Base Pairs Research Articles

First record of the blue-and-yellow grouper Epinephelus flavocaeruleus (Lacepède 1802) (Perciformes: Epinephelidae) from the Borneo waters, Malaysia

Groupers of the family Epinephelidae constitute a diverse and commercially valuable group of reef fishes globally. They comprise an assemblage of carnivorous marine fishes, comprising more than 177 species across 16 genera. The epinephelid genus Epinephelus, which consists of over 90 species, is found worldwide in the tropics and subtropics. To date, the ichthyofauna of Malaysia has documented a total of 43 epinephelid species. Apart from these, Epinephelus flavocaeruleus (Lacepède, 1802), commonly known as the blue-and-yellow grouper, is rarely reported in the Indo-Pacific Ocean. The present study extends the documented distribution range of E. flavocaeruleus eastwards from the Andaman Sea to the Borneo waters of Sabah, Malaysia. Five specimens of the blue-and-yellow grouper were collected from a local fish market. Species identification was confirmed by the color patterns and DNA barcoding of 630 base pairs of the cytochrome C oxidase I gene for all E. flavocaeruleus specimens, Epinephelus cyanopodus (Richardson, 1846), and 10 closely related Epinephelus species. The interspecies genetic distance ranged from 0.002–0.168. Results from the Templeton, Crandall, and Sing (TCS) haplotype network analysis and maximum likelihood phylogeny based on the COI marker indicate a close genetic relationship between E. flavocaeruleus and E. cyanopodus. However, we refrain from proposing any taxonomic revisions given that more in-depth studies using multiple molecular markers or phylogenomic analysis on a larger sample size are necessary to confirm the taxonomic status of both species. This study significantly contributes to a better understanding of the taxonomy, phylogenetic relationship, and genetic diversity of E. flavocaeruleus.

The triple code model for advancing research in rare and undiagnosed diseases beyond the base pairs.

Rare and undiagnosed diseases pose significant challenges for understanding their mechanisms, diagnosis, and treatment. The Triple Code Model, an integrative paradigm described here, considers the combined influence of the genetic code, epigenetic code, and nuclear structure (an emerging code), as fundamental biochemical mechanisms underlying many rare diseases. Studies demonstrate dysfunctional membrane and cytoplasmic signals instruct the epigenome to ultimately impact the 3D structure and dynamics of the nucleus, highlighting their close interrelationships. Consequently, this model offers a holistic perspective on rare and undiagnosed diseases by moving beyond a solely genetic view. We propose that this integrated framework will efficiently guide rare disease research by taking it 'Beyond the Base Pairs,' leading to improved diagnostics and personalized treatments.

Comparative chloroplast genomes analysis of nine Primulina (Gesneriaceae) rare species, from karst region of southwest China

Guizhou Province is one of the most important karst regions of southwest China, with 22 Primulina species. These species are highly habitat-specialized and dependent on the soils of the karst region, and many inter-species classifications are unclear. Therefore, studying the chloroplast genomes and estimating the divergence times of there species can not only provide a better understanding of interspecific relationships but also help to know the species speciation and divergence in the karst environment. So, we sequenced and assembled the chloroplast genomes of nine Primulina species (including six endemic species of Guizhou) and conducted chloroplast genome comparison analysis and phylogenetic study. The chloroplast genome structures of the nine Primulina species were quadripartite with total lengths of 152,869–153,364 base pairs (bp) and GC content of 37.55–37.64%. There are 132 functional genes annotated, respectively. A total of 375 simple sequence repeats and 375 interspersed nuclear elements were identified. The 30 highly preferred codons identified were used at similar frequencies in different species, respectively. A phylogenetic tree constructed on the basis of the 38 chloroplast genomes showed that Primulina species form a monophyletic group. Eleven mutational hotspot regions that could serve as potential molecular markers were identified, of which two regions near the 3′ and 5′ ends of the ycf1 gene were of appropriate size and can serve as molecular markers for phylogenetic studies of Primulina. The results of molecular clock analyses indicate that the three major branches of Primulina begin to diverge in the Miocene, and the number of species proliferated in the Pliocene and Pleistocene. Most of the species of Primulina in Guizhou Province were formed in the Pleistocene and rapidly diverged within a short period of time. This research study enriches the genetic resource information of Primulina and deepens the understanding of the phylogenetic relationships of the genus.

