of the leading causes of death among children in low- and high-income countries. Human metapneumovirus (hMPV) is a key contributor to respiratory illnesses commonly reported among children and causes serious clinical complications ranging from mild respiratory infections to severe lower respiratory tract anomalies mainly in the form of bronchiolitis and pneumonia. However, due to the lack of a national surveillance system, the clinical significance of hMPV remains obscure in the Pakistani population. This study was conducted to screen throat swabs samples collected from 127 children reported with respiratory symptoms at a tertiary care hospital in Islamabad. Out of 127, 21 (16.5%) samples were positive for hMPV with its genotype distribution as A2a (10%), A2b (20%), B1 (10%), and B2 (60%). Phylogenetic analysis showed that the hMPV viruses were closely related to those reported from neighboring countries including India and China. This work will contribute to a better understanding of this virus, its diagnosis, and the handling of patients in clinical setups. Further studies at a large-scale are warranted for a better understanding of the disease burden and epidemiology of hMPV in Pakistan.

Objective: To compare the efficacy of double-dose oral Terbinafine and Itraconazole in treating dermatophytic infections. Study Design: Randomized controlled trial (ClinicalTrials.gov: NCT04880980). Place and Duration of Study: Department of Dermatology, Pak Emirates Military Hospital, Rawalpindi Pakistan, from Mar 2021 to Mar 2022. Methodology: One hundred and twenty patients with dermatophyte infections of the skin (i.e., tinea corporis and cruris) diagnosed by clinical presentation and KOH mount were included in this study. After randomization, patients were divided into two groups. Group-A was managed with double-dose oral Terbinafine, while Group-B was managed with double-dose oral Itraconazole. Clinical response and side effects were seen and recorded initially at two weeks and then at four weeks.Efficacy and adverse effects were compared in both groups at the end of four weeks. Results: Out of 120 patients with dermatophyte infections of the skin included in the study, 59(49.2%) took double-dose oral Terbinafine while 61(50.8%) took double-dose oral Itraconazole after randomization. It was revealed that Itraconazole was more efficacious in achieving cure at the end of 4-weeks as compared to double dose Terbinafine (p-value=0.001), while adverse effects studied were not statistically significant or different in both the groups (p-value>0.005). Conclusion: This randomized controlled trial showed that double dose oral Itraconazole was more efficacious in treating dermatophyte infections of the skin (i.e., Tinea corporis and cruris) compared to double dose oral Terbinafine. Adverse effects were minimal in both groups and were not specially related to any of these medications.

COVID-19 resulted in extensive morbidity and mortality worldwide. SARS-CoV-2 evolved rapidly, with increasing transmission due to Variants of Concern (VOC). Identifying VOC became important but genome submissions from low-middle income countries (LMIC) remained low leading to gaps in genomic epidemiology. We demonstrate the use of a specific mutation RT-PCR based approach to identify VOC in SARS-CoV-2 positive samples through the pandemic in Pakistan. We selected 2150 SARS-CoV-2 PCR positive respiratory specimens tested between April 2021 and February 2022, at the Aga Khan University Hospital Clinical Laboratories, Karachi, Pakistan. Commercially available RT-PCR assays were used as required for mutations in Spike protein (N501Y, A570D, E484K, K417N, L452R, P681R and deletion69_70) to identify Alpha, Beta, Gamma, Delta, and Omicron variants respectively. Three pandemic waves associated with Alpha, Delta and Omicron occurred during the study period. Of the samples screened, VOC were identified in 81.7% of cases comprising mainly; Delta (37.2%), Alpha (29.8%) and Omicron (17.1%) variants. During 2021, Alpha variants were predominant in April and May; Beta and Gamma variants emerged in May and peaked in June; the Delta variant peaked in July and remained predominant until November. Omicron (BA.1) emerged in December 2021 and remained predominant until February 2022. The CT values of Alpha, Beta, Gamma and Delta were all significantly higher than that of Omicron variants (p

