Search results for: gene annotation

Authors: Helmi Baazaoui, Mounir Zrigui

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This study addresses the challenge of hate speech detection in Tunisian Arabic text, a critical issue for online safety and moderation. Leveraging the strengths of the AraBERT model, we fine-tuned and evaluated its performance against the Bi-LSTM model across four distinct datasets: T-HSAB, TNHS, TUNIZI-Dataset, and a newly compiled dataset with diverse labels such as Offensive Language, Racism, and Religious Intolerance. Our experimental results demonstrate that AraBERT significantly outperforms Bi-LSTM in terms of Recall, Precision, F1-Score, and Accuracy across all datasets. The findings underline the robustness of AraBERT in capturing the nuanced features of Tunisian Arabic and its superior capability in classification tasks. This research not only advances the technology for hate speech detection but also provides practical implications for social media moderation and policy-making in Tunisia. Future work will focus on expanding the datasets and exploring more sophisticated architectures to further enhance detection accuracy, thus promoting safer online interactions.

Keywords: hate speech detection, Tunisian Arabic, AraBERT, Bi-LSTM, Gemini annotation tool, social media moderation

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Authors: Seyed Esmail Seyedi Bariran, Khairul Salleh Mohamed Sahari

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In multi-echelon supply chain networks, optimal supplier selection significantly depends on the accuracy of suppliers’ performance prediction. Different methods of multi criteria decision making such as ANN, GA, Fuzzy, AHP, etc have been previously used to predict the supplier performance but the “black-box” characteristic of these methods is yet a major concern to be resolved. Therefore, the primary objective in this paper is to implement an artificial intelligence-based gene expression programming (GEP) model to compare the prediction accuracy with that of ANN. A full factorial design with %95 confidence interval is initially applied to determine the appropriate set of criteria for supplier performance evaluation. A test-train approach is then utilized for the ANN and GEP exclusively. The training results are used to find the optimal network architecture and the testing data will determine the prediction accuracy of each method based on measures of root mean square error (RMSE) and correlation coefficient (R2). The results of a case study conducted in Supplying Automotive Parts Co. (SAPCO) with more than 100 local and foreign supply chain members revealed that, in comparison with ANN, gene expression programming has a significant preference in predicting supplier performance by referring to the respective RMSE and R-squared values. Moreover, using GEP, a mathematical function was also derived to solve the issue of ANN black-box structure in modeling the performance prediction.

Keywords: Supplier Performance Prediction, ANN, GEP, Automotive, SAPCO

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Authors: Mohammed Abdelsabour-Khalaf, F. Yusuf , B Brand-Saberi

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Purpose: Applications of nanotechnology nowadays extended to include a wide range of scientific areas such electron micrscopy and gene expression. The aim of the current study was to investigate the developmental expression pattern of genes involved in human glomerulo-nephropathies associated with massive proteinuria and podocyte differentiation using the chicken mesonephros as a model system. Method: We performed in situ hybridization using chicken specific mRNA probes for genes expressed in the early nephron and slit diaphragm genes. The probes used were cNeph1, cNeph2, cSim1, cLmx1b, and cAtoh8. Chicken embryos from Hamburger Hamilton developmental stage HH19 (E3) to HH 34 (E9) were used for the in situ hybridization (ISH). ISH was performed on whole mount embryos which were sectioned by vibratome. Results: Our result show that Neph1, Neph2, Sim1. Lmx1b and Atoh8 genes are dynamically expressed during nephron morphogenesis and Neph1 and Atoh8 are also specifically expressed in the podocytes during late stages of differentiation. Conclusion: We conclude from our results that the genes implicated in congenital and acquired glomerulo-nephropathies like Neph1 and Neph2 are dynamically expressed during mesonephros development pointing towards a role in the formation of the filtration barrier and the differentiation of the mesonephric podocytes. Thus the avian mesonephros could serve as a model to study human kidney diseases.

Keywords: mesonephros, chicken embryo, gene expression, immunohistochemistry

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Authors: Dani Sukkar, Ali Kanso, Philippe Laval-Gilly, Jairo Falla-Angel

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The honeybees' immune system is challenged by different risk factors that induce various responses. However, complex scenarios where bees are exposed to different pesticides simultaneously with immune activation are not well evaluated. The Toll pathway is one of the main signaling pathways studied in invertebrate immune responses, and it is a good indicator of the effect of such complex interactions in addition to key signaling elements of other pathways like Relish of the immune deficiency (IMD) pathway or Eater, the phagocytosis receptor or vitellogenin levels. Honeybee hemocytes extracted from 5th instar larvae were exposed to imidacloprid and/or amitraz with or without the presence of the zymosan a as an immune activator. The gene expression of multiple immune related genes were studied, including spaetzle, Toll, myD88, relish, eater and vitellogenin, by real-time polymerase chain reaction after RNA extraction. The results demonstrated that the Toll pathway is mainly affected by the pesticides; imidacloprid and amitraz, especially by their different combinations. Furthermore, immune activation by zymosan A, a fungal cell-wall component, acts to mitigate to some extent the effect of pesticides on the different levels of the Toll pathway. In addition, imidacloprid, amitraz, and zymosan A have complex and context-specific interactions depending on the levels of immune activation and the pathway evaluated affecting immune-gene expression differently.

Keywords: toll pathway, immune modulation, β-glucan, imidacloprid, amitraz, honeybees, immune genes

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Authors: Aswini M. S.

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Tomato (Solanum lycopersicum L.) is one of the most significant vegetable crops in terms of its economic benefits. Both fresh and processed tomatoes are consumed. Tomatoes have a limited genetic base, which makes breeding extremely challenging. Plant breeding has become much simpler and more effective with genome editing tools of CRISPR and CRISPR-associated 9 protein (CRISPR/Cas9), which address the problems with traditional breeding, chemical/physical mutagenesis, and transgenics. With the use of CRISPR/Cas9, a number of tomato traits have been functionally distinguished and edited. These traits include plant architecture as well as flower characters (leaf, flower, male sterility, and parthenocarpy), fruit ripening, quality and nutrition (lycopene, carotenoid, GABA, TSS, and shelf-life), disease resistance (late blight, TYLCV, and powdery mildew), tolerance to abiotic stress (heat, drought, and salinity) and resistance to herbicides. This study explores the potential of CRISPR/Cas9 genome editing for enhancing yield in tomato plants. The study utilized the CRISPR/Cas9 genome editing technology to functionally edit various traits in tomatoes. The de novo domestication of elite features from wild cousins to cultivated tomatoes and vice versa has been demonstrated by the introgression of CRISPR/Cas9. The CycB (Lycopene beta someri) gene-mediated Cas9 editing increased the lycopene content in tomato. Also, Cas9-mediated editing of the AGL6 (Agamous-like 6) gene resulted in parthenocarpic fruit development under heat-stress conditions. The advent of CRISPR/Cas has rendered it possible to use digital resources for single guide RNA design and multiplexing, cloning (such as Golden Gate cloning, GoldenBraid, etc.), creating robust CRISPR/Cas constructs, and implementing effective transformation protocols like the Agrobacterium and DNA free protoplast method for Cas9-gRNAs ribonucleoproteins (RNPs) complex. Additionally, homologous recombination (HR)-based gene knock-in (HKI) via geminivirus replicon and base/prime editing (Target-AID technology) remains possible. Hence, CRISPR/Cas facilitates fast and efficient breeding in the improvement of tomatoes.

