Concurrent Typhoid Fever and Dengue Hemorrhagic Fever: A Case Report.

Dengue virus can co-infect with a number of viruses, bacteria, and parasites of which dengue malaria co-infection is most well-known. We report a rare case of dengue virus co-infection with typhoid fever and the development of dengue hemorrhagic fever (DHF) during a dengue outbreak. The second spike of high-grade fever following initial defervescence with antibiotic therapy, hemorrhagic manifestations, new onset leucopenia and thrombocytopenia, and evidence of plasma leakage raised suspicion of DHF. Diagnosis of dengue co-infection was made by seroconversion for anti-dengue immunoglobulin M (IgM) antibodies by enzyme-linked immunosorbent assay (ELISA) on the seventh day of new-onset fever. Early recognition and judicious use of fluid therapy prevented the patient from developing shock and its complications. Prompt diagnosis, early recognition of plasma leakage, and appropriate management of DHF can reduce morbidity and mortality.

Utility of teleconsultation in accessing eye care in a developing country during COVID-19 pandemic.

OBJECTIVE: To evaluate the utility of teleconsultation in the provision of eye care services during the COVID-19 lockdown. Disparities in the consultation burden of sub-specialities and socio-demographic differences in teleconsultation utilization were also assessed. METHODS: Al-Shifa Trust Eye Hospital Rawalpindi began audio and video teleconsultation using broadband telecommunication services during the lockdown. Patients' and consultations' data gathered during the first three weeks after the commencement of this programme were compared with data from the four weeks prior to lockdown. The weekly consultation ratio and overall consultation burden of sub-specialities were measured. Chi-Square tests of association determined the relationship between different variables (socioeconomic status and consultation characteristics) and consultation modality (on-site vs online). RESULTS: In total, 17507 on-site consultations (4377/week) were conducted compared to 1431 teleconsultations (477/week), which maintained 10.89% of the weekly pre-lockdown eye care services. The post-lockdown teleconsultation programme saw a relatively higher percentage of service utility among female (47.09% vs 44.71%), younger-age (31.33±19.45 vs 41.25±23.32 years) and higher-socioeconomic-status (32.21% vs 0.30%) patients compared to pre-lockdown on-site consultations. The most common indication for teleconsultation was red-eye (16.70%). While cornea and glaucoma clinics maintained most of the pre-lockdown services (30.42% and 29% respectively), the highest dropout was seen in optometric and vitreoretinal services supporting only 5.54% and 8.28% of pre-lockdown services, respectively. CONCLUSION: Digital initiatives could partially maintain eye care services during the lockdown. Focused strategies to improve teleconsultation utilization are required during the pandemic and beyond.

A missense allele of PEX5 is responsible for the defective import of PTS2 cargo proteins into peroxisomes.

Peroxisomes, single-membrane intracellular organelles, play an important role in various metabolic pathways. The translocation of proteins from the cytosol to peroxisomes depends on peroxisome import receptor proteins and defects in peroxisome transport result in a wide spectrum of peroxisomal disorders. Here, we report a large consanguineous family with autosomal recessive congenital cataracts and developmental defects. Genome-wide linkage analysis localized the critical interval to chromosome 12p with a maximum two-point LOD score of 4.2 (θ = 0). Next-generation exome sequencing identified a novel homozygous missense variant (c.653 T > C; p.F218S) in peroxisomal biogenesis factor 5 (PEX5), a peroxisome import receptor protein. This missense mutation was confirmed by bidirectional Sanger sequencing. It segregated with the disease phenotype in the family and was absent in ethnically matched control chromosomes. The lens-specific knockout mice of Pex5 recapitulated the cataractous phenotype. In vitro import assays revealed a normal capacity of the mutant PEX5 to enter the peroxisomal Docking/Translocation Module (DTM) in the presence of peroxisome targeting signal 1 (PTS1) cargo protein, be monoubiquitinated and exported back into the cytosol. Importantly, the mutant PEX5 protein was unable to form a stable trimeric complex with peroxisomal biogenesis factor 7 (PEX7) and a peroxisome targeting signal 2 (PTS2) cargo protein and, therefore, failed to promote the import of PTS2 cargo proteins into peroxisomes. In conclusion, we report a novel missense mutation in PEX5 responsible for the defective import of PTS2 cargo proteins into peroxisomes resulting in congenital cataracts and developmental defects.

