Genomic Coding Sequences Research Articles

Olive Leaf Mottling Virus: A New Member of the Genus Olivavirus.

Studies of the virome of olive trees with symptoms of leaf mottling by high-throughput sequencing (HTS) revealed the presence of a new virus. Full coding genome sequences of two isolates were determined and consisted of a single RNA segment of 16,516 nt and 16,489, respectively. The genomic organization contained 10 open reading frames (ORFs) from 5' to 3': ORF1a, ORF1b (RdRp), ORF2 (p22), ORF3 (p7), ORF4 (HSP70h), ORF5 (HSP90h), ORF6 (CP), ORF7 (p19), ORF8 (p12), ORF9 (p23) and ORF10 (p9). Phylogenetic analyses clustered this virus in the genus Olivavirus, family Closteroviridae, with the closest species being Olivavirus flaviolae, commonly named olive leaf yellowing-associated virus (OLYaV). However, amino acid sequences of all taxonomically relevant proteins showed, in all cases, a divergence higher than 25% between OLYaV and the new virus, indicating that it represents a new species in the genus Olivavirus for which the common name of olive leaf mottling virus (OLMV) is proposed. This study represents an advance in the genus Olivavirus and provides new insights into the olive virome.

Molecular evolution of Hokkaido virus, a genotype of Orthohantavirus puumalaense, among Myodes rodents

Viruses in the genus Orthohantavirus within the family Hantaviridae cause human hantavirus infections and represent a threat to public health. Hokkaido virus (HOKV), a genotype of Orthohantavirus puumalaense (Puumala virus; PUUV), was first identified in Tobetsu, Hokkaido, Japan. Although it is genetically related to the prototype of PUUV, the evolutionary pathway of HOKV is unclear. We conducted a field survey in a forest in Tobetsu in 2022 and captured 44 rodents. Complete coding genome sequences of HOKVs were obtained from five viral-RNA-positive rodents (four Myodes rufocanus bedfordiae and one Apodemus speciosus). Phylogenetic analysis revealed a close relationship between the phylogenies and geographical origins of M. rufocanus-related orthohantaviruses. Comparison of the phylogenetic trees of the S segments of orthohantaviruses and the cytochrome b genes of Myodes species suggested that Myodes-related orthohantaviruses evolved in Myodes rodent species as a result of genetic isolation and host switching.

The complete coding sequence of Influenza A/Unknown/Chelyabinsk/206/H7N4.

An influenza virus strain was obtained during a bird surveillance study in 2023 near Lake Chebarkul in the Chelyabinsk region, Russia. This complete coding genome sequence of the virus sampled from the Ural region significantly expands the knowledge about the spread of the H7N4 subtype of the influenza A virus.

Bacterial hypothetical proteins may be of functional interest

Genomic analysis is part of the daily routine for many microbiology researchers. These analyses frequently unveil genes that encode proteins with uncertain functions, and for many bacterial species, these unknown genes constitute a significant proportion of their genomic coding sequences. Because these genes do not have defined functions, they are often overlooked in analyses. Experimentally determining the function of a gene can be challenging; however, ongoing advancements in bioinformatics tools, especially in protein structural analysis, are making it progressively easier to assign functions to hypothetical sequences. Leveraging various complementary tools and automated pipelines for annotating hypothetical sequences could ultimately enhance our comprehension of microbial functions and provide direction for new laboratory experiments.

Complete coding genome sequence of a Teschovirus A genotype strain.

This study reports the complete coding genome sequence of a novel Teschovirus A genotype strain, SG2, isolated from the fecal sample of an infected indigenous pig in western Jiangxi, China.

Genomic Analysis of Romanian Lycium Genotypes: Exploring BODYGUARD Genes for Stress Resistance Breeding.

