Translated Amino Acid Sequence Research Articles

New record of Carnidae (Diptera) from Taiwan and potential challenges in DNA barcode amplification due to pseudogene

The genus Carnus Nitzsch, 1818 comprises small ectoparasites that feed on the blood of juvenile avians. They are characterised by dealated adults with setose abdominal intersegmental membranes. Carnus orientalis Maa, 1968 was previously recorded in Malaysia and the Ryukyu Islands of Japan, parasitising two owl species: Ketupa ketupu (Horsfield, 1821) and Otus elegans (Cassin, 1852). This study confirms the occurrence of C. orientalis in Taiwan and presents a new host record, along with COI barcode sequences. Additionally, the study also elucidates the difficulties posed by blood meal contamination and pseudogene amplification as confounding factors intrinsic to the molecular taxonomic delineation of C. orientalis via universal DNA barcoding primers. The following new information regarding C. orientalis is provided in this study: Carnus orientalis is first recorded in Taiwan, filling the gap in its East Asian distribution. This is also the first record of Carnidae from Taiwan. Otus lettia (Hodgson, 1836) (Aves, Strigidae) is reported as a new host for C. orientalis, identified on a fallen fledgling. Co-amplification of the host's COI is reported in this study using the universal PCR primer set LCO1490/HCO2198. Additionally, the amplification of a COI-like pseudogene using a newly-designed primer set is detected through abnormal translated amino acid sequences and the occurrence of a stop codon. New specific primers for the COI gene of Carnus were designed in this study. Carnus orientalis is first recorded in Taiwan, filling the gap in its East Asian distribution. This is also the first record of Carnidae from Taiwan. Otus lettia (Hodgson, 1836) (Aves, Strigidae) is reported as a new host for C. orientalis, identified on a fallen fledgling. Co-amplification of the host's COI is reported in this study using the universal PCR primer set LCO1490/HCO2198. Additionally, the amplification of a COI-like pseudogene using a newly-designed primer set is detected through abnormal translated amino acid sequences and the occurrence of a stop codon. New specific primers for the COI gene of Carnus were designed in this study. The new distribution and ecological data of C. orientalis enhance our understanding of this species. The provision of new COI primers is anticipated to contribute to future studies employing DNA barcoding in bird-parasitic flies. The following new information regarding C. orientalis is provided in this study: Carnus orientalis is first recorded in Taiwan, filling the gap in its East Asian distribution. This is also the first record of Carnidae from Taiwan. Otus lettia (Hodgson, 1836) (Aves, Strigidae) is reported as a new host for C. orientalis, identified on a fallen fledgling. Co-amplification of the host's COI is reported in this study using the universal PCR primer set LCO1490/HCO2198. Additionally, the amplification of a COI-like pseudogene using a newly-designed primer set is detected through abnormal translated amino acid sequences and the occurrence of a stop codon. New specific primers for the COI gene of Carnus were designed in this study. Carnus orientalis is first recorded in Taiwan, filling the gap in its East Asian distribution. This is also the first record of Carnidae from Taiwan. Otus lettia (Hodgson, 1836) (Aves, Strigidae) is reported as a new host for C. orientalis, identified on a fallen fledgling. Co-amplification of the host's COI is reported in this study using the universal PCR primer set LCO1490/HCO2198. Additionally, the amplification of a COI-like pseudogene using a newly-designed primer set is detected through abnormal translated amino acid sequences and the occurrence of a stop codon. New specific primers for the COI gene of Carnus were designed in this study. The new distribution and ecological data of C. orientalis enhance our understanding of this species. The provision of new COI primers is anticipated to contribute to future studies employing DNA barcoding in bird-parasitic flies.

