Infection of plant cells by viral pathogens triggers RNA silencing, an innate antiviral defense mechanism. In response to infection, small RNAs (sRNAs) are produced that associate with Argonaute (AGO)-containing silencing complexes which act to inactivate viral genomes by posttranscriptional gene silencing (PTGS). Deep sequencing was used to compare virus-derived small RNAs (vsRNAs) in cassava genotypes NASE 3, TME 204 and 60444 infected with the positive sense single-stranded RNA (+ssRNA) viruses cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV), the causal agents of cassava brown streak disease (CBSD). An abundance of 21-24nt vsRNAs was detected and mapped, covering the entire CBSV and UCBSV genomes. The 21nt vsRNAs were most predominant, followed by the 22 nt class with a slight bias toward sense compared to antisense polarity, and a bias for adenine and uracil bases present at the 5'-terminus. Distribution and frequency of vsRNAs differed between cassava genotypes and viral genomes. In susceptible genotypes TME 204 and 60444, CBSV-derived sRNAs were seen in greater abundance than UCBSV-derived sRNAs. NASE 3, known to be resistant to UCBSV, accumulated negligible UCBSV-derived sRNAs but high populations of CBSV-derived sRNAs. Transcript levels of cassava homologues of AGO2, DCL2 and DCL4, which are central to the gene-silencing complex, were found to be differentially regulated in CBSV- and UCBSV-infected plants across genotypes, suggesting these proteins play a role in antiviral defense. Irrespective of genotype or viral pathogen, maximum populations of vsRNAs mapped to the cytoplasmic inclusion, P1 and P3 protein-encoding regions. Our results indicate disparity between CBSV and UCBSV host-virus interaction mechanisms, and provide insight into the role of virus-induced gene silencing as a mechanism of resistance to CBSD.

Coevolution between hosts and pathogens is thought to occur between interacting molecules of both species. This results in the maintenance of genetic diversity at pathogen antigens (or so-called effectors) and host resistance genes such as the major histocompatibility complex (MHC) in mammals or resistance (R) genes in plants. In plant-pathogen interactions, the current paradigm posits that a specific defense response is activated upon recognition of pathogen effectors via interaction with their corresponding R proteins. According to the "Guard-Hypothesis," R proteins (the "guards") can sense modification of target molecules in the host (the "guardees") by pathogen effectors and subsequently trigger the defense response. Multiple studies have reported high genetic diversity at R genes maintained by balancing selection. In contrast, little is known about the evolutionary mechanisms shaping the guardee, which may be subject to contrasting evolutionary forces. Here we show that the evolution of the guardee RCR3 is characterized by gene duplication, frequent gene conversion, and balancing selection in the wild tomato species Solanum peruvianum. Investigating the functional characteristics of 54 natural variants through in vitro and in planta assays, we detected differences in recognition of the pathogen effector through interaction with the guardee, as well as substantial variation in the strength of the defense response. This variation is maintained by balancing selection at each copy of the RCR3 gene. Our analyses pinpoint three amino acid polymorphisms with key functional consequences for the coevolution between the guardee (RCR3) and its guard (Cf-2). We conclude that, in addition to coevolution at the "guardee-effector" interface for pathogen recognition, natural selection acts on the "guard-guardee" interface. Guardee evolution may be governed by a counterbalance between improved activation in the presence and prevention of auto-immune responses in the absence of the corresponding pathogen.

Hepatitis viral infection is a leading cause of chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). Over one million people are estimated to be persistently infected with hepatitis C virus (HCV) worldwide. As capsid core protein is the key element in spreading HCV; hence, it is considered to be the superlative target of antiviral compounds. Novel drug inhibitors of HCV are in need to complement or replace the current treatments such as pegylated interferon's and ribavirin as they are partially booming and beset with various side effects. Our study was conducted to predict 3D structure of capsid core protein of HCV from northern part of India. Core, the capsid protein of HCV, handles the assembly and packaging of HCV RNA genome and is the least variable of all the ten HCV proteins among the six HCV genotypes. Therefore, we screened four phytochemicals inhibitors that are known to disrupt the interactions of core and other HCV proteins such as (a) epigallocatechin gallate (EGCG), (b) ladanein, (c) naringenin, and (d) silybin extracted from medicinal plants; targeted against active site of residues of HCV-genotype 3 (G3) (Q68867) and its subtypes 3b (Q68861) and 3g (Q68865) from north India. To study the inhibitory activity of the recruited flavonoids, we conducted a quantitative structure-activity relationship (QSAR). Furthermore, docking interaction suggests that EGCG showed a maximum number of hydrogen bond (H-bond) interactions with all the three modeled capsid proteins with high interaction energy followed by naringenin and silybin. Thus, our results strongly correlate the inhibitory activity of the selected bioflavonoid. Finally, the dynamic predicted capsid protein molecule of HCV virion provides a general avenue to target structure-based antiviral compounds that support the hypothesis that the screened inhibitors for viral capsid might constitute new class of potent agents but further confirmation is necessary using in vitro and in vivo studies.