Goji berries, long valued in Traditional Chinese Medicine and Asian cuisine for their wide range of medicinal benefits, are now considered a 'superfruit' and functional food worldwide. Because of growing demand, Europe and North America are increasing their goji berry production, using goji berry varieties that are not originally from these regions. European breeding programs are focusing on producing Lycium varieties adapted to local conditions and market demands. By 2023, seven varieties of goji berries were successfully registered in Romania, developed using germplasm that originated from sources outside the country. A broader project focused on goji berry breeding was initiated in 2014 at USAMV Bucharest. In the present research, five cultivated and three wild L. barbarum genotypes were compared to analyse genetic variation at the whole genome level. In addition, a case study presents the differences in the genomic coding sequences of BODYGUARD (BDG) 3 and 4 genes from chromosomes 4, 8, and 9, which are involved in cuticle-related resistance. All three BDG genes show distinctive differences between the cultivated and wild-type genotypes at the SNP level. In the BDG 4 gene located on chromosome 8, 69% of SNPs differentiate the wild from the cultivated genotypes, while in BDG 3 on chromosome 4, 64% of SNPs could tell the difference between the wild and cultivated goji berry. The research also uncovered significant SNP and InDel differences between cultivated and wild genotypes, in the entire genome, providing crucial insights for goji berry breeders to support the development of goji berry cultivation in Romania.

The first genomic characterization of a stable, hemin-dependent small colony variant strain of Staphylococcus epidermidis isolated from a prosthetic-joint infection

Phenotype switching from a wild type (WT) to a slow-growing subpopulation, referred to as small colony variants (SCVs), supports an infectious lifestyle of Staphylococcus epidermidis, the leading cause of medical device-related infections. Specific mechanisms underlying formation of SCVs and involved in the shaping of their pathogenic potential are of particular interest for stable strains as they have been only rarely cultured from clinical specimens. As the SCV phenotype stability implies the existence of genetic changes, the whole genome sequence of a stable, hemin-dependent S. epidermidis SCV strain (named 49SCV) involved in a late prosthetic joint infection was analyzed. The strain was isolated in a monoculture without a corresponding WT clone, therefore, its genome was compared against five reference S. epidermidis strains (ATCC12228, ATCC14990, NBRC113846, O47, and RP62A), both at the level of the genome structure and coding sequences. According to the Multilocus Sequence Typing analysis, the 49SCV strain represented the sequence type 2 (ST2) regarded as the most prominent infection-causing lineage with a worldwide dissemination. Genomic features unique to 49SCV included the absence of the Staphylococcal Cassette Chromosome (SCC), ~12 kb deletion with the loss of genes involved in the arginine deiminase pathway, and frameshift-generating mutations within the poly(A) and poly(T) homopolymeric tracts. Indels were identified in loci associated with adherence, metabolism, stress response, virulence, and cell wall synthesis. Of note, deletion in the poly(A) of the hemA gene has been considered a possible trigger factor for the phenotype transition and hemin auxotrophy in the strain. To our knowledge, the study represents the first genomic characterization of a clinical, stable and hemin-dependent S. epidermidis SCV strain. We propose that previously unreported indels in the homopolymeric tracts can constitute a background of the SCV phenotype due to a resulting truncation of the corresponding proteins and their possible biological dysfunction. Streamline of genetic content evidenced by the loss of the SCC and a large genomic deletion can represent a possible strategy associated both with the SCV phenotype and its adaptation to chronicity.

Multi-environment ecogenomics analysis of the cosmopolitan phylum Gemmatimonadota.

