A study of association between presence or absence of GSTT1 and GSTM1 and/or single nucleotide polymorphism in FABP2 and GSTP1 with incidence of diabetes type 2: A case-control study.

OBJECTIVE: To assess the association of single nucleotide polymorphisms in fatty acid binding protein-2 (rs1799883) and glutathione S-transferase pi (rs1695) genes with presence/absence of glutathione S-transferase mu and glutathione S-transferase theta genes in type 2 diabetes. METHODS: The cross-sectional case-control study was conducted at Institute of Molecular Biology and Biotechnology during March till September 2019 and comprised type 2 diabetes patients and non-diabetic controls from two districts in southern Punjab. Polymerase chain reaction, polymerase chain reaction-restriction fragment length polymorphism and tetra-primer amplification refractory mutation system-polymerase chain reaction were applied to investigate glutathione S-transferase theta, mu and pi genes as well as fatty acid binding protein-2, as appropriate. The association of single nucleotide polymorphisms in all genes with the disease were studied either individually or in various combinations. Data was analysed using Minitab 18. RESULTS: Of the 448 subjects, 248(55.4%) were patients and 200(44.6%) were controls. Overall there were 213(47.5%) males and 235(52.5) were females, and 141(31.5%) were aged 30-46 years. The presence of rs1799883 in fatty acid binding protein-2 (p=0.03) and the absence of glutathione S-transferase mu gene (p<0.001) had significant association with type 2 diabetes, while the presence of glutathione S-transferase theta and rs1695 in glutathione S-transferase pi genes were not associated with the disease. Individuals with glutathione S-transferase mu gene null and Ileu/Ileu or Ileu/Val genotype of rs1695 in glutathione S-transferase pi gene have potential to develop type 2 diabetes in their lifetime (p<0.05). CONCLUSIONS: The presence of rs1799883 in fatty acid binding protein-2 and the absence of glutathione S-transferase mu gene were found to play significantly in the development of type 2 diabetes.

Prevalence, epidemiology, seasonality, and phylogeny of Anaplasma marginale in blood samples of goats collected from Punjab, Pakistan.

PURPOSE: In Pakistan, a major constrain to goat farming is the tick and tick-borne diseases that results in financial losses to livestock farmers. This study was conducted to report the molecular prevalence of Anaplasma (A.) marginale in goat blood samples collected during four seasons from Khanewal district in Punjab (Pakistan). METHODS AND RESULTS: The mps1 gene of A. marginale was targeted in 900 blood samples that were collected on seasonal basis (n = 225 per season) and 6.6% (61/900) goats were found positive with A. marginale. Anaplasma marginale positive PCR products were sequenced and submitted to the GenBank. Prevalence of A. marginale varied with sampling season (P = 0.002) and it was highest in the summer (11.5%) followed by the autumn (7.6%), spring (5.3%), and winter seasons (2.7%) respectively. Anaplasma marginale prevalence varied significantly between goat breeds during the autumn (p = 0.01) and summer seasons (p = 0.02). Goats more than 2 years old and livestock farms where only goats were kept and dogs were associated with herds were risk factors for ovine anaplasmosis during different seasons. White and red blood cell counts and parameters associated with their counts were affected in A. marginale infected goats while studied serum parameters remained unaffected. CONCLUSION: PCR is a reliable tool for the detection of A. marginale in goat blood samples. A relatively low prevalence of A. marginale in goats of Khanewal district was observed and the parasite prevalence in goats was higher in the summer (May until September) and autumn (October and November) seasons. Control measures are required to prevent tick-borne diseases in ruminants from Pakistan.

Metal-tolerant and siderophore producing Pseudomonas fluorescence and Trichoderma spp. improved the growth, biochemical features and yield attributes of chickpea by lowering Cd uptake.