Identification of Novel Drug Compound for the Mutated Protein (PKBB-Protein Kinase B, Beta) Responsible for Endometrial Cancer Using Advanced Insilico Drug Designing Technique

Currently, one of the most important health problems affecting individuals worldwide is endometrial cancer. Numerous studies have been conducted by oncologists in an effort to create preventive medications that will lower the cancer's death rate. Using 3D Insilico drug docking techniques, we investigate the potential interactions between the mutant target protein, PKBB-Protein Kinase B, Beta, and the anti-cancer pharmaceutical (control drug), Mercaptopurine, and the proposed de novo drug derivative. To perform drug docking procedures, the translated amino acid sequence and three-dimensional chemical compound were acquired from the NCBI database. The most sophisticated 3D molecular imaging instruments were used to conduct post-docking studies. The de novo medicine directly blocks amino acid mutational sites, as the results of the docking investigation clearly showed (PKBB). The suggested de novo drug has a higher binding score than the control medication, mercaptopurine. Using molecular dynamics techniques, a three-dimensional image of the H-bond contact force between PKBB and the de novo drug is produced. Therefore, we deduce that the suggested anti-cancer de novo drug may aid in the treatment of endometrial cancer. Thus, we suggest the de novo medicine, which lessens discomfort while reducing the signs and symptoms of malignancy.

Heightened incidence of adverse events associated with a live attenuated varicella vaccine strain that lacks critical genetic polymorphisms in open reading frame 62

ObjectivesThis study aimed to identify the specific vaccine strain associated with herpes zoster (HZ) in children following a series of diagnosed cases and to explore whether differences in single nucleotide polymorphisms (SNPs) among various vaccine strains are linked to an increased incidence of herpes zoster after vaccination. MethodsFrom February 2021 to March 2024, children <12 years old suspected of vaccine-related varicella-like rash or HZ were included. Varicella zoster virus DNA isolated from the patients were sequenced to differentiate vaccine type versus wild-type. 3D protein structures of pORF62 were simulated using open reading frame 62 sequences extracted from whole genome sequencing of vOka, MAV/06, Oka/SK vaccines, and pOka reference. ResultsA total of 27 children with a median age of 2.1 (interquartile range, 1.5–3.4) years old presented with vaccine-related varicella-like rash (n = 4/27, 14.8%) or HZ (n = 23/27, 85.2%). One patient with varicella-like rash and 34.8% (n = 8/23) with HZ had disseminated skin involvement. All were immunized with the Oka/SK strain varicella vaccine. Genotyping showed 88.2% (n = 15/17) had SNPs specific to the Oka/SK strain, and two had SNPs considered pOka type contained within the Oka/SK vaccine. Despite accumulations of SNPs in ORF 62 of Oka/SK, the translated amino acid sequence and 3D protein structure were identical to wild-type pOka's pORF62. In vOKA and MAV/06, changes in amino acids occurred at two positions, S628G and R958G, within pORF62. The predicted 3D protein structure of vOka and MAV/06's pORF62 showed that the α helical structure within region I undergoes conformational change, potentially increasing difficulties in interactions with infection-related proteins and thereby decreasing virulence. pORF62 in pOka and Oka/SK exhibited more stable structure complex of the α helical structure. DiscussionLack of structural alternations in region I of pORF62 due to the absence of critical genetic polymorphisms in open reading frame 62 could be associated with the heightened incidence of adverse events.

Genetic Diversity Analysis of Rhynocoris marginatus Fabricius Based on 18S Ribosomal RNA Gene (Heteroptera: Reduviidae)

The R. marginatus are essential components of ecosystem, but also important in the biological control of insect pest, infesting a varity of agro ecosystem and medicine. The present investigation was carried out in the insect molecular genetic variation of 18S Ribosomal RNA gene from R. marginatus. The study was represent by the reduviid insect R. marginatus nucleotide gene sequences were translate amino acid sequence and obtained hydropathy, Domain, Transmembranes of proteins were calculated. The multiple gene sequence alignment of in-silico translated amino acid sequence of the partial ribosomal genes protein of R. marginatus were generated and the phylogenetic relationships were observed.