Plant disease management using nanotechnology is evolving continuously across the world. The purpose of this study was to determine the effect of different concentrations of green synthesized zinc oxide nanoparticles (ZnO NPs) using Trachyspermum ammi seed extract on Cercospora leaf spot disease in mung bean plants under in-vitro and in-planta conditions. Additionally, the effects on mung bean agronomic and physiological parameters were also assessed. The green synthesized ZnO NPs were characterized using UV-visible spectroscopy, Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and Scanning electron microscopy (SEM). Green synthesized NPs were tested for their ability to inhibit fungal growth at five different concentrations under in-vitro experiment. After 7 days of inoculation, ZnO NPs (1200 ppm) inhibited mycelial growth substantially (89.86% ± 0.70). The in-planta experiment showed statistically significant result of disease control (30% ± 11.54) in response to 1200 ppm ZnO NPs. The same treatment showed statistically significant improvements in shoot length, root length, number of leaves, number of pods, shoot fresh weight (28.62%), shoot dry weight (85.18%), root fresh weight (38.88%), and root dry weight (38.88%) compared to the control. Our findings show that green synthesized ZnO NPs can control Cercospora canescens in mung bean, pointing to their use in plant disease control and growth enhancement.

Objective: To identify electrocardiographic changes associated with parenteral meglumine antimoniate and to estimate frequency of these changes. Study Design: Prospective observational study. Place and Duration of Study: Dermatology department, Combined Military Hospital Quetta and Multan, from Apr 2018 to Feb 2019. Methodology: Total of 87 patients was recruited by consecutive sampling technique. Cases of Leishmaniasis were confirmed by Leishmania donovani bodies in smear or skin biopsy. Base line electrocardiograph was done and then repeated weekly. All patients were given 15mg/kg body weight intramuscular meglumine antimoniate (Glucantime). Any change in electrocardiograph was documented. Follow up electrocardiograph was repeated two weeks after hospital discharge. Data was analyzed using SPSS-20. Results: Changes in electrocardiograph appeared between 7-27 days mean 16 SD ± 4.58. T-wave inversion was recorded in 41 (47.12%), ST elevation in 1 (1.14%), prolonged QT interval in 1 (1.14%). Meglumine antimoniate therapy was discontinued after ECG change for 5 ± SD 3 days of therapy. Follow up electrocardiograph two weeks after hospital discharge was normal. Conclusion: Early repolarization defects of T wave inversion and ST segment deviation were found with standard doses of meglumine antimoniate therapy.

Objective: To determine population-based estimates of coronavirus disease 2019 (COVID-19) in a densely populated urban community of Karachi, Pakistan. Methods: Three cross-sectional surveys were conducted in April, June and August 2020 in low- and high-transmission neighbourhoods. Participants were selected at random to provide blood for Elecsys immunoassay for detection of anti-severe acute respiratory syndrome coronavirus-2 antibodies. A Bayesian regression model was used to estimate seroprevalence after adjusting for the demographic characteristics of each district. Results: In total, 3005 participants from 623 households were enrolled in this study. In Phase 2, adjusted seroprevalence was estimated as 8.7% [95% confidence interval (CI) 5.1–13.1] and 15.1% (95% CI 9.4–21.7) in low- and high-transmission areas, respectively, compared with 0.2% (95% CI 0–0.7) and 0.4% (95% CI 0–1.3) in Phase 1. In Phase 3, it was 12.8% (95% CI 8.3–17.7) and 21.5% (95% CI 15.6–28) in low- and high-transmission areas, respectively. The conditional risk of infection was 0.31 (95% CI 0.16–0.47) and 0.41 (95% CI 0.28–0.52) in low- and high-transmission neighbourhoods, respectively, in Phase 2. Similar trends were observed in Phase 3. Only 5.4% of participants who tested positive for COVID-19 were symptomatic. The infection fatality rate was 1.66%, 0.37% and 0.26% in Phases 1, 2 and 3, respectively. Conclusion: Continuing rounds of seroprevalence studies will help to improve understanding of secular trends and the extent of infection during the course of the pandemic.