Industrialization and human urbanization have led to an increase in heavy metal (HM) pollution which often cause negative/toxic effect on agricultural crops. The soil-HMs cannot be degraded biologically however, microbe-mediated detoxification of toxic HMs into lesser toxic forms are reported. Considering the potentiality of HMs-tolerant soil microbes in metal detoxification, Pseudomonas fluorescence PGPR-7 and Trichoderma sp. T-4 were recovered from HM-affected areas. Under both normal and cadmium stress, the ability of both microorganisms to produce different plant hormones and biologically active enzymes was examined. Strains PGPR-7 and T-4 tolerated cadmium (Cd) an up-to 1800 and 2000 µg mL-1, respectively, and produced various plant growth regulating substances (IAA, siderophore, ACC deaminase ammonia and HCN) in Cd-stressed condition. The growth promoting and metal detoxifying ability of both strains were evaluated (either singly/combined) by applying them in chickpea (Cicer arietinum L.) plants endogenously contaminated with different Cd levels (0-400 µg kg-1 soils). The higher Cd concentration (400 µg kg-1 soils) negatively influenced the plant parameters which, however, improved following single/combined inoculation of P. fluorescence PGPR-7 and Trichoderma sp. T-4. Both microbial strains increased the growth of Cd-treated chickpeas however, their combined inoculation (PGPR-7 + T-4) caused the most positive effect. For instance, 25 µg Cd Kg-1 + PGPR-7 + T4 treatment caused maximum increase in germination percentage (10%), root dry biomass (71.4%) and vigour index (33%), chl-a (38%), chl-b (41%) and carotenoid content (52%). Furthermore, combined inoculation of P. fluorescence PGPR-7 and Trichoderma sp. T-4 maximally decreased the proline, MDA content, POD and CAT activities by 50%, 43% and 62%, respectively following their application in 25 µg Cd kg-1 soils-treated chickpea. Additionally, microbial strains lowered the plant uptake of Cd. For example, Cd-uptake in root tissues was decreased by 42 and 34% when 25 µg Cd Kg-1- treated chickpea plants were inoculated with P. fluorescence PGPR-7, Trichoderma sp. T-4 and co-inoculation (PGPR-7 + T4) of both strains, respectively. Therefore, from the current observation, it is suggested that dual inoculation of metal tolerant P. fluorescence and Trichoderma sp. may potentially be used in detoxification and reclamation of metal-contaminated soils.

Pancreatic regenerative potential of manuka honey evidenced through pancreatic histology and levels of transcription factors in diabetic rat model.

Background: Diabetes mellitus is a commonly occurring metabolic disorder accompanied by high morbidity and alarming mortality. Besides various available therapies, induction of pancreatic regeneration has emerged as a promising strategy for alleviating the damaging effect of diabetes. Honey, a potent antioxidative and anti-inflammatory agent, has been reported in the literature archive to exhibit favourable results in the regeneration process of several organ systems. Design: The current research work was intended to explore the potential role of manuka honey in pancreatic regeneration in alloxan-induced diabetic rats by accessing the pancreatic histology and levels of relevant transcription factors, including MAFA, PDX-1, INS-1, INS-2, NEUROG3, NKX6-1, and NEUROD. An equal number of rats were allocated to all four experimental groups: normal, negative control, positive control, and treatment group. Diabetes was induced in all groups except normal through a single intraperitoneal dose of alloxan monohydrate. No subsequent treatment was given to the negative control group, while the positive control and treatment groups were supplemented with metformin (150 mg/kg/day) and manuka honey (3 g/kg/day), respectively. Results: Statistical comparison of glucose and insulin levels, oxidative stress indicators, changes in the architecture of pancreatic islets, and expression levels of regeneration-associated transcription factors advocated the potential role of manuka honey in ameliorating the alloxan-induced hyperglycaemia, hyperinsulinemia, oxidative stress, and necrotic changes in islets along with significant upregulation of relevant transcription factors. Conclusion: This suggests to us the auspicious role of antioxidants in honey in pancreatic regeneration and advocates the favourable role of manuka honey in combating diabetes mellitus.

Seasonality, epidemiology and phylogeny of Theileria ovis with a note on hematological and biochemical changes in asymptomatic infected goats from Pakistan.

