Antimicrobial Efficacy of Biogenic Cobalt and Copper Nanoparticles against Pathogenic Isolates.

Biogenic synthesis of cobalt (Co) and copper (Cu) nanoparticles (NPs) was performed using the bacterial strains Escherichia coli and Bacillus subtilis. Prepared NPs were confirmed by a color change to maroon for CoNPs and green for CuNPs. The NPs characterization using FTIR showed the presence of functional groups, i.e., phenols, acids, protein, and aromatics present in the Co and CuNPs. UV-vis spectroscopy of E. coli and B. subtilis CuNPs showed peaks at 550 and 625 nm, respectively. For E. coli and B. subtilis CoNPs, peaks were observed at 300 nm and 350 nm, respectively. Antibacterial and antifungal activity of B. subtilis and E. coli Co and CuNPs was determined at 100 mg/mL concentration against two bacterial strains at 5, 2.5, and 1.5 mg/mL against fungal two strains F. oxysporum and T. viridi, respectively. B. subtilis CuNPs showed significantly higher inhibition zones (ZOI=25.7-29.7 mm) against E. coli and B. subtilis compared to other biogenic NPs. Likewise, B. Subtilis CuNPs showed lower MIC (4.3 ± 6.3) and MBC (5.3 mg/mL) values against both tested isolates. Antifungal activity of B. subtilis and E. coli CuNPs and CoNPs showed a concentration-dependent decrease in ZOI. Among all biogenic NPs, B. subtilis CoNPs showed the highest ZOI (25-30 mm) against F. oxysporum followed by E. coli CuNPs with maximum ZOI (20-27 mm) against T. viridi. Again, B. subtilis CoNPs and E. coli CuNPs showed lowest MIC and MFC values against both fungal isolates. In conclusion, the current study showed that biogenically synthesized B. subtilis Cu or CoNPs can be used as effective antimicrobial agents due to their potential antibacterial and antifungal potential.

Diversity and Abundance of Delineated Earthworm (Annelida: Clitellata) in Pakistan: A Review.

Earthworm, a ubiquitous (but neglected) macro-invertebrate, is found in terrestrial vicinity of Pakistan. Moreover, the occurrence of earthworms is often diverse with fluctuating quantity depending upon abiotic factors and land usage patterns. The aim of this study was to summarize all the reported information related to earthworm diversity in different areas of Pakistan. Almost all the data published from year 2001 to 2021 were collected. Following data organization, total 42 earthworm's species including five families (Acanthodrilidae, Lumbricidae, Moniligastridae, Octochaetidae and Megascolecidae) were reported from various researchers. Among five families, family Acanthodrilidae was found to have only one specie (Ramiella bishambari), Lumbricidae consist of 10 species (Apporactodea rosea, Allolobophora trapezoids, Allolobophora chloroticaa, Aporrectodea longa, A. caliginosa, Bimastus parvus, Eisenia fetida, Helodrilus foetidus, Lumbricus terrestris and L. rubillus), Moniligastridae has two species (Drawida nepalensis and D. pellucida) while Octochaetidae possess only one specie (Eutyphoeus incommodus). The most abundant and diverse family Megascolecidae consist of 28 earthworm species in all habitats of different regions of Pakistan. Among geographical areas, Faisalabad was found as the richest territory with most reported earthworm species (i.e. 28). The current study suggests further in depth research to explore the unidentified and/ missing species of earthworms in Pakistan.

Phenotypic and Molecular Characterization of CTX-M Type B-Lactamases in Gram Negative Bacterial Strains Isolated from Hospitals, Lahore, Pakistan.

One of the principal mechanisms that contribute resistance to antibiotics is the production of extended spectrum beta lactamase (ESBL) in Gram negative bacteria. In the present study, molecular methods were used to evaluate the prevalence of the extended-spectrum beta-lactamase (ESBL)-encoding CTX-M gene among Gram negative bacterial strains. In total, 148 clinical samples were collected from different tertiary care hospitals of Lahore, Pakistan. Disc synergy diffusion method was used to detect the presence of ESBL production. Moreover, antibiotic resistance patterns and molecular detection of bla CTX-M ESBLs, were also studied. The pathogens isolated from the 148 samples included Escherichia coli (43%) followed by Klebsiella sp. (28%), Proteus sp. (18%) and Pseudomonas sp. (11%). In all 148 strains, 95 (64%) were ESBL producers while 53 (36%) were non ESBL producers. The strains which were phenotypically ESBL producers, bla CTX-M were found in 46% E. coli strains, while 50% Klebsiella sp. were harboring the gene. A high resistance rate was observed against cephalosporins (cefopodoxime 67%, cefoperazone 73%, cephalexin 63% sparaxin 61%). Lower resistance was observed against meropenem among all isolated bacterial strains. Genotypic detection of bla CTX-M genes by PCR revealed 46% of E. coli and 50% of Klebsiella strains harbored bla CTX-M gene. The present study showed that ESBLs producers were resistant to commonly used antibiotics. Similarly, bla CTX-M ESBL production is more prevalent in our clinical isolates.