Background: Data on the viral etiology of acute lower respiratory infections are scarce in Pakistan. Human respiratory syncytial virus (RSV) is an important cause of morbidity in children but no effective vaccine or antiviral therapy is currently available. As vaccines are expected to become available in the future, it is important to understand the epidemiology of locally prevalent RSV subtypes. This study aimed to define the molecular epidemiology of RSV (A and B) genotypes in Pakistani children under 5 years. Methods: World Health Organization case definitions for influenza-like illness (ILI) and severe acute respiratory illness (SARI) were used for case selection. Children under 5 years who presented with ILI or SARI at tertiary care hospitals from all provinces/regions, including the eight influenza sentinel sites, during October–April each year between 2010 and 2013 were enrolled. Demographic and clinical data of the children were recorded and nasopharyngeal/throat swabs taken for analysis. All samples were tested for RSV A and B using real-time polymerase chain reaction for non-influenza respiratory viruses. Specific oligonucleotide primers for RSV A and B were used for subtyping and sequencing of the G protein, followed by phylogenetic analysis. Results: A total of 1941 samples were included. RSV was detected in 472 (24%) children, with RSV A detected in 367 (78%) and RSV B in 105 (22%). The G protein of all RSV A strains clustered in the NA1/GA2 genotype while RSV B strains carried the signature 60 nucleotide duplication and were assigned to three BA genotypes: BA-9, BA-10 and the new BA-13 genotype. Conclusions: This study highlights the importance of RSV as a viral etiologic agent of acute respiratory infections in children in Pakistan, and the diversity of RSV viruses. Continued molecular surveillance for early detection of prevalent and newly emerging genotypes is needed to understand the epidemiology of RSV infections in Pakistan. Keywords: Respiratory syncytial virus, Human, Child, Molecular epidemiology, Genotype, Pakistan

Let G be a simple connected graph. The second Gourava index of graph G is defined as GO2G=∑θϑ∈EGdθ+dϑdθdϑ where dθ denotes the degree of vertex θ. If removal of a vertex of G forms a tree, then G is called an apex tree. Let L⊂VG with ∣L∣=q. If removal of L from VG forms a tree and any other subset of VG whose cardinality is less than ∣L∣ does not form a tree, then G is known as q-apex tree. In this paper, we have calculated upper bound for 2nd Gourava index with respect to q-apex trees.

In this paper, the extended simple equation method (ESEM) and the generalized Riccati equation mapping (GREM) method are applied to the nonlinear third-order Gilson–Pickering (GP) model to obtain a variety of new exact wave solutions. With the suitable selection of parameters involved in the model, some familiar physical governing models such as the Camassa–Holm (CH) equation, the Fornberg–Whitham (FW) equation, and the Rosenau–Hyman (RH) equation are obtained. The graphical representation of solutions under different constraints shows the dark, bright, combined dark–bright, periodic, singular, and kink soliton. For the graphical representation, 3D plots, contour plots, and 2D plots of some acquired solutions are illustrated. The obtained wave solutions motivate researchers to enhance their theories to the best of their capacities and to utilize the outcomes in other nonlinear cases. The executed methods are shown to be practical and straightforward for approaching the considered equation and may be utilized to study abundant types of NLEEs arising in physics, engineering, and applied sciences.