Caprine theileriosis, caused by Theileria ovis is a serious production issue, especially in the areas that depend on goats and sheep for milk, meat, and other economic benefits. Pakistan has a large goat population, but few reports have been documented from this country regarding PCR-based detection of T. ovis. The molecular prevalence of T. ovis, on a seasonal basis, in various goat breeds enrolled from Muzaffar Garh district of Punjab in Pakistan was determined from October 2018 to September 2019. In this study, 1084 goat blood samples were screened for the detection of T. ovis DNA through PCR-based amplification of 18S rRNA gene. Out of 1084 goats, 12 (1.11%) were infected with T. ovis. The parasite prevalence varied with the sampling seasons (Chi square test, P = 0.008), and the parasite prevalence was highest in goat blood samples collected in summer (2.39%) followed by winter (1.88%). DNA sequencing and BLAST analysis confirmed the presence of T. ovis, and the amplified isolates from the 18S rRNA gene of T. ovis were found to be highly conserved during phylogenetic analysis. Young goats (Fischer exact test, P = 0.022) were found more infected with T. ovis during the winter season. Infected goats had elevated white blood cell counts (Two-sample t-test, P = 0.04), blood urea nitrogen to Creatinine ratio (Two-sample t-test, P = 0.02) and decreased serum Creatinine (Two-sample t-test, P = 0.001) as compared to T. ovis negative goats. We report a relatively low molecular prevalence of T. ovis in goats from the Muzaffar Garh district. However, it is recommended that control measures to eradicate T. ovis infection in goats in this area should be taken.

Zinc Foliar Application Mitigates Cadmium-Induced Growth Inhibition and Enhances Wheat Growth, Chlorophyll Contents, and Yield.

Cadmium (Cd) is a toxic heavy metal that significantly threatens plants and the environment. Its toxicity in plants can result in various adverse effects, including reduced growth, altered metabolism, and cell damage. Cadmium can also interfere with nutrient uptake, particularly zinc (Zn), leading to Zn deficiency and further exacerbating Cd toxicity. On the other hand, foliar application of zinc might be a useful strategy to mitigate cadmium (Cd) toxicity in plants. Hence, a pot experiment was conducted with three replications. The wheat plants were treated with various concentrations of Zn as a foliar spray (control, 0.1, 0.2, 0.4, and 0.5%) in Cd-spiked soil in pots. The results showed that foliar use of Zn at 0.4 or 0.5% resulted in higher plant height, grain yield, and dry matter yield than the control group. Using Zn as foliar spray enriched shoot and grain Zn content while reducing Cd content in the shoot and grain. The leaf's electrolyte leakage (EL) decreased by 15.4, 29.8, 40.7, and 45.9% in the Zn 0.1%, Zn 0.2%, Zn 0.4%, and Zn 0.5% treatments, respectively, compared to the control treatment. Regarding superoxide dismutase (SOD) activity, Zn 0.5% treatment showed a decrease of 42.9% over control. Specifically, the Zn 0.1% showed a 27.2%, Zn 0.2% showed a 56.8%, Zn 0.4% showed a 91.1%, and Zn 0.5% showed a 133.7% increase in total chlorophyll content than control. Based on the results, it is recommended that 0.4% Zn solution may be used for foliar application for enhancing crop productivity and Zn concentration in plants under high Cd stress. Additionally, continued research on the mechanisms of cadmium uptake, transport, and detoxification in plants may lead to the identification of new targets for intervention.

First report on molecular epidemiology, seasonality and phylogeny of Toxoplasma gondii infecting goats from Khanewal district in Punjab, Pakistan.

Raising small ruminants is the main source of income for farmers in Pakistan. Economic losses caused by Toxoplasma gondii to small ruminants have been reported worldwide, however reports on molecular detection of T. gondii are lacking in Pakistan despite a large goat population. The current study was carried out from March 2019 till February 2020 to report the seasonal and molecular prevalence of T. gondii in different breeds of goats located in Khanewal district of Punjab, Pakistan. A total of 898 blood goat samples were collected during the four seasons and screened for T. gondii DNA by using PCR based on the amplification of ITS-1 partial sequence. Out of 898 goats, 48 (5.3%) were found positive to T. gondii. The prevalence of T. gondii varied according to season (Chi square test,P = 0.016) and the highest prevalence was observed in goats tested during the summer (8.8%) followed by the spring (5.7%), the winter (4.4%) and the autumn season (2.2%). PCR products positive to T. gondii were confirmed by DNA sequencing and BLAST analysis. Phylogenetic study based on ITS 1 gene provided evidence that the amplified isolates of T. gondii were highly conserved in Pakistani goats. Buck (Fischer exact test, P = 0.002) and farms containing other dairy animals next to goats (Fischer exact test, P = 0.001) and farms with a water supply from pools (Fischer exact test, P = 0.001) were more infected with T. gondii. Infected goats had a reduction on red blood cell count (Two-sample t test, P = 0.01) and hemoglobin concentration (Two-sample t test, P = 0.03) and an increase in the number of monocytes (%) (Two-sample t test, P = 0.05), mean cell hemoglobin (Two-sample t test, P = 0.01) and serum creatinine (Two-sample t test, P = 0.01) as compared to T. gondii uninfected goats. In conclusion, we report a relatively low PCR based prevalence of T. gondii in goats from Khanewal district as previously the serum ELISA test based prevalence of T. gondii in Pakistani goats varied between 19-52%. Control measures should be taken to eradicate T. gondii infection in goats of the study area.