Cassava brown streak disease (CBSD) threatens food and economic security for smallholder farmers throughout East and Central Africa, and poses a threat to cassava production in West Africa. CBSD is caused by two whitefly-transmitted virus species: Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) (Genus: Ipomovirus, Family Potyviridae). Although varying levels of tolerance have been achieved through conventional breeding, to date, effective resistance to CBSD within East African cassava germplasm has not been identified. RNAi technology was utilized to integrate CBSD resistance into the Ugandan farmer-preferred cassava cultivar TME 204. Transgenic plant lines were generated expressing an inverted repeat construct (p5001) derived from coat-protein (CP) sequences of CBSV and UCBSV fused in tandem. Northern blots using probes specific for each CP sequence were performed to characterize 169 independent transgenic lines for accumulation of CP-derived siRNAs. Transgenic plant lines accumulating low, medium and high levels of siRNAs were bud graft challenged with the virulent CBSV Naliendele isolate alone or in combination with UCBSV. Resistance to CBSD in the greenhouse directly correlated to levels of CP-derived siRNAs as determined by visual assessment of leaf and storage root symptoms, and RT-PCR diagnosis for presence of the pathogens. Low expressing lines were found to be susceptible to CBSV and UCBSV, while medium to high accumulating plant lines were resistant to both virus species. Absence of detectable virus in the best performing p5001 transgenic lines was further confirmed by back-inoculation via sap or graft challenge to CBSD susceptible Nicotiana benthamiana and cassava cultivar 60444, respectively. Data presented shows robust resistance of transgenic p5001 TME 204 lines to both CBSV and UCBSV under greenhouse conditions. Levels of resistance correlated directly with levels of transgene derived siRNA expression such that the latter can be used as predictor of resistance to CBSD.

The current analyses of vegetation were aimed to study the different effects of environmental variables and plant species and communities interaction to these variables, identified threats to local vegetation and suggestion for remedial measures in the Mount Eelum, Swat, Pakistan. For assessment of environmental variability quantitative ecological techniques were used through quadrats having sizes of 2 × 2, 5 × 5 and 10 × 10 m2 for herbs, shrubs and trees respectively. Result of the present study revealed 124 plant species in the study area. Canonical Correspondence Analysis (CCA) was used to analyze the ecological gradient of vegetation. The environmental data and species abundance were used in CANOCO software version 4.5. The presence absence data of plant species were elaborated with Cluster and Two Way Cluster Analysis techniques using PC-ORD version 5 to show different species composition that resulted in five plant communities. Findings indicate that elevation, aspect and soil texture are the strongest variables that have significant effect on species composition and distribution of various communities shown with P value 0.0500. It is recommended to protect and use sensibly whole of the Flora normally and rare species particularly in the region.

Elucidation of key regulators in Arabidopsis fruit patterning has facilitated knowledge-translation into crop species to address yield loss caused by premature seed dispersal (pod shatter). In the 1980s, plant scientists descended on a small weed Arabidopsis thaliana (thale cress) and developed it into a powerful model system to study plant biology. The massive advances in genetics and genomics since then have allowed us to obtain incredibly detailed knowledge on specific biological processes of Arabidopsis growth and development, its genome sequence and the function of many of the individual genes. This wealth of information provides immense potential for translation into crops to improve their performance and address issues of global importance such as food security. Here, we describe how fundamental insight into the genetic mechanism by which seed dispersal occurs in members of the Brassicaceae family can be exploited to reduce seed loss in oilseed rape (Brassica napus). We demonstrate that by exploiting data on gene function in model species, it is possible to adjust the pod-opening process in oilseed rape, thereby significantly increasing yield. Specifically, we identified mutations in multiple paralogues of the INDEHISCENT and GA4 genes in B. napus and have overcome genetic redundancy by combining mutant alleles. Finally, we present novel software for the analysis of pod shatter data that is applicable to any crop for which seed dispersal is a serious problem. These findings highlight the tremendous potential of fundamental research in guiding strategies for crop improvement.