Protein-Protein Docking Studies on Magnesium Chelatase of Ulva fasciata Against the Human TSHR Protein

The course and prognosis of hypothyroidism in humans are affected by the most common mutation in the hypothyroid protein, TSHR. To facilitate the potential mutant TSHR's interaction with Ulva fasciata's magnesium-chelatase, we utilize 3D Insilico drug docking approaches. The purpose of the entire research work is to introduce the seaweed protein against the humanTSHR protein using Insilico protocols. The translated amino acid sequence and three-dimensional chemical compound were taken from the NCBI database to perform drug docking procedures. In post-docking tests, advanced 3D molecular visualization capabilities were utilized. In post-docking tests, advanced 3D molecular visualization capabilities were utilized. The findings of the docking investigation indisputably demonstrate that amino acid mutational sites are directly suppressed by magnesium chelatase. The H-bond interaction between TSHR and magnesium chelatase is shown in three dimensions utilizing concepts from molecular dynamics techniques. Ultimately, we found that Ulva fasciata's pharmaceutical ingredient, magnesium chelatase, aids in the treatment of hypothyroidism. Hence, we finally conclude that one of the main endocrine hormonal problems is hypothyroidism, and our research serves to demonstrate how well the seaweed Ulva fasciata works as a unique medicinal agent to treat this disorder.

Protein-protein Interaction Studies between TSHR Protein in Human and Photosystem II Protein D1 of Ulva fasciata using In silico Protocols

The most prevalent mutation in Hypothyroid Protein, TSHR impacts the course and prognosis of Hypothyroidism in human. We employ 3D in silico drug docking techniques to make the possible mutant TSHR interact with Photosystem II Protein D1 of Ulva fasciata. To carry out drug docking techniques, the translated amino acid sequence and three-dimensional chemical compound were obtained from the NCBI database. The use of sophisticated 3D molecular visualization tools was employed in post-docking experiments. The use of sophisticated 3D molecular visualization tools was employed in post-docking experiments. The docking study results unequivocally show that Photosystem II Protein D1 directly suppresses amino acid mutational sites. TSHR and Photosystem II Protein D1’s electrostatic force is depicted in a three-dimensional manner using notions from molecular dynamics techniques. In the end, we determined that Photosystem II Protein D1, a medicinal component of Ulva fasciata, helps in treating hypothyroidism. Hypothyroidism, being one of the major Endocrinological disorders, our research work helps to prove that the sea weed, Ulva fasciata can effectively act a novel therapeutic agent for treating this disorder.

Molecular Interaction Study between Gentamicin and the Cancer Protein Target (TP53) using in silico Tools

The most prevalent mutation in hepatocellular carcinoma (HCC), TP53 mutations impacts the course and prognosis of HCC. 3D in silico drug docking techniques were employed to make the possible mutant target tumor protein 53 (TP53) interact with benzene acetic acid (Gentamicin) and non-steroidal anti-inflammatory drugs (NSAIDs). To carry out drug docking techniques, the translated amino acid sequence and three-dimensional chemical compound were obtained from the NCBI database. The use of sophisticated 3D molecular visualization tools was employed in post-docking experiments. The docking study results unequivocally show that gentamicin directly suppresses amino acid mutational sites. TP53 and Gentamicin's electrostatic force is depicted in a three-dimensional manner using notions from molecular dynamics techniques. In the end, it was determined that gentamicin, an Antibiotic medication, helps treat liver cancer. Most chemotherapy medications cause pain. Therefore, Gentamicin shall be used to lessen discomfort while minimizing the symptoms of cancer.

The first detection of coccidia (Conoidasida: Eimeriidae) DNA in Godlewski’s sculpin Abyssocottus (Limnocottus) godlewskii (Dybowski, 1874)

For the first time, fragments of the cox1 gene of a representative of the family Eimeriidae were obtained by high-throughput sequencing in the digestive tract of Godlewski’s sculpin Abyssocottus (Limnocottus) godlewskii (Dybowski, 1874). The nucleotide sequences of the coccidia, which accounted for less than 0.01% of the total data set, belonged to a single genotype and were significantly different from all previously known. Phylogenetic reconstruction based on the translated amino acid sequences reliably revealed the basal location of branches belonging to representatives of the family Eimeriidae among fishes. The question of the genus of the detected organism remains unresolved due to the limited nucleotide data for representatives of the genera Eimeria, Calyptospora, and Goussia from fish.