Bacillus licheniformis (MN900686) Mediated Synthesis, Characterization and Antimicrobial Potential of Silver Nanoparticles.

The use of bacteria in the synthesis of silver nanoparticles (AgNPs) emerges as an ecofriendly and exciting approach. In the present study, we reported the biosynthesis of AgNPs by using culture supernatant of the bacteria Bacillus licheniformis (MN900686). The biogenically synthesized AgNPs were confirmed by the change in the color of the culture filtrate from yellow to brown after the addition of AgNO3. Further characterization performed by means of UV vis-spectroscopy showed absorption peak at 414 nm which confirmed the formation of AgNPs. Fourier Transfer infrared (FTIR) confirmed the involvement of biological molecules in the formation of nanoparticles (NPs). The SEM revealed that the NPs have approximately 38 nm size. The agar well diffusion assay was used to determine antibacterial activity while tube dilution method was used to determine minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The human pathogenic bacterial strains i.e., P. aeruginosa (MN900691) and B. subtilis (MN900684), were used as test strains. The anti-bacterial assay against test strains revealed that these NPs showed concentration dependent increased zone of inhibition (ZOI). The maximum ZOI at 25 µL of AgNPs was 20 mm against B. subtilis after 24 hours of incubation. One-way ANOVA test showed significant ZOI (p ≤ 0.05) against B. subtilis. The MIC was ranged from 4.3-6.6 µg/mL while MBC ranged from 8.3 to 6.6 µg/mL. Overall, this study suggested that the biogenically synthesized NPs are an effective alternative source of antimicrobials against pathogenic bacteria.

Photosynthesis and Salt Exclusion Are Key Physiological Processes Contributing to Salt Tolerance of Canola (Brassica napus L.): Evidence from Physiology and Transcriptome Analysis.

Plant salt tolerance is controlled by various physiological processes such as water and ion homeostasis, photosynthesis, and cellular redox balance, which are in turn controlled by gene expression. In the present study, plants of six canola cultivars (DGL, Dunkled, Faisal Canola, Cyclone, Legend, and Oscar) were evaluated for salt tolerance by subjecting them to 0 or 200 mM NaCl stress. Based on growth, cultivars DGL, Dunkled, and Faisal Canola were ranked as salt tolerant, while cultivars Cyclone, Legend, and Oscar were ranked as salt-sensitive ones. Differential salt tolerance in these canola cultivars was found to be associated with a relatively lower accumulation of Na+ and greater accumulation of K+ in the leaves, lower oxidative damage (MDA), and better antioxidative defense system (Superoxide dismutase, SOD; peroxidase, POD, and catalase, CAT). Cultivar Oscar was the poorest to discriminate Na+ and K+ uptake and accumulation in leaves and had poor antioxidant potential to scavenge ROS. Salt stress did not affect the structural stability of photosystem-II (PSII) till three weeks, thereafter it caused a significant decrease. Salt stress increased the performance index (PIABS) by increasing the density of active reaction centers in Oscar. Salt stress decreased the antenna size thereby lowering the absorption and trapping energy flux, and maintaining the electron transport with an increase in heat dissipation. This may represent a potential mechanism to cope with salt stress. Transcriptome analysis of salt-sensitive cultivar Oscar further revealed that salt stress down-regulated DEGs related to hormonal signal transduction pathways, photosynthesis, and transcription factors, while DEGs related to the biosynthesis of amino acid and ion transport were up-regulated. In conclusion, salt tolerance in canola cultivars was associated with ion exclusion and maintenance of photosynthesis. Salt stress sensitivity in cultivar Oscar was mainly associated with poor control of ion homeostasis which caused oxidative stress and reduced photosynthetic efficiency.

Foliar fertigation of ascorbic acid and zinc improves growth, antioxidant enzyme activity and harvest index in barley (Hordeum vulgare L.) grown under salt stress.