Combination of preconditioned adipose-derived mesenchymal stem cells and platelet-rich plasma improves the repair of osteoarthritis in rat.

Aim: To observe the combined effect of platelet-rich plasma (PRP) and preconditioned adipose-derived mesenchymal stem cells (ADMSCs) on the injured articular cartilage of the rat. Materials & methods: Animals in the study received an intra-articular injection of PRP and preconditioned ADMSCs, both in combination and separately. The response to therapeutic intervention was evaluated by inflammatory markers, proteoglycans content, chondrogenesis and gene expression analyses. Results: The combined therapy resulted in a reduction of IL-6 and TNF-α, increased proteoglycan content of the articular cartilage, upregulation of Acan, Col2a1 and PCNA genes. Downregulation of Col1a1, Col10a1 and Casp3 genes was observed as compared with the untreated osteoarthritis rat model. Conclusion: PRP potentiates the effects of ADMSCs on the repair of damaged articular cartilage.

Echocardiographic Evaluation of Left Atrial Volume Index in Patients with Chronic Kidney Disease.

Chronic kidney disease (CKD) patients are at high risk of developing cardio vascular disease. Left atrial volume index (LAVi) is an indicator of left ventricular diastolic dysfunction. We conducted this study to find out the correlation of LAVi and other echocardio- graphic parameters with estimated glomerular filtration rate (eGFR). We prospectively enrolled 170 individuals: 69 patients with CKD and 101 controls. Echocardiographic parameters including systolic and diastolic volumes of left ventricle, LAVi, ejection fraction (EF), pulmonary artery systolic pressure (PASP), and E/e ratio were measured in all participants. The demographic, clinical, and echocardiographic parameters were examined. From the total of 170 individuals, 69 (40.5%) patients had CKD and 101 (59.5%) had normal renal profile. There were 38 (55.07%) males in the CKD group and 71 (70.29%) in the control group. Patients with CKD had higher median LAVi [33.33 mL/m2 ± 11.71 vs. 22.54 mL/m2 ± 5.82; P < 0.001], higher median E/e ratio [10.41 ± 6.28 vs. 7.48 ± 2.28; P < 0.001], higher median PASP [42.47 ± 13.64 vs. 33.59 ± 12.51; P < 0.001], and lower median EF [52.79% ± 14.37 vs. 60.7% ± 8; P < 0.001]. There was a statistically significant negative correlation of eGFR with LAVi (r = -0.515, P < 0.001), PASP (r = -0.44, P = 0.001), and E/e ratio (r = -0.331, P = 0.001). Patients with CKD have higher LAVi, PASP, and E/e ratio and lower EF as compared to individuals without CKD. There is a significant negative correlation between eGFR and LAVi.

Mutations in CERKL and RP1 cause retinitis pigmentosa in Pakistani families.

This study was conducted to identify the genetic basis of retinal dystrophies in consanguineous Pakistani families. We recruited two families with retinitis pigmentosa (RP) displaying visual difficulties, including nyctalopia and constricted visual fields. Linkage analysis and Sanger sequencing resulted in the identification of a previously reported nonsense mutation, c.847C > T, in exon 5 of CERKL in one family and a novel four-base pair deletion in exon 4 of RP1, c.delAGAA4218_4221, leading to premature protein termination in the second family. Here, we report two RP-causing mutations extending the genetic heterogeneity of the disease.

Bi-allelic Variants in METTL5 Cause Autosomal-Recessive Intellectual Disability and Microcephaly.