Molecular survey of Toxoplasma gondii in cattle and buffaloes and phylogenetic position of Pakistani isolates based on ITS-1 gene.

Toxoplasma gondii (T. gondii) is a facultative heterogeneous parasite that belongs to Apicomplexa and can infect almost all warm-blooded animals, including ruminants, birds and humans. To date, no information is available about the molecular investigation of T. gondii in large ruminants from Pakistan. In the present study, prevalence, risk factors and genetic diversity of this parasite were evaluated by using PCR based on ITS-1 gene followed by sequencing of three selected positive PCR products. A total of 310 blood samples from cattle (N = 190) and buffaloes (N = 120) were collected from randomly selected farms located in Rajanpur district in Punjab (Pakistan). The overall infection rates of T. gondii were 12.2% (23/190) and 0% (0/120), respectively, in cattle and buffaloes. All studied epidemiological factors were not found associated with T. gondii infection in cattle. Sequence analysis of our T. gondii isolates infecting cattle revealed only one sequence considered as the most represented genetic variant (GV1) among T. gondii isolates around the world. Phylogenetic analysis revealed that ITS-1 partial sequences of our isolates clustered with those from T. gondii isolates infecting goats and birds from Pakistan and other isolates found in several animal species from different worldwide countries like China, Thailand, Poland, Canada, USA and Brazil. Our report indicates a natural infection with T. gondii of cattle for the first time in Pakistan by using molecular method. This study is important to the design of control strategy against this parasite in order to improve the output of livestock sector which is the main income source of the population in Pakistan.

Proteomic changes in various plant tissues associated with chromium stress in sunflower.

Heavy metal stress is one of the major abiotic stresses that cause environmental pollution in recent decades. An elevated concentration of these heavy metals is highly toxic to plant. Chromium (Cr) is one of the heavy metals whose concentration in the environment is still increasing alarmingly. It is harmful for plant growth and achene yield. To check out the growth and protein alternation towards pollutants, two sunflower varieties (RA-713 and AHO-33) were subjected to different chromium concentrations (control, 200 ppm, 400 ppm) by soil application. This study has elaborated that 400 ppm Cr resulted in a reduction of various growth parameters. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) was used to enhance the understanding of plant proteomic modulation under Cr stress. Different protein bands like 48 and 49, 26 kDa have newly appeared, and three 60, 47, and 42 kDa, and two protein bands 49 and 13 kDa were up-regulated in seeds of RA-713 and AHO-33, respectively. Some proteins (52, 16 kDa) are down-regulated in leaf tissues of both varieties. Only 6 and 81 kDa protein showed up-regulation and 154 kDa down-regulation behavior in the shoot in response to stress. The finding s of study might support the selection of tolerant genotype under Cr contamination and the discovery of new protein biomarkers that can use as monitoring tools in heavy metal stress biology.

Genome-wide association studies of seedling quantitative trait loci against salt tolerance in wheat.

