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Advancements in Life Sciences, volume 7, issue 3Cover; Volume 7, Issue 3
Published online: 25 May 2020
ISSN 2310-5380 

IN THIS ISSUE

 

Short Communication

Domestic animals’ identification using PCR-RFLP analysis of cytochrome b gene
Murad Ali Rahat, Muhammad Haris, Zaib Ullah, Saba Gul Ayaz, Mian Nouman, Akhtar Rasool, Muhammad Israr, pages 113-116
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 Background: Species identification is an important process to identify the origin of meat, adulteration and for  cooked and processed meat. The present study was conducted to identify cattle (Bos taurus) and buffalo (Bubalus bubalis) by using mitochondrial cytochrome-b (Cyt-b) gene. Size of the gene is 1140 bp, but we amplified 359 bp that is cleaved by specific restriction endonucleases. The aim of this study was species identification through Cyt-b gene by using PCR-RFLP analysis.

Methods: For this study, 55 blood samples were collected from different species of domestic animals. The DNA was extracted from the whole blood through blood extraction kit. The DNA of these samples were amplified through PCR using universal Cyt-b primers. The amplified product was treated with restriction enzymes Alu I. The resultant fragments were viewed on 3.0 % agarose gel.

Results: Cyt-b gene was amplified of all included animals. Different bands were observed as compared with 50 bp DNA ladder. Animals were identified on the base RFLP mediated by Alu1 restriction enzyme.

Conclusion: We identified domestic animals on the basis of Mitochondrial Cyt-b gene by the process of PCR-RFLP. To identify specific animals through RFLP, a larger sample size and confirmation by gene sequence analysis may be helpful. 

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Full Length Research Articles


Risk factors associated with relapse of drug dependence after treatment and rehabilitation in areas under the influence of war on terror
Muhammad Abdullah, Muhammad Imran Khan, Faiza Mumtaz, Faridullah Shah, Rosana C Ximenes, Vahid Nikoui, Abdul Wahab,
 pages 117-121
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 Background: People exposed to war on terror are more prone to neuropsychiatric disorders and drug addiction. The present study was aimed to investigate the risk factors associated with relapse of drug dependence after treatment and rehabilitation in areas under the influence of war and terrorism.

Methods: Total 57 individuals who had a relapse from the war affected area were included in current study. Retrospective data were collected from Drug Detoxification and Health Welfare Research Center, Bannu, KPK, Pakistan. Along with demographic characteristics, current study also focused on the prevalence of psychological problems due to war, in association with prevalence of relapse.

Results: A total of 93% of Individuals with relapse have concurrent stress. Similarly, depression was also common in 78.9% of relapsed individuals. Relapse was more common in Polydrug abusers (66.7%) as compared to single drug abusers. As far as abstinence duration is concerned, subjects having depression and stress due to war, relapsed in less time as compared to individuals with no psychological disorders.

Conclusion: Wars are related with mental and psychological problems like stress and depression and these factors significantly contribute to the relapse of drug dependence as suggested by results of current study. 

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Molecular cloning and expression of recombinant Trichoderma harzianum chitinase in Pichia pastoris
Muhammad Saleem Iqbal Khan, Anwar Khan, Olawale Samuel Adeyinka, Iqra Yousaf, Saman Riaz, Bisma Bashir, Muhammad Tariq, Bushra Tabassum, pages 122-128

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 Background: The importance of chitinases over the years had attracted huge biotechnological attention because its usage cut across wide range of field. It plays a significant role in the defensive mechanism against fungal pathogens.

Methods: In this study, an endochitinase gene was isolated from Trichoderma harzianum, and characterized in-silico by using various bioinformatics tools. Further, the gene was cloned in eukaryotic expression vector (pPICZA) under the control of AOX1 promoter for recombinant expression in Pichia pastoris GS115 host strain.

Results: The chitinase cDNA was ~1000 bp long, while in-silico studies revealed an open reading frame of 888 bp encoding 295 amino acids with a calculated molecular mass of 37332.76 Da and an estimated isoelectric point of 4.07. Recombinant chitinase protein expressed intracellularly and revealed high expression in P. pastoris host. The 37 kDa recombinant chitinase protein developed with antigen antibody confirmed its expression in P. pastoris.

Conclusion: Conclusively, T. harzianum derived chitinase gene was successfully over expressed in P. pastoris where recombinant protein was expressed intracellular in the form of inclusion bodies. 

