Genome wide analysis of carotenoid cleavage oxygenases (CCO) gene family in Arachis hypogaea (peanut) under biotic stress.

Carotenoid cleavage oxygenases (CCOs) enzymes play a vital role in plant growth and development through the synthesis of apocarotenoids and their derivative. These chemicals are necessary for flower and fruit coloration, as well as the manufacture of plant hormones such as abscisic acid (ABA) and strigolactones, which control a variety of physiological processes. The CCOs gene family has not been characterized in Arachis hypogaea. Genome mining of A. hypogaea identifies 24 AhCCO gene members. The AhCCO gene family was divided into two subgroups based on the recent study of the Arabidopsis thaliana CCO gene family classification system. Twenty-three AhCCO genes, constituting 95.8% of the total, were regulated by 29 miRNAs, underscoring the significance of microRNAs (miRNAs) in governing gene expression in peanuts. AhCCD19 is the only gene that lacks a miRNA target site. The physicochemical characteristics of CCO genes and their molecular weights and isoelectric points were studied further. The genes were then characterized regarding chromosomal distribution, structure, and promoter cis-elements. Light, stress development, drought stress, and hormone responsiveness were discovered to be associated with AhCCO genes, which can be utilized in developing more resilient crops. The investigation also showed the cellular location of the encoded proteins and discovered that the peanut carotenoid oxygenase gene family's expansion was most likely the result of tandem, segmental, and whole-genome duplication events. The localization expresses the abundance of genes mostly in the cytoplasm and chloroplast. Expression analysis shows that AhCCD7 and AhCCD14 genes show the maximum expression in the apical meristem, lateral leaf, and pentafoliate leaf development, while AhNCED9 and AhNCED13 express in response to Aspergillus flavus resistance. This knowledge throws light on the evolutionary history of the AhCCO gene family and may help researchers better understand the molecular processes behind gene duplication occurrences in plants. An integrated synteny study was used to find orthologous carotenoid oxygenase genes in A. hypogaea, whereas Arabidopsis thaliana and Beta vulgaris were used as references for the functional characterization of peanut CCO genes. These studies provide a foundation for future research on the regulation and functions of this gene family. This information provides valuable insights into the genetic regulation of AhCCO genes. This technology could create molecular markers for breeding programs to develop new peanut lines.

Neurological disorders and disability in Pakistan: A cross-sectional multicenter study.

BACKGROUND/OBJECTIVE: The aim of this study is to identify frequencies of various neurological disorders (NDs) and associated disability in patients attending neurologic clinics in rural and urban centers in Pakistan. METHODS: This is an observational study conducted in 39 neurological centers in both rural and urban areas, public and private health sectors all over Pakistan. This study was conducted between august 2017 to December 2019. RESULTS: A total of 28,845 adults were enrolled. Mean age of the study participants was 46.2 ± 17.2 years, 15,252 (52.9%) were men and 13,593 (47.1%) were women. Most common comorbid medical condition was hypertension 7622(26.4%) followed by Diabetes 3409(11.8%). Among neurological diagnoses, vascular diseases (20%) were the most common followed by Headache disorders (18.6%), Epilepsy (12.5%), nerve and root diseases (12.4%), Psychiatric diseases (10%), Dementias (8%) and movement disorders (7.9%). Half of the patients 15,503(53.7%) had no neurological disability, while minor disability was present in 10,442(36.2%) of cases. Moderate to severe disability was present in 2876(10%) cases. Headache disorders, psychiatric diseases, muscle pain/muscle related disorders and demyelinating diseases were more common in women. Vascular diseases, movement disorders and Dementias were more common in 46 years and above age group whereas headache disorders, Epilepsy and Psychiatric disorders were more prevalent in <46 years age groups. CONCLUSION: Vascular diseases are the most common presentation of patients in neurology clinics followed by headache disorders and epilepsies. Minor disability was present in 36% while moderate to severe disability was present in 10% cases.