A Novel Polymerase Chain Reaction (PCR)-Based Method for the Rapid Identification of Chrysodeixis includens and Rachiplusia nu

Chrysodeixis includens and Rachiplusia nu are two species belonging to the Plusiinae subfamily within the Noctuidae family. Due to their morphological similarity, the identification of their larvae is difficult and time-consuming. A rapid and accurate identification of these two species is essential for their management as these species exhibit differential susceptibilities to insecticides and crops engineered to express Bacillus thuringiensis (Bt) proteins, and a molecular tool can easily provide this differentiation. Currently, molecular analysis can identify these species through genetic sequencing, an expensive and time-consuming process. In our study, after sequencing part of the mtDNA cytochrome c oxidase I (COI) gene and based on the differences found in the gene of each species, a set of species-specific primers was developed: one reverse primer common to both species and two forward primers, specific to each species, amplifying fragments of 199 base pairs (bp) for C. includens and 299 bp for R. nu. Our results indicate that the primers were specific for these species, enabling the identification of individuals directly through agarose gel. The new methodology proved to be accurate, rapid, and reliable for the correct identification of these two species of loopers.

A new species of Myxobolus (Cnidaria: Myxosporea: Myxobolidae) from the gills of the green sunfish, Lepomis cyanellus (Perciformes: Centrarchidae), from the Ouachita River Drainage of Western Arkansas.

Forty-two species of myxozoans, including 31 species of Myxobolus Bütschli, 1882, have been described from centrarchid fishes. One species, the green sunfish (Lepomis cyanellus), has been reported to host at least three species of Myxobolus. Between March 2023 and June 2024, 42 L. cyanellus were collected from watersheds in Montgomery and Polk counties, Arkansas, and their gills, gallbladders, urinary bladders, fins, integument, other major organs, and musculature were examined for myxozoans. Two (5%) L. cyanellus from Polk County were found to harbor a new species of Myxobolus infecting the gill lamellae. A qualitative and quantitative morphological description was based on formalin-fixed preserved plasmodia and myxospores. Plasmodia of Myxobolus polkensis n. sp. are 133 µm long × 123 µm wide, and myxospores are 19.8 µm long × 6.5 µm wide, with two narrowly pyriform unequal polar capsules, one usually longer (9.2 µm) than the other (8.5 µm). Molecular data consisted of a 2,025 base pair sequence of the partial small subunit rRNA gene (SSU). Variably sized, polysporic plasmodia were randomly distributed throughout the gill lamellae in intralamellar locations. Large plasmodia displaced adjacent lamellae inducing mild epithelial proliferation but minimal inflammatory changes. Phylogenetic analysis revealed that M. polkensis n. sp. is a member of a clade of myxozoan species that predominately infect centrarchids from North America. This is the fourth report of a Myxobolus from L. cyanellus but the first report of a species infecting the gill lamellae. This article was registered in the Official Register of Zoological Nomenclature (ZooBank) as urn:lsid:zoobank.org:pub:E9BA08E0-0A7E-487B-80AE-C5C723DBB661.

Characterization of Guest DNA Transport and Adsorption within Host Porous Protein Crystals.