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Introduction to Galaxy Platform for NGS Variant Calling Pipeline
Rashid Saif, Aniqa Ejaz, Tania Mehmood, Fatima Asif, Suliman Mohammad Alghanem, Talha Saleem Ahmad, pages 129-134

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 Background: Galaxy web-based platform for Next Generation Sequence (NGS) data analysis provides unprecedented opportunities to characterize, analyze and computationally visualize genomic landscapes with limited-resources. An initiative was taken to explore this pipeline for NGS data-analysis by using Galaxy platform, for its relative accessibility, reproducibility, transparency and scalability.  

Methods: Variant calling and associated workflows were executed on NGS pooled-seq data of 12 Pakistani Teddy goats. Different tools used in this pipeline are FastQC for quality checks, Trimmomatic for trimming data, SAM/BAM tools for conversion of file formats, Picard tools for marking deduplicates, VCFtools/FreeBayes for genomic variant detection and SnpSift to annotate the variants.

Results: Highly associated functionally untrivial 43,712 loci were percolated having 87,510 alleles. Besides, 1,548 variants with 1,134 SNPs, 23 mixed variants, 76 MNP, 183 insertions and 132 deletions were observed in Teddy breed using San Clement ARS1 reference genome. Furthermore, 1,283 homozygous and 265 heterozygous variant were also divulged out of 43,447 loci. These variants are likely to be liable for general phenotypic traits of Teddy with smaller body-size, tender meat quality and agility along with other breed specific traits. 

Conclusion: Galaxy fulfills the core function of reproducibility and easy accessibility by removing the gaps between large data analysis and its interpretations. This variant calling pipeline reveals the genomic differences of Teddy specific characteristics as compare to ARS1 reference genome. 

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Competence of Benzoil Tree (Moringa Oleifera L.) as Antibacterial and Antifungal Agent
Khushnood ur Rehman, Muhammad Hamayun, Sumera Afzal Khan, Shahab Saeed Khan, Sher Wali, pages 135-139

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 Background: A plant’s activity towards biological properties is the first step to consider it for medicinal and therapeutic purposes. To evaluate the medicinal properties, we have determined the anti-bacterial and anti-fungal potential of Moringa oleifera L. (Benzoil tree). The focus was to obtain and isolate certain chemical substances that can neutralize the effect of common and pathogenic selected bacterial and fungal species collected from local hospitals of Khyber Pakhtunkhwa.

Methods: A total of five fractions were selected i.e. crude methanolic extracts, n-hexane, chloroform, ethyl acetate and aqueous extracts were prepared and their activity checked against four bacterial strains including Streptococcus mutans, Staphylococcus aurous, MRSA (methicillin resistance Staphylococcus aurous) and Serratia marcescens; and four fungal strains i.e. Fusarium oxysporum, Aspergillus flavus, Polysphondylium pallidum and Alternaria alternata.

Results: The highest anti-bacterial activity shown by crude methanolic extract fraction i.e. (48-38%) and the lowest activity was exhibited by aqueous extract (15-0%) against all the selected bacterial strains. Similarly, the highest anti-fungal activity indicated by crude methanolic extracts (60-45%) against the four selected fungal species and lowest activity shown by the aqueous fractions (26-0%). Reasonable activity was also exhibited by others fractions as well.

Conclusion: The outcomes of our experiment strongly supports that Moringa oleifera has significant antibacterial and antifungal activities, so the plant is effective antibacterial and antifungal agent. 

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Rhizobacterial mediated antimetabolites and their significant effect on human pathogens
Unaiza Malik, Atia Iqbal, Sadia Batool, pages 140-145

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 Background: The emergence of antibiotic resistant bacteria is increasing globally. Therefore, new strategies and exploration of new metabolites are need of the day to combat various diseases.  The aim of the current study is to isolate the rhizobacterial strains from medicinal plants to examine their antibacterial activity against human pathogens.

Methods: The total 239 rhizobacterial isolates were screened with agar plug and well diffusion methods, characterized biochemically and identified by Bergey’s manual scheme. In chemical screening, metabolic extracts of rhizobacterial isolates were separated in thin layer chromatography (TLC) and further identified by HPLC. The ability of rhizobacterial strains for optimum metabolite production was checked in vitro under different environmental conditions.

