The aging process, or senescence, is characterized by age-specific decline in physical and physiological function, and increased frailty and genomic changes, including mutation accumulation. However, the mechanisms through which changes in genomic architecture influence human longevity have remained obscure. Copy number variants (CNVs), an abundant class of genomic variants, offer unique opportunities for understanding age-related genomic changes. Here we report the spectrum of CNVs in a cohort of 670 Ashkenazi Jewish centenarians, their progeny, and unrelated controls. The average ages of these groups were 97.4 ± 2.8, 69.2 ± 9.2, and 66.5 ± 7.0 respectively. For the first time, we compared different size classes of CNVs, from 1 kB to 100 MB in size. Using a high-resolution custom Affymetrix array, targeting 44,639 genomic regions, we identified a total of 12,166, 22,188, and 10,285 CNVs in centenarians, their progeny, and control groups, respectively. Interestingly, the offspring group showed the highest number of unique CNVs, followed by control and centenarians. While both gains and losses were found in all three groups, centenarians showed a significantly higher average number of both total gains and losses relative to their controls (p < 0.0327, 0.0182, respectively). Moreover, centenarians showed a lower total length of genomic material lost, suggesting that they may maintain superior genomic integrity over time. We also observe a significance fold increase of CNVs among the offspring, implying greater genomic integrity and a putative mechanism for longevity preservation. Genomic regions that experienced loss or gains appear to be distributed across many sites in the genome and contain genes involved in DNA transcription, cellular transport, developmental pathways, and metabolic functions. Our findings suggest that the exceptional longevity observed in centenarians may be attributed to the prolonged maintenance of functionally important genes. These genes are intrinsic to specific genomic regions as well as to the overall integrity of the genomic architecture. Additionally, a strong association between longer CNVs and differential gene expression observed in this study supports the notion that genomic integrity could positively influence longevity.

The sheer number of gene variants and the extent of the observed clinical and molecular heterogeneity recorded in neuropsychiatric disorders (NPDs) could be due to the magnified downstream effects initiated by a smaller group of genomic higher-order alterations in response to endogenous or environmental stress. Chromosomal common fragile sites (CFS) are functionally linked with microRNAs, gene copy number variants (CNVs), sub-microscopic deletions and duplications of DNA, rare single-nucleotide variants (SNVs/SNPs), and small insertions/deletions (indels), as well as chromosomal translocations, gene duplications, altered methylation, microRNA and L1 transposon activity, and 3-D chromosomal topology characteristics. These genomic structural features have been linked with various NPDs in mostly isolated reports and have usually only been viewed as areas harboring potential candidate genes of interest. The suggestion to use a higher level entry point (the \'fragilome\' and associated features) activated by a central mechanism (\'stress\') for studying NPD genetics has the potential to unify the existing vast number of different observations in this field. This approach may explain the continuum of gene findings distributed between affected and unaffected individuals, the clustering of NPD phenotypes and overlapping comorbidities, the extensive clinical and molecular heterogeneity, and the association with certain other medical disorders.

Mycoplasma felis has been isolated from diseased cats and horses, but to date only a single fully assembled genome of this species, of an isolate from a horse, has been characterized. This study aimed to characterize and compare the completely assembled genomes of four clinical isolates of M. felis from three domestic cats, assembled with the aid of short- and long-read sequencing methods. The completed genomes encoded a median of 759 ORFs (range 743-777) and had a median average nucleotide identity of 98.2 % with the genome of the available equid origin reference strain. Comparative genomic analysis revealed the occurrence of multiple horizontal gene transfer events and significant genome reassortment. This had resulted in the acquisition or loss of numerous genes within the Australian felid isolate genomes, encoding putative proteins involved in DNA transfer, metabolism, DNA replication, host cell interaction and restriction modification systems. Additionally, a novel mycoplasma phage was detected in one Australian felid M. felis isolate by genomic analysis and visualized using cryo-transmission electron microscopy. This study has highlighted the complex genomic dynamics in different host environments. Furthermore, the sequences obtained in this work will enable the development of new diagnostic tools, and identification of future infection control and treatment options for the respiratory disease complex in cats.