Background: The Middle East and North Africa (MENA) region faces a dual challenge with regard to influenza infection due to severe zoonotic influenza outbreaks episodes and the circulation of Northern Hemisphere human influenza viruses among pilgrims. Methods: The MENA Influenza Stakeholder Network (MENA-ISN) was set-up with the aim of increasing seasonal influenza vaccination coverage by (i) enhancing evidence-based exchanges, and (ii) increasing awareness on the safety and benefits of seasonal vaccination. During the 7th MENA-ISN meeting, representatives from 8 countries presented their influenza surveillance, vaccination coverage and actions achieved and provided a list of country objectives for the upcoming 3 years. Results: MENA-ISN countries share the goal to reduce influenza related morbidity and mortality. Participants admitted that lack of knowledge about influenza, its consequences in terms of morbidity, mortality and economy are the major barrier to attaining higher influenza vaccination coverage in their countries. The cost of the vaccine is another key barrier that could contribute to low vaccination coverage. Participants drew a list of strategic interventions to bridge gaps in the knowledge of influenza burden in this region. Conclusions: Participating countries concluded that despite an increase in vaccine uptake observed during the last few years, influenza vaccination coverage remains relatively low. Priority areas should be identified and action plans tailored to each country situation set-up to investigate the best way to move forward. Keywords: Influenza, Vaccination coverage, Surveillance, Middle East and North Africa

BackgroundInfluenza is known to have a specific pattern of seasonality the reasons for which are yet to be fully ascertained. Temperate zones show influenza epidemic during the winter months. The tropical and subtropical regions show more diverse influenza outbreak patterns. This study explores the seasonality of influenza activity and predicts influenza peak based on historical surveillance time series data in Islamabad and Multan, Pakistan.MethodsThis is a descriptive study of routinely collected monthly influenza sentinel surveillance data and meteorological data from 2012-16 in two sentinel sites of Pakistan: Islamabad (North) and Multan (Central).ResultsMean number of cases of influenza and levels of precipitation were higher in Islamabad compared to Multan. Mean temperature and humidity levels were similar in both the cities. The number of influenza cases rose with decrease in precipitation and temperature in Islamabad during 2012-16, although the same cannot be said about humidity. The relationship between meteorological parameters and influenza incidence was not pronounced in case of Multan. The forecasted values in both the cities showed a significant peak during the month of January.ConclusionThe influenza surveillance system gave a better understanding of the disease trend and could accurately forecast influenza activity in Pakistan.

Despite substantial interventions in the understanding and case management of acute gastroenteritis, diarrheal diseases are still responsible for a notable amount of childhood deaths. Although the rotavirus is known to cause a considerable burden of pediatric diarrheal cases, the roles of other viruses remain undefined for the Pakistani population. This study was based on tertiary care hospital surveillance, from January 2009 to December 2010, including the detection of rotavirus, norovirus, astrovirus, and human parechovirus in children under the age of five using serological or molecular assays. Rotavirus, human parechovirus, norovirus, and astrovirus were detected in 66%, 21%, 19.5%, and 8.5% subjects, respectively. Human parechovirus genotypes, determined through analysis of VP1 gene sequences, showed a great diversity among co-circulating strains. Eighty percent of hospitalized children had dual or multiple viral infections, while 98% parechovirus positive cases were co-infected with rotavirus. The remarkable diversity of viruses associated with the childhood diarrhea in Pakistan calls for large-scale epidemiological surveys, coupled with case control studies, to ascertain their role in clinical manifestations. In addition, these findings also highlight the need for the implementation of up-to-date health interventions, such as the inclusion of a rotavirus vaccine in routine immunization programs for the improvement of quality in child health care.