Gemmatimonadota is a diverse bacterial phylum commonly found in environments such as soils, rhizospheres, fresh waters, and sediments. So far, the phylum contains just six cultured species (five of them sequenced), which limits our understanding of their diversity and metabolism. Therefore, we analyzed over 400 metagenome-assembled genomes (MAGs) and 5 culture-derived genomes representing Gemmatimonadota from various aquatic environments, hydrothermal vents, sediments, soils, and host-associated (with marine sponges and coral) species. The principal coordinate analysis based on the presence/absence of genes in Gemmatimonadota genomes and phylogenomic analysis documented that marine and host-associated Gemmatimonadota were the most distant from freshwater and wastewater species. A smaller genome size and coding sequences (CDS) number reduction were observed in marine MAGs, pointing to an oligotrophic environmental adaptation. Several metabolic pathways are restricted to specific environments. For example, genes for anoxygenic phototrophy were found only in freshwater, wastewater, and soda lake sediment genomes. There were several genomes from soda lake sediments and wastewater containing type IC/ID ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO). Various genomes from wastewater harbored bacterial type II RuBisCO, whereas RuBisCO-like protein was found in genomes from fresh waters, soil, host-associated, and marine sediments. Gemmatimonadota does not contain nitrogen fixation genes; however, the nosZ gene, involved in the reduction of N2O, was present in genomes from most environments, missing only in marine water and host-associated Gemmatimonadota. The presented data suggest that Gemmatimonadota evolved as an organotrophic species relying on aerobic respiration and then remodeled its genome inventory when adapting to particular environments. IMPORTANCE Gemmatimonadota is a rarely studied bacterial phylum consisting of a handful of cultured species. Recent culture-independent studies documented that these organisms are distributed in many environments, including soil, marine, fresh, and waste waters. However, due to the lack of cultured species, information about their metabolic potential and environmental role is scarce. Therefore, we collected Gemmatimonadota metagenome-assembled genomes (MAGs) from different habitats and performed a systematic analysis of their genomic characteristics and metabolic potential. Our results show how Gemmatimonadota have adapted their genomes to different environments.

Visualizing genomic data: The mixing perspective

We report on a novel way to visualize genomic data. By considering genome coding sequences, cds, as sets of the N=61 non-stop codons, one obtains a partition of the total number of codons in each cds. Partitions exhibit a statistical property known as mixing character which characterizes how mixed the partition is. Mixing characters have been shown mathematically to exhibit a partial order known as majorization (Ruch, 1975). In previous work (Seitz and Kirwan, 2022) we developed an approach that combined mixing and entropy that is visualized as a scatter plot. If we consider all 1,121,505 partitions of 61 codons, this produces a plot we call the theoretical mixing space, TGMS. A normalization procedure is developed here and applied to real genomic data to produce the genome mixing signature, GMS. Example GMS’s of 19 species, including Homo sapiens, are shown and discussed.

Complete Coding Genome Sequences of Five Foot-and-Mouth Disease Viruses Belonging to Serotype O, Isolated from Cattle in Uganda in 2015 to 2016.

ABSTRACTComplete coding genome sequences of five foot-and-mouth disease virus (FMDV) serotype O strains that were isolated from the field between 2015 and 2016 showed five lineages within the EA-2 topotype circulating in four different regions (northern, western, eastern, and central) of Uganda. The genomic diversity may help in devising FMDV control strategies for Uganda.

Genomic and In Vitro Phenotypic Comparisons of Epidemic and Non-Epidemic Getah Virus Strains.

Getah virus is an emerging mosquito-borne animal pathogen. Four phylogenetic groups of GETV, Group I (GI), GII, GIII and GIV, were identified. However, only the GETV GIII was associated with disease epidemics suggesting possible virulence difference in this virus group. Here, we compared the genetic and in vitro phenotypic characteristics between the epidemic and non-epidemic GETV. Our complete coding genome sequence analyses revealed several amino acid substitutions unique to the GETV GIII and GIV groups, which were found mainly in the hypervariable domain of nsP3 and E2 proteins. Replication kinetics of the epidemic (GIII MI-110 and GIII 14-I-605) and non-epidemic GETV strains (prototype GI MM2021 and GIV B254) were compared in mammalian Vero cells and mosquito C6/36 and U4.4 cells. In all cells used, both epidemic GETV GIII MI-110 and GIII 14-I-605 strains showed replication rates and mean maximum titers at least 2.7-fold and 2.3-fold higher than those of GIV B254, respectively (Bonferroni posttest, p < 0.01). In Vero cells, the epidemic GETV strains caused more pronounced cytopathic effects in comparison to the GIV B254. Our findings suggest that higher virus replication competency that produces higher virus titers during infection may be the main determinant of virulence and epidemic potential of GETV.