Keywords: CRISPR-Cas, biotic and abiotic stress, flower and fruit traits, genome editing, polygenic trait, tomato and trait introgression

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Authors: O. Ubani, H. I. Atagana, M. S. Thantsha, R. Adeleke

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The oil sludge components (polycyclic aromatic hydrocarbons, PAHs) have been found to be cytotoxic, mutagenic and potentially carcinogenic and microorganisms such as bacteria and fungi can degrade the oil sludge to less toxic compounds such as carbon dioxide, water and salts. In the present study, we isolated different bacteria with PAH-degrading potentials from the co-composting of oil sludge and different animal manure. These bacteria were isolated on the mineral base medium and mineral salt agar plates as a growth control. A total of 31 morphologically distinct isolates were carefully selected from 5 different compost treatments for identification using polymerase chain reaction (PCR) of the 16S rDNA gene with specific primers (16S-P1 PCR and 16S-P2 PCR). The amplicons were sequenced and sequences were compared with the known nucleotides from the gene bank database. The phylogenetical analyses of the isolates showed that they belong to 3 different clades namely Firmicutes, Proteobacteria and Actinobacteria. These bacteria identified were closely related to genera Bacillus, Arthrobacter, Staphylococcus, Brevibacterium, Variovorax, Paenibacillus, Ralstonia and Geobacillus species. The results showed that Bacillus species were more dominant in all treated compost piles. Based on their characteristics these bacterial isolates have high potential to utilise PAHs of different molecular weights as carbon and energy sources. These identified bacteria are of special significance in their capacity to emulsify the PAHs and their ability to utilize them. Thus, they could be potentially useful for bioremediation of oil sludge and composting processes.

Keywords: bioaugmentation, biodegradation, bioremediation, composting, oil sludge, PAHs, animal manures

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Authors: Dedy Pratama, Akhmadu Muradi, Hilman Ibrahim, Patrianef Darwis, Alexander Jayadi Utama, Raden Suhartono, D. Suryandari, Luluk Yunaini, Tom Ch Adriani

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Objective: Diabetic Foot Ulcer (DFU) is one of the complications of Type 2 Diabetes Mellitus (T2DM) that can lead to disability and death. Inadequate vascularization condition will affect healing process of DFU. Therefore, we investigated the expression of polymorphism TGF- β1 in the relation of the occurrence of DFU in T2DM. Methods: We designed a case-control study to investigate the polymorphism TGF- β1 gene 1800469 C > T and 1982073 C > T in T2DM in Cipto Mangunkusumo National Hospital (RSCM) Jakarta from June to December 2016. We used PCR techniques and compared the results in a group of T2DM patients with DFU as the case study and without DFU as the control group. Results: There were 203 patients, 102 patients with DFU and 101 patients control without DFU. 49,8% is male and 50,2% female with mean age about 56 years. Distribution of wild-type genotype TGF-B1 1800469 C > T wild type CC was found in 44,8%, the number of mutant heterozygote CT was 10,8% and mutant homozygote is 11,3%. Distribution of TGF-B1 1982073 C>T wild type CC was 32,5%, mutant heterozygote is 38,9% and mutant homozygote 25,1%. Conclusion: Distribution of alleles from TGF-B1 1800469 C > T is C 75% and T 25% and from TGF-B1 1982073 C > T is C53,8% and T 46,2%. In the other word polymorphism TGF- β1 plays a role in the occurrence and healing process of the DFU in T2DM patients.

Keywords: diabetic foot ulcers, diabetes mellitus, polymorphism, TGF-β1

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Authors: Loubna Safar, Lama Youssef, Majd Aljamali

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Age-related macular degeneration (AMD) is one of the leading causes of blindness worldwide. Complement factor H polymorphism (Y402H) is thought to play a potential role in the predisposition to AMD and response of patients with exudative AMD to treatment with anti-Vascular Endothelial Growth Factor (anti-VEGF). This study aimed to investigate the frequency of Y402H among Syrians, its impact on their susceptibility to AMD, and the hypothesized role of Y402H in patients' response to intravitreal anti-VEGF (i.e.,, bevacizumab). Our case-control study encompassed unrelated 54 AMD cases and 44 controls. Genotyping was determined by standard sequencing of PCR products. Frequency was compared between patients and controls, and correlation between genotype and response to treatment was assessed in 20 patients with wet AMD who received a therapeutic course of three intravitreal bevacizumab injections (once monthly). Our results revealed a significantly higher prevalence of the risk allele C among AMD cases (51.9%) in comparison with controls (37.5%) (P= 0.04, OR= 1.386, CI= 0.999- 1.923). Patients with the TT genotype (no risk allele) exhibited a significantly better primary response rate, reached 87.5% compared to only 41.7% in patients carrying the risk allele C (TC + CC), (P= 0.04, OR= 9.8, CI=0.899- 106.84). The findings of this study prove the importance of investigating Y402H polymorphism as a prognostic marker for predicting response to bevacizumab in AMD patients.

Keywords: age-related macular degeneration, bevacizumab, complement factor H gene, polymorphism, Y402H

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Authors: D. P. Houhoula, J. Papaparaskevas, S. Konteles, A. Dargenta, A. Farka, C. Spyrou, M. Ziaka, S. Koussisis, E. Charvalos