Salinity is one of the significant factors in decreasing wheat yield and quality. To counter this, it is necessary to develop salt-tolerant wheat varieties through conventional and advanced molecular techniques. The current study identified quantitative trait loci in response to salt stress among worldwide landraces and improved varieties of wheat at the seedling stage. A total of 125 landraces and wheat varieties were subjected to salt treatment (50, 100, and 150 mM) with control. Morphological seedling traits, i.e., shoot length, root length, and fresh and dry shoot and root weights for salinity tolerance were observed to assess salt tolerance and genetic analysis using SNP data through DArT-seq. The results showed that, at the seedling stage, 150 mM NaCl treatment decreased shoot length, root length, and fresh and dry weights of the shoot and root. The root length and dry root weight were the most affected traits at the seedling stage. Effective 4417 SNPs encompassing all the chromosomes of the wheat genome with marker density, i.e., 37%, fall in genome B, genome D (32%), and genome A (31%). Five loci were found on four chromosomes 6B, 6D, 7A, and 7D, showing strong associations with the root length, fresh shoot weight, fresh root weight, and dry root weight at the p < 0.03 significance level. The positive correlation was found among all morphological traits under study.

Retrospective study of frequency of ABO and Rhesus blood group among population of Safdarabad and Faisalabad cities of Pakistan.

OBJECTIVE: The current study aimed to investigate the ABO and rhesus (Rh) blood group frequency in the people of District Faisalabad and Sheikhupura, Punjab Province, Pakistan. The retrospective study was conducted on more than thirty thousand people including both male and female patients admitted to the Tehsil Headquarter Hospital, Safdarabad and The Best Hospital, Faisalabad. Blood samples were taken from each subject and subsequently ABO and Rh blood groups were evaluated separately. The antigen antibody agglutination slide test for blood grouping (ABO) and Rh were used to assess the blood group frequencies. RESULTS: The frequencies of ABO blood group distribution indicated that blood group B was predominant in the people of Safdarabad followed by O, A and AB respectively. While, among people of Faisalabad, blood group O was predominant followed B, A and AB respectively. Rh negative phenotype was found lesser distributed as compared to the positive Rh phenotype.

Mitochondrial Transcriptome Control and Intercompartment Cross-Talk During Plant Development.

We address here organellar genetic regulation and intercompartment genome coordination. We developed earlier a strategy relying on a tRNA-like shuttle to mediate import of nuclear transgene-encoded custom RNAs into mitochondria in plants. In the present work, we used this strategy to drive trans-cleaving hammerhead ribozymes into the organelles, to knock down specific mitochondrial RNAs and analyze the regulatory impact. In a similar approach, the tRNA mimic was used to import into mitochondria in Arabidopsis thaliana the orf77, an RNA associated with cytoplasmic male sterility in maize and possessing sequence identities with the atp9 mitochondrial RNA. In both cases, inducible expression of the transgenes allowed to characterise early regulation and signaling responses triggered by these respective manipulations of the organellar transcriptome. The results imply that the mitochondrial transcriptome is tightly controlled by a "buffering" mechanism at the early and intermediate stages of plant development, a control that is released at later stages. On the other hand, high throughput analyses showed that knocking down a specific mitochondrial mRNA triggered a retrograde signaling and an anterograde nuclear transcriptome response involving a series of transcription factor genes and small RNAs. Our results strongly support transcriptome coordination mechanisms within the organelles and between the organelles and the nucleus.

Organellar non-coding RNAs: emerging regulation mechanisms.

Originally focused on the nuclear and cytosolic compartments, the concept of regulation driven by non-coding RNAs (ncRNAs) is extending to mitochondria and chloroplasts. These organelles have distinct genetic systems that need coordination with cellular demands. In mammals, nuclear-encoded microRNAs were found associated with the mitochondria. Some of these contribute to the regulation of mitochondrial transcription and translation. Others were proposed to be stored in the organelles and to be released for regulation of nuclear transcripts. Further ncRNAs of various sizes derive from the mitochondrial genome and it was speculated that organelles host antisense or RNA interference pathways. Long ncRNAs mapping to the mitochondrial DNA seem to operate in the nucleus. Altogether, the origin and trafficking of ncRNAs categorized as mitochondrial in mammals raise questions far beyond the current knowledge. In protozoa, hundreds of guide RNAs specify editing events needed to generate functional messenger RNAs. Only few ncRNAs have been reported in plant mitochondria, but editing sites were revealed in non-coding regions of the organellar genome, suggesting that the corresponding transcripts have a function. Conversely, numerous ncRNA candidates were identified in chloroplasts, essentially mapping to the plastid genome. A synthetic view of the data with their functional implications is given here.