Gram staining has guided microbiology for over a century by coloring cells purple or pink, a read-out thought to distinguish monoderms (single membrane, thick peptidoglycan) from diderms (inner and outer membranes with thin wall). Here we show this rule fails repeatedly across Bacillaceae lineages historically deemed "Gram-positive". By combining light and transmission-electron microscopy, antibiotic-sensitivity assays and comparative genomics across 57 strains, we identify "Gram-negative-staining monoderms" lacking outer membrane yet retaining thick peptidoglycan walls. These bacteria lack lipopolysaccharide- and β-barrel assembly-genes and remain highly susceptible to vancomycin and lysozyme (agents normally excluded by diderm envelopes), demonstrating functional monoderm status. Surprisingly, teichoic-acid biosynthetic pathways are patchily distributed and do not predict staining behavior. This discovery calls into question the textbook purple-or-pink dichotomy, decoupling stain color from membrane architecture. Clinically, misidentifying pink-staining Bacillaceae (including emerging pathogens such as Bacillus infantis) risks inappropriate therapy, whereas genome-guided diagnostics enable precise antibiotic stewardship.

This study employed genomic analysis to elucidate the hybrid origin of Rosa del Peru type 2, a significant patrimonial grapevine in the Americas. Twelve accessions from a historical vineyard in Ensenada, Mexico, were characterized using simple sequence repeats (SSR), with three representative samples—Listan Prieto and two Rosa del Peru type 2 accessions—selected for whole-genome resequencing (WGS). SSR profiling revealed a high prevalence of Listan Prieto within the historical vineyard. Neighbor-net analysis, based on 109,529 high-quality single nucleotide polymorphisms (SNPs), clustered Listan Prieto and Rosa del Peru type 2 with Listan Negro and Manai, a Muscat cultivar of Chinese origin. ADMIXTURE analysis (k = 9, 64,175 SNPs) demonstrated that while Listan Prieto shares its genetic profile with Listan Negro, Rosa del Peru type 2 exhibits a hybrid composition derived from Listan Prieto and Manai. Subsequent PhyloNet modeling further supported that Rosa del Peru type 2 originated from a hybridization event between the lineages of Listan Prieto and Manai. Notably, we identified 7,585 SNPs in these patrimonial cultivars not present in their immediate ancestors Listan Negro and Manai, suggesting potential local adaptation, clonal divergence or incomplete lineage sorting. By providing the first WGS datasets for these cultivars, this work offers a high-resolution genomic resource and clarifies the complex historical migration and diversification of New World grapevines.

Understanding the organization and evolution of metabolic networks is essential for uncovering how organisms adapt to changing environments. Whereas free-living bacteria typically maintain robust and redundant metabolic systems, endosymbiotic bacteria undergo extreme genome reduction during their adaptation to intracellular life. This process results in highly streamlined and interconnected metabolic networks, in some cases smaller than the theoretical minimum required for sustaining independent cellular function. Using a large-scale comparative framework, we analyzed 101 genomes of insect endosymbiotic bacteria by computing two metabolic network models: metabolite- and reaction-based. We found strong correlations between genome size and key topological properties, including clustering coefficient, network diameter, and number of nodes, indicating that genome reduction directly constrains metabolic network architecture. Despite extensive gene loss, endosymbiotic metabolic networks retain scale-free organization, suggesting the preservation of essential connectivity and robustness. Furthermore, clustering analyses revealed that network topology reflects phylogenetic relationships across bacterial taxa, demonstrating that metabolic organization retains evolutionary signals even in the most reduced genomes. Our findings show that the metabolic networks of insect endosymbiotic bacteria preserve clear evolutionary imprints, revealing a deep connection between genomic reduction, network structure, and phylogenetic history. The complementary use of metabolite- and reaction-based models provide a powerful framework for exploring how symbiotic evolution reshapes metabolic systems while maintaining essential biological organization.