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Objectives: Nanotechnology is providing revolutionary opportunities for the rapid and simple diagnosis of many infectious diseases. Staphylococcus aureus, Listeria monocytogenes and Salmonella enteritis are important human pathogens. Diagnostic assays for bacterial culture and identification are time consuming and laborious. There is an urgent need to develop rapid, sensitive, and inexpensive diagnostic tests. In this study, a gold nanoprobe strategy developed and relies on the colorimetric differentiation of specific DNA sequences based approach on differential aggregation profiles in the presence or absence of specific target hybridization. Method: Gold nanoparticles (AuNPs) were purchased from Nanopartz. They were conjugated with thiolated oligonucleotides specific for the femA gene for the identification of members of Staphylococcus aureus, the mecA gene for the differentiation of Staphylococcus aureus and MRSA Staphylococcus aureus, hly gene encoding the pore-forming cytolysin listeriolysin for the identification of Listeria monocytogenes and the invA sequence for the identification of Salmonella enteritis. DNA isolation from Staphylococcus aureus Listeria monocytogenes and Salmonella enteritis cultures was performed using the commercial kit Nucleospin Tissue (Macherey Nagel). Specifically 20μl of DNA was diluted in 10mMPBS (pH5). After the denaturation of 10min, 20μl of AuNPs was added followed by the annealing step at 58oC. The presence of a complementary target prevents aggregation with the addition of acid and the solution remains pink, whereas in the opposite event it turns to purple. The color could be detected visually and it was confirmed with an absorption spectrum. Results: Specifically, 0.123 μg/μl DNA of St. aureus, L.monocytogenes and Salmonella enteritis was serially diluted from 1:10 to 1:100. Blanks containing PBS buffer instead of DNA were used. The application of the proposed method on isolated bacteria produced positive results with all the species of St. aureus and L. monocytogenes and Salmonella enteritis using the femA, mecA, hly and invA genes respectively. The minimum detection limit of the assay was defined at 0.2 ng/μL of DNA. Below of 0.2 ng/μL of bacterial DNA the solution turned purple after addition of HCl, defining the minimum detection limit of the assay. None of the blank samples was positive. The specificity was 100%. The application of the proposed method produced exactly the same results every time (n = 4) the evaluation was repeated (100% repeatability) using the femA, hly and invA genes. Using the gene mecA for the differentiation of Staphylococcus aureus and MRSA Staphylococcus aureus the method had a repeatability 50%. Conclusion: The proposed method could be used as a highly specific and sensitive screening tool for the detection and differentiation of Staphylococcus aureus Listeria monocytogenes and Salmonella enteritis. The use AuNPs for the colorimetric detection of DNA targets represents an inexpensive and easy-to-perform alternative to common molecular assays. The technology described here, may develop into a platform that could accommodate detection of many bacterial species.

Keywords: gold nanoparticles, pathogens, nanotechnology, bacteria

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Authors: Damien P. Fevre, Peter K. Dearden

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Honeybee population sustainability is a critical concern for environmental stability and human food security. The success of a colony relies heavily on the egg-laying capacity of the queen, as it determines the production of thousands of worker bees who, in turn, perform essential functions in foraging and transforming food to make it digestible for the colony. The main sources of nutrition for honeybees are nectar, providing carbohydrates, and pollen, providing protein. This study delves into the impact of the proportion of these macronutrients on the food consumption patterns of nurse bees responsible for feeding the queen and how it affects the characteristics of the eggs produced. Using nutritional geometry, qRT-PCR, and RNA-seq analysis, this study sheds light on the pivotal role of nutrition in influencing gene expression in nurse bees, honeybee queen egg-laying capacity and embryonic development. Interestingly, while nutrition is crucial, the queen's genotype plays an even more significant role in this complex relationship, highlighting the importance of genotype-by-environment interactions. Understanding the interplay between genotype and nutrition is key to optimizing beekeeping management and strategic queen breeding practices. The findings from this study have significant implications for beekeeping practices, emphasizing the need for an appropriate nutrition to support the social nutrition of Apis mellifera. Implementing these insights can lead to improved colony health, increased productivity, and sustainable honeybee conservation efforts.

Keywords: honeybee, egg-laying, nutrition, transcriptomics

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Authors: Maddalena Arigoni, Maria Luisa Ratto, Raffaele A. Calogero, Luca Alessandri

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The presence of tumor heterogeneity, where distinct cancer cells exhibit diverse morphological and phenotypic profiles, including gene expression, metabolism, and proliferation, poses challenges for molecular prognostic markers and patient classification for targeted therapies. Understanding the causes and progression of cancer requires research efforts aimed at characterizing heterogeneity, which can be facilitated by evolving single-cell sequencing technologies. However, analyzing single-cell data necessitates computational methods that often lack objective validation. Therefore, the establishment of benchmarking datasets is necessary to provide a controlled environment for validating bioinformatics tools in the field of single-cell oncology. Benchmarking bioinformatics tools for single-cell experiments can be costly due to the high expense involved. Therefore, datasets used for benchmarking are typically sourced from publicly available experiments, which often lack a comprehensive cell annotation. This limitation can affect the accuracy and effectiveness of such experiments as benchmarking tools. To address this issue, we introduce omics benchmark experiments designed to evaluate bioinformatics tools to depict the heterogeneity in single-cell tumor experiments. We conducted single-cell RNA sequencing on six lung cancer tumor cell lines that display resistant clones upon treatment of EGFR mutated tumors and are characterized by driver genes, namely ROS1, ALK, HER2, MET, KRAS, and BRAF. These driver genes are associated with downstream networks controlled by EGFR mutations, such as JAK-STAT, PI3K-AKT-mTOR, and MEK-ERK. The experiment also featured an EGFR-mutated cell line. Using 10XGenomics platform with cellplex technology, we analyzed the seven cell lines together with a pseudo-immunological microenvironment consisting of PBMC cells labeled with the Biolegend TotalSeq™-B Human Universal Cocktail (CITEseq). This technology allowed for independent labeling of each cell line and single-cell analysis of the pooled seven cell lines and the pseudo-microenvironment. The data generated from the aforementioned experiments are available as part of an online tool, which allows users to define cell heterogeneity and generates count tables as an output. The tool provides the cell line derivation for each cell and cell annotations for the pseudo-microenvironment based on CITEseq data by an experienced immunologist. Additionally, we created a range of pseudo-tumor tissues using different ratios of the aforementioned cells embedded in matrigel. These tissues were analyzed using 10XGenomics (FFPE samples) and Curio Bioscience (fresh frozen samples) platforms for spatial transcriptomics, further expanding the scope of our benchmark experiments. The benchmark experiments we conducted provide a unique opportunity to evaluate the performance of bioinformatics tools for detecting and characterizing tumor heterogeneity at the single-cell level. Overall, our experiments provide a controlled and standardized environment for assessing the accuracy and robustness of bioinformatics tools for studying tumor heterogeneity at the single-cell level, which can ultimately lead to more precise and effective cancer diagnosis and treatment.

Keywords: single cell omics, benchmark, spatial transcriptomics, CITEseq

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Authors: Ozhan Simsek, Dicle Donmez, Burhanettin Imrak, Ahsen Isik Ozguven, Yildiz Aka Kacar

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Pomegranate (Punica granatum L.) is known to be one of the oldest edible fruit tree species, with a wide geographical global distribution. Fruits from the two defined varieties (Hicaznar and 33N26) were taken at intervals after pollination and fertilization at different sizes. Seed samples were used for transcriptome sequencing. Primary sequencing was produced by Illumina Hi-Seq™ 2000. Firstly, we had raw reads, and it was subjected to quality control (QC). Raw reads were filtered into clean reads and aligned to the reference sequences. De novo analysis was performed to detect genes expressed in seeds of pomegranate varieties. We performed downstream analysis to determine differentially expressed genes. We generated about 27.09 gb bases in total after Illumina Hi-Seq sequencing. All samples were assembled together, we got 59,264 Unigenes, the total length, average length, N50, and GC content of Unigenes are 84.547.276 bp, 1.426 bp, 2,137 bp, and 46.20 %, respectively. Unigenes were annotated with 7 functional databases, finally, 42.681(NR: 72.02%), 39.660 (NT: 66.92%), 30.790 (Swissprot: 51.95%), 20.212 (COG: 34.11%), 27.689 (KEGG: 46.72%), 12.328 (GO: 20.80%), and 33,833 (Interpro: 57.09%) Unigenes were annotated. With functional annotation results, we detected 42.376 CDS, and 4.999 SSR distribute on 16.143 Unigenes.