Results: Results showed that 31% isolates had antibacterial activity against Bacillus cereusBacillus subtilisE. coccusE. coliSalmonellaKlebsiella, Staphylococcus aureus ATCC20737, Staphylococcus aureus ATCC25923 and Neisseria. Gonorrhea ATCC19424, methicillin resistant Staphylococcus aureus 1, methicillin resistant Staphylococcus aureus 6 and methicillin resistant Staphylococcus aureus 8.  Maximum rhizobacterial strains showed best antibacterial activity at 35C and at pH7. ST5 strain showed significant results against E. coli ETCC and methicillin resistant Staphylococcus aureus-7 (25mm zone of inhibition). The bands of ST5 and ST2 extracts on a TLC plate were detected at Rf value 4.7 and 4.8 respectively. In HPLC, different peaks at different retention times with maximum peak at 2.90 and 2.92 indicating the production of antimetabolites.

Conclusion: These rhizobacterial isolates are valuable inputs in natural consortium to produce specific antibacterial metabolites. Demonstrating the structure and purification of these bioactive compounds may enhance the antimicrobial activities which further can be helpful in development of antibiotics. 

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Evaluation of Antibiotics Pattern of Extended Spectrum Beta-Lactamase Producing Multi-Drug Resistant Pseudomonas aeruginosa
Anees Muhammad, Ihsan Ali, Muhammad Owais, Sadiq Noor Khan, Irfan Qadir Afridi, Nasir Ali, pages 146-150

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 Background: Pseudomonas aeruginosa (Ps. aeruginosa) is considered as an opportunistic pathogen and the leading cause of morbidity and mortality in immunocompromised individuals. Globally, approximately 10-15% of the nosocomial infections are caused by Ps. aeruginosa. The Ps. aeruginosa can acquire resistance against broad-spectrum antibiotics. According to recent studies increased mortality has been observed due to infection with extended-spectrum-beta-lactamase (ESBL) producing Ps. aeruginosa strains. This study was designed to determined antibiogram of ESBL producing multi-drug resistant Ps. aeruginosa in Khyber Pakhtunkhwa.

Methods: The clinical confirmed Ps. aeruginosa samples were collected according to the standard protocol, at Khyber Teaching Hospital (KTH), Peshawar. All collected samples were sub- cultured on appropriate culture media. After isolation and identification, the antibiotics susceptibility testing was performed. The detection of ESBL was carried out by the double-disc diffusion method. Carbapenemase-producing bacteria was confirmed by the modified Hodge test. Descriptive analysis was performed for statistical analysis of collected data.

Results: A total of one hundred and sixty-two (n=162) Ps. aeruginosa confirmed isolates were collected, in which 59.3% were male and 40.7% were from female patients. The percentages of ESBL and carbapenemase producing Ps. aeruginosa isolates were 5.5% and 23.5%, respectively. The multidrug resistance was observed against 27.2% isolates. Among tested antibiotics highest percentages of resistance was observed against ciprofloxacin (43%) and ceftazidime (39.5%). 

Conclusion: We observed highest level of drug resistance in Ps. aeruginosa clinical isolates against tested antibiotics and majority of the isolates were Multi-drug resistant (MDR). 

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Benefit Cost Ratio of Buds Chips Planting and its Effects on Yield and Quality of Sugarcane
Abdul Khaliq, Arshad Mahmood, Hafiz Bashir Ahmad, Muhammad Ashfaq Nadeem, Naeem Ahmad, Rashad ul Sher, Muhammad Rizwan Khursheed, pages 151-156

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 Background: High price and poor quality of seed material is a great concern to Farmers. Sugarcane seed is 20% of total cost of production. Cane Growers used setts having 2-3 buds with 8-10 t ha-1 cane stalk as Seed. The large quantity of seed carriages a great problematic in handling, transport, seed treatment for diseases, storage ,  viability of buds and their germination. The use of bud chips is less bulky, easily handling in seed treatment, storage and transportation.

Methods: In this view, a research experiment was designed at Sugarcane Research Institute, Faisalabad during 2016 and 2017, to study the feasibility of bud chips as an alternate to conventional planting in sugarcane crop. The experiment was planted with sugarcane variety CPF-252 in randomized complete block design with split plot arrangement having three replications.

Results: The results of study revealed that setts planting on 15 April gave maximum cane (61 t ha-1) and sugar yield (7.15 t ha-1). While bud chips sown on 15 April produced higher yield. The benefit cost ratio (BCR) was high in bud chips planting (1.63) than setts planting of sugarcane.

Conclusion: The study concludes that bud chip planting on 15 April is an improved technique with greater potential to provide disease free seed, improve the yield with minimum quantity of seed than conventional planting. 