Background: Respiratory syncytial virus (RSV) infection is an important cause of pneumonia mortality in young children. However, clinical data for fatal RSV infection are scarce. We aimed to identify clinical and socioeconomic characteristics of children aged younger than 5 years with RSV-related mortality using individual patient data. Methods: In this retrospective case series, we developed an online questionnaire to obtain individual patient data for clinical and socioeconomic characteristics of children aged younger than 5 years who died with community-acquired RSV infection between Jan 1, 1995, and Oct 31, 2015, through leading research groups for child pneumonia identified through a comprehensive literature search and existing research networks. For the literature search, we searched PubMed for articles published up to Feb 3, 2015, using the key terms “RSV”, “respiratory syncytial virus”, or “respiratory syncytial viral” combined with “mortality”, “fatality”, “death”, “died”, “deaths”, or “CFR” for articles published in English. We invited researchers and clinicians identified to participate between Nov 1, 2014, and Oct 31, 2015. We calculated descriptive statistics for all variables. Findings: We studied 358 children with RSV-related in-hospital death from 23 countries across the world, with data contributed from 31 research groups. 117 (33%) children were from low-income or lower middle-income countries, 77 (22%) were from upper middle-income countries, and 164 (46%) were from high-income countries. 190 (53%) were male. Data for comorbidities were missing for some children in low-income and middle-income countries. Available data showed that comorbidities were present in at least 33 (28%) children from low-income or lower middle-income countries, 36 (47%) from upper middle-income countries, and 114 (70%) from high-income countries. Median age for RSV-related deaths was 5·0 months (IQR 2·3–11·0) in low-income or lower middle-income countries, 4·0 years (2·0–10·0) in upper middle-income countries, and 7·0 years (3·6–16·8) in high-income countries. Interpretation: This study is the first large case series of children who died with community-acquired RSV infection. A substantial proportion of children with RSV-related death had comorbidities. Our results show that perinatal immunisation strategies for children aged younger than 6 months could have a substantial impact on RSV-related child mortality in low-income and middle-income countries. Funding: Bill & Melinda Gates Foundation.

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.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.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.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.

BACKGROUND: Influenza is a viral infection that can lead to serious complications and death(s) in vulnerable groups if not diagnosed and managed in a timely manner. This study was conducted to improve the accuracy of predicting influenza through various clinical and statistical models. METHODOLOGY: A retrospective cross sectional analysis was done on demographic and epidemiological data collected from March 2009 to March 2010. Patients were classified as ILI or SARI using WHO case definitions. Respiratory specimens were tested by RT-PCR. Clinical symptoms and co-morbid conditions were analyzed using binary logistic regression models. RESULTS: In the first approach, analysis compared children (≤12) and adults (>12). Of 1,243 cases, 262 (21%) tested positive for A(H1N1)pdm09 and the proportion of children (≤12) and adults (>12) were 27% and 73% respectively. Four symptoms predicted influenza in children: fever (OR 2.849, 95% CI 1.931-8.722), cough (OR 1.99, 95% CI 1.512-3.643), diarrhea (OR 2.100, 95% CI 2.040-3.25) and respiratory disease (OR 3.269, 95% CI 2.128-12.624). In adults, the strongest clinical predictor was fever (OR 2.80, 95% CI 1.025-3.135) followed by cough (OR 1.431, 95% CI 1.032-2.815). In the second instance, patients were separated into two groups: SARI 326 (26%) and ILI 917 (74%) cases. Male to female ratio was 1.41∶1.12 for SARI and 2∶1.5 for ILI cases. Chi-square test showed that fever, cough and sore throat were significant factors for A(H1N1)pdm09 infections (p = 0.008). CONCLUSION: Studies in a primary care setting should be encouraged focused on patients with influenza-like illness to develop sensitive clinical case definition that will help to improve accuracy of detecting influenza infections. Formulation of a standard "one size fits all" case definition that best correlates with influenza infections can help guide decisions for additional diagnostic testing and also discourage unjustified antibiotic prescription and usage in clinical practice.

Respiratory syncytial virus (RSV) is the major cause of acute lower respiratory tract infections in young children, but very little is known about its epidemiology and circulating genotypes in Pakistan. This study analyzed the epidemiological and molecular characteristics of RSV genotypes detected in Pakistani children less than 2 years of age with acute respiratory tract infections (ARIs) in a tertiary care hospital in Gilgit Baltistan (GB) province during 2011-12 winter season. RSV was detected in 75 out of 105 children presenting with acute respiratory infection. Male infants between 2-6 months age made up the highest percentage of RSV positive cases. Epidemiological factors such as pre-maturity, mean weight, clinical features and diagnosis when compared between RSV positive and negative groups were found to be statistically insignificant. Phylogenetic analysis classified all 75 of the RSV strains into 71 strains of subgroups A and 4 strains of subgroup B, respectively. Strains belonging to subgroups A and B were further subdivided into NA1/GA2 and BA, respectively. The nucleotide and deduced amino acid sequence identities were relatively high among these strains (>90%). Both RSV-A and RSV-B isolates had two potential N-glycosylation sites in HVR2 of G protein and with heavy O-glycosylation of serine and threonine residues (G scores of 0.5-0.7). This report highlights the significance of RSV as a dominant viral etiologic agent of pediatric ARIs, and need for continued molecular epidemiological surveys for early detection of prevalent strains and newly emerging genotypes to understand epidemiology of RSV infections in various regions of Pakistan.