Genome-wide development of lncRNA-derived-SSR markers for Dongxiang wild rice (Oryza rufipogon Griff.)

AbstractDongxiang wild rice (Oryza rufipogon Griff.) (DXWR) is the northernmost distributed wild rice found in the world. Similar to other populations of O. rufipogon, DXWR contains a large number of agronomically valuable genes, which makes it a natural gene pool for rice breeding. Molecular markers, especially simple repeat sequence (SSR) markers, play important roles in plant breeding. Although a large number of SSR markers have been developed, most of them are derived from the genome coding sequences, rarely from non-coding sequences. Meanwhile, long non-coding RNAs (lncRNAs), which are derived from the transcription of non-coding sequences, play vital roles in plant growth, development and stress responses. In our previous study, we obtained 1655 lncRNA transcripts from DXWR using strand-specific RNA sequencing. In this study, 1878 SSR loci were detected from the lncRNA sequences of DXWR, and 1258 lncRNA-derived-SSR markers were developed on the genome-wide scale. To verify the validity and applicability of these markers, 72 pairs of primers were randomly selected to test 44 rice accessions. The results showed that 42 (58.33%) pairs of primers have abundant polymorphism among these rice materials; the polymorphism information content values ranged from 0.04 to 0.87 with an average of 0.50; the genetic diversity index of SSR loci varied from 0.04 to 0.88 with an average of 0.56; and the number of alleles per marker ranged from 2 to 11 with an average of 4.36. Thus, we concluded that these lncRNA-derived-SSR markers are a very useful source for future basic and applied research.

Complete Coding Genome Sequence of a Novel Porcine Reproductive and Respiratory Syndrome Virus 2 Restriction Fragment Length Polymorphism 1-4-4 Lineage 1C Variant Identified in Iowa, USA.

ABSTRACTA porcine reproductive and respiratory syndrome virus 2 strain was identified in lung samples from nursery piglets associated with a 17.15% mortality rate on a swine farm in Iowa. Open reading frame 5 (ORF5) sequencing indicated that this strain is a restriction fragment length polymorphism (RFLP) 1-4-4 lineage 1C variant strain, and its complete coding genome sequence was determined.

Genotyping atypical porcine pestivirus using NS5a

Atypical porcine pestivirus (APPV) is an emerging virus discovered in 2014 and it can cause congenital tremors in pigs. Molecular epidemiology serves as an essential tool in monitoring and controlling the disease. Virus epidemiology mainly relies on genome sequencing and phylogenetic characterization. Previous molecular epidemiology studies have been using different genes/regions for phylogeny, namely whole genome, Npro, and E2 coding sequences. However, with increasing number of APPV sequences available in GenBank, no systemic studies have been performed for detailed classification of APPV strains around the globe. The goal of this study is to propose a classification strategy or taxonomy of APPV strains at genotype, subgenotype, and isolate levels. A total of 76 whole genomes and 16 partial polyprotein coding sequences were analyzed for genetic variability and suitability of all individual genes for viral phylogenies. Our results revealed that, among all the viral genes, NS5a coding sequences were proved to be the most suitable alternative for tracing APPV strains supported by its capability of reproducing the same phylogenetic and evolutionary information as the whole viral genome did. Also, a reliable cutoff to accurately classify APPV at different levels is established. We propose a genotyping scheme with three well-defined genotypes (1-3) and 7 subgenotypes for genotype 1 (1.1-1.7). For whole genome analysis, a threshold value of 84%-91% pairwise identity allows separation of all APPV subgenotypes, whereas 80% identity clearly segregate the three major APPV genotypes. For NS5a gene analysis, 82%-91% identity allows subgenotype separation and 76% identity segregate APPV genotypes. Additionally, genetic distance of whole genome exhibits ≤8% in isolate level, 9%-14% in subgenotype level, and 17%-22% in genotype level, while for NS5a encoding sequences the genetic distance displays ≤9% in isolate level, 9.9%-19.1% in subgenotype level, and 21.6%-29.7% in genotype level. These allow a clear segregation among APPV genotypes, subgenotypes, and isolates. Therefore, the proposed strategy in this study provides a solid and improved basis for molecular phylogenetics to understand APPV genetic diversity, trace the origins and control the spread of new disease outbreaks.