Nucleic acid transport through protein-based pores is a well-characterized phenomenon due in part to advancements in nanopore sequencing. A less studied area is nucleic acid transport through extended protein-based channels, where the additional surface area and increased contact time allow for the study of prolonged binding interactions. Porous protein crystals composed of "CJ", a putative polyisoprenoid-binding protein from Campylobacter jejuni, represent a favorable, highly ordered material for studying DNA transport and binding/unbinding along protein-based channels. These crystals adopt a hexagonal prism habit and contain a densely packed hexagonal array of 13 nm diameter axial nanopores that run from the top to the bottom of the crystal. After cross-linking, the crystals are easily manipulated for experimentation. An adsorption isotherm between host crystals and guest double-stranded 8 base pair DNA (8mer) revealed a high equilibrium adsorption constant of 206 ± 30 L/g. Fluorescence confocal microscopy tracked the loading of guest DNA into host crystals predominately along the major axial crystal nanopores. Four different computational models based on the finite volume (FV) method were assessed to model the transport process for guest 8mer and 15mer dsDNA loading into empty host crystals in terms of fundamental parameters, such as the intrapore diffusion constant. Fitting the models to the data revealed that the most basic FV model sufficed to describe the observed loading behavior, characterized by a single effective diffusion coefficient. Leveraging Fick's first law, we more directly fit a numerical range for the observed intrapore diffusion coefficient as a function of time, position within the crystal, and relative guest concentration. This new transport analysis strategy was applied to both out-of-equilibrium loading and fluorescence recovery after photobleaching (FRAP) experiments. The intrapore diffusion constants are comparable between 8mer and 15mer dsDNA and were found to be 2 orders of magnitude faster for DNA loading into empty crystals than that observed in FRAP experiments, which averaged (10 ± 4) × 10-11 cm2/s.

Thrombin Nanochannel Logic Gate Inspired by BioMemory.

The process of "reading" and "writing" in biomemory involves the transmission of electrical signals between neurons, with ligand-gated ion channels assuming a key role. The solid-state nanochannels exhibit certain similarities with neurons. Information transmission can be achieved by controlling the flow of ions within nanochannels, rendering them potentially suitable for simulating neuron behavior. Herein, thrombin (Thr) was chosen as the target protein, and a functionalized nanochannel sensing system was successfully constructed using DNA aptamers, enabling a highly sensitive Thr response with a detection limit of 0.221 fM. Simultaneously, based on Watson-Crick base pairing and programmable chain displacement reactions, controlled release and cyclic response of the target molecule were further achieved. This mechanism elucidates the rules governing specific input-output relationships, innovatively linking them with memory storage and recognition through the Thr-nanochannel logic gate, thereby realizing the reading of biomemory at the hardware level. In summary, the biological hybrid nanofluidic control device of this invention converts molecular events into electrical signals, providing potential avenues for establishing connections between the mechanisms of biomemory and solid-state nanochannel biosensing and recognition in the future.

Identification of Genes Encoded Toxin-Antitoxin System in Mycobacterium Tuberculosis Strains from Clinical Sample.

The toxin-antitoxin system is a genetic element that is highly present in Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis. The toxin-antitoxin system comprises toxin protein and antitoxin protein or non-encoded RNA interacting with each other and inhibiting toxin activity. M. Tuberculosis has more classes of TA loci than non-tubercle bacilli and other microbes, including VapBC, HigBA, MazEF, ParDE, RelBE, MbcTA, PemIK, DarTG, MenTA, one tripartite type II TAC chaperone system, and hypothetical proteins. The study aims to demonstrate the genes encoded toxin-antitoxin system in mycobacterium tuberculosis strains from clinical samples. The pulmonary and extra-pulmonary tuberculosis clinical samples were collected, and smear microscopy (Ziehl-Neelsen staining) was performed for the detection of high bacilli (3+) count, followed by nucleic acid amplification assay. Bacterial culture and growth assay, genomic DNA extraction, and polymerase chain reaction were also carried out. The positive PTB and EPTB samples were determined by 3+ in microscopy smear and the total count of tubercle bacilli determined by NAAT assay was 8.0×1005 in sputum and 1.3×1004 CFU/ml in tissue abscess. Moreover, the genomic DNA was extracted from culture, and the amplification of Rv1044 and Rv1045 genes in 624 and 412 base pairs (between 600-700 and 400-500 in ladder), respectively, in the H37Rv and clinical samples was observed. It has been found that Rv1044 and Rv1045 are hypothetical proteins with 624 and 882 base pairs belonging to the AbiEi/AbiEii family of toxin-antitoxin loci. Moreover, the significant identification of TA-encoded loci genes may allow for the investigation of multidrugresistant and extensively drug-resistant tuberculosis.