People around the world are currently affected by Major Depressive Disorder (MDD). Despite its many aspects, symptoms, manifestations and impacts, efforts have been made to identify the root causes of the disorder. In particular, genetic studies have concentrated on identifying candidate genes for MDD and exploring associations between these genes and some specific group of individuals. The aim of this research was to find out the association between single nucleotide polymorphisms in 6 candidate genes linked to the neurobiology of major depressive disorder in the North-Western population of Pakistan. We performed a case-control analysis, with 400 MDD and 232 controls. A trained psychiatrist or clinical psychologists evaluated the patients. Six polymorphisms were genotyped and tested for allele and genotype association with MDD. There were no statistical variations between MDD patients and healthy controls for genotypic and allelic distribution of all the polymorphisms observed. Thus, our analysis does not support the major role of these polymorphisms in contributing to MDD susceptibility, although it does not preclude minor impact. The statistically significant correlation between six polymorphisms and major depressive disorder in the studied population was not observed. There are inconsistencies in investigations around the world. Future research, including GWAS and association analysis on larger scale should be addressed for further validation and replication of the present findings.

Inflammatory bowel disease (IBD) is a persistent, lifelong inflammation of the digestive system. Dextran sulfate sodium is commonly used to induce colitis in experimental animal models, which causes epithelial damage, intestinal inflammation, mucin depletion, and dysbiosis of the gut microbiota. Various prebiotics, polysaccharides, and polypeptides are used for IBD treatment. In this study, we used a murine model utilizing BALB/c mice, with 10 mice per group, to investigate the treatment effect of sea conch peptide hydrolysate (CPH) on DSS-induced colitis mice. Colitis was induced through the administration of 2.5% DSS in drinking water over a seven-days period. Furthermore, on the eighth day of the experiment, sea conch peptide hydrolysate (CPH) at low (100 mg/kg), medium (200 mg/kg), and high (400 mg/kg) doses, which were continued for 14 days, were assessed for medicinal purposes in DSS-induced colitis mice. Our results showed that CPH treatment significantly alleviated the severity and symptoms of colitis. The epithelial integrity and histological damage were improved. Intestinal inflammation and inflammatory cell infiltration were improved. Furthermore, the expression of pro-inflammatory cytokines was reduced, and intestinal barrier integrity was restored by elevating the tight junction proteins. Moreover, 16s RNA sequencing revealed dysbiosis of the gut microbiota was observed upon DSS treatment, which was reinstated after CPH treatment. An increased level of Firmicutes and Lactobacillus was observed in the treatment groups. Finally, our results suggest that CPH would be recommended as a functional food source and also have the potential to be used as a medicinal product for different gastrointestinal disorders.

Falls are the third-leading cause of disability among the elderly population worldwide. It is multifactorial, and the occurrence of falls depends on different factors, which can be different from context to context, and individual to individual. Therefore, regular assessment of fall risk factors is required to develop a strategy for fall prevention. The study aimed to identify fall-related risk factors in Pakistani healthy older adults at risk of developing physical disabilities. It also aimed to create a risk-predictive model for fall occurrence, offering evidence for preventive strategies.

A sentinel plot case study was carried out to identify and map the distribution of begomovirus-betasatellite complexes in sentinel plots and commercial cotton fields over a four-year period using molecular and high-throughput DNA 'discovery' sequencing approaches. Samples were collected from 15 study sites in the two major cotton-producing areas of Pakistan. Whitefly- and leafhopper-transmitted geminiviruses were detected in previously unreported host plant species and locations. The most prevalent begomovirus was cotton leaf curl Kokhran virus-Burewala (CLCuKoV-Bu). Unexpectedly, a recently recognized recombinant, cotton leaf curl Multan virus-Rajasthan (CLCuMuV-Ra) was prevalent in five of 15 sites. cotton leaf curl Alabad virus (CLCuAlV) and cotton leaf curl Kokhran virus-Kokhran, 'core' members of CLCuD-begomoviruses that co-occurred with CLCuMuV in the 'Multan' epidemic were detected in one of 15 sentinel plots. Also identified were chickpea chlorotic dwarf virus and 'non-core' CLCuD-begomoviruses, okra enation leaf curl virus, squash leaf curl virus, and tomato leaf curl New Delhi virus. Cotton leaf curl Multan betasatellite (CLCuMuB) was the most prevalent CLCuD-betasatellite, and less commonly, two 'non-core' betasatellites. Recombination analysis revealed previously uncharacterized recombinants among helper virus-betasatellite complexes consisting of CLCuKoV, CLCuMuV, CLCuAlV and CLCuMuB. Population analyses provided early evidence for CLCuMuV-Ra expansion and displacement of CLCuKoV-Bu in India and Pakistan from 2012-2017. Identification of 'core' and non-core CLCuD-species/strains in cotton and other potential reservoirs, and presence of the now predominant CLCuMuV-Ra strain are indicative of ongoing diversification. Investigating the phylodynamics of geminivirus emergence in cotton-vegetable cropping systems offers an opportunity to understand the driving forces underlying disease outbreaks and reconcile viral evolution with epidemiological relationships that also capture pathogen population shifts.