In late 2019, a new virus, SARS-CoV-2, emerged in Wuhan, China, causing COVID-19 and the subsequent global pandemic. As of 30 April 2023, more than 774 million cases of COVID-19 had been reported worldwide, including over 7.5 million in Mexico. Despite advances in vaccination, epidemic surges of COVID-19 continued to occur globally, highlighting the importance of sharing and disseminating the experiences gained during these first years to better understand the virus’s evolution and respond accordingly. For this reason, the National Council for Science and Technology (CONACYT) organized the meeting “Challenges and Opportunities for Genomic Surveillance of SARS-CoV-2 in Mexico” from 15 to 17 August 2022, to present the efforts and results accumulated over more than two years of the pandemic. In this meeting report, we summarize the key findings of each participant and provide their contact information.

Hanseniaspora species stand out among yeasts for having the smallest genomes, marked by extensive gene family contractions. Consequently, they have been proposed as model organisms for studying the evolution of free-living cells lacking genes otherwise considered essential. Here, we show that Hanseniaspora yeasts are prevalent in agave fermentations used to produce traditional spirits across Mexico. We sequenced the genomes of 15 strains, unambiguously identifying them as H. lachancei, H. pseudoguilliermondii, H. guilliermondii , and H. opuntiae . Comparative genomic analyses revealed dynamic shifts in gene family sizes and compositions, suggesting ongoing gene loss within the fast-evolving branch of this genus. Notably, gene losses varied across functional categories, even among isolates of the same species. Growth assays across diverse stress conditions and carbon sources indicated that these genomic disparities do not directly translate into phenotypic differences. Together, our findings suggest that differential gene loss within species is an active evolutionary process shaping Hanseniaspora genomes, highlighting agave-fermentation populations as valuable models for studying genome reduction in a natural context. ### Competing Interest Statement The authors have declared no competing interest. Secretaría de Ciencia, Humanidades, Tecnología e Innovación del Gobierno de México (SECIHTI), FORDECYT-PRONACES/103000/2020, CF-2023-G-695, CBF-2025-G-838, 4133922, I0200/111/2024 UNAM-PAPIIT, IN212524, UK BBSRC under the Global Challenges Research Fund (GCRF) Growing Research Capability call through the CABANA Innovation Fund, BB/P027849/1

The ultimate consequence of Darwin’s theory of common descent implies that all life on earth descends ultimately from a common ancestor. Biochemistry and molecular biology now provide sufficient evidence of shared ancestry of all extant life forms. However, the nature of the Last Universal Common Ancestor (LUCA) has been a topic of much debate over the years. This review offers a historical perspective on different attempts to infer LUCA’s nature, exploring the debate surrounding its complexity. We further examine how different methodologies identify sets of ancient protein that exhibit only partial overlap. For example, different bioinformatic approaches have identified distinct protein subunits from the ATP synthetase identified as potentially inherited from LUCA. Additionally, we discuss how detailed molecular evolutionary analysis of reverse gyrase has modified previous inferences about an hyperthermophilic LUCA based mainly on automatic bioinformatic pipelines. We conclude by emphasizing the importance of developing a database dedicated to studying genes and proteins traceable back to LUCA and earlier stages of cellular evolution. Such a database would house the most ancient genes on earth.

Societal Impact StatementThe cultural significance of the grapevine is undeniable. However, we fail to acknowledge how the grapevine has and continues to influence the most pressing political questions of our time. From the beginning of the Conquest, Indigenous peoples were forced to plant the vine, Spain burned the vines Miguel Hidalgo used to teach the poor, and César Chávez and the Delano grape strike demanded justice for agricultural laborers. From the Grito de Dolores to Sí se puede, we demonstrate how the continuing relationship between Mexico and the grapevine influences debates surrounding labor, immigration, and human rights in the United States and throughout the world. To enhance the reach of this work, a Spanish language version of the paper is available in the Supporting Information (see Translation_ES).SummaryThe wild grapevine species (Vitis spp.) that comprise the pedigrees of rootstocks, the Americas as the source (and solution) to the Phylloxera crisis that decimated European vineyards, and California as a premier wine‐growing region are the topics that usually frame the history of grapes in North America. This Anglo‐American perspective ignores that domesticated grape varieties were first introduced to North America in what is now Mexico and the singular contributions of Mexican labor to the California wine economy that continue to influence politics. Here, we highlight the neglected history of grapevines in Mexico and argue that the politics of labor that played out during the Conquest never ceased and still shape debates surrounding immigration. Beginning with Hernán Cortés, Indigenous peoples were forced to plant grapevines and when they were successful, they were abruptly forbidden by Spain to grow grapes. This interference influenced Miguel Hidalgo, who taught the poor viticulture as a trade and who would lead the Mexican War of Independence and pay with his life. The grapevine continued its journey north to California, where Franciscans established the missions and cultivated the Mission grapes, which had lasting impacts on the genetics of grapevine varieties. Finally, it was the Delano grape strike that coalesced César Chávez and the United Farm Workers to demand justice for agricultural laborers that is the foundation of the California wine economy and still shapes the current political debate of immigration, labor, and human rights between the United States and Mexico.