Keywords: next generation sequencing, SSR, RNA-Seq, Illumina

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Authors: Naman Solanki, Souraj Mondal

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With the world moving towards online communication, the video datastore has exploded in the past few years. Consequently, it has become crucial to analyse participant’s engagement levels in online communication videos. Engagement prediction of people in videos can be useful in many domains, like education, client meetings, dating, etc. Video-level or frame-level prediction of engagement for a user involves the development of robust models that can capture facial micro-emotions efficiently. For the development of an engagement prediction model, it is necessary to have a widely-accepted standard dataset for engagement analysis. DAiSEE is one of the datasets which consist of in-the-wild data and has a gold standard annotation for engagement prediction. Earlier research done using the DAiSEE dataset involved training and testing standard models like CNN-based models, but the results were not satisfactory according to industry standards. In this paper, a multi-level classification approach has been introduced to create a more robust model for engagement analysis using the DAiSEE dataset. This approach has recorded testing accuracies of 0.638, 0.7728, 0.8195, and 0.866 for predicting boredom level, engagement level, confusion level, and frustration level, respectively.

Keywords: computer vision, engagement prediction, deep learning, multi-level classification

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Authors: I. Wiedemann, A. R. Sharifi, A. Mählmeyer, C. Knorr

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A requirement for sperm motility is a morphologically intact flagellum with a central axoneme. The flagellar beating is caused by the varying activation and inactivation of dynein molecules which are located in the axoneme. DNAL4 (dynein, axonemal, light chain 4) is regarded as a possible functional candidate gene encoding a small subunit of the dyneins. In the present study, 5814bp of the porcine DNAL4 (GenBank Acc. No. AM284696.1, 6097 bp, 4 exons) were comparatively sequenced using three boars with a high motility (>68%) and three with a low motility (<60%). Primers were self-designed except for those covering exons 1, 2 and 3. Prior to sequencing, the PCR products were purified. Sequencing was performed with an ABI PRISM 3100 Genetic Analyzer using the BigDyeTM Terminator v3.1 Cycle Sequencing Reaction Kit. Finally, 23 SNPs were described and genotyped for 82 AI boars representing the breeds Piétrain, German Large White and German Landrace. The genotypes were used to assess possible associations with standard spermatological parameters (ejaculate volume, density, and sperm motility (undiluted (Motud), 24h (Mot1) and 48h (Mot2) after semen collection) that were regularly recorded on the AI station. The analysis included a total of 8,833 spermatological data sets which ranged from 2 to 295 sets per boar in five years. Only SNP g.1007A>G had a significant effect. Finally, the gene substitution effect using the following statistical model was calculated: Yijk= µ+αi+βj+αβij+b1Sijk+b2Aijk+b3T ijk + b4Vijk+b5(α*A)ijk +b6(β*A)ijk+b7(A*T)ijk+Uijk+eijk where Yijk is the semen characteristics, µ is the general mean, α is the main effect of breed, β is the main effect of season, S is the effect of SNP (g.1007A > G), A is the effect of age at semen collection, V is the effect of diluter, αβ, α*A, β*A, A*T are interactions between the fixed effects, b1-b7 are regression coefficients between y and the respective covariate, U is the random effect of repeated observation on animal and e is the random error. The results from the single marker regression analysis revealed highly significant effects (p < 0.0001) of SNP g.1007A > G on Mot1 resp. on Mot2, resulting in a marked reduction by 11.4% resp. 15.4%. Furthermore a loss of Motud by 4.6% was detected (p < 0.0178). Considering the SNP g.1007A > G as a main factor (dominant-recessive model), significant differences between genotypes AA and AG as well as AA and GG for Mot1 and Mot2 exist. For Motud there was a significant difference between AA and GG.

Keywords: association, DNAL4, porcine, sperm traits

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Authors: Asma Karim

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Fish taxonomy plays a fundamental role in the study of biodiversity. However, traditional methods of fish taxonomy rely on morphological features, which can lead to confusion due to great similarities between closely related species. To overcome this limitation, modern taxonomy employs DNA barcoding as a species identification method. This involves using a short standardized mitochondrial DNA region as a barcode, specifically a 658 base pair fragment near the 5′ ends of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene, to exploit the diversity in this region for identification of species. To test the effectiveness and reliability of DNA barcoding, 25 fish specimens from nine different fish species found in various rivers of Pakistan were identified morphologically using a dichotomous key at the start of the study. Comprising nine freshwater fish species, including Mystus cavasius, Mystus bleekeri, Osteobrama cotio, Labeo rohita, Labeo culbasu, Labeo gonius, Cyprinus carpio, Catla catla and Cirrhinus mrigala from different rivers of Pakistan were used in the present study. DNA was extracted from one of the pectoral fins and a partial sequence of CO1 gene was amplified using the conventional PCR method. Analysis of the barcodes confirmed that genetically identified fishes were the same as those identified morphologically at the beginning of the study. The sequences were also analyzed for biodiversity and phylogenetic studies. Based on the results of the study, it can be concluded that DNA barcoding is an effective and reliable method for studying biodiversity and conducting phylogenetic analysis of different fish species in Pakistan.

Keywords: DNA barcoding, fresh water fishes, taxonomy, biodiversity, Pakistan

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Authors: Akeem Akinboro, Bala Alhaj Kegun

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Haemoglobinopathy is a genetic disorder that has the potentiality to cause death of individuals in whom both the alpha (α) and beta (β) globin chains of the haemoglobin molecule are defective due to mutations in their genes. The haemoglobin genotype variants among some residents of Niger state, Nigeria, were determined using the secondary data available at Bida, Minna and Kotangora general hospitals of the state. A total of 1,639 data, representing 434, 655 and 550, collected from the outside patients who visited the medical laboratory units of the three general hospitals, respectively, over five years period (2015-2020) were analyzed into gene frequency, sex and age to determine their haemoglobin genotypes status. More males (51.6 – 58.7%) than females (41.3 – 48.4%) visited the three hospitals during the period covered and most of the patients were between 11 - 20 years old. The frequency of HbA allele in the human population was 0.72, 0.65, 0.68 for Bida, Minna and Kotangora, respectively, while it was 0.25, 0.29 and 0.28 for HbS allele. The HbC allele was prevalent at 0.03, 0.06 and 0.05 among the human population in Bida, Minna and Kotangora cities of Niger state. In overall, the prevalence of HbA, HbS and HbC alleles in Niger state of Nigeria was 0.68, 0.28 and 0.05. Minna being the capital city of Niger state and the most populous among the three cities in the state seems to have influx of more people who are carriers of abnormal haemoglobin genotypes which has resulted to higher frequency of HbS and HbC than those of the other two cities in this study. These results show that the pattern of haemoglobin genotypes frequency of Kontagora could be a prediction for the whole of Niger state. It is therefore necessary and important to take screening of blood for haemoglobin genotype serious among intending couples to prevent and reduce the possibility of having increase in the number of people with abnormal haemoglobin genotypes in the state.