Macrophages respond against Mycobacterium leprae infection through interacting with vitamin D and vitamin D receptor (VDR). There has been no study analyzing the association between vitamin D and VDR with bacteriological index (BI) in leprosy patients in Indonesia. To analyze the serum level of 25- hydroxyvitamin D (25(OH)D) and plasma level of VDR as well as their association with BI in leprosy patients in Indonesia. This is a cross-sectional study. Serum level of 25(OH)D was assessed with in vitro chemiluminescent immunoassay. Plasma level of VDR was assessed with enzyme linked immunosorbent assay method. Median serum level of 25(OH)D was 12.68 ng/mL. There was no correlation between serum level of 25(OH)D and BI (r=0.033; p=0.869). Median plasma level of VDR was 1.36 ng/mL. There was no correlation between plasma level of VDR and BI (r=- 0.063; p=0.749) and no significant association between BI and serum level of 25(OH) and plasma level of VDR (R2=0.055). There was no association between serum level of 25(OH)D and plasma level of VDR with BI in leprosy patients.

Hepatitis C virus roots a chronic liver disease. Currently approved treatment strategy includes administration of alpha interferon and ribavirin combined therapy for 24-48 weeks. One of the predictor of sustained virological response is an early virological response to treatment characterized as rapid response. Hyper variable region 1 (HVR1) of E2 protein is responsible for viral entry and acts as a target for neutralizing antibodies. Any mutation in this region would effect virus interaction with target cell and viral persistence.

At least five begomoviral species that cause leaf curl disease of cotton have emerged recently in Asia and Africa, reducing fiber quality and yield. The potential for the spread of these viruses to other cotton-vegetable growing regions throughout the world is extensive, owing to routine, global transport of alternative hosts of the leaf curl viruses, especially ornamentals. The research reported here describes the design and validation of polymerase chain reaction (PCR) primers undertaken to facilitate molecular detection of the two most-prevalent leaf curl-associated begomovirus-betasatellite complexes in the Indian Subcontinent and Africa, the Cotton leaf curl Kokhran virus-Burewala strain and Cotton leaf curl Gezira virus, endemic to Asia and Africa, respectively. Ongoing genomic diversification of these begomoviral-satellite complexes was evident based on nucleotide sequence alignments, and analysis of single nucleotide polymorphisms, both factors that created new challenges for primer design. The additional requirement for species and strain-specific, and betasatellite-specific primer design, imposes further constraints on primer design and validation due to the large number of related species and strains extant in 'core leaf curl virus complex', now with expanded distribution in south Asia, the Pacific region, and Africa-Arabian Peninsula that have relatively highly conserved coding and non-coding regions, which precludes much of the genome-betasatellite sequence when selecting primer 'targets'. Here, PCR primers were successfully designed and validated for detection of cloned viral genomes and betasatellites for representative 'core leaf curl' strains and species, distant relatives, and total DNA isolated from selected plant species. The application of molecular diagnostics to screen plant imports prior to export or release from ports of entry is expected to greatly reduce the likelihood of exotic leaf curl virus introductions that could dramatically affect the production of cotton as well as vegetable and ornamental crop hosts.

The Plasmodium falciparum apical membrane antigen-1 (PfAMA1) is considered as an ideal vaccine candidate for malaria control due to its high level of immunogenicity and essential role in parasite survival. Among the three domains of PfAMA1 protein, hyper-variable region (HVR) of domain I is the most immunogenic. The present study was conducted to evaluate the extent of genetic diversity across HVR domain I of the pfama1 gene in P. falciparum isolates from Hazara division of Pakistan.