Intellectual disability (ID) is a genetically and clinically heterogeneous disorder, characterized by limited cognitive abilities and impaired adaptive behaviors. In recent years, exome sequencing (ES) has been instrumental in deciphering the genetic etiology of ID. Here, through ES of a large cohort of individuals with ID, we identified two bi-allelic frameshift variants in METTL5, c.344_345delGA (p.Arg115Asnfs∗19) and c.571_572delAA (p.Lys191Valfs∗10), in families of Pakistani and Yemenite origin. Both of these variants were segregating with moderate to severe ID, microcephaly, and various facial dysmorphisms, in an autosomal-recessive fashion. METTL5 is a member of the methyltransferase-like protein family, which encompasses proteins with a seven-beta-strand methyltransferase domain. We found METTL5 expression in various substructures of rodent and human brains and METTL5 protein to be enriched in the nucleus and synapses of the hippocampal neurons. Functional studies of these truncating variants in transiently transfected orthologous cells and cultured hippocampal rat neurons revealed no effect on the localization of METTL5 but alter its level of expression. Our in silico analysis and 3D modeling simulation predict disruption of METTL5 function by both variants. Finally, mettl5 knockdown in zebrafish resulted in microcephaly, recapitulating the human phenotype. This study provides evidence that biallelic variants in METTL5 cause ID and microcephaly in humans and highlights the essential role of METTL5 in brain development and neuronal function.

Cardiovascular Autonomic Neuropathy and its Association with Cardiovascular and All-cause Mortality in Patients with End-stage Renal Disease.

Background End-stage renal disease frequently leads to increased cardiovascular mortality. Cardiovascular autonomic neuropathy (CAN) may be predictive of cardiac arrhythmias and sudden cardiac death in patients with end-stage renal disease. Methods A total of 70 patients with end-stage renal disease were included in the study. The assessment of cardiac dysautonomia was based on the four standardized tests performed at the baseline and, again, at the end of the study. The criteria for CAN included at least two abnormal test results. Results Fifty of 70 patients completed the study and were followed-up after one year. Out of the 50 patients, 44 (88%) had CAN at baseline. Twelve (24%) patients died at the one-year follow-up. Sudden cardiac death was reported in seven out of 12 (58%) patients. All seven patients who died had high dysautonomia scores (three abnormal tests) at the baseline. There was a significantly higher percentage of patients with all four abnormal tests amongst patients who died of any cause (56% vs. 17%; RR 6.07, 95% CI 1.29-28.49; p-value 0.02) or due to sudden cardiac death (43% vs. 10.5%; RR 6.37, 95% CI 1.03-39.36; p-value 0.04). All five patients who did not have CAN at the baseline developed this abnormality on repeat testing after one year. Conclusion The prevalence of CAN in patients with end-stage renal disease on maintenance hemodialysis was significantly higher. CAN was an independent predictor of all-cause and cardiovascular mortality, which highlights it as a risk stratification tool in patients with end-stage renal disease.

Whole-Exome Sequencing Identifies Novel Variants that Co-segregates with Autosomal Recessive Retinal Degeneration in a Pakistani Pedigree.

PURPOSE: To identify the molecular basis of inherited retinal degeneration (IRD) in a familial case of Pakistani origin using whole-exome sequencing. METHODS: A thorough ophthalmic examination was completed, and genomic DNA was extracted using standard protocols. Whole exome(s) were captured with Agilent V5 + UTRs probes and sequenced on Illumina HiSeq genome analyzer. The exomeSuite software was used to filter variants, and the candidate causal variants were prioritized, examining their allele frequency and PolyPhen2, SIFT, and MutationTaster predictions. Sanger dideoxy sequencing was performed to confirm the segregation with disease phenotype and absence in ethnicity-matched control chromosomes. RESULTS: Ophthalmic examination confirmed retinal degeneration in all affected individuals that segregated as an autosomal recessive trait in the family. Whole-exome sequencing identified two homozygous missense variants: c.1304G > A; p.Arg435Gln in ZNF408 (NM_024741) and c.902G > A; p.Gly301Asp in C1QTNF4 (NM_031909). Both variants segregated with the retinal phenotype in the PKRD320 and were absent in ethnically matched control chromosomes. CONCLUSION: Whole-exome sequencing coupled with bioinformatics analysis identified potential novel variants that might be responsible for IRD.

Biallelic variants in LINGO1 are associated with autosomal recessive intellectual disability, microcephaly, speech and motor delay.