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DNA-based Eye Color Prediction of Pakhtun Population Living in District Swat KP Pakistan
Murad Ali Rahat, Hakim Khan, Ishtiaq Hassan, Muhammad Haris, Muhammad Israr, pages 157-163

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 Background: Forensic DNA Phenotyping (FDP) or the prediction of Externally Visible Characteristics (EVCs) from a DNA sample has gained importance in the last decade or so in the forensic community. If and when the traditional forensic DNA typing via Short Tandem Repeats (STR) fails due to the absence of a reference sample, an individual can be traced by a DNA sample using FDP. Amongst the many available EVCs, eye color is one such character that can be predicted by employing previously developed IrisPlex system using Single Nucleotide Polymorphism (SNP) assay. In this study, we applied the IrisPlex system to samples collected from population of District Swat for prediction of eye colours from DNA.

Method: Eye colour digital photographs and buccal swab samples were collected from 267 Pakhtun individuals of District Swat. Any person with eye disease was excluded from the study. Genomic DNA was extracted through Phenol-Chloroform extraction method. The amplified SNPs were typed using Multiplexed Single Base Extension (SBE). The genotypes were checked for eye color phenotypes through IrisPlex online tool and correlation were checked between SNPs, Gender, pie score and eye color.

Result: Brown eye color was found prevalent as compared to intermediate and blue. Females have highly brown eye color compared to males while males have intermediate and blue. Three SNPs rs12913832 (in the HERC2), rs1393350 (TYR gene), rs1800407 (OCA2 gene) were strongly significant to eye color. Pie score was also significant to eye color and rs12913832 SNP. IrisPlex analysis in 20 individuals of District Swat was performed. The prediction accuracy of IrisPlex for blue or brown was 100% in the studied individuals. However, the IrisPlex tool predicted the intermediate phenotype incorrectly as brown or blue.

Conclusion: It is concluded from the data that intermediate eye colour was not predicted accurately, therefore, inclusion of more SNPs in the IrisPlex system is needed to predict intermediate eye colour accurately. 

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CT analysis of subcutaneous and visceral adipose tissue in normal BMI subjects: association with level of physical activity and hypertension
Hafiz Muhammad Rizwan, Zareen Fatima, Shoaib Rasool, Iqra Ilyas, Khalid Idrees, Taiba Zulfiqar, Maria Mohsan, Arshad Jamal, Ahmed Bilal Waqar, pages 156-169

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 Background: The distribution of adipose tissue, complex factors affecting it and its pathological consequences are among the hot topics in medical research nowadays. Most of the studies reported in the literature however describe the association of factors affecting the fat distribution in overweight and obese individuals. This particular study was however planned to find out the same in subjects having normal basal metabolic index (BMI). The objectives of the study were to analyze total adipose tissue (TAT), subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) in the abdomen volumetrically using CT, to establish the association of these to the levels of physical activity, presence or absence of hypertension and to compare these associations in both the genders.

Methods: A prospective study was carried out on seventy five, normal BMI subjects aged between 20–50 years. CT imaging was used for volumetric measurement of TAT, SAT and VAT. Pearson’s correlation of these were then found out with age. Kruskal Wallis test was also performed to compare these in hypertensive and non-hypertensive subjects and in those with different physical activity levels (PAL).

Results: Women showed significantly higher volumes of TAT and SAT. Men showed statistically significant correlations of TAT and VAT with age. SAT volumes had significant negative association with the PAL in both genders. Men showed higher responsiveness of fat deposition in all compartments to the presence of hypertension.

Conclusion: In conclusion, factors such as gender, age, level of physical activity and hypertension affect the site specific deposition of fat even in those individuals who aren’t over-weight or obese. 

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Antagonistic, biofilm-forming rhizospheric Pseudomonas spp. isolated from Hail province
Mousa M. Alreshidi, Abdel Moneim E. Sulieman, Vajid N. Veettil, Mejdi Snoussi, pages 170-176

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 Background: The objectives of this study were to characterize Pseudomonas rhizospheric strains, that have a biocontrol activity, in rhizosphere soil in Hail province and study their ability to form biofilm.

Methods: Rhizosphere soil samples were collected from rhizosphere of soil plantation areas, to be used for bacteria isolation. The identified bacteria were qualitatively tested for their ability to produce slime and subsequently develop biofilm.

Results: The cultural and biochemical identification techniques, including morphological, biochemical and molecular methods revealed that the antagonistic bacteria- from the distinctive rhizosphere soil samples belong to Pseudomonas genus in particular, P. aeruginosa (PF1a, pf2a, PF-8) and P. putida (PF-7). These identified isolates inhibited Aspergillus niger development with percentage of parasitic growth inhibition greater than (48.095 ± 2.182)% for P. aeruginosa (pf-8). In addition, these identified isolates were significantly shown to be able to produce exopolysaccharide and subsequently develop biofilm on polystyrene and glass surfaces.