A recurrent rare intronic variant in CAPN3 alters mRNA splicing and causes autosomal recessive limb-girdle muscular dystrophy-1 in three Pakistani pedigrees.

Autosomal recessive limb-girdle muscular dystrophy-1 (LGMDR1) is an autosomal recessive disorder characterized by progressive weakness of the proximal limb and girdle muscles. Biallelic mutations in CAPN3 are reported frequently to cause LGMDR1. Here, we describe 11 individuals from three unrelated consanguineous families that present with typical features of LGMDR1 that include proximal muscle wasting, weakness of the upper and lower limbs, and elevated serum creatine kinase. Whole-exome sequencing identified a rare homozygous CAPN3 variant near the exon 2 splice donor site that segregates with disease in all three families. mRNA splicing studies showed partial retention of intronic sequence and subsequent introduction of a premature stop codon (NM_000070.3: c.379 + 3A>G; p.Asp128Glyfs*15). Furthermore, we observe reduced CAPN3 expression in primary dermal fibroblasts derived from an affected individual, suggesting instability and/or nonsense-mediated decay of mutation-bearing mRNA. Genome-wide homozygosity mapping and single-nucleotide polymorphism analysis identified a shared haplotype and supports a possible founder effect for the CAPN3 variant. Together, our data extend the mutational spectrum of LGMDR1 and have implications for improved diagnostics for individuals of Pakistani origin.

A novel deletion mutation in EPM2A underlies progressive myoclonic epilepsy (Lafora body disease) in a Pakistani family.

Lafora body disease (MIM-254780), a glycogen storage disease, characterized by Lafora bodies (deformed glycogen molecules) accumulating in multiple organs, is a rare form of myoclonic epilepsy. It manifests in early adolescent years, initially with seizures and myoclonus, followed by dementia and progressive cognitive decline, ultimately culminating in death within 10 years. In Pakistan so far 5 cases have been reported. Here, we report a new case of Lafora body disease belonging to a consanguineous family from Pakistan. Histopathological analysis confirmed presence of lafora bodies in the patient`s skin. Sanger sequencing revealed novel homozygous 5bp deletion mutation (NM_005670.4; c.359_363delGTGTG) in exon 2 of the EPM2A gene, which was truly segregated in the family. These results will increase our understanding regarding the aetiology of this disorder and will further add to the mutation spectrum of EPM2A gene.

Post-COVID Neuromyelitis Optica Spectrum Disorder.

After novel coronavirus pandemic that emerged from Wuhan, China in December 2019, several cases of inflammatory and immune-mediated disorders have been reported, thought to be triggered by SARS-CoV-2 infection. Neuromyelitis optica spectrum disorder (NMOSD) is one of the autoimmune demyelinating disorders, which is thought to be triggered by viral infection. Herein, we describe a case of NMOSD in a pediatric patient with a previous SARS-CoV-2 infection, acting as a possible triggering factor. Key Words: Neuromyelitis optica spectrum disorder (NMOSD), Aquaporin 4 (AQP-4), Severe acute respiratory syndrome (SARS).

Structural and functional annotation of PR/SET Domain (PRDM) protein family: In-silico study elaborating role of PRDM12 mutation in congenital insensitivity to pain.

Congenital insensitivity to pain (CIP), classified as a type of hereditary sensory and autonomic neuropathies, is a rare disease in which the affected individuals fail to perceive sensation of pain. One of the PR/SET Domain Proteins, PRDM12, has been identified in recent past as a candidate gene for congenital insensitivity to pain. In the present study, we performed whole exome sequencing in a Pakistani family with CIP phenotype to ascertain the causative mutation. We identified a previously described alanine repeat duplication in PRDM12 (Ala353_Ala359dup) in this family. After this, we performed structural annotations for PR/SET Domain (PRDM) containing protein family to prognosticate the potential hypothetical structure of PRDM proteins with physical and chemical parameters. Out of nineteen members of this family, four members (PRDM5, PRDM8, PRDM12 and PRDM13) were specially focused because of their role in neurological disorders. Predictions about structure and interactions of these proteins revealed novel interacting molecules and pathways. Detailed in silico analysis of PRDM12 was performed to elaborate importance of its domain structure in interaction with other proteins and its role in pain insensitivity phenotype. These results have substantially enhanced our understanding regarding the etiology of congenital pain insensitivity and would stimulate further research on therapy and prevention.