Genome-scale approach to reconstructing the phylogenetic tree of psyllids (superfamily Psylloidea) with account of systematic bias

Psyllids (class Insecta: order Hemiptera: superfamily Psylloidea) are a taxonomically and phylogenetically challenging clade. Recent studies have largely advanced the phylogeny of this group, yet the family-level relationships among Aphalaridae, Carsidaridae, and others remain unresolved. Genome-scale phylogenetic analysis is known to provide a finer resolution for problems like that. However, such phylogenomics also introduces new problems: incorrect trees with high confidence yielded due to systematic error (bias). Here we addressed these issues using hundreds of single-copy orthologous (SCO) genes in psyllid transcriptomes and genomes. Our analyses revealed conflicts between the nucleotide-based and amino-acid-based phylogenetic trees. While the nucleotide-based phylogeny strongly supported the (Aphalaridae + Carsidaridae) + Others relationship, the amino-acid-based one recovered Aphalaridae + (Carsidaridae + Others) with 100% support. Further inspection revealed significant compositional heterogeneity in nucleotide sequences for 67% of SCO genes, but not in the corresponding translated amino acid sequences. We then used different strategies to combat this compositional bias, and found that using the RY-coding strategy (coding the standard nucleotides as purines and pyrimidines) the nucleotide-based phylogeny became consistent with the amino-acid-based one. We further applied RY-coding to a published concatenated nucleotide dataset and recovered the Aphalaridae monophyly (which is refuted by the original literature on non-recoded sequences) at the base of psyllid tree. Moreover, it was found that variations in evolutionary rate could lead to errors in nucleotide-based phylogeny. The fast-evolving Heteropsylla cubana (Psyllidae: Ciriacreminae) was incorrectly placed within the subfamily Psyllinae. This bias can be avoided by using data removal or RY-coding strategies. Together, our results strongly support the family relationship of Aphalaridae + (Carsidaridae + Others), and show that the amino-acid-based concatenation analysis is more robust than nucleotide-based one. Future phylogenomic analysis of psyllid nucleotide sequences should take into account methods such as the RY-coding scheme to address potential systematic biases arising from composition and rate heterogeneities.

In silico analysis of xylanase in Bacillus coagulans ST-6

Bacillus coagulans ST-6 is a thermophile isolated from a local hot spring in Malaysia. It expresses xylanase activity and has potential industrial applications. In silico structure prediction and modeling of a 210 amino acid sequence (XYNBC) translated from 872 bp Bacillus coagulans ST-6 xylanase gene was performed. The predicted molecular weight of the translated amino acid sequence is 20 kDa with theoretical isoelectric point value at 9.10 predicting that the xylanase belongs to the GH11 xylanase family. The protein is predicted to be hydrophilic with Grand average of hydropathicity (GRAVY) value of -0.6555 and possibly has better interaction with water. Predicted motif for XYNBC was revealed to be from Glycosyl hydrolases family 11(IPR00137) with signature motif 1 and 2 and only one known activity, xylanase. A three-dimensional (3D) model was constructed using PDB ID 2DCZ|A (http://www.rcsb. org) as the template as it has the highest similarity with XYNBC protein sequence. The 2DCZ|A xylanase sequence is from Bacillus Subtilis family-11 and is 185 amino acid long. The predicted 3D model consists of eleven beta sheets and one alpha helix. The stereochemical quality of protein structure revealed by Ramachandran Plot showed acceptable model with 99.4% residues fall in the most favored regions. Ten residues were predicted to be involved in active sites where residues Tyr7, Val15 and Asn16 are located at beta sheet while Asp118, Gly119, Thr120 are located at the loop. The catalytic residues E78 and E172 common to other G1H1 xylanases were also revealed.