Pakistan harbors high disease burden of gastro-enteric infections with majority of these caused by rotavirus. Unfortunately, lack of proper surveillance programs and laboratory facilities have resulted in scarcity of available data on rotavirus associated disease burden and epidemiological information in the country. We investigated 1306 stool samples collected over two years (2008-2009) from hospitalized children under 5 years of age for the presence of rotavirus strains and its genotypic diversity in Lahore. The prevalence rate during 2008 and 2009 was found to be 34% (n = 447 out of 1306). No significant difference was found between different age groups positive for rotavirus (p>0.05). A subset of EIA positive samples was further screened for rotavirus RNA through RT-PCR and 44 (49.43%) samples, out of total 89 EIA positive samples, were found positive. G and P type prevalence was found as follows: G1P [4] = 3(6.81%); G1P [6] = 9(20.45%); G1P [8] = 1(2.27%); G2P [4] = 21(47.72%); G2P [8] = 1(2.27%); G9P [4] = 1(2.27%); G9P [6] = 1(2.27%) and G9P [8] = 7(15.90%). Phylogenetic analysis revealed that the VP7 and VP4 sequences clustered closely with the previously detected strains in the country as well as Belgian rotaviruses. Antigenic characterization was performed by analyzing major epitopes in the immunodominant VP7 and VP4 gene segments. Although the neutralization conferring motifs were found variable between the Pakistani strains and the two recommended vaccines strains (Rotarix™ and RotaTeq™), we validate the use of rotavirus vaccine in Pakistan based on the proven and recognized vaccine efficacy across the globe. Our findings constitute the first report on rotavirus' genotype diversity, their phylogenetic relatedness and epidemiology during the pre-vaccination era in Lahore, Pakistan and support the immediate introduction of rotavirus vaccine in the routine immunization program of the country.

This paper describes the use of trigonometric spline to visualize the given planar data. The goal of this work is to determine the smoothest possible curve that passes through its data points while simultaneously satisfying the shape preserving features of the data. Positive, monotone, and constrained curve interpolating schemes, by using a C1 piecewise rational cubic trigonometric spline with four shape parameters, are developed. Two of these shape parameters are constrained and the other two are set free to preserve the inherited shape features of the data as well as to control the shape of the curve. Numerical examples are given to illustrate the worth of the work.

There is little information about influenza among the Pakistani population. In order to assess the trends of Influenza-like-Illness (ILI) and to monitor the predominant circulating strains of influenza viruses, a country-wide lab-based surveillance system for ILI and Severe Acute Respiratory Illness (SARI) with weekly sampling and reporting was established in 2008. This system was necessary for early detection of emerging novel influenza subtypes and timely response for influenza prevention and control.Five sentinel sites at tertiary care hospitals across Pakistan collected epidemiological data and respiratory samples from Influenza-like illness (ILI) and severe acute respiratory illness (SARI) cases from January 2008 to December 2011. Samples were typed and sub-typed by Real-Time RT-PCR assay.A total of 6258 specimens were analyzed; influenza virus was detected in 1489 (24%) samples, including 1066 (72%) Influenza type A and 423 (28%) influenza type B viruses. Amongst influenza A viruses, 25 (2%) were seasonal A/H1N1, 169 (16%) were A/H3N2 and 872 (82 %) were A(H1N1)pdm09. Influenza B virus circulation was detected throughout the year along with few cases of seasonal A/H1N1 virus during late winter and spring. Influenza A/H3N2 virus circulation was mainly observed during summer months (August-October).The findings of this study emphasize the need for continuous and comprehensive influenza surveillance. Prospective data from multiple years is needed to predict seasonal trends for vaccine development and to further fortify pandemic preparedness.