In eukaryotic genomes, rDNA generally resides as a highly repetitive and dynamic structure, making it difficult to study. Here, a synthetic rDNA array on chromosome III in budding yeast was constructed to serve as the sole source of rRNA. Utilizing the loxPsym site within each rDNA repeat and the Cre recombinase, we were able to reduce the copy number to as few as eight copies. Additionally, we constructed strains with two or three rDNA arrays and found that the presence of multiple arrays did not affect the formation of a single nucleolus. Although alteration of the position and number of rDNA arrays did impact the three-dimensional genome structure, the additional rDNA arrays had no deleterious influence on cell growth or transcriptomes. Overall, this study sheds light on the high plasticity of rDNA organization and opens up opportunities for future rDNA engineering.

A key step for comparative genomics is to group open reading frames into functionally and evolutionarily meaningful gene clusters. Gene clustering is complicated by intraspecific duplications and horizontal gene transfers that are frequent in prokaryotes. In consequence, gene clustering methods must deal with a trade-off between identifying vertically transmitted representatives of multicopy gene families, which are recognizable by synteny conservation, and retrieving complete sets of species-level orthologs. We studied the implications of adopting homology, orthology, or synteny conservation as formal criteria for gene clustering by performing comparative analyses of 125 prokaryotic pangenomes.
Clustering criteria affect pangenome functional characterization, core genome inference, and reconstruction of ancestral gene content to different extents. Species-wise estimates of pangenome and core genome sizes change by the same factor when using different clustering criteria, allowing robust cross-species comparisons regardless of the clustering criterion. However, cross-species comparisons of genome plasticity and functional profiles are substantially affected by inconsistencies among clustering criteria. Such inconsistencies are driven not only by mobile genetic elements, but also by genes involved in defense, secondary metabolism, and other accessory functions. In some pangenome features, the variability attributed to methodological inconsistencies can even exceed the effect sizes of ecological and phylogenetic variables.
Choosing an appropriate criterion for gene clustering is critical to conduct unbiased pangenome analyses. We provide practical guidelines to choose the right method depending on the research goals and the quality of genome assemblies, and a benchmarking dataset to assess the robustness and reproducibility of future comparative studies.

Halophilic archaea (haloarchaea) are known to exhibit multiple chromosomes, with one main chromosome and one or several smaller secondary chromosomes or megaplasmids. Halorubrum lacusprofundi, a model organism for studying cold adaptation, exhibits one secondary chromosome and one megaplasmid that include a large arsenal of virus defense mechanisms. We isolated a virus (Halorubrum tailed virus DL1, HRTV-DL1) infecting Hrr. lacusprofundi, and present an in-depth characterization of the virus and its interactions with Hrr. lacusprofundi. While studying virus-host interactions between Hrr. lacusprofundi and HRTV-DL1, we uncover that the strain in use (ACAM34_UNSW) lost the entire megaplasmid and about 38% of the secondary chromosome. The loss included the majority of virus defense mechanisms, making the strain sensitive to HRTV-DL1 infection, while the type strain (ACAM34_DSMZ) appears to prevent virus replication. Comparing infection of the type strain ACAM34_DSMZ with infection of the laboratory derived strain ACAM34_UNSW allowed us to identify host responses to virus infection that were only activated in ACAM34_UNSW upon the loss of virus defense mechanisms. We identify one of two S-layer proteins as primary receptor for HRTV-DL1 and conclude that the presence of two different S-layer proteins in one strain provides a strong advantage in the arms race with viruses. Additionally, we identify archaeal homologs to eukaryotic proteins potentially being involved in the defense against virus infection.