Keywords: haemoglobin, genotype, niger state, gene frequency, general hospitals

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Authors: Aidin Foroutan, David S. Wishart, Leluo L. Guan, Carolyn Fitzsimmons

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Maximizing efficiency and growth potential of beef cattle requires not only genetic selection (i.e. residual feed intake (RFI)) but also adequate nutrition throughout all stages of growth and development. Nutrient restriction during gestation has been shown to negatively affect post-natal growth and development as well as fertility of the offspring. This, when combined with RFI may affect progeny traits. This study aims to investigate the impact of selection for divergent genetic potential for RFI and maternal nutrition during early- to mid-gestation, on bull calf traits such as fertility and muscle development using multiple ‘omics’ approaches. Comparisons were made between High-diet vs. Low-diet and between High-RFI vs. Low-RFI animals. An epigenetics experiment on semen samples identified 891 biomarkers associated with growth and development. A gene expression study on Longissimus thoracis muscle, semimembranosus muscle, liver, and testis identified 4 genes associated with muscle development and immunity of which Myocyte enhancer factor 2A [MEF2A; induces myogenesis and control muscle differentiation] was the only differentially expressed gene identified in all four tissues. An initial metabolomics experiment on serum samples using nuclear magnetic resonance (NMR) identified 4 metabolite biomarkers related to energy and protein metabolism. Once all the biomarkers are identified, bioinformatics approaches will be used to create a database covering all the ‘omics’ data collected from this project. This database will be broadened by adding other information obtained from relevant literature reviews. Association analyses with these data sets will be performed to reveal key biological pathways affected by RFI and maternal nutrition. Through these association studies between the genome and metabolome, it is expected that candidate biomarker genes and metabolites for feed efficiency, fertility, and/or muscle development are identified. If these gene/metabolite biomarkers are validated in a larger animal population, they could potentially be used in breeding programs to select superior animals. It is also expected that this work will lead to the development of an online tool that could be used to predict future traits of interest in an animal given its measurable ‘omics’ traits.

Keywords: biomarker, maternal nutrition, omics, residual feed intake

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Authors: Bavneet Kaur Sidhu, Manoj Tiwari

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Global Developmental Delay (GDD) is a common pediatric condition. Etiologies of GDD might, however, differ in developing countries. In the last decade, sporadic families are being reported in various countries. As to the author’s best knowledge, many risk factors and their correlation with the prevalence of GDD have been studied but its statistical correlation has not been done. Thus we propose the present study by targeting the risk factor, prevalence and their statistical correlation with GDD. FMR1 gene was studied to confirm the disease and its penetrance. A complete questionnaire-based performance was designed for the statistical studies having a personal, past and present medical history along with their socio-economic status as well. Methods: We distributed the children’s age in 4 different age groups having 5-year intervals and applied structural equation modeling (SEM) techniques, Spearman’s rank correlation coefficient, Karl Pearson correlation coefficient, and chi-square test.Result: A total of 1100 families were enrolled for this study; among them, 330 were clinically and biologically confirmed (radiological studies) for the disease, 204 were males (61.8%), 126 were females (38.18%). We found that 27.87% were genetic and 72.12 were sporadic, out of 72.12 %, 43.277% cases from urban and 56.72% from the rural locality, the mothers' literacy rate was 32.12% and working women numbers were 41.21%. Conclusions: There is a significant association between mothers' age and GDD prevalence, which is also followed by mothers' literacy rate and mothers' occupation, whereas there was no association between fathers' age and GDD.

Keywords: global developmental delay, FMR1 gene, spearman’ rank correlation coefficient, structural equation modeling

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Authors: Claudia Patricia Beltran Miranda, Mónica López Anaya, Mónica Navarro Meza, Maria Leonor Valderrama Chairez

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Background: Mexico occupies the second world-wide place of morbid obese people (10- 12 million). Factors that predispose the development of MO are genetic, environmental, physiological, psycho-social and behavioral (nourishing habits). Objective: Associate Gln223Arg polymorphism of gene LEPR, levels of leptina and nourishing habits with the presence of morbid obesity in adolescents of the south of Jalisco (México). Methods: In 41 adolescents (18 normal weight and 23 morbid obesity) of 12 to 19 years of age, both sexes that were measure size and weight with tanita scale and stadimeter to determine IMC. Morbid obesity was determined by tables of the WHO and was established with a standard deviation >3. The Gln223Arg polymorphism have been identify by PCR and leptina levels by ELISA. Nourishing habits were evaluate by the questionnaire the Adolescent Food Habits Checklist. The statistical analysis was performed to compare mean scores obtained from the questionnaire when we compare morbid obesity vs. normal weight adolescents with p=0.03 and a significance of 95%. Results: frequencies alellics and genics were not stadistics significatives p= 0,011 and p=0,279 respectly when were compared between normal weight adolescents and morbib obesity Leptin levels and nourishing habits were associated with morbid obesity. The polymorphism not shown significance with morbid obesity. Conclusions: Dietary habits and leptin levels in adolescents are important factor that predisposes the development of obesity in adolescents. The presence of the polymorphism are not associated with morbid obesity in these subjects.

Keywords: leptin, nourishing habits, morbid obesity, polymorphism

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Authors: Amal Saafan, Kareem Al Sofy, Sameh AbdelGhani, Magdy Amin

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Background: Acinetobacter spp. resistance towards β-lactam antibiotics is mediated mainly by different classes of β-lactamases production; detection of some genes responsible for production of β-lactamases is the objective of the study. Methods: One hundred fifty bacterial isolates were recovered from blood, sputum, and urine specimens from different hospitals in Egypt. Sixty-nine isolate were identified as Acinetobacter baumannii using traditional biochemical tests, CHROM agar, MicroScan and PCR amplification of blaoxa-51like gene. Acinetobacterbaumannii isolates were grouped into carbapenem resistant group (GP1), cefotaxime, ceftazidime and cefoxitin resistant group (GP2) and carbapenem and cephalosporin non-resistant group (GP3). Carbapenemase activity was screened using modified Hodge test (MHT) for GP1.Metallo-β-lactamases screening was performed for MHT positive isolates using double disk synergy test (DDST) and combined disk test (CDT). Amp C activity was screened using Amp C disk test with Tris-EDTA, DDST, and CDT for GP2. Finally, PCR amplification of blaoxa-51like, blaoxa-23like, blaIMP-like, blaVIM-like, and blaADC-like genes was performed for isolates that showed, at least, two positive results of three for both AmpC and carbapenemases phenotypic screening tests (obvious activity), in addition to GP3 (for comparison). Detection of blaoxa-51like and blaADC-like genes preceded by ISAba1 was also performed. Results: Antibiogram of 69 pure Acinetobacter baumannii isolates resulted in 57, 64, and 2 isolates enrolled into GP1, GP2, and GP3, respectively. Carbapenemase activity was shown by 49(85.9%) isolate using MHT. Metallo-β-lactamases screening revealed 32(65.3%) and 35(71.4%) using DDST and CDT, respectively.AmpC activity was shown by 43(67.2%) and 50 (78.1%) isolates using AmpC disk test with Tris-EDTA, and both DDST and CDT, respectively. Twenty-seven isolates showed obvious activity, all of them (100%) were harboring blaoxa-51like and blaADC-like genes, while blaoxa-23like, blaIMP-like andblaVIM-like genes were harbored by 23(85.2%), 9 (33.%) and no isolate respectively. Only 12 (44.4%) isolates harbored blaoxa-51like and blaADC-like genes preceded by ISAba1. GP3 isolates showed only positive blaoxa-51like and blaADC-like genes. Conclusion: It is not possible to correlate resistance with presence of blaoxa-51like and blaADC-like genes and presence of ISAba1 was immediate as transcriptional promoter. A blaoxa-23like gene played an important role in carbapenem resistance when compared with blaIMP-like and blaVIM-like gene.