PURPOSE: To elucidate the novel molecular cause in two unrelated consanguineous families with autosomal recessive intellectual disability. METHODS: A combination of homozygosity mapping and exome sequencing was used to locate the plausible genetic defect in family F162, while only exome sequencing was followed in the family PKMR65. The protein 3D structure was visualized with the University of California-San Francisco Chimera software. RESULTS: All five patients from both families presented with severe intellectual disability, aggressive behavior, and speech and motor delay. Four of the five patients had microcephaly. We identified homozygous missense variants in LINGO1, p.(Arg290His) in family F162 and p.(Tyr288Cys) in family PKMR65. Both variants were predicted to be pathogenic, and segregated with the phenotype in the respective families. Molecular modeling of LINGO1 suggests that both variants interfere with the glycosylation of the protein. CONCLUSION: LINGO1 is a transmembrane receptor, predominantly found in the central nervous system. Published loss-of-function studies in mouse and zebrafish have established a crucial role of LINGO1 in normal neuronal development and central nervous system myelination by negatively regulating oligodendrocyte differentiation and neuronal survival. Taken together, our results indicate that biallelic LINGO1 missense variants cause autosomal recessive intellectual disability in humans.

Homozygous Truncating Variants in TBC1D23 Cause Pontocerebellar Hypoplasia and Alter Cortical Development.

Pontocerebellar hypoplasia (PCH) is a heterogeneous group of rare recessive disorders with prenatal onset, characterized by hypoplasia of pons and cerebellum. Mutations in a small number of genes have been reported to cause PCH, and the vast majority of PCH cases are explained by mutations in TSEN54, which encodes a subunit of the tRNA splicing endonuclease complex. Here we report three families with homozygous truncating mutations in TBC1D23 who display moderate to severe intellectual disability and microcephaly. MRI data from available affected subjects revealed PCH, small normally proportioned cerebellum, and corpus callosum anomalies. Furthermore, through in utero electroporation, we show that downregulation of TBC1D23 affects cortical neuron positioning. TBC1D23 is a member of the Tre2-Bub2-Cdc16 (TBC) domain-containing RAB-specific GTPase-activating proteins (TBC/RABGAPs). Members of this protein family negatively regulate RAB proteins and modulate the signaling between RABs and other small GTPases, some of which have a crucial role in the trafficking of intracellular vesicles and are involved in neurological disorders. Here, we demonstrate that dense core vesicles and lysosomal trafficking dynamics are affected in fibroblasts harboring TBC1D23 mutation. We propose that mutations in TBC1D23 are responsible for a form of PCH with small, normally proportioned cerebellum and should be screened in individuals with syndromic pontocereballar hypoplasia.

Molecular Genetic Analysis of Pakistani Families With Autosomal Recessive Congenital Cataracts by Homozygosity Screening.

Purpose: To identify the genetic origins of autosomal recessive congenital cataracts (arCC) in the Pakistani population. Methods: Based on the hypothesis that most arCC patients in consanguineous families in the Punjab areas of Pakistan should be homozygous for causative mutations, affected individuals were screened for homozygosity of nearby highly informative microsatellite markers and then screened for pathogenic mutations by DNA sequencing. A total of 83 unmapped consanguineous families were screened for mutations in 33 known candidate genes. Results: Patients in 32 arCC families were homozygous for markers near at least 1 of the 33 known CC genes. Sequencing the included genes revealed homozygous cosegregating sequence changes in 10 families, 2 of which had the same variation. These included five missense, one nonsense, two frame shift, and one splice site mutations, eight of which were novel, in EPHA2, FOXE3, FYCO1, TDRD7, MIP, GALK1, and CRYBA4. Conclusions: The above results confirm the usefulness of homozygosity mapping for identifying genetic defects underlying autosomal recessive disorders in consanguineous families. In our ongoing study of arCC in Pakistan, including 83 arCC families that underwent homozygosity mapping, 3 mapped using genome-wide linkage analysis in unpublished data, and 30 previously reported families, mutations were detected in approximately 37.1% (43/116) of all families studied, suggesting that additional genes might be responsible in the remaining families. The most commonly mutated gene was FYCO1 (14%), followed by CRYBB3 (5.2%), GALK1 (3.5%), and EPHA2 (2.6%). This provides the first comprehensive description of the genetic architecture of arCC in the Pakistani population.