ProsmORF-pred: a machine learning-based method for the identification of small ORFs in prokaryotic genomes.

Small open reading frames (smORFs) encoding proteins less than 100amino acids (aa) are known to be important regulators of key cellular processes. However, their computational identification remains a challenge. Based on a comprehensive analysis of known prokaryotic small ORFs, we have developed the ProsmORF-pred resource which uses a machine learning (ML)-based method for prediction of smORFs in the prokaryotic genome sequences. ProsmORF-pred consists of two ML models, one for initiation site recognition in nucleic acid sequences upstream of putative start codons and the other uses translated amino acid sequences to decipher functional protein like sequences. The nucleotide sequence-based initiation site recognition model has been trained using longer ORFs (>100aa) in the same genome while the ML model for identification of protein like sequences has been trained using annotated smORFs from Escherichia coli. Comprehensive benchmarking of ProsmORF-pred reveals that its performance is comparable to other state-of-the-art approaches on the annotated smORF set derived from 32 prokaryotic genomes. Its performance is distinctly superior to other tools like PRODIGAL and RANSEPS for prediction of newly identified smORFs which have a length range of 10-30aa, where prediction of smORFs has been a major challenge. Apart from identification of smORFs in genomic sequences, ProsmORF-pred can also aid in functional annotation of the predicted smORFs based on sequence similarity and genomic neighbourhood similarity searches in ProsmORFDB, a well-curated database of known smORFs. ProsmORF-pred along with its backend database ProsmORFDB is available as a user-friendly web server (http://www.nii.ac.in/prosmorfpred.html).

Intragenomic rearrangements involving 5′-untranslated region segments in SARS-CoV-2, other betacoronaviruses, and alphacoronaviruses

BackgroundVariation of the betacoronavirus SARS-CoV-2 has been the bane of COVID-19 control. Documented variation includes point mutations, deletions, insertions, and recombination among closely or distantly related coronaviruses. Here, we describe yet another aspect of genome variation by beta- and alphacoronaviruses that was first documented in an infectious isolate of the betacoronavirus SARS-CoV-2, obtained from 3 patients in Hong Kong that had a 5′-untranslated region segment at the end of the ORF6 gene that in its new location translated into an ORF6 protein with a predicted modified carboxyl terminus. While comparing the amino acid sequences of translated ORF8 genes in the GenBank database, we found a subsegment of the same 5′-UTR-derived amino acid sequence modifying the distal end of ORF8 of an isolate from the United States and decided to carry out a systematic search.MethodsUsing the nucleotide and in the case of SARS-CoV-2 also the translated amino acid sequence in three reading frames of the genomic termini of coronaviruses as query sequences, we searched for 5′-UTR sequences in regions other than the 5′-UTR in SARS-CoV-2 and reference strains of alpha-, beta-, gamma-, and delta-coronaviruses.ResultsWe here report numerous genomic insertions of 5′-untranslated region sequences into coding regions of SARS-CoV-2, other betacoronaviruses, and alphacoronaviruses, but not delta- or gammacoronaviruses. To our knowledge this is the first systematic description of such insertions. In many cases, these insertions would change viral protein sequences and further foster genomic flexibility and viral adaptability through insertion of transcription regulatory sequences in novel positions within the genome. Among human Embecorivus betacoronaviruses, for instance, from 65% to all of the surveyed sequences in publicly available databases contain inserted 5′-UTR sequences.ConclusionThe intragenomic rearrangements involving 5′-untranslated region sequences described here, which in several cases affect highly conserved genes with a low propensity for recombination, may underlie the generation of variants homotypic with those of concern or interest and with potentially differing pathogenic profiles. Intragenomic rearrangements thus add to our appreciation of how variants of SARS-CoV-2 and other beta- and alphacoronaviruses may arise.