Anti NMDA receptor antibody encephalitis in Pakistan: Clinicopathological features and treatment outcomes.

Anti-NMDA receptor antibody encephalitis (anti-NMDAR Encephalitis) is the most common subtype of autoimmune encephalitis in which IgG antibodies directed against NR1 subunit of NMDA receptors are present. It is a potentially lethal encephalitis which responds favourably to timely immunosuppressive therapy. If untreated, its progression leads from delusions, paranoia, movement disorder, memory deficit and seizures into a state of unresponsiveness with autonomic instability and even death. We present clinicopathological features, treatment and outcomes of eight autoantibodyproven cases of anti-NMDAR Encephalitis. There were 7 females and 1 male with a mean age of 15 years (age range: 1 to 28 years). Clinical features included seizures, altered consciousness, memory deficit, delusions, paranoia and hallucinations. Hyperactivity and irritability were prominent features among the children. Patients treated with immunosuppressive therapy including steroids, IVIg, plasmapheresis and Rituximab, recovered completely within a month of therapy. Whereas patients who received only steroids as immunosuppressive therapy suffered from residual brain damage.

CT pattern of Infarct location and not infarct volume determines outcome after decompressive hemicraniectomy for Malignant Middle Cerebral Artery Stroke.

Malignant middle cerebral artery [MMCA] infarction has a different topographic distribution that might confound the relationship between lesion volume and outcome. Retrospective study to determine the multivariable relationship between computerized tomographic [CT] infarct location, volume and outcomes in decompressive hemicraniectomy [DHC] for MMCA infarction. The MCA infarctions were classified into four subgroups by CT, subtotal, complete MCA [co-MCA], Subtotal MCA with additional infarction [Subtotal MCAAI] and co-MCA with additional infarction [Co-MCAAI]. Maximum infarct volume [MIV] was measured on the pre-operative CT. Functional outcome was measured by the modified Rankin Scale [mRS] dichotomized as favourable 0-3 and unfavourable ≥4, at three months. In 137 patients, from least favourable to favourable outcome were co-MCAAI, subtotal MCAAI, co-MCA and subtotal MCA infarction. Co-MCAAI had the worst outcome, 56/57 patients with additional infarction had mRS ≥ 4. Multiple comparisons Scheffe test showed no significant difference in MIV of subtotal infarction, co-MCA, Subtotal MCAAI but the outcome was significantly different. Multivariate analysis confirmed MCAAI [7.027 (2.56-19.28), p = 0.000] as the most significant predictor of poor outcomes whereas MIV was not significant [OR, 0.99 (0.99-01.00), p = 0.594]. Other significant independent predictors were age ≥ 55 years 12.14 (2.60-56.02), p = 0.001 and uncal herniation 4.98(1.53-16.19), p = 0.007]. Our data shows the contribution of CT infarction location in determining the functional outcome after DHC. Subgroups of patients undergoing DHC had different outcomes despite comparable infarction volumes.

Clinical efficacy and tolerability of Gabapentinoids with current prescription patterns in patients with Neuropathic pain.