BackgroundIn early 2009, a novel influenza A(H1N1) virus that emerged in Mexico and United States rapidly disseminated worldwide. The spread of this virus caused considerable morbidity with over 18000 recorded deaths. The new virus was found to be a reassortant containing gene segments from human, avian and swine influenza viruses.Methods/resultsThe first case of human infection with A(H1N1)pdm09 in Pakistan was detected on 18(th) June 2009. Since then, 262 laboratory-confirmed cases have been detected during various outbreaks with 29 deaths (as of 31(st) August 2010). The peak of the epidemic was observed in December with over 51% of total respiratory cases positive for influenza. Representative isolates from Pakistan viruses were sequenced and analyzed antigenically. Sequence analysis of genes coding for surface glycoproteins HA and NA showed high degree of high levels of sequence identity with corresponding genes of regional viruses circulating South East Asia. All tested viruses were sensitive to Oseltamivir in the Neuraminidase Inhibition assays.ConclusionsInfluenza A(H1N1)pdm09 viruses from Pakistan form a homogenous group of viruses. Their HA genes belong to clade 7 and show antigenic profile similar to the vaccine strain A/California/07/2009. These isolates do not show any amino acid changes indicative of high pathogenicity and virulence. It is imperative to continue monitoring of these viruses for identification of potential variants of high virulence or drug resistance.

Background and Aims: COVID‐19 morbidity and mortality varied globally through the pandemic. We studied the relationship of SARS‐CoV‐2 variants of concern (VOC) with COVID‐19 severity and mortality among hospitalized patients in Pakistan. Methods: A retrospective review of clinical, laboratory, and vaccination data of 197 COVID‐19 adult patients at the Aga Khan University Hospital, Karachi between April 2021, and February 2022 was performed. SARS‐CoV‐2 VOC identified in respiratory samples were analyzed. Univariate and multivariate analysis was conducted to identify factors associated with COVID‐19 outcomes. Results: The median age of cases was 55 years and 51.8% were males. Twenty‐four percent of females were pregnant. Of COVID‐19 cases, 48.2% had nonsevere disease, while 52.8% had severe/critical disease. Hypertension (48%) and diabetes mellitus (41%) were common comorbids. SARS‐CoV‐2 VOC identified comprised; Omicron (55.3%), Beta (14.7%), Alpha (13.7%), Delta (12.7%), and Gamma (3.6%) variants. Most (59.7%) study subjects were unvaccinated. Of vaccines, 88% had received inactivated virus COVID‐19 vaccines. Increased risk of severe disease was associated with age ≥50 years (odds ratio [OR]: 5.73; 95% confidence interval [CI]: [2.45–13.7]), as well as with diabetes mellitus (OR: 4.24; 95% CI: [1.82–9.85]). Full vaccination (OR: 0.25; 95% CI: [0.11–0.58]) or infection with Omicron (OR: 0.42; 95% CI: [0.23–0.74]) was associated with reduced disease severity. The risk of mortality increased with age ≥50 years (OR: 5.07; 95% CI: [1.92–13.42]) and a history of myocardial infarction (OR: 5.11; 95% CI: [1.45–17.93]) whilst, infection with Omicron was found to reduce the risk (OR: 0.22; 95% CI: [0.10–0.53]). Conclusion: Our study describes the relationship between the severity of COVID‐19, in‐hospital mortality in relation to SARS‐CoV‐2 variants, and the impact of COVID‐19 vaccination in Pakistan. Outcomes were more favorable in younger individuals, after vaccinations and with Omicron variant infections. Most cases received inactivated virus vaccines therefore these data highlight the protection provided against severe COVID‐19. [ABSTRACT FROM AUTHOR]