Crop productivity is limited by several environmental constraints. Among these, salt stress plays a key role in limiting the growth and yield production of economically important agricultural crops. However, the exogenous fertigation of vitamins and minerals could serve as a "shot-gun" approach for offsetting the deleterious effects of salts present in the rhizosphere. Therefore, an experiment was conducted to quantify the efficacy of foliar fertigation of ascorbic acid (vitamin-C) and zinc (Zn) on the physio-biochemical attributes of barley (Hordeum vulgare L. Genotype B-14011) grown in a saline environment. The salt stress resulted in a reduced biological yield associated with a decrease in chlorophyll pigment, while a significant enhancement in Na+ and Zn2+ was observed under salinity stress. Similarly, the contents of total soluble proteins, total free amino acids, lipid peroxidation, and H2O2 and the activities of antioxidative enzymes (SOD, POD, CAT, APX and proline) were significantly enhanced under salinity stress. Moreover, salinity negatively affected the yield attributes and ion uptake of plants. However, foliar fertigation with AsA +0.03% Zn enhanced vegetative growth, photosynthetic pigments, synchronized ion uptake, the synthesis of enzymatic and non-enzymatic antioxidants, and the harvest index. It is inferred from this study that among all treatments, the effect of foliar fertigation with the AsA+0.03% Zn combination not only improved the salt stress tolerance but also improved the yield attributes, which will aid in the improvement in barley seed yield and is a step to solve the problem of malnutrition through biofortification of vitamin-C and zinc.

RNAseq Analysis Reveals Altered Expression of Key Ion Transporters Causing Differential Uptake of Selective Ions in Canola (Brassica napus L.) Grown under NaCl Stress.

Salinity is one of the major abiotic stresses prevailing throughout the world that severely limits crop establishment and production. Every crop has an intra-specific genetic variation that enables it to cope with variable environmental conditions. Hence, this genetic variability is a good tool to exploit germplasms in salt-affected areas. Further, the selected cultivars can be effectively used by plant breeders and molecular biologists for the improvement of salinity tolerance. In the present study, it was planned to identify differential expression of genes associated with selective uptake of different ions under salt stress in selected salt-tolerant canola (Brassica napus L.) cultivar. For the purpose, an experiment was carried out to evaluate the growth response of different salt-sensitive and salt-tolerant canola cultivars. Plants were subjected to 200 mM NaCl stress. Canola cultivars-Faisal Canola, DGL, Dunkled, and CON-II-had higher growth than in cvs Cyclone, Ac-EXcel, Legend, and Oscar. Salt-tolerant cultivars were better able to maintain plant water status probably through osmotic adjustment as compared to salt-sensitive cultivars. Although salt stress increased shoot Na+ and shoot Cl- contents in all canola cultivars, salt-tolerant cultivars had a lower accumulation of these toxic nutrients. Similarly, salt stress reduced shoot K+ and Ca2+ contents in all canola cultivars, while salt-tolerant cultivars had a higher accumulation of K+ and Ca2+ in leaves, thereby having greater shoot K+/Na+ and Ca2+/Na+ ratios. Nutrient utilization efficiency decreased significantly in all canola cultivars due to the imposition of salt stress; however, it was greater in salt-tolerant cultivars-Faisal Canola, DGL, and Dunkled. Among four salt-tolerant canola cultivars, cv Dunkled was maximal in physiological attributes, and thus differentially expressed genes (DEGs) were assessed in it by RNA-seq analysis using next-generation sequencing (NGS) techniques. The differentially expressed genes (DEG) in cv Dunkled under salt stress were found to be involved in the regulation of ionic concentration, photosynthesis, antioxidants, and hormonal metabolism. However, the most prominent upregulated DEGs included Na/K transporter, HKT1, potassium transporter, potassium channel, chloride channel, cation exchanger, Ca channel. The RNA-seq data were validated through qRT-PCR. It was thus concluded that genes related to the regulation of ionic concentrate are significantly upregulated and expressed under salt stress, in the cultivar Dunkled.

Microbial assessment of pathogenic bacterial growth in ice cream and kulfa.

Current study was focused to examine the total bacterial count in packed and unpacked ice cream and kulfa collected from 12 different localities of Lahore. The bacterial colonies were isolated and grown on agar-broth media under sterilized conditions. Serial dilution technique was used to compose the replicates to get total viable count of bacteria. Results in case of packed ice cream samples indicated maximum (618 × 10-6 CFU/g) and minimum (79 × 10-6 CFU/g) bacterial count while in case of unpacked ice cream samples maximum and minimum bacterial count was 163 × 10-6 CFU/g and 71 × 10-6 CFU/g, respectively. Whereas in case of packed kulfa samples, maximum and minimum recorded bacterial count was 163 × 10-6 CFU/g and 72 × 10-6 CFU/g, respectively. The LM and SEM of the isolated bacteria were also performed for correct identification. Results indicated that the total bacterial count recorded in the samples exceeded the standard tolerable range which can lead to serious health damage of consumers.