OBJECTIVE: To investigate the current dosing regimens of gabapentinoids in Pakistani patients with neuropathic pain and to compare their clinical efficacy and tolerability in terms of pain relief and adverse effects using difference in pain score as a treatment outcome. METHODS: This observational, prospective study was conducted in 320 patients with neuropathic pain from August 2016 to March 2018 at Basic Medical Sciences Institute (BMSI), Karachi in collaboration with Shifa International Hospital and Benazir Bhutto Hospital, Islamabad. Demographic data, treatment-related adverse effects and pain intensity was documented at recruitment and follow up visits at two, four and eight weeks. Discontinuation due to adverse effects and lack of efficacy were also recorded. Data was entered and analyzed using SPSS version 22. RESULTS: Mean age of patients was 52.57±12.47 and the most common ethnicity were Punjabi speaking population (66%). Diabetic neuropathy (51%) was the most common etiology followed by radicular pain (25%). Mean dosages of pregabalin and gabapentin were 114mg and 470mg respectively. Mean pain score was significantly reduced by gabapentinoids (<0.001). Dizziness, drowsiness and somnolence were frequent adverse effects. Common dosages for pregabalin and gabapentin were 75 mg/day and 300 mg/day respectively. CONCLUSION: Current dosing regimens of gabapentinoids in Pakistani patients with neuropathic pain were found to be efficacious at low dosages in comparison to international recommended dosages. Gabapentin and pregabalin were both similar in terms of reducing pain score but onset of pain relief was relatively faster with pregabalin. Dizziness, drowsiness and somnolence were frequently reported with both gabapentinoids; however, visual blurring, ataxia and weight gain were observed only with the use of pregabalin. Adverse effects are frequently observed with gabapentinoids which necessitates reverting back to low dosages or switching to other drugs for pain relief.

Clinical Efficacy and Safety of Tacrolimus in Pakistani Living Donor Liver Transplant Recipients.

OBJECTIVE: To evaluate the association between tacrolimus trough levels and dosage in Pakistani patients undergoing live donor liver transplantation (LDLT), and the efficacy and adverse effects at different tacrolimus trough levels and dosages. STUDY DESIGN: An observational study. PLACE AND DURATION OF STUDY: Shifa International Hospital, Shifa Tameer-e-Millat University, Islamabad and Basic Medical Sciences Institute, Karachi, from September 2016 to October 2018. METHODOLOGY: Sixty liver transplant recipients were included. Demographics, clinical data, tacrolimus trough levels and doses were monitored as per routine protocol. Electrochemiluminescence immunoassay (ECLIA) was used to measure tacrolimus trough levels. Acute cellular rejection (ACR), sepsis and other adverse events were monitored at different tacrolimus trough levels in early post-transplantation period. RESULTS: Mean age of transplant recipients was 49.1 ± 10.6 years. Mean tacrolimus trough levels were 6.1 ± 2.2 ng/ml and mean dose was 0.94 ± 0.3 mg. Sepsis (27%) psychosis (20%), seizures (10%), and renal insufficiency (13%) were the most common adverse effects. Acute cellular rejection (ACR) was observed in 15% patients. Patients with sepsis had significantly high mean tacrolimus levels of 7.7 ± 2.5 ng/ml versus 5.5 ± 1.9 ng/ml (p=0.001). Mean tacrolimus trough levels in patients with ACR were significantly lower (4.05 ± 1.6 ng/ml vs. 6.43 ± 2.2ng/ml, p=0.003). None of the patients with a single tacrolimus trough level >10 ng/ml experienced ACR. CONCLUSION: A tacrolimus trough level between 5 to 7.5 ng/ml appears to be safe in Pakistani liver transplant recipients significantly minimising the risk of ACR and other adverse events.

Recurrent aseptic meningitis: A rare clinical presentation of Sjogren's syndrome.

Sjogren's syndrome most commonly presents with dry eyes, dry mouth, joint pain and fatigue. However, recurrent aseptic meningitis, reported as the most uncommon initial symptom, was the presenting feature in our case. We present the case of a 19-year-old female with recurrent episodes of aseptic meningitis. She presented with fever, headache, vomiting and photophobia. Neurological examination showed neck stiffness. Fundoscopy was normal. On two previous occasions her cerebrospinal fluid analysis was consistent with meningitis; however, it was normal at this presentation. Review of system revealed history of fatigue and sicca symptoms since early childhood. Autoimmune workup showed antinuclear antibodies with a titer of 1:400 and positive anti SSA (Ro) antibodies that led to the diagnosis of Sjogren's syndrome. She responded well to intravenous steroids, followed by oral prednisolone and hydroxychloroquine. To conclude, diagnosis of Sjogren's syndrome may also be considered in a patient presenting with recurrent aseptic meningitis.