A simple integrated primary health care based model for detection of diabetic retinopathy in resource-limited settings in Pakistani population.

OBJECTIVE: To find out prevalence of Diabetic Retinopathy in general population of three districts in Pakistan. METHODS: A community based cross-sectional survey was conducted in three large districts of Pakistan namely Rawalpindi in Punjab, Peshawar in Khyber Pakhtoonkhwa and Hyderabad in Sindh between January 2013 and August 2015. Lady Health Workers identified individuals at high risk for diabetes based on pre-defined criteria. High risk population was tested for dysglycemia. Fundoscopic evaluation for evidence of DR was performed in all individuals with a random blood glucose >190mg/dl. Individuals with the evidence of DR were referred to affiliated tertiary care ophthalmology departments. RESULTS: A total of 42,629 individuals reported at the project sites and 63% (n=26,859) were female. Fifty one percent (n=21,989) individuals met high risk criteria. Out of these 21,989 individuals, dysglycemia was found in 3,869 (17.6%). Fundoscopy showed evidence of DR in 1,042 (27%) individuals. Amongst high risk population, dysglycemia was significantly more common in females as compared to males. The frequency of DR in dysglycemic patients was comparable across both gender groups. CONCLUSION: The prevalence of DR in Pakistani population is alarmingly high. This preventable cause of blindness is largely undiagnosed in our population and a simple integrated model based on primary health care facilities can help identify and treat a large population of DR patients.

Pathogenic mutations in TULP1 responsible for retinitis pigmentosa identified in consanguineous familial cases.

PURPOSE: To identify pathogenic mutations responsible for autosomal recessive retinitis pigmentosa (arRP) in consanguineous familial cases. METHODS: Seven large familial cases with multiple individuals diagnosed with retinitis pigmentosa were included in the study. Affected individuals in these families underwent ophthalmic examinations to document the symptoms and confirm the initial diagnosis. Blood samples were collected from all participating members, and genomic DNA was extracted. An exclusion analysis with microsatellite markers spanning the TULP1 locus on chromosome 6p was performed, and two-point logarithm of odds (LOD) scores were calculated. All coding exons along with the exon-intron boundaries of TULP1 were sequenced bidirectionally. We constructed a single nucleotide polymorphism (SNP) haplotype for the four familial cases harboring the K489R allele and estimated the likelihood of a founder effect. RESULTS: The ophthalmic examinations of the affected individuals in these familial cases were suggestive of RP. Exclusion analyses confirmed linkage to chromosome 6p harboring TULP1 with positive two-point LOD scores. Subsequent Sanger sequencing identified the single base pair substitution in exon14, c.1466A>G (p.K489R), in four families. Additionally, we identified a two-base deletion in exon 4, c.286_287delGA (p.E96Gfs77*); a homozygous splice site variant in intron 14, c.1495+4A>C; and a novel missense variation in exon 15, c.1561C>T (p.P521S). All mutations segregated with the disease phenotype in the respective families and were absent in ethnically matched control chromosomes. Haplotype analysis suggested (p<10(-6)) that affected individuals inherited the causal mutation from a common ancestor. CONCLUSIONS: Pathogenic mutations in TULP1 are responsible for the RP phenotype in seven familial cases with a common ancestral mutation responsible for the disease phenotype in four of the seven families.

Loss of function mutations in RP1 are responsible for retinitis pigmentosa in consanguineous familial cases.