Keywords: acinetobacter, beta-lactams, resistance, antimicrobial agents

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Authors: Kamellia Reshadi, Pranav Ragji, Theodoros Soldatos

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Recent advancements in cloud computing and storage, along with rapid progress in artificial intelligence (AI), have transformed approaches to developing efficient, scalable applications. However, integrating AI with cloud computing poses challenges as these fields are often disjointed, and many advancements remain difficult to access, obscured in complex documentation or scattered across research reports. For this reason, we share experiences from prototype projects combining these technologies. Specifically, we focus on Google Cloud Platform (GCP) functionalities and describe vision and speech activities applied to labeling, subtitling, and urban traffic flow tasks. We describe challenges, pricing, architecture, and other key features, considering the goal of real-time performance. We hope our demonstrations provide not only essential guidelines for using these functionalities but also enable more similar approaches.

Keywords: artificial intelligence, cloud computing, real-time applications, case studies, knowledge management, research and development, text labeling, video annotation, urban traffic analysis, public safety, prototyping, Google Cloud Platform

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Authors: Sirikwan Ponprateep, Anchalee Tassanakajon, Vichien Rimphanitchayakit

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A Kazal-type serine proteinase inhibitor, SPIPm2, was abundantly expressed in the hemocytes and secreted into shrimp plasma has anti-viral property against white spot syndrome virus (WSSV). To discover the molecular mechanism of antiviral activity, the binding assay showed that SPIPm2 bind to the components of viral particle and shrimp hemocyte. From our previous report, viral target protein of SPIPm2 was identified, namely WSV477 using yeast two-hybrid screening. WSV477 is an early gene product of WSSV and involved in viral propagation. In this study, the co-immunoprecipitation technique and Tandem Mass Spectrometry (LC-MS/MS) was used to identify the target protein of SPIPm2 from shrimp hemocyte. The target protein of SPIPm2 was cyclophilin A. In vertebrate, cyclophilin A or peptidylprolyl isomerase A was reported to be the immune suppressor interacted with cyclosporin A involved in immune defense response. The recombinant cyclophilin A from Penaeus monodon (rPmCypA) was produced in E.coli system and purified using Ni-NTA column to confirm the protein-protein interaction. In vitro pull-down assay showed the interaction between rSPIPm2 and rPmCypA. To study the biological function of these proteins, the expression analysis of immune gene in shrimp defense pathways will be investigated after rPmCypA administration.

Keywords: cyclophilin A, protein-protein interaction, Kazal-type serine proteinase inhibitor, Penaeus monodon

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Authors: Zhaoxu Lu, Yufeng Ma, Linying Gao, Li Wang, Mei Qiang

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Mercury (Hg) is a well-recognized environmental pollutant known by its toxicity of development and neurotoxicity, which may result in adverse health outcomes. However, the mechanisms underlying the teratogenic effects of Hg are not well understood. Imprinting genes are emerging regulators for fetal development subject to environmental pollutants impacts. In this study, we examined the association between paternal preconception Hg exposures and the alteration of DNA methylation of imprinting genes in human sperm DNA. A total of 618 men aged from 22 to 59 was recruited from the Reproductive Medicine Clinic of Maternal and Child Care Service Center and the Urologic Surgery Clinic of Shanxi Academy of Medical Sciences during April 2015 and March 2016. Demographic information was collected using questionnaires. Urinary Hg concentrations were measured using a fully-automatic double-channel hydride generation atomic fluorescence spectrometer. And methylation status in the DMRs of imprinting genes H19, Meg3 and Peg3 of sperm DNA were examined by bisulfite pyrosequencing in 243 participants. Spearman’s rank and multivariate regression analysis were used for correlation analysis between sperm DNA methylation status of imprinting genes and urinary Hg levels. The median concentration of Hg for participants overall was 9.09μg/l (IQR: 5.54 - 12.52μg/l; range = 0 - 71.35μg/l); no significant difference was found in median concentrations of Hg among various demographic groups (p > 0.05). The proportion of samples that a beyond intoxication criterion (10μg/l) for urinary Hg was 42.6%. Spearman’s rank correlation analysis indicates a negative correlation between urinary Hg concentrations and average DNA methylation levels in the DMRs of imprinted genes H19 (rs=﹣0.330, p = 0.000). However, there was no such a correlation found in genes of Peg3 and Meg3. Further, we analyzed of correlation between methylation level at each CpG site of H19 and Hg level, the results showed that three out of 7 CpG sites on H19 DMR, namely CpG2 (rs =﹣0.138, p = 0.031), CpG4 (rs =﹣0.369, p = 0.000) and CpG6 (rs=﹣0.228, p = 0.000), demonstrated a significant negative correlation between methylation levels and the levels of urinary Hg. After adjusting age, smoking, drinking, intake of aquatic products and education by multivariate regression analysis, the results have shown a similar correlation. In summary, mercury nonoccupational environmental exposure in reproductive-aged men associated with altered DNA methylation outcomes at DMR of imprinting gene H19 in sperm, implicating the susceptibility of the developing sperm for environmental insults.

Keywords: epigenetics, genomic imprinting gene, DNA methylation, mercury, transgenerational effects, sperm

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Authors: Meenakshi Srivastava, A. K. Mishra

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This study characterizes the metabolic versatility of denitrifying bacterial communities residing in the paddy soil using the GC-MS based Phospholipid Fatty Acid (PLFA) analyses simultaneously with nosZ gene based PCR-DGGE (Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis) and real time Q-PCR analysis. We have analyzed the abundance of nitrous oxide reductase (nosZ) genes, which was subsequently related to soil PLFA profile and DGGE based denitrifier community structure. Soil denitrifying bacterial community comprised majority or dominance of Ochrobactrum sp. following Cupriavidus and uncultured bacteria strains in paddy soil of selected sites. Initially, we have analyzed the abundance of the nitrous oxide reductase gene (nosZ), which was found to be related with PLFA based lipid profile. Chandauli of Eastern UP, India represented greater amount of lipid content (C18-C20) and denitrifier’s diversity. This study suggests the positive co-relation between soil PLFA profiles, DGGE, and Q-PCR data. Thus, a close networking among metabolic abilities and taxonomic composition of soil microbial communities existed, and subsequently, such work at greater extent could be helpful in managing nutrient dynamics as well as microbial dynamics of paddy soil ecosystem.