Awareness of Stroke and Health-seeking Practices among Hypertensive Patients in a Tertiary Care Hospital: A Cross-sectional Survey.

INTRODUCTION: Stroke is a major cause of death with hypertension being identified as an important modifiable risk factor. Prompt identification of stroke symptoms and timely management is noted to be significant in lowering both morbidity and mortality. Baseline stroke knowledge in hypertensive patients is crucial to develop effectively targeted, and appropriate health promotion campaigns; thus, the main objectives of this study are to assess the awareness of stroke and to determine health-seeking practices among hypertensive patients. MATERIALS AND METHODS: A standardized questionnaire survey regarding awareness and practices about stroke among hypertensive patients was conducted in a tertiary care hospital of Islamabad. The sample size was calculated as 384. RESULTS: Out of 384 patients evaluated, 80.5% had heard about stroke, 71.6% knew someone with stroke, and 76% identified the brain as the organ affected. Sudden onset numbness of limb (66.9%) and hypertension (93.5%) were common warning symptom and risk factor identified. 87.5% would take stroke patients to a hospital. Only 45.1% of the patients took their medications regularly, and 38% checked their blood pressure. CONCLUSION: Majority of hypertensive patients were aware of stroke but the awareness of risk factors and warning signs was poor. Stroke prevention practices were also sub-optimal. There is a need to increase knowledge regarding risk factors, which will benefit the community at large.

Expression of PAX-5 in B Cell Hodgkin and Non Hodgkin Lymphoma

Background: B-cell malignancies including Precursor B-cell lymphoblastic lymphoma/leukemia and Hodgkin Lymphoma show a wide spectrum of B-cell differentiation from early stage B-cell precursors to mature B-cells ending in terminal differentiation to plasma cells. Pan-B-cell antigens routinely used for the diagnosis of B-cell lymphoma, include CD19, CD20, CD22 and CD79a.PAX-5 protein, also known as B-cell-specific activation protein is a B-cell-specific transcription factor; essential for commitment and functional maintenance used in the diagnosis of B cell Hodgkin and non-Hodgkin lymphoma. PAX-5 show nuclear positivity in B cell lymphomas and moderate (dim) positivity in Hodgkinlymphoma Reed Sternberg cells make this marker ideal for diagnosing B cell malignances. Objective: To determine the expression of PAX-5 in B cell Hodgkin and non-Hodgkin Lymphoma in order to improve the diagnosis of B-cell lymphomas. Methods: In this Prospective study, all the cases of B cell lymphoma diagnosed at The Aga Khan University Hospital, Karachi from July 2010 to July 2011were included. A panel of Immunohistochemical stain was performed in all cases along with additional PAX- 5 stain with appropriate controls. Results: Total 125 cases were included. Hodgkin Lymphoma (Mixed cellularity) was the commonest B-cell lymphoma subtype, 32 (25%) cases. Other common subtypes included Hodgkin lymphoma (Nodular sclerosis subtype), diffuse large B-cell lymphoma and B lymphoblastic lymphoma. Conclusion: This study demonstrates that PAX-5 is the most sensitive and reliable immuhohistochemicalmarker in the diagnosis of B cell Hodgkin and non-Hodgkin lymphoma.

Non-motor Symptoms in Patients with Parkinson's Disease: A Cross-sectional Survey.