The spike protein determines the host-range specificity of coronaviruses. In particular, the Receptor-Binding Motif in the spike protein from SARS-CoV-2 contains the amino acids involved in molecular recognition of the host Angiotensin Converting Enzyme 2. Therefore, to understand how SARS-CoV-2 acquired its capacity to infect humans it is necessary to reconstruct the evolution of this important motif. Early during the pandemic, it was proposed that the SARS-CoV-2 Receptor-Binding Domain was acquired via recombination with a pangolin infecting coronavirus. This proposal was challenged by an alternative explanation that suggested that the Receptor-Binding Domain from SARS-CoV-2 did not originated via recombination with a coronavirus from a pangolin. Instead, this alternative hypothesis proposed that the Receptor-Binding Motif from the bat coronavirus RaTG13, was acquired via recombination with an unidentified coronavirus. And as a consequence of this event, the Receptor-Binding Domain from the pangolin coronavirus appeared as phylogenetically closer to SARS-CoV-2. Recently, the genomes from coronaviruses from Cambodia (bat_RShST182/200) and Laos (BANAL-20-52/103/247) which are closely related to SARS-CoV-2 were reported. However, no detailed analysis of the evolution of the Receptor-Binding Motif from these coronaviruses was reported. Here we revisit the evolution of the Receptor-Binding Domain and Motif in the light of the novel coronavirus genome sequences. Specifically, we wanted to test whether the above coronaviruses from Cambodia and Laos were the source of the Receptor-Binding Domain from RaTG13. We found that the Receptor-Binding Motif from these coronaviruses is phylogenetically closer to SARS-CoV-2 than to RaTG13. Therefore, the source of the Receptor-Binding Domain from RaTG13 is still unidentified. In accordance with previous studies, our results are consistent with the hypothesis that the Receptor-Binding Motif from SARS-CoV-2 evolved by vertical inheritance from a bat-infecting population of coronaviruses.

The coronavirus SARS-CoV-2 is the most sequenced pathogen ever, with several million genome copies deposited in the GISAID database. This large amount of genomic information poses non-trivial bioinformatic challenges for those interested in studying the evolution of SARS-CoV-2. One common problem when studying the phylogeny of the coronavirus in its geographical context is to count with accurate information of the location of the samples. However, this information is filled by hand by research groups all over the world and sometimes typos and inconsistencies are introduced in the metadata when submitting the sequences to GISAID. Correcting these errors is laborious and time-consuming. Here, we provide a suite of Perl scripts designated to facilitate the curation of this vital information and perform a random sampling of genome sequences if necessary. The scripts provided here can be used to curate geographic information in the metadata and sample the sequences from any country of interest to ease the preparation of files for Nextstrain and Microreact, thus accelerating evolutionary studies of this important pathogen. CurSa scripts are accessible via: https://github.com/luisdelaye/CurSa/.