Conclusion: Superior strains of these bio-control and plant growth promoting rhizobacteria will enable for better biological control of fungal and bacterial plant diseases and may reduce chemical pesticide usage. The indigenous strains isolated could potentially have a great impact on controlling plant diseases, in particular, those caused by microorganisms, and could be used as an alternative bio control agents instead of harmful chemical pesticides. Most of the tried microbes produced exopolysaccharides as well as formed biofilm on polystyrene and glass surfaces. 

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Study of diagnostic and prognostic parameters in Breast cancer patients without metastasis
Aqsa Neelam, Ahmad Ali, Rabail Alam, Ghulam Mujtaba, Imran Riaz Malik, pages 177-180

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 Background: Carcinoma of the breast is the most prevalent cause of mortality from cancer in women aged 40-69 years. The aim of the present study was to examine any alteration in the status of lactate dehydrogenase, ferritin, gamma glutamyl transferase, Platelets, hemoglobin, alkaline phosphatase, aspartate transaminase, alanine transferase and bilirubin in the plasma of breast cancer patients without distant metastasis, to establish their diagnostic and prognostic values.

Methods: Current reading describes a study conducted on 50 breast cancer patients from Institute of Nuclear Medicine and Oncology (Lahore). The patients were clinically categorized as stage II (19 patients) and stage III (29 patients) respectively. Most of the patients were diseased with infiltrative ductal carcinoma (48) of the breast. Blood samples of all patients were collected after forty days of chemotherapy course besides 10 healthy subjects.

Results: There was a significant rise in LDH (221.48 ± 18.44) and Ferritin (356.46 ± 12.28) levels as compared to control groups. Concentration of GGT (34.12 ± 1.61) was also elevated but not significantly raised. There was no significant rise in ALP (279.27 ± 26.60), AST (49.44 ± 8.510), ALT (33.40 ± 2.83), bilirubin (4.87 ± 3.96), Hb (11.16 ± .299) and platelets (032.54 ± 21.68) levels in comparison to normal control subjects. An elevation of LDH and ferritin levels in cases of carcinoma breast signifies its importance as a biomarker of disease. A serial measurement of these enzymes would have diagnostic and prognostic significance and help treatment decisions.

Conclusion: The ferritin along with lactate dehydrogenase can be used as a valuable biomarker for breast cancer diagnosis and prognosis. 

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Review Article


Alternative approaches for the treatment of Asthma and COPD: Focus on Cell-based therapies, Epigenetics, and Gene silencing approaches
Sibtain Afzal, Khushnooda Ramzan, Ahmed Bilal Waqar, pages 181-189

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 Despite many organized health initiatives and critically acclaimed guidelines for proper management of asthma therapy, there is still a large population of severe asthmatics having an uncontrolled disease. Severe persistent asthma, characterized by chronic airway inflammation, increased eosinophils and serum IgE is currently managed by using inhaled corticosteroids. It is quite challenging to get the best treatment guidelines for bronchial asthma in severe asthmatics, particularly in the presence of steroid resistance and the non-responsiveness to β-agonists. For that purpose, other methodologies are required to reverse the uncontrolled airway remodeling in steroid-resistant severe asthma. These advanced alternative approaches should be able to treat asthma symptoms and to improve the inflammatory conditions underlying characteristic pathological features of asthma. The current review focuses and summarizes the alternative approaches used in severe asthma patients. Agents targeting inflammatory cytokines, phosphodiesterase inhibitors, antibodies, oligonucleotides, stem cells, and target drug delivery using gene silencing, offer promise in treating severe asthma. 

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Scope of the Journal

Advancements in Life Sciences (ISSN 2310-5380) provides free platform for the fast peer-reviewed publication of articles covering novel and innovative aspects of biological sciences. Including full length research articles and short communications, we accept material in other categories too view other submission materials. This Journal publishes articles falling in the scope of “life sciences” ranging from molecular biological or genetic discoveries to biochemical systems and new microbiological concepts in human, veterinary and plant sciences. Multidisciplinary research articles, providing novel insight and original information, that would not be publishable in single discipline devoted journals are welcomed particularly in this journal for publication. The following research areas are covered in the Journal:

* Molecular Biology
* Biochemistry
* Plant-biotechnology 
* Genomics
* Veterinary sciences
* Microbiology
* Health and Epidemiological studies

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