Deletion of the RGD motif in LON-2/glypican is associated with morphological abnormalities.

The lon-2 gene in Caenorhabditis elegans encodes a heparan sulfate proteoglycan family glypican that negatively regulates the BMP signaling pathway responsible for controlling body length. LON-2 contains multiple functional domains, including an RGD (Arg-Gly-Asp) motif at amino acid number from 348 to 350. A novel mutant allele of lon-2 was investigated in this study. In this mutant allele, lon-2(kq348ΔRGD), the RGD motif at position 348 was deleted. Another pre-existing mutant allele, lon-2(e678), contains a ~9kb deletion and lacks most of the genomic coding sequence. The lon-2(e678) line was used as a reference allele. The novel mutant line was significantly shorter than wild-type animals, suggesting that removal of the RGD motif in LON-2 may improve its ability to inhibit BMP signaling.

Natural infection of parvovirus in wild fishing cats (Prionailurus viverrinus) reveals extant viral localization in kidneys.

Carnivore protoparvovirus-1 (CPPV-1), a viral species containing feline panleukopenia virus (FPV) and canine parvovirus (CPV) variants, are widely spread among domestic and wild carnivores causing systemic fatal diseases. Wild fishing cats (Prionailurus viverrinus), a globally vulnerable species, have been found dead. Postmortem examination of the carcasses revealed lesions in intestine, spleen and kidney. CPPV-1 antigen identification in these tissues, using polymerase chain reaction (PCR) and immunohistochemistry (IHC), supported the infection by the virus. PCR- and IHC-positivity in kidney tissues revealed atypical localization of the virus while in situ hybridization (ISH) and transmission electron microscopy (TEM) with the pop-off technique confirmed the first description of viral localization in kidneys. Complete genome characterization and deduced amino acid analysis of the obtained CPPV-1 from the fishing cats revealed FPV as a causative agent. The detected FPV sequences showed amino acid mutations at I566M and M569R in the capsid protein. Phylogenetic and evolutionary analyses of complete coding genome sequences revealed that the fishing cat CPPV-1 genomes are genetically clustered to the FPV genomes isolated from domestic cats in Thailand. Since the 1970s, these genomes have also been shown to share a genetic evolution with Chinese FPV strains. This study is the first evidence of CPPV-1 infection in fishing cats and it is the first to show its localization in the kidneys. These findings support the multi-host range of this parvovirus and suggest fatal CPPV-1 infections may result in other vulnerable wild carnivores.

Sediminibacterium soli sp. nov., isolated from soil.

A Gram-stain-negative, facultative anaerobic strain, designated WSJ-3T, was isolated from soil. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain WSJ-3T belongs to genus Sediminibacterium and exhibits the highest sequence similarities to Sediminibacterium roseum SYL130T (97.0%), Sediminibacterium goheungense DSM 28323T (96.9%), Sediminibacterium aquarii AA5T (96.7%), and Sediminibacterium salmoneum NBRC 103935T (95.2%). The average nucleotide identity values of strain WSJ-3T/S. roseum SYL130T and strain WSJ-3T/S. goheungense DSM 28323T are 72.2% and 70.4%, respectively, and digital DNA-DNA hybridization values for these are 19.2% and 19.1%, respectively. Strain WSJ-3T has a genome size of 3.88Mb, with a DNA G + C content of 50.1mol% and comprises of 3263 predicted genes. A phylogenetic tree constructed using the genomic core protein coding sequences revealed that strain WSJ-3T clusters with S. roseum SYL130T. Strain WSJ-3T has menaquinone-7 as the only respiratory quinone and phosphatidylethanolamine, three unidentified phospholipids, four unidentified aminophospholipids, two unidentified aminolipids, and three unidentified lipids as the polar lipids. The major fatty acids of strain WSJ-3T are iso-C15:0, iso-C17:0 3-OH, and iso-C15:1 G. On the basis of the polyphasic results, the isolate represents a novel species of the genus Sediminibacterium, for which the name Sediminibacterium soli sp. nov. is proposed. The type strain is WSJ-3T (= KCTC 72839T = CCTCC AB 2019408T).