Yeasts are a diverse group of fungal microorganisms that are widely used to produce fermented foods and beverages. In Mexico, open fermentations are used to obtain spirits from agave plants. Despite the prevalence of this traditional practice throughout the country, yeasts have only been isolated and studied from a limited number of distilleries. To systematically describe the diversity of yeast species from open agave fermentations, here we generate the YMX-1.0 culture collection by isolating 4524 strains from 68 sites with diverse climatic, geographical, and biological contexts. We used MALDI-TOF mass spectrometry for taxonomic classification and validated a subset of the strains by ITS and D1/D2 sequencing, which also revealed two potential novel species of Saccharomycetales. Overall, the composition of yeast communities was weakly associated with local variables and types of climate, yet a core set of six species was consistently isolated from most producing regions. To explore the intraspecific variation of the yeasts from agave fermentations, we sequenced the genomes of four isolates of the nonconventional yeast Kazachstania humilis. The genomes of these four strains were substantially distinct from a European isolate of the same species, suggesting that they may belong to different populations. Our work contributes to the understanding and conservation of an open fermentation system of great cultural and economic importance, providing a valuable resource to study the biology and genetic diversity of microorganisms living at the interface of natural and human-associated environments.

Over 200 different SARS-CoV-2 lineages have been observed in Mexico by November 2021. To investigate lineage replacement dynamics, we applied a phylodynamic approach and explored the evolutionary trajectories of five dominant lineages that circulated during the first year of local transmission. For most lineages, peaks in sampling frequencies coincided with different epidemiological waves of infection in Mexico. Lineages B.1.1.222 and B.1.1.519 exhibited similar dynamics, constituting clades that likely originated in Mexico and persisted for >12 months. Lineages B.1.1.7, P.1 and B.1.617.2 also displayed similar dynamics, characterized by multiple introduction events leading to a few successful extended local transmission chains that persisted for several months. For the largest B.1.617.2 clades, we further explored viral lineage movements across Mexico. Many clades were located within the south region of the country, suggesting that this area played a key role in the spread of SARS-CoV-2 in Mexico.

Although recombination is a feature of coronavirus evolution, previously detected recombinant lineages of SARS-CoV-2 have shown limited circulation thus far. Here, we present a detailed phylogenetic analysis of four SARS-CoV-2 lineages to investigate the possibility of virus recombination among them. Our analyses reveal well-supported phylogenetic differences between the Orf1ab region encoding viral non-structural proteins and the rest of the genome, including Spike (S) protein and remaining reading frames. By accounting for several deletions in NSP6, Orf3a, and S, we conclude that the B.1.628 major cluster, now designated as lineage XB, originated from a recombination event between viruses of B.1.631 and B.1.634 lineages. This scenario is supported by the spatiotemporal distribution of these lineages across the USA and Mexico during 2021, suggesting that the recombination event originated in this geographical region. This event raises important questions regarding the role and potential effects of recombination on SARS-CoV-2 evolution.

The ascomycetous yeast Kazachstania humilis is an active species in backslopped sourdough and in the spontaneous fermentation of several traditional foods and beverages. Here, we report the draft genome sequence of a K. humilis strain isolated from agave must from a traditional distillery in Mexico.

Codon usage is the outcome of different evolutionary processes and can inform us about the conditions in which organisms live and evolve. Here, we present R_ENC’, which is an improvement to the original S index developed by dos Reis et al. (2004). Our index is less sensitive to G+C content, which greatly affects synonymous codon usage in prokaryotes, making it better suited to detect selection acting on codon usage. We used R_ENC’ to estimate the extent of selected codon usage bias in 1800 genomes representing 26 prokaryotic phyla. We found that Gammaproteobacteria, Betaproteobacteria, Actinobacteria, and Firmicutes are the phyla/subphyla showing more genomes with selected codon usage bias. In particular, we found that several lineages within Gammaproteobacteria and Firmicutes show a similar set of functional terms enriched in genes under selected codon usage bias, indicating convergent evolution. We also show that selected codon usage bias tends to evolve in genes coding for the translation machinery before other functional GO terms. Finally, we discuss the possibility to use R_ENC’ to predict whether lineages evolved in copiotrophic or oligotrophic environments.

Evolution has long been considered to be a conservative process in which new genes arise from pre-existing genes through gene duplication, domain shuffling, horizontal transfer, overprinting, retrotransposition, etc. However, this view is changing as new genes originating from non-genic sequences are discovered in different organisms. Still, rather limited functional information is available. Here, we have identified TWISTED1 (TWT1), a possible de novo-originated protein-coding gene that modifies microtubule arrangement and causes helicoidal growth in Arabidopsis thaliana when its expression is increased. Interestingly, even though TWT1 is a likely recent gene, the lack of TWT1 function affects A. thaliana development. TWT1 seems to have originated from a non-genic sequence. If so, it would be one of the few examples to date of how during evolution de novo genes are integrated into developmental cellular and organismal processes.