Identification and in silico structural and functional analysis of a histone H2A-derived antimicrobial peptide from teleost fishes

BackgroundAntimicrobial peptides (AMPs) are small peptides that form the first line of defence in all living organisms. In addition to conventional AMPs, there exist molecules that are often not regarded as part of the innate defences, but possess antimicrobial activity. Histone H2A-derived peptides are a group of unconventional AMPs that represent very ancient defence factors, re-used in new ways during evolution. The present study attempts to compare histone H2A-derived AMPs from four commercially important teleost fishes inhabiting various habitats, viz. Asian sea bass (Lates calcarifer), tank goby (Glossogobius giuris), grey mullet (Mugil cephalus), and Malabar trevally (Carangoides malabaricus). Methods and resultsRT-PCR amplification of the mRNAs extracted from these fishes yielded H2A-derived AMP genes with slight differences in the nucleotide sequences, nevertheless with similar translated amino acid sequences. The amino acid sequence was found to possess marked similarity at the N-terminal region and showed antimicrobial motif sequence characteristic of histone H2A-derived AMPs. Phylogenetic analysis showed an evolutionary relationship with invertebrate ones suggesting a common ancestral origin for the vertebrate and invertebrate histone H2A. The secondary structure of histone H2A-derived AMP showed the presence of α-helices and intermittent coils having a proline hinge connecting the N- and C-terminus. The peptide was also predicted to be a cell-penetrating one. ConclusionIn silico structural and functional characterization of histone H2A-derived AMP revealed its potential antibacterial property and point towards the possibility of being an efficient AMP with the potential of being made use of in therapeutics.

A non-synonymous SNP in homolog of BADH2 gene is associated with fresh pod fragrance in dolichos bean (Lablab purpureus var. lignosus (Prain) Kumari).

Fresh pods are harvestable and marketable economic product in dolichos bean. Fresh pod fragrance is one of the 'farmers' and 'consumers' preferred traits in dolichos bean varieties. The pods with high fragrance fetch a premium price in the market. In breeding programmes, pod fragrance is routinely assessed by organoleptic (sensory) means, which is highly relative and subjective. Identification of linked DNA markers not only offer an objective means but also enable selection of fragment genotypes at seedling stage itself. Betaine aldehyde dehydrogenase (BADH) is known to be the key gene responsible for fragrance in other legumes such as vegetable soybean and mung bean. Taking cues from highly conserved domains in proteins coded by BADH genes, we isolated dolichos bean homolog (LpBADH2) of soybean GmBADH2 gene using reported degenerate primers designed to conserved domains. Analysis of the translated amino acid sequence of LpBADH2 showed high degree of similarity (97.30%) with those of soybean homolog (GmBADH2). Conserved amino acid sequence of aldehyde dehydrogenase-super family were also identified in LpBADH2. Multiple sequence alignment of nucleotide sequences of LpBADH2 with those of related legumes using "ClustalW" revealed the presence of a single non-synonymous single nucleotide polymorphic (SNPs) and three synonymous SNP sites in LpBADH2. The substitution of the amino acid tyrosine in (fragrant genotypes) with phenyl alanine (non-fragrant genotypes) in protein coded by LpBADH2 appeared to be the cause for switch over from fragrance to non-fragrance in dolichos bean. These results are discussed in relation to strategies to breed dolichos bean cultivars with desired level of pod fragrance.

Longer Duration of Active Oil Biosynthesis during Seed Development Is Crucial for High Oil Yield-Lessons from Genome-Wide In Silico Mining and RNA-Seq Validation in Sesame.

Sesame, one of the ancient oil crops, is an important oilseed due to its nutritionally rich seeds with high protein content. Genomic scale information for sesame has become available in the public databases in recent years. The genes and their families involved in oil biosynthesis in sesame are less studied than in other oilseed crops. Therefore, we retrieved a total of 69 genes and their translated amino acid sequences, associated with gene families linked to the oil biosynthetic pathway. Genome-wide in silico mining helped identify key regulatory genes for oil biosynthesis, though the findings require functional validation. Comparing sequences of the SiSAD (stearoyl-acyl carrier protein (ACP)-desaturase) coding genes with known SADs helped identify two SiSAD family members that may be palmitoyl-ACP-specific. Based on homology with lysophosphatidic acid acyltransferase (LPAAT) sequences, an uncharacterized gene has been identified as SiLPAAT1. Identified key regulatory genes associated with high oil content were also validated using publicly available transcriptome datasets of genotypes contrasting for oil content at different developmental stages. Our study provides evidence that a longer duration of active oil biosynthesis is crucial for high oil accumulation during seed development. This underscores the importance of early onset of oil biosynthesis in developing seeds. Up-regulating, identified key regulatory genes of oil biosynthesis during early onset of seed development, should help increase oil yields.