Background In multiple studies around the globe, non-motor symptoms (NMS) have been identified as a source of immense disability in patients with Parkinson's disease (PD). However, there is a scarcity of data from Asia. This is the first study of the Pakistani population to assess the impact of NMS in PD on patients. Objectives To determine the frequency of NMS of PD in the Pakistani population and compare it with existing data. Methods In this cross-sectional survey, patient demographics were retrospectively collected from a tertiary care hospital neurology database. This study population comprised 97 patients at different stages of PD who presented to the neurology outpatient department. Disease severity was assessed using the Hoehn and Yahr scale. The NMS questionnaire was employed to identify the presence of NMS. Medical records were reviewed for demographic data and recent treatment history. Results The mean age was 67 years (76.3% of patients had adult onset PD and 23.7% had young onset PD). The NMS with the highest frequencies were nocturia (77.3%), urinary urgency (61.9%), constipation (59.8%), dementia (58.8%), insomnia (52.6%), and orthostatic hypotension (52.6%). The earliest manifestations of NMS were nocturia, forgetfulness, low mood, and orthostatic hypotension. Sleep abnormalities, falling episodes, and hallucinations are prevalent among patients with advanced disease. Conclusion There is a higher frequency of NMS present in the Pakistani population as compared to existing data in other populations.

Hemiconvulsion Hemiplegia Syndrome in a young boy with developmental delay.

Hemiconvulsion hemiplegia epilepsy (HHE) syndrome is a rare complication of prolonged focal seizures in children upto 4 years of age. It is usually idiopathic and seen in the setting of febrile seizures in otherwise normal children but less commonly is also associated with structural, infective, traumatic and degenerative diseases that predispose to seizures. It has 3 stages, the first of prolonged focal seizures, then the development of hemiplegia and then followed by final stage of development of epilepsy after a variable latent period. Early recognition and seizure control is important to prevent the development of hemiplegia and intractable epilepsy. We report a child with developmental delay and epilepsy who developed HH syndrome after prolonged unrecognized and difficult to control partial status epilepticus.

Chorea-acanthocytosis: Homozygous 1-kb deletion in VPS13A detected by whole-genome sequencing.

OBJECTIVE: To determine a molecular diagnosis for a large multigenerational family of South Asian ancestry with seizures, hyperactivity, and episodes of tongue biting. METHODS: Two affected individuals from the family were analyzed by whole-genome sequencing on the Illumina HiSeq X platform, and rare variants were prioritized for interpretation with respect to the phenotype. RESULTS: A previously undescribed, 1-kb homozygous deletion was identified in both individuals sequenced, which spanned 2 exons of the VPS13A gene, and was found to segregate in other family members. CONCLUSIONS: VPS13A is associated with autosomal recessive chorea-acanthocytosis, a diagnosis consistent with the phenotype observed in this family. Whole-genome sequencing presents a comprehensive and agnostic approach for detecting diagnostic mutations in families with rare neurologic disorders.

Identification of novel L2HGDH mutation in a large consanguineous Pakistani family- a case report.

BACKGROUND: L-2-hydroxyglutaric aciduria (L2HGA) is a progressive neurometabolic disease of brain caused by mutations of in L-2-hydroxyglutarate dehydrogenase (L2HGDH) gene. Cardinal clinical features include cerebellar ataxia, epilepsy, neurodevelopmental delay, intellectual disability, and other clinical neurological deficits. CASE PRESENTATION: We describe an index case of the family presented with generalised tonic-clonic seizure, developmental delay, intellectual disability, and ataxia. Initially, the differential diagnosis was difficult to be established and a SNP genome wide scan identified the candidate region on chromosome 14q22.1. DNA sequencing showed a novel homozygous mutation in the candidate gene L2HGDH (NM_024884.2: c.178G > A; p.Gly60Arg). The mutation p.Gly60Arg lies in the highly conserved FAD/NAD(P)-binding domain of this mitochondrial enzyme, predicted to disturb enzymatic function. CONCLUSIONS: The combination of homozygosity mapping and DNA sequencing identified a novel mutation in Pakistani family with variable clinical features. This is second report of a mutation in L2HGDH gene from Pakistan and the largest family with L2HGA reported to date.