Keywords: denaturing gradient gel electrophoresis, DGGE, nitrifying and denitrifying bacteria, PLFA, Q-PCR

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Authors: M. Ottenbrite, G. Yilmaz, J. Devenish, M. Kang, H. Dan, M. Lin, C. Lau, C. Carrillo, K. Bessonov, J. Nash, E. Topp, J. Guan

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Consumption of food contaminated by antibiotic-resistant (AR) bacteria may lead to the transmission of AR genes in the gut microbiota and cause AR bacterial infection, a significant public health concern. However, information is limited on if and how background microbes from the food matrix (food microbes) may influence resistance transmission. Thus, we assessed the colonization of a β-lactam resistant Salmonella Heidelberg strain (donor) and a β-lactam susceptible S. Typhimurium strain (recipient) and the transfer of the resistance genes in the mouse gut in the presence or absence of food microbes that were derived from washing freshly-harvested carrots. Mice were pre-treated with streptomycin and then inoculated with both donor and recipient bacteria or recipient only. Fecal shedding of the donor, recipient, and transconjugant bacteria was enumerated using selective culture techniques. Transfer of AR genes was confirmed by whole genome sequencing. Gut microbial composition was determined by 16s rRNA amplicon sequencing. Significantly lower numbers of donors and recipients were shed from mice that were inoculated with food microbes compared to those without food microbe inoculation. S. Typhimurium transconjugants were only recovered from mice without inoculation of food microbes. A significantly higher survival rate was in mice with vs. without inoculation of food microbes. The results suggest that the food microbes may compete with both the donor and recipient Salmonella, limit their growth and reduce transmission of the β-lactam resistance gene in the mouse gut.

Keywords: antibiotic resistance, gene transfer, gut microbiota, Salmonella infection

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Authors: Wissam Saliba, Mandy Nicholson

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Secretory carcinoma (SC) is a rare type of salivary gland cancer. It was first realized as a distinct type of malignancy in 2010and wasinitially termed “mammary analogue secretory carcinoma” because of similarities with secretory breast cancer. The name was later changed to SC. Most SCs originate in parotid glands, and most harbour a rare gene mutation: ETV6-NTRK3. This mutation is rare in common cancers and common in rare cancers; it is present in most secretory carcinomas. Disease outcomes for SC are usually described as favourable as many cases of SC are lowgrade (LG), and cancer growth is slow. In early stages, localized therapy is usually indicated (surgery and/or radiation). Despitea favourable prognosis, a sub-set of casescan be much more aggressive.These cases tend to be of high-grade(HG).HG casesare associated with a poorer prognosis.Management of such cases can be challenging due to limited evidence for effective systemic therapy options. This case report describes the clinical management of a 46-year-oldmale patient with a unique case of SC. He was initially diagnosed with a low/intermediate grade carcinoma of the left parotid gland in 2009; he was treated with surgery and adjuvant radiation. Surgical pathology favoured primary salivary adenocarcinoma, and 2 lymph nodes were positive for malignancy. SC was not yet realized as a distinct type of cancerat the time of diagnosis, and the pathology reportvalidated this gap by stating that the specimen lacked features of the defined types of salivary carcinoma.Slow-growing pulmonary nodules were identified in 2017. In 2020, approximately 11 years after the initial diagnosis, the patient presented with malignant pleural effusion. Pathology from a pleural biopsy was consistent with metastatic poorly differentiated cancer of likely parotid origin, likely mammary analogue secretory carcinoma. The specimen was sent for Next Generation Sequencing (NGS); ETV6-NTRK3 gene fusion was confirmed, and systemic therapy was initiated.One cycle ofcarboplatin/paclitaxel was given in June 2020. He was switched to Larotrectinib (NTRK inhibitor (NTRKi)) later that month. Larotrectinib continued for approximately 9 months, with discontinuation in March 2021 due to disease progression. A second-generation NTRKi (Selitrectinib) was accessed and prescribedthrough a single patient study. Selitrectinib was well tolerated. The patient experienced a complete radiological response within~4 months. Disease progression occurred once again in October 2021. Progression was slow, and Selitrectinib continuedwhile the medical team performed a thorough search for additional treatment options. In January 2022, a liver lesion biopsy was performed, and NGS showed an NTRKG623R solvent-front resistance mutation. Various treatment pathways were considered. The patient pursuedanother investigational NTRKi through a clinical trial, and Selitrectinib was discontinued in July 2022. Excellent performance status was maintained throughout the entire course of treatment.It can be concluded that NTRK inhibitors provided satisfactory treatment efficacy and tolerance for this patient with high-grade transformation and NTRK gene fusion cancer. In the future, more clinical research is needed on systemic treatment options for high-grade transformations in NTRK gene fusion SCs.

Keywords: secretory carcinoma, high-grade transformations, NTRK gene fusion, NTRK inhibitor

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Authors: Franco Van De Velde, Debora Esposito, Maria E. Pirovani, Mary A. Lila

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The physiology of various inflammatory diseases is a complex process mediated by inflammatory and immune cells such as macrophages and monocytes. Chronic inflammation, as observed in many cardiovascular and autoimmune disorders, occurs when the low-grade inflammatory response fails to resolve with time. Because of the complexity of the chronic inflammatory disease, major efforts have focused on identifying novel anti-inflammatory agents and dietary regimes that prevent the pro-inflammatory process at the early stage of gene expression of key pro-inflammatory mediators and cytokines. The ability of the extracts of three blackberry cultivars (‘Jumbo’, ‘Black Satin’ and ‘Dirksen’), and one strawberry cultivar (‘Camarosa’) to inhibit four well-known genetic biomarkers of inflammation: inducible nitric oxide synthase (iNOS), cyclooxynase-2 (Cox-2), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in an in vitro lipopolysaccharide-stimulated murine RAW 264.7 macrophage model were investigated. Moreover, the effect of latter extracts on the intracellular reactive oxygen species (ROS) and nitric oxide (NO) production was assessed. Assay was conducted with 50 µg/mL crude extract concentration, an amount that is easily achievable in the gastrointestinal tract after berries consumption. The mRNA expression levels of Cox-2 and IL-6 were reduced consistently (more than 30%) by extracts of ‘Jumbo’ and ‘Black Satin’ blackberries. Strawberry extracts showed high reduction in mRNA expression levels of IL-6 (more than 65%) and exhibited moderate reduction in mRNA expression of Cox-2 (more than 35%). The latter behavior mirrors the intracellular ROS production of the LPS stimulated RAW 264.7 macrophages after the treatment with blackberry ‘Black Satin’ and ‘Jumbo’, and strawberry ‘Camarosa’ extracts, suggesting that phytochemicals from these fruits may play a role in the health maintenance by reducing oxidative stress. On the other hand, effective inhibition in the gene expression of IL-1β and iNOS was not observed by any of blackberry and strawberry extracts. However, suppression in the NO production in the activated macrophages among 5–25% was observed by ‘Jumbo’ and ‘Black Satin’ blackberry extracts and ‘Camarosa’ strawberry extracts, suggesting a higher NO suppression property by phytochemicals of these fruits. All these results suggest the potential beneficial effects of studied berries as functional foods with antioxidant and anti-inflammatory roles. Moreover, the underlying role of phytochemicals from these fruits in the protection of inflammatory process will deserve to be further explored.