PURPOSE: This study was undertaken to identify causal mutations responsible for autosomal recessive retinitis pigmentosa (arRP) in consanguineous families. METHODS: Large consanguineous families were ascertained from the Punjab province of Pakistan. An ophthalmic examination consisting of a fundus evaluation and electroretinography (ERG) was completed, and small aliquots of blood were collected from all participating individuals. Genomic DNA was extracted from white blood cells, and a genome-wide linkage or a locus-specific exclusion analysis was completed with polymorphic short tandem repeats (STRs). Two-point logarithm of odds (LOD) scores were calculated, and all coding exons and exon-intron boundaries of RP1 were sequenced to identify the causal mutation. RESULTS: The ophthalmic examination showed that affected individuals in all families manifest cardinal symptoms of RP. Genome-wide scans localized the disease phenotype to chromosome 8q, a region harboring RP1, a gene previously implicated in the pathogenesis of RP. Sanger sequencing identified a homozygous single base deletion in exon 4: c.3697delT (p.S1233Pfs22*), a single base substitution in intron 3: c.787+1G>A (p.I263Nfs8*), a 2 bp duplication in exon 2: c.551_552dupTA (p.Q185Yfs4*) and an 11,117 bp deletion that removes all three coding exons of RP1. These variations segregated with the disease phenotype within the respective families and were not present in ethnically matched control samples. CONCLUSIONS: These results strongly suggest that these mutations in RP1 are responsible for the retinal phenotype in affected individuals of all four consanguineous families.

High prevalence of preobesity and obesity among medical students of Lahore and its relation with dietary habits and physical activity.

OBJECTIVE: The objective of this study was to determine the prevalence of obesity among students of medical colleges of Lahore and to study its correlation with high-caloric diet intake and physical inactivity. STUDY DESIGN: A cross-sectional survey was conducted at four medical colleges of Lahore, Pakistan between March and June 2012. METHODS: A total of 244 medical students (85 males, 159 females) of the median age of 20 years (range: 18-25) were randomly included in the study. Anthropometric measures were obtained. High-caloric diet intake and physical profile were assessed through a self-reported questionnaire. The relationships between obesity indices (body mass index [BMI], waist-to-hip ratio) were investigated and correlated with the studied dietary and physical activity factors. RESULTS: Approximately, 30.5% males and 16% females had BMI ≥25.0 kg/m(2) overall affecting 21% of total medical students. Central obesity was found in 46% of male and 31.4% of female students. Central obesity was associated with a higher total daily caloric intake, studying at private medical college and male gender. Overall, 197 of 244 (80.7%) students played no sports in college. Median time to watch television or work on the computer was 120 min a day (range: 30-420). Only 70 (28.7%) students had regular walk or jogging. CONCLUSION: A substantial proportion of Pakistani medical students were overweight or obese. Higher total daily caloric intake was associated with central obesity but not a BMI >25. Physical activity parameters favored an overall sedentary aptitude for medical students.

Causes of short stature in Pakistani children found at an Endocrine Center.

BACKGROUND AND OBJECTIVE: Short stature is defined as height below 3rd centile. Causes of short stature can range from familial, endocrine disorders, chronic diseases to chromosomal disorders. Most common cause in literature being idiopathic short stature. Early detection and management of remedial disorders like malnutrition and vitamin D deficiency, Endocrine disorders like growth hormone deficiency & hypothyroidism can lead to attainment of expected height. Pakistani data shows idiopathic short stature as the most common cause of short stature. Our study aimed at detecting causes of short stature in children/adolescents at an Endocrine referral center. METHODS: A retrospective study was conducted at WILCARE Center for Diabetes, Endocrinology & Metabolism, Lahore on 70 well-nourished children/adolescents. The patients had been evaluated clinically, biochemically and radiologically as needed. Biochemical testing included hormonal testing as well to detect endocrine causes. Data was entered and analyzed in SPSS 20.0. RESULTS: Leading cause of short stature in our population was Growth Hormone (GH) deficiency seen in 48 out of 70 (69%) patients. Second most common endocrine abnormality seen in these patients was Vitamin D deficiency [44 out of 70 patients (63%)]. Primary hypothyroidism; pan-hypopituitarism & adrenal insufficiency were other endocrine causes. The weight for age was below 3rd percentile in 57 (81%) patients, with no association with other major causes. CONCLUSION: Growth hormone and Vitamin D deficiency constitute one of the major causes of short stature among well-nourished children with short stature in Pakistan.