The complete mitochondrial genome of Proedromys bedfordi Thomas 1911 (Arvicolinae, Rodentia)

The Duke of Bedford’s vole (Proedromys bedfordi Thomas 1911) is distributed only at the border areas of the Sichuan and Gansu Provinces, China. In this study, the first complete mitochondrial genome of P. bedfordi is generated and characterized. The assembled genome is 16,262 base pairs in length and the base compositions present clearly the A-T bias (60.84%). Its genetic constitution and arrangement are consistent with the taxon of the voles, including 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 2 main non-coding regions, D-loop region and OL region. This mitochondrial genome will be a significant supplement for the genus Proedromys and whole mitogenome phylogenetic analysis provided insights into further evolutionary research of the subfamily Arvicolinae.

The complete chloroplast genome and phylogenetic analysis of Rubus sumatranus Miq 1861 (Roseaceae)

The wild raspberry species Rubus sumatranus Miq 1861 is a promising resource for breeding thermotolerant cultivars. Its complete chloroplast genome spans 155,935 base pairs (bp), featuring the classic quadripartite structure: an 18,729 bp small single-copy region, an 85,662 bp large single-copy region, and two 25,772 bp inverted repeats. A total of 130 genes were identified, including 86 protein-coding, 36 tRNA genes, and 8 rRNA genes. A maximum likelihood phylogenetic tree based on chloroplast genomes shows that R. sumatranus, within the subgenus Idaeobatus, is sister to the subgenus Batothamnus. This confirms the polyphyletic nature of the subgenus Idaeobatus. The chloroplast genome assembly of R. sumatranus enhances our understanding of its evolutionary history.

Molecular self-assembly of stable and small branched DNA nanostructures: Higher than a helical turn is enough for hybridization

The Watson-Crick base pairing property of DNA is widely used for fabricating DNA nanostructures with well-defined geometry. Moreover, DNA nanostructures can be easily modified in terms of shape, size and function at the nanoscale level. Therefore, investigation on smaller and stable branched DNA (bDNA) is of critical significance for biomedical applications. In the present communication, we report smaller and stable branched DNA (bDNA) which is of critical significance for biomedical applications. In the present study, a novel strategy has been used in identifying stable bDNA nanostructures with a minimum number of Watson-Crick base pairings. The importance of hybridizing regions and helical twists between multiple oligonucleotides has been explored using various biophysical techniques. The electrophoretic analysis demonstrated that hybridizing regions with ≥12 nt nucleotides can form stable bDNA structures. Substantial negative enthalpic contributions determine the significance of base stacking and the length of oligonucleotides in the hybridization process. Finally, thermal melting investigations confirmed the creation of bDNA nanostructures with ≥12 nt long hybridizing regions. In general, our findings indicate that bDNA oligonucleotides do not undergo hybridization if the number of base pairs is lesser for a single helical turn. Furthermore, the yield and stability of smaller bDNA nanostructures in physiological conditions are comparable with the earlier reported higher-order structures. Hence, smaller bDNAs are more stable which may be preferred over conventional bDNA nanostructures for advanced biomedical applications.

Isolation, characterization, and potential application of Acinetobacter baumannii phages against extensively drug-resistant strains.

One of the significant issues in treating bacterial infections is the increasing prevalence of extensively drug-resistant (XDR) strains of Acinetobacter baumannii. In the face of limited or no viable treatment options for extensively drug-resistant (XDR) bacteria, there is a renewed interest in utilizing bacteriophages as a treatment option. Three Acinetobacter phages (vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln) were identified from hospital sewage and analyzed for their morphology, host ranges, and their genome sequences were determined and annotated. These phages and vB_AbaS_SA1 were combined to form a phage cocktail. The antibacterial effects of this cocktail and its combinations with selected antimicrobial agents were evaluated against the XDR A. baumannii strains. The phages exhibited siphovirus morphology. Out of a total of 30 XDR A. baumannii isolates, 33% were sensitive to vB_AbaS_Ftm, 30% to vB_AbaS_Gln, and 16.66% to vB_AbaS_Eva. When these phages were combined with antibiotics, they demonstrated a synergistic effect. The genome sizes of vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln were 48487, 50174, and 50043 base pairs (bp), respectively, and showed high similarity. Phage cocktail, when combined with antibiotics, showed synergistic effects on extensively drug-resistant (XDR) strains of A. baumannii. However, the need for further study to fully understand the mechanisms of action and potential limitations of using these phages is highlighted.