Keywords: cyclooxygenase-2, functional foods, interleukin-6, reactive oxygen species

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Authors: Foluke E. Sola-Ojo, Ibraheem A. Abubakar, Semiu F. Bello, Isiaka H. Fatima, Sule Bisola, Adesina M. Olusegun, Adeniyi C. Adeola

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The domesticated pigeon, Columba livia domestica, has many valuable characteristics, including high nutritional value and fast growth rate. There is a lack of information on its genetic diversity in Nigeria; thus, the genetic variability in mitochondrial cytochrome oxidase subunit I (COI) sequences of 150 domestic pigeons from four different locations was examined. Three haplotypes (HT) were identified in Nigerian populations; the most common haplotype, HT1, was shared with wild and domestic pigeons from Europe, America, and Asia, while HT2 and HT3 were unique to Nigeria. The overall haplotype diversity was 0.052± 0.025, and nucleotide diversity was 0.026± 0.068 across the four investigated populations. The phylogenetic tree showed significant clustering and genetic relationship of Nigerian domestic pigeons with other global pigeons. The median-joining network showed a star-like pattern suggesting population expansion. AMOVA results indicated that genetic variations in Nigerian pigeons mainly occurred within populations (99.93%), while the Neutrality tests results suggested that the Nigerian domestic pigeons’ population experienced recent expansion. This study showed a low genetic diversity and population differentiation among Nigerian domestic pigeons consistent with a relatively conservative COI sequence with few polymorphic sites. Furthermore, the COI gene could serve as a candidate molecular marker to investigate the genetic diversity and origin of pigeon species. The current data is insufficient for further conclusions; therefore, more research evidence from multiple molecular markers is required.

Keywords: Nigeria pigeon, COI, genetic diversity, genetic variation, conservation

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Authors: Minhas Alam, Muhammad Hidayat Rasool, Mohsin Khurshid, Bilal Aslam

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Acinetobacter baumannii is a notorious bacterial pathogen that is an emerging nightmare in clinical settings and is mainly involved in severe nosocomial infections. However, the data related to carbapenem-resistant A. baumannii (CRAB) from veterinary settings is limited, especially in developing countries like Pakistan. To investigate the genetic diversity and molecular basis of carbapenem resistance in Acinetobacter baumannii isolates from Cattle, a total of 1960 samples were collected from cattle from Punjab, Pakistan. The isolates were analyzed by routine microbiological procedures and confirmed by polymerase chain reaction (PCR). The isolates were further screened for antimicrobial susceptibility and the presence of multiple antimicrobial-resistant determinants by PCR. Multilocus sequence typing (MLST) was performed. The results of the current study revealed that the overall prevalence of A. baumannii in cattle was 3.28% (65/1980). Among cattle 27.7% (18/65) were found CRAB strains. The CRAB isolates harbor class D β- lactamases genes, e-g, blaOXA-23 and blaOXA-51, 94.4% (17/18). CRAB isolates carry class B β- lactamases gene blaIMP, and only one isolate carries the blaNDM-1 gene. The MLST results of CRAB isolates from cattle demonstrated 5 STs and one new ST. The commonly found sequence types in CRAB isolates were ST2 (n=10, 55.5%), followed by ST642 (n=5, 27.8%) and ST600 & ST889 (n=1, 5.55%). The presence of CRAB isolates in cattle indicates an alarming situation in Punjab, Pakistan. Immediate control measures should be taken to stop the transmission of CRAB isolates within cattle, to the environment, and to clinical settings.

Keywords: acinetobacter baumannii, carbapenemases, veterinary, drug resistance

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Authors: Okolie Pius Ifeanyi, Emerenini Emilymary Chima

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Aims: The study investigated the diversity and identities of Lactic Acid Bacteria (LAB) isolated from different fresh fruits using Molecular Nested PCR analysis and the efficacy of cell free supernatants from Lactic Acid Bacteria (LAB) isolated from fresh fruits for in vitro control of some tomato pathogens. Study Design: Nested PCR approach was used in this study employing universal 16S rRNA gene primers in the first round PCR and LAB specific Primers in the second round PCR with the view of generating specific Nested PCR products for the LAB diversity present in the samples. The inhibitory potentials of supernatant obtained from LAB isolates of fruits origin that were molecularly characterized were investigated against some tomato phytopathogens using agar-well method with the view to develop biological agents for some tomato disease causing organisms. Methodology: Gram positive, catalase negative strains of LAB were isolated from fresh fruits on Man Rogosa and Sharpe agar (Lab M) using streaking method. Isolates obtained were molecularly characterized by means of genomic DNA extraction kit (Norgen Biotek, Canada) method. Standard methods were used for Nested Polymerase Chain Reaction (PCR) amplification targeting the 16S rRNA gene using universal 16S rRNA gene and LAB specific primers, agarose gel electrophoresis, purification and sequencing of generated Nested PCR products (Macrogen Inc., USA). The partial sequences obtained were identified by blasting in the non-redundant nucleotide database of National Center for Biotechnology Information (NCBI). The antimicrobial activities of characterized LAB against some tomato phytopathogenic bacteria which include (Xanthomonas campestries, Erwinia caratovora, and Pseudomonas syringae) were obtained by using the agar well diffusion method. Results: The partial sequences obtained were deposited in the database of National Centre for Biotechnology Information (NCBI). Isolates were identified based upon the sequences as Weissella cibaria (4, 18.18%), Weissella confusa (3, 13.64%), Leuconostoc paramensenteroides (1, 4.55%), Lactobacillus plantarum (8, 36.36%), Lactobacillus paraplantarum (1, 4.55%) and Lactobacillus pentosus (1, 4.55%). The cell free supernatants of LAB from fresh fruits origin (Weissella cibaria, Weissella confusa, Leuconostoc paramensenteroides, Lactobacillus plantarum, Lactobacillus paraplantarum and Lactobacillus pentosus) can inhibits these bacteria by creating clear zones of inhibition around the wells containing cell free supernatants of the above mentioned strains of lactic acid bacteria. Conclusion: This study shows that potentially LAB can be quickly characterized by molecular methods to specie level by nested PCR analysis of the bacteria isolate genomic DNA using universal 16S rRNA primers and LAB specific primer. Tomato disease causing organisms can be most likely biologically controlled by using extracts from LAB. This finding will reduce the potential hazard from the use of chemical herbicides on plant.

Keywords: nested pcr, molecular characterization, 16s rRNA gene, lactic acid bacteria

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