Fungal species have dynamic genomes and often exhibit genomic plasticity in response to stress. This genome plasticity often comes with phenotypic consequences that affect fitness and resistance to stress. Fungal pathogens exhibit genome plasticity in both clinical and agricultural settings and often during adaptation to antifungal drugs, posing significant challenges to human health. Therefore, it is important to understand the rates, mechanisms, and impact of large genomic changes. This review addresses the prevalence of polyploidy, aneuploidy, and copy number variation across diverse fungal species, with special attention to prominent fungal pathogens and model species. We also explore the relationship between environmental stress and rates of genomic changes and highlight the mechanisms underlying genotypic and phenotypic changes. A comprehensive understanding of these dynamic fungal genomes is needed to identify novel solutions for the increase in antifungal drug resistance.

The Insertion Sequence 711 (IS711) is linked to the Brucella genus. Mapping the genomic distribution of IS711 can help understand this insertion element\'s biological and evolutionary role. This work aimed to delineate the genomic distribution of the IS711 element and to study its association with Brucella evolution. A total of 124 genomes representing 9 Brucella species were searched using BLASTn sequence alignment tool to identify complete and truncated copies of IS711. Based on the genomic context, each IS711 locus was assigned a code using the initial letters of its neighboring genes. Various tools were used to annotate the neighboring genes and determine the shared synteny around orthologous IS711 loci. The tool Islandviewer 4 was used to scan for genomic islands. The Codon Tree method was used to build phylogenetic trees of B. melitensis, B. abortus, and B. suis genomes. The phylogenetic trees of the three species were analyzed, taking into account the genomic distribution patterns of IS711. The result of IS711 frequency analysis showed a relatively conserved number of copies/genome for the different species and for some biovars. The analysis showed that Brucella species with a relatively low IS711 copy number (4-8 copies/genome) are linked to domestic animals as primary hosts and have potential for zoonotic transmission. However, species with a relatively higher copy number (12-30 copies/genome) are less zoonotic and tend to be linked with wild animals as primary hosts. Analyzing the genomic distribution map of IS711 loci showed several unique patterns of IS711 distribution that are correlated with the evolution of Brucella species and biovars. The results also showed that 46.2% of the conserved IS711 elements are located within genomic islands. Based on our results and previous data, we postulate a model explaining the IS711 role in Brucella evolution. We assume that during the transition from a free-living to an intracellular lifestyle, a descendant of the Brucella genus had acquired a progenitor sequence of the IS711. Subsequently, a burst in IS711 transposition occurred. This parasitic expansion can be deleterious and has to be counteracted by evolutionary forces to prevent lineage extension and to promote adaptation to host. Similar to other plasmid-free pathogenic α-Proteobacteria bacteria, the balance of expansion and reduction of insertion elements could be one of the mechanisms to control genome reduction and streamlining. We hypothesize that the IS711-mediated genomic changes and other small sequence nucleotide changes in specific orthologous genes could significantly contribute to Brucella\'s evolution and adaptation to different animal hosts.

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Deinococcus radiopugnans DY59 (formerly Deinococcus swuensis DY59) is a radiation-resistant bacterium isolated from soil. From the 3.5 Mb genomic DNA sequence of strain DY59 (December 2014), 31 insertion sequence (IS) elements of six IS families including IS1, IS4, IS5, IS66, IS630, and IS701 and five unclassified IS elements were detected. Upon induction of oxidative stress with 80 and 100 mM H2O2, the unique ISs of the IS4 family member were actively translocated into a carotenoid biosynthesis gene phytoene desaturase (QR90_10400), resulting in non-pigment phenotypic selection. Therefore, these active transpositions of a specific IS family member were induced by oxidative stress at 80 and 100 mM H2O2. Furthermore, D. radiopugnans DY59 exhibited extremely higher MIC values against H2O2 treatment. To explain this phenomenon, qRT-PCR was conducted to assess the expression levels of catalase and three LysR family regulators. Our findings indicated that the ISDrpg2 and ISDrpg3 elements of the IS4 family were actively transposed into the phytoene desaturase gene by H2O2 treatment via replicative transposition. However, high H2O2 resistance did not originate from H2O2-induced expression of catalase and LysR family regulators.