Molecular Characterization of TLR4 Gene of Swamp and Riverine Type of Water Buffaloes

Molecular characterization of TLR4 genes of swamp and riverine types of water buffaloes was conducted to determine unique genotypic characteristics specific to each type of water buffalo and provide baseline information for explaining differences in disease resistance between each type. The nucleotide sequence of the riverine TLR4 gene produced 2,526 bp, whereas the swamp TLR4 sequence produced 2,322 bp. Riverine and swamp buffalo TLR4 nucleotide sequences were 99% similar to that of B. bubalis sequence from GenBank. The translated amino acid sequence of TLR4 showed that the riverine buffalo TLR4 sequence had a higher percentage similarity to B. bubalis sequence as compared to the swamp buffalo TLR4 amino acid sequence. Phylogenetic analysis confirmed the degree of relationship between the bubaline species and showed the distinctness of each breed. The predicted functional domains identified were 13 LRRs and a single LRRCT. Transmembrane protein and globular cytoplasmic domain TIR1 were found to be conserved regions.

Phylogenetic analyses of the mitochondrial, plastid, and nuclear genes of Babesia sp. Mymensingh and its naming as Babesia naoakii n. sp.

BackgroundThe recently discovered Babesia sp. Mymensingh, which causes clinical bovine babesiosis, has a wide geographical distribution. We investigated the phylogenetic position of Babesia sp. Mymensingh using its mitochondrial, plastid, and nuclear genes. Based on morphological and molecular data, Babesia sp. Mymensingh is a unique species and we named it as Babesia naoakii n. sp.MethodsA blood DNA sample from a Babesia sp. Mymensingh-infected cow was subjected to genome sequencing to obtain the sequences of mitochondrial, plastid, and nuclear genes. Six phylogenetic trees were then constructed with (1) concatenated amino acid sequences of cytochrome oxidase subunit I, cytochrome oxidase subunit III, and cytochrome b genes of the mitochondrial genome; (2) 16S rRNA of the plastid genome; (3) nucleotide sequences of the elongation factor Tu gene of the plastid genome; (4) ITS1-5.8S rRNA-ITS2; (5) concatenated nucleotide sequences of 89 nuclear genes; and (6) concatenated amino acid sequences translated from the 89 nuclear genes.ResultsIn all six phylogenetic trees, B. naoakii n. sp. formed a sister clade to the common ancestor of Babesia bigemina and B. ovata. The concatenated nuclear genes of B. naoakii n. sp. and their translated amino acid sequences shared lower identity scores with the sequences from B. bigemina (82.7% and 84.7%, respectively) and B. ovata (83.5% and 85.5%, respectively) compared with the identity scores shared between the B. bigemina and B. ovata sequences (86.3% and 87.9%, respectively).ConclusionsOur study showed that B. naoakii n. sp. occupies a unique phylogenetic position distinct from existing Babesia species. Our findings, together with morphological differences, identify B. naoakii n. sp. as a distinct parasite species.Graphical

Analysis of sequence variations in the GABA gated chloride gene associated with resistance to fipronil in Rhipicephalus (Boophilus) microplus ticks from Argentina