Molecular characterization of selected endophytic fungi isolate IDGG 3 leaf galing galing (cayratia trifolia L.) with the polymerase chain reaction method

Background: Endophytic fungi live in plant tissues and are usually not harmful to their host plants. One of the host plants for endophytic fungi is bush grape leaves (Cayratia trifolia L.). Objective: The research aimed to determine the molecular characteristics of the endophytic fungi isolates selected IDGG 3 of bush grape leaves. Methods: The test used the polymerase chain reaction method to determine the species-level characteristics using internal transcribed spacers (ITS) 1 and 4. The DNA band was successfully amplified with 500 base pairs with 3000 markers. Results: The fungi isolates selected IDGG 3 of bush grape leaves in molecular identification based on the Basic Local Alignment Search Tool (BLAST) analysis on Genebank NCBI that the IDGG 3 samples of bush grape leaves had a similarity level of 99%, namely Fusarium incarnatum JL5-2, Fusarium incarnatum JL3-4- 1, Fusarium incarnatum CBB-2, Fusarium incarnatum JL3-3, Fusarium incarnatum CBA-3, Fusarium incarnatum CBB-1, Fusarium incarnatum CBA-2, Fusarium chlamydospore, Fusarium cf. Incarnatum, and Fusarium sp. Conclusion: The results of the molecular characteristics of the selected endophytic fungi isolate IDGG 3 galing-galing leaves (Cayratia trifoliata L.) have the closest degree of kinship with the species Fusarium incarnatum JL3-4-1. The importance of the PCR method in the molecular characterization of endophytic fungi, as well as opening opportunities for further exploration of the biotechnological potential of endophytic fungi from Cayratia trifolia L.

SERS aptasensor detection of aflatoxin B1 based on silicon-au-ag Janus nanocomposites

Aflatoxin B1 (AFB1) is a prevalent contaminant in maize, posing significant threats to human health. This study designed AuAg Janus NPs with intrinsic Raman signals as signal probes and SiO2@AgNPs as capture probes. The two were coupled through complementary base pairing to ensure the ordered, controlled distribution of noble metal nanoparticles. The AuAg Janus NPs and the highly stable SiO2 carrier is expected to avoid the adverse effects on stability caused by using signal molecules and the formation of random aggregates when using the noble metal nanoparticle gap effect to concentrate on the electromagnetic field. This study improved the negative impact of AgNPs' high surface energy on their uniformity, while enhancing the pH adaptability of AuAg Janus NPs. In the presence of AFB1, the composite disintegrates, and the SERS intensity showed a negative correlation with AFB1 concentration, enabling highly sensitive and stable detection of AFB1.

Endogenous HiBiT-tagging combined with affinity complementation: A new strategy for small open reading frame-encoded polypeptide detection in bacteria

Bacterial genome annotations are continuously refined with the advent of novel techniques. Ribosome profiling, or Ribo-seq, utilizing next-generation sequencing to link genomic regions with translational activity, has uncovered numerous small open reading frames (sORFs) - arbitrarily defined as ORFs no longer than 300 base pairs - as a generally under-annotated class of genomic elements in both eukaryotic and prokaryotic genomes. While sORFs can function as regulatory elements, they may also translate into small proteins (equal to or shorter than 100 amino acids), classified as sORF-encoded polypeptides (SEPs). The inherent limitations of ribosome profiling necessitate the experimental validation of predicted (s)ORFs at the protein expression level. However, the small size and unique biochemical characteristics of SEPs pose significant challenges for traditional protein detection methods, like mass spectrometry and immunoblotting. In this study, we introduce HiBiT blotting, a luminescent, complementation-based protein detection method, as a highly sensitive alternative to antibody-based immunoblotting for investigating SEP expression in Salmonella enterica (serovar) Typhimurium (S. Typhimurium) at endogenous levels. We demonstrate its complementarity to mass spectrometry as an expression validation tool. Additionally, employing a biochemical fractionation approach, we determined the subcellular localization of validated S. Typhimurium SEPs, initiating exploration into the functional aspects of these SEPs, and proposed enrichment strategies that may facilitate future experimental validation of Ribo-seq-predicted sORFs.