Evolution of Transmissible Gastroenteritis Virus (TGEV): A Codon Usage Perspective.

Transmissible gastroenteritis virus (TGEV) is a coronavirus associated with diarrhea and high mortality in piglets. To gain insight into the evolution and adaptation of TGEV, a comprehensive analysis of phylogeny and codon usage bias was performed. The phylogenetic analyses of maximum likelihood and Bayesian inference displayed two distinct genotypes: genotypes I and II, and genotype I was classified into subtypes Ia and Ib. The compositional properties revealed that the coding sequence contained a higher number of A/U nucleotides than G/C nucleotides, and that the synonymous codon third position was A/U-enriched. The principal component analysis based on the values of relative synonymous codon usage (RSCU) showed the genotype-specific codon usage patterns. The effective number of codons (ENC) indicated moderate codon usage bias in the TGEV genome. Dinucleotide analysis showed that CpA and UpG were over-represented and CpG was under-represented in the coding sequence of the TGEV genome. The analyses of Parity Rule 2 plot, ENC-plot, and neutrality plot displayed that natural selection was the dominant evolutionary driving force in shaping codon usage preference in genotypes Ia and II. In addition, natural selection played a major role, while mutation pressure had a minor role in driving the codon usage bias in genotype Ib. The codon adaptation index (CAI), relative codon deoptimization index (RCDI), and similarity index (SiD) analyses suggested that genotype I might be more adaptive to pigs than genotype II. Current findings contribute to understanding the evolution and adaptation of TGEV.

Prediction, isolation, overexpression and antifungal activity analysis of Medicago truncatula var. truncatula putative thaumatin like proteins (TLP-1, -2, -3, -4 and -5).

Pathogenesis-related proteins (PR-proteins) are induced in response to environmental stresses such as osmotic and drought stress, wounding, microbial infections and treatment with specific plant hormones and elicitors. These proteins are classified into several groups (PR-1 through PR-17) based on their amino acid sequence and biochemical functions. The present study focuses on prediction, isolation, over-expression and analysis of the antifungal activities of the thaumatin-like proteins (i.e. PR-5) in the model legume M. truncatula var. truncatula. Analysis of M. truncatula genome sequence, available freely on the NCBI website, indicated the presence of at least 15 PR-5 Open Reading Frames (ORFs), 5 of them (dubbed TLP-1, -2, -3, -4 and -5) were selected for this study. Using gene-specific primers, the genomic coding sequences were isolated, sequenced and all confirmed to match with those reported in the database. All the fragments were, then, cloned in Escherichia coli isolate BL21 (DE3), using pET-21c(+) plasmids for subsequent overexpression (overexpression). All 5 genes were expressed as inclusion bodies (IBs) with masses, estimated by SDS PAGE, corresponding to the theoretical values. As expected, none of the protein IBs had no detectable effect on the phytopathogenic fungi Rhizoctonia solani, Alternaria alternata, Fusarium graminearum, Fusarium solani, Verticillium sp. and Phytophtora spp. However, when the in vitro refolded IB preparations were applied, all displayed comparable strong antifungal activities against the tested fungi. The current study is the first report of overexpression and evaluation of antifungal activities of PR-5 family of proteins from M. truncatula Var. truncatula, and provides experimental evidence that all investigated proteins have the potential for enhancing resistance against some important fungal pathogens.

Complete Coding Genome Sequence of an Avian Influenza A/H3N8 Virus Strain Detected in North Kazakhstan in 2018.

We report the complete coding genome sequence of the influenza A/H3N8 virus, isolated from Anas querquedula in northern Kazakhstan in 2018. Phylogenetic analysis of the surface antigens of strain A/garganey/North-Kazakhstan/45/2018 showed that its hemagglutinin belonged to the Asian line, while its neuraminidase was assigned to the Eurasian group.