This study aimed to evaluate the sequence variations of the GABA-Cl gene of Rhipicephalus (Boophilus) microplus ticks from Argentina with different resistance levels to fipronil. Genomic DNA was obtained from engorged females (n = 50) of fipronil-susceptible and resistant tick field populations from five localities in northern Argentina. Tick populations were tested for fipronil resistance by in vitro larval packet test. DNA encoding amino acid residues 264–360 of the GABA-Cl was amplified by PCR. PCR products were sequenced and the translated amino acid sequences were aligned and compared among each other and with sequences of the GABA-Cl belonging to the Brazilian and Uruguayan reference strains fipronil-susceptible (Mozo) and resistant (RFSan and Juarez). All the GABA-Cl amino acid sequences obtained from the R. microplus specimens from Argentina were identical, regardless of the fipronil resistance-susceptibility status of each population. Mutations present in GABA-Cl sequence of RFSan and Juarez strains were not present in any of the populations from Argentina analyzed. These results indicate the failure of this gene as a molecular marker of resistance to fipronil, at least in the R. microplus populations from northern Argentina.

Biodiversity of N-acyl homoserine lactonase (aiiA) gene from Bacillus subtilis

Microorganisms rely on the benefit of using chemical signals called autoinducers (AIs) as a connection matter in term of population, this mechanism is known as quorum sensing (QS). Quorum sensing is responsible for formation of biofilm together with virulence in bacteria. The most known QS molecule is N-acyl homoserine lactones (AHLs). A lot of degrading enzymes including lactonases that open the AHL ring and acylases that breakdown its acyl side chain can degrade or inactivate AHL. Due to similarity in lactone ring structure among AHLs it is susceptible to most of lactonases. Bacillus species are among the most promising bacteria producing AHL-lactonase. The aim of the work is to identify and study the diversity of the AHL-Lactonase gene among different Bacillus subtilis as a promising Quorum Quenching (QQ) strategy to prevent bacterial infections and biofilm formation. The AHL-lactonase (aiiA) gene of 64 B. subtilis isolates was amplified and sequenced followed by multiple sequence alignment of the translated amino acid sequences, homology modeling and docking study. An expected PCR product of about 750 base pair was detected in 22 B. subtilis isolates, and the results revealed that the isolates' sequences showed identity ranged between 97.61% and 99.47% with those in the NCBI GenBank database with 100% query coverage and 0.0 E-value. In addition, the results revealed high level of identity between many aiiA gene sequences of our isolates as they were closely related to the same sequences to many sequences of the NCBI GenBank database. The alignment of the amino acid sequences from the 22 B. subtilis isolates indicated that 84.4% of the amino acid residues were conserved between the aligned sequences. Docking of the co-crystalized ligand to wildtype and H109Y mutated protein showed a significant reduction of docking score for the mutated protein. This result indicate that this mutation might affect recognition or at least kinetics of these enzymes and hence their roles in quorum-quenching.

Molecular Characterization and Polymorphism of Inhibin (INHβA) Gene in Water Buffalo (Bubalus bubalis) Bulls

The study characterized inhibin-βA (INHβA) gene of 12 water buffalo (Bubalus bubalis) bulls [two Philippine Carabao (PC), five Bulgarian Murrah (BM), and five Italian Murrah (IM)] using DNA extracted from semen. Using MEGA 7.0 software and Signal P® version 4, the sequences were assembled and aligned and amino acid sequences were determined, respectively. BM bull sequence showed 100% similarity with B. bubalis from the National Center for Biotechnology Information (NCBI) database. However, a lower similarity was seen on both PC and IM at 98%. The translated amino acid sequence of PC, BM, and IM had 95, 97, and 93% similarity with B. bubalis sequence from NCBI, respectively. The phylogeny tree revealed that both PC and IM were closely similar in their gene sequence, while BM was more similar with B. bubalis from NCBI. Their similarities in nucleotide sequence suggest that INHβA gene was conserved in bulls. The signal peptide was observed in BM and B. bubalis from NCBI. BM has an amino acid exchanged from threonine to alanine, while in B. bubalis has an amino acid exchanged from histidine to aspartic acid. This polymorphism from their cleaving site could cause an interaction in the reproduction, growth, and maintenance of the bull.