Multivariate analysis of morphological variation, biochemical content, antioxidant activity, and molecular characteristics of autochthonous Berberis crataegina DC. genotypes in Central Türkiye

BackgroundOne of the wild fruit species with a natural distribution in Türkiye, and historically used for medicinal purposes due to its rich composition, is Berberis crataegina DC. Various parts of the plant, including its roots, bark, leaves, flowers, and fruits, have been utilized in traditional medicine, while its fruits are also consumed in various forms as food. This study aimed to characterize the morphological, biochemical, and molecular traits of B. crataegina genotypes naturally growing in the Kayseri region, located in central Türkiye.ResultsThe fruit weight of the genotypes ranged from 0.047 to 0.137 g, fruit width from 3.06 to 4.64 mm, and fruit length from 5.80 to 9.05 mm. Similarly, the leaf traits of the genotypes exhibited wide variation. The total phenolic content ranged from 190.53 to 297.55 mg GAE/100 g, total flavonoid content from 82.03 to 203.89 mg QE/100 g, total anthocyanin content from 4.54 to 11.76 mg cyn-3 gluc/100 g, and total antioxidant capacity between 57.76 and 87.93%. A principal component analysis (PCA) of 11 traits identified four principal components with eigenvalues greater than 1. The first four components accounted for 71.89% of the total variation, with PCA1 explaining 23.48%, PCA2 18.68%, PCA3 16.39%, and PCA4 13.34%. ISSR molecular analysis using nine markers revealed a band count ranging from 4 to 13, polymorphic band count between 3 and 10, and polymorphism rates from 61.54 to 100%, with band lengths ranging from 200 to 1000 base pairs. According to the UPGMA dendrogram based on molecular analyses, the genetic similarity between the genotypes ranged from 0.64 to 1.00, with B2 and B13 being the most similar genotypes.ConclusionsIn conclusion, the Kayseri region is rich in B. crataegina genotypes with wide genetic variation. The genotypes identified in this region may serve as valuable genetic resources for future studies.

Molecular basis of mRNA delivery to the bacterial ribosome.

Protein synthesis begins with the formation of a ribosome-messenger RNA (mRNA) complex. In bacteria, the small ribosomal subunit (30S) is recruited to many mRNAs through base pairing with the Shine-Dalgarno (SD) sequence and RNA binding by ribosomal protein bS1. Translation can initiate on nascent mRNAs, and RNA polymerase (RNAP) can promote the recruitment of the pioneering 30S. Here, we examined 30S recruitment to nascent mRNAs using cryo-electron microscopy, single-molecule fluorescence colocalization, and in-cell cross-linking mass spectrometry. We show that bS1 delivers the mRNA to the ribosome for SD duplex formation and 30S activation. Additionally, bS1 and RNAP stimulate translation initiation. Our work provides a mechanistic framework for how the SD duplex, ribosomal proteins, and RNAP cooperate in 30S recruitment to mRNAs and establish transcription-translation coupling.

Whole Genome Sequencing of Brucella melitensis Isolates from Kuwait for the Identification of Biovars, Variants and Relationship within a Biovar.