Globally, 15 million infants are born preterm and another 23.2 million infants are born small for gestational age (SGA). Determining burden of preterm and SGA births, is essential for effective planning, modification of health policies and targeting interventions for reducing these outcomes for which accurate estimation of gestational age (GA) is crucial. Early pregnancy ultrasound measurements, last menstrual period and post-natal neonatal examinations have proven to be not feasible or inaccurate. Proposed algorithms for GA estimation in western populations, based on routine new-born screening, though promising, lack validation in developing country settings. We evaluated the hypothesis that models developed in USA, also predicted GA in cohorts of South Asia (575) and Sub-Saharan Africa (736) with same precision.

To detect single nucleotide polymorphism in toll-like receptor 2 (TLR2) gene in complicated cases of measles, in order to understand the genetic basis of complex human immune responses against measles complications.

Different abiotic stresses induce OsTZF1, a tandem CCCH-type zinc finger domain gene, in rice. Here, we report that transgenic rice plants overexpressing OsTZF1 under own promoter (POsTZF1:OsTZF1-OX [for overexpression]) transferred to soil showed normal growth similar to vector control plants. The POsTZF1:OsTZF1-OX produced normal leaves without any lesion mimic phenotype and exhibited normal seed setting. The POsTZF1:OsTZF1-OX plants showed significantly increased tolerance to salt and drought stresses and enhanced post stress recovery. Microarray analysis revealed a total of 846 genes up-regulated and 360 genes down-regulated in POsTZF1:OsTZF1-OX salt-treated plants. Microarray analysis of POsTZF1:OsTZF1-OX plants showed the regulation of many abiotic stress tolerance genes. These results suggest that OsTZF1-OX under own promoter show abiotic stress tolerance and produces no pleiotropic effect on phenotype of transgenic rice plant.

Babies born early and/or small for gestational age in Low and Middle-income countries (LMICs) contribute substantially to global neonatal and infant mortality. Tracking this metric is critical at a population level for informed policy, advocacy, resources allocation and program evaluation and at an individual level for targeted care. Early prenatal ultrasound examination is not available in these settings, gestational age (GA) is estimated using new-born assessment, last menstrual period (LMP) recalls and birth weight, which are unreliable. Algorithms in developed settings, using metabolic screen data, provided GA estimates within 1-2 weeks of ultrasonography-based GA. We sought to leverage machine learning algorithms to improve accuracy and applicability of this approach to LMICs settings.

Pakistan covers a key geographic area in human history, being both part of the Indus River region that acted as one of the cradles of civilization and as a link between Western Eurasia and Eastern Asia. This region is inhabited by a number of distinct ethnic groups, the largest being the Punjabi, Pathan (Pakhtuns), Sindhi, and Baloch.

Cassava brown streak disease (CBSD) presents a serious threat to cassava production in East and Central Africa. Currently, no cultivars with high levels of resistance to CBSD are available to farmers. Transgenic RNAi technology was employed to combat CBSD by fusing coat protein (CP) sequences from Ugandan cassava brown streak virus (UCBSV) and Cassava brown streak virus (CBSV) to create an inverted repeat construct (p5001) driven by the constitutive Cassava vein mosaic virus promoter. Twenty-five plant lines of cultivar TME 204 expressing varying levels of small interfering RNAs (siRNAs) were established in confined field trials (CFTs) in Uganda and Kenya. Within an initial CFT at Namulonge, Uganda, non-transgenic TME 204 plants developed foliar and storage root CBSD incidences at 96-100% by 12 months after planting. In contrast, 16 of the 25 p5001 transgenic lines showed no foliar symptoms and had less than 8% of their storage roots symptomatic for CBSD. A direct positive correlation was seen between levels of resistance to CBSD and expression of transgenic CP-derived siRNAs. A subsequent CFT was established at Namulonge using stem cuttings from the initial trial. All transgenic lines established remained asymptomatic for CBSD, while 98% of the non-transgenic TME 204 stake-derived plants developed storage roots symptomatic for CBSD. Similarly, very high levels of resistance to CBSD were demonstrated by TME 204 p5001 RNAi lines grown within a CFT over a full cropping cycle at Mtwapa, coastal Kenya. Sequence analysis of CBSD causal viruses present at the trial sites showed that the transgenic lines were exposed to both CBSV and UCBSV, and that the sequenced isolates shared >90% CP identity with transgenic CP sequences expressed by the p5001 inverted repeat expression cassette. These results demonstrate very high levels of field resistance to CBSD conferred by the p5001 RNAi construct at diverse agro-ecological locations, and across the vegetative cropping cycle.