The identification of Brucella genotypes is essential for epidemiological studies. The whole genome sequencing is emerging as a novel tool for genetic characterization of infectious microbes. The aim of this study was to genotype B. melitensis isolates from Kuwait using whole genome sequencing and variant analysis of the sequence data. DNA were purified from 15 heat-inactivated B. melitensis isolates and used to prepare sequencing libraries employing Nextera XT DNA Sample Preparation Kit (Illumina San Diego, CA, USA) and sequenced on a MiSeq (Illumina). The sequence files were aligned to three biovars of B. melitensis, i.e., biovar 1 str. 16M, biovar 2 str. 63/9, and biovar 3 str. Ether. The alignment and variant calling were performed using 'bwa mem' and SAMtools/VCFtools, respectively. The genome size of all the isolates was around 3.3 mega base pairs and resembled B. melitensis biovar 2. Single nucleotide polymorphisms (SNPs), insertions and deletions, (indels) were spread all over the genome; but 138 SNPs were common among the 14 isolates, supporting the same ancestral origin. A neighbor-joining tree analysis identified isolate 2 as an outlier. In addition, SNPs (2 - 478) specific to each isolate were also identified, which divided the B. melitensis biovar 2 into two major groups/genotypes. A further analysis showed that the Kuwaiti isolates of the present study shared phylogeny mainly with strains from the Middle Eastern countries. Among the 15 isolates studied in Kuwait, biovar 2 is the most prevalent biovar of B. melitensis. Furthermore, isolate-specific genetic variations were identified, which may be useful in epidemiological investigations.

First report of Hepatozoon and Lankesterella spp. infections in wild rodents from Pakistan, and their potential impact on blood parameters and oxidative stress markers in vital organs.

Pakistan is home to a rich rodent fauna, yet no investigations have been conducted on the presence of Haemogregarines in these animals, leaving their epidemiology and genetic diversity in this subtropical region unexplored. In this study, blood samples from four wild rodent species, Rattus (R.) rattus (n = 122), Mus (M.) musculus (n = 64), Rattus norvegicus (n = 57), and Dryomys nitedula (n = 1), were collected between May 2022 and July 2023 from three districts in Punjab and three in Khyber Pakhtunkhwa. PCR targeting a fragment of 600 base pairs from the 18S rDNA gene revealed a 2.86% (7/244) prevalence of Haemogregarines in the rodents. DNA sequencing and BLAST analysis confirmed the presence of Hepatozoon spp. and Lankesterella spp. in the blood samples. Phylogenetic analysis indicated genetic diversity in the Pakistani sequences, which clustered with sequences found in reptiles, amphibians, birds, and mammals from various countries. The prevalence of the parasite varied among rodent species, with R. rattus showing the highest infection rate, followed by R. norvegicus and M. musculus. Female rodents were more frequently infected than males. Infected R. rattus exhibited significant disruptions in white blood cells, red blood cells, and platelet counts. Oxidative stress markers indicated elevated superoxide dismutase in the kidney, catalase in the heart, and malondialdehyde in the liver and lungs of infected rodents compared to uninfected ones. This study is an important contribution towards science as it the first report of Haemogregarines and Lankesterella spp. infections among Pakistani rodents.

WITHDRAWN: Mechanism of genome editing tools and their application on genetic inheritance disorders

In the fields of medicine and bioscience, gene editing is increasingly recognized as a promising therapeutic approach for treating pathogenic variants in humans and other living organisms. With advancements in technology and knowledge, it is now understood that most genetic defects are caused by single-base pair variants. The ability to substitute genes using genome editing tools enables scientists and doctors to cure genetic diseases and disorders. Starting with CRISPR/Cas, the technology has evolved to become more efficient and safer, leading to the development of Base and Prime Editors. Furthermore, various approaches are used to treat genetic disorders such as Hemophilia, Cystic Fibrosis, and Duchenne Muscular Dystrophy. As previously mentioned, most genetic defects leading to specific diseases are caused by single base pair variants, which can occur at many locations in corresponding gene, potentially causing the same disease. This means that, even when using the same genome editing tool, results in terms of editing efficiency or treatment effectiveness may differ. Therefore, different approaches may need to be applied to different types of diseases. Prevalently, due to the safety of AAV vectors in gene therapy, most clinical trials of gene therapy are based on AAV delivery methods. However, despite their safety and non-integration into the host genome, their limitations, such as confined capacity, dosage-dependent viral toxicity, and immunogenicity, necessitate the development of new approaches to enhance treatment effects. This review provides the structure and function of each CRISPR-based gene editing tool and focuses on introducing new approaches in gene therapy associated with improving treatment efficiency.