SARS-CoV-2 variants surveillance is a worldwide task that has been approached with techniques such as Next Generation Sequencing (NGS); however, this technology is not widely available in developing countries because of the lack of equipment and limited funding in science. An option is to deploy a RT-qPCR screening test which aids in the analysis of a higher number of samples, in a shorter time and at a lower cost. In this study, variants present in samples positive for SARS-CoV-2 were identified with a RT-qPCR mutation screening kit and were later confirmed by NGS. A sample with an abnormal result was found with the screening test, suggesting the simultaneous presence of two viral populations with different mutations. The DRAGEN Lineage analysis identified the Delta variant, but there was no information about the other three mutations previously detected. When the sequenced data was deeply analyzed, there were reads with differential mutation patterns, that could be identified and classified in terms of relative abundance, whereas only the dominant population was reported by DRAGEN software. Since most of the software developed to analyze SARS-CoV-2 sequences was aimed at obtaining the consensus sequence quickly, the information about viral populations within a sample is scarce. Here, we present a faster and deeper SARS-CoV-2 surveillance method, from RT-qPCR screening to NGS analysis.

During the coronavirus disease 2019 (COVID-19) pandemic, the emergence and rapid increase of the B.1.1.7 (Alpha) lineage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), first identified in the United Kingdom in September 2020, was well documented in different areas of the world and became a global public health concern because of its increased transmissibility. The B.1.1.7 lineage was first detected in Mexico during December 2020, showing a slow progressive increase in its circulation frequency, which reached its maximum in May 2021 but never became predominant. In this work, we analyzed the patterns of diversity and distribution of this lineage in Mexico using phylogenetic and haplotype network analyses. Despite the reported increase in transmissibility of the B.1.1.7 lineage, in most Mexican states, it did not displace cocirculating lineages, such as B.1.1.519, which dominated the country from February to May 2021. Our results show that the states with the highest prevalence of B.1.1.7 were those at the Mexico-U.S. border. An apparent pattern of dispersion of this lineage from the northern states of Mexico toward the center or the southeast was observed in the largest transmission chains, indicating possible independent introduction events from the United States. However, other entry points cannot be excluded, as shown by multiple introduction events. Local transmission led to a few successful haplotypes with a localized distribution and specific mutations indicating sustained community transmission. IMPORTANCE The emergence and rapid increase of the B.1.1.7 (Alpha) lineage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) throughout the world were due to its increased transmissibility. However, it did not displace cocirculating lineages in most of Mexico, particularly B.1.1.519, which dominated the country from February to May 2021. In this work, we analyzed the distribution of B.1.1.7 in Mexico using phylogenetic and haplotype network analyses. Our results show that the states with the highest prevalence of B.1.1.7 (around 30%) were those at the Mexico-U.S. border, which also exhibited the highest lineage diversity, indicating possible introduction events from the United States. Also, several haplotypes were identified with a localized distribution and specific mutations, indicating that sustained community transmission occurred in the country.

Understanding the evolution of the SARS-CoV-2 virus in various regions of the world during the Covid-19 pandemic is essential to help mitigate the effects of this devastating disease. We describe the phylogenomic and population genetic patterns of the virus in Mexico during the pre-vaccination stage, including asymptomatic carriers. A real-time quantitative PCR screening and phylogenomic reconstructions directed at sequence/structure analysis of the spike glycoprotein revealed mutation of concern E484K in genomes from central Mexico, in addition to the nationwide prevalence of the imported variant 20C/S:452R (B.1.427/9). Overall, the detected variants in Mexico show spike protein mutations in the N-terminal domain (i.e. R190M), in the receptor-binding motif (i.e. T478K, E484K), within the S1-S2 subdomains (i.e. P681R/H, T732A), and at the basis of the protein, V1176F, raising concerns about the lack of phenotypic and clinical data available for the variants of interest we postulate: 20B/478K.V1 (B.1.1.222 or B.1.1.519) and 20B/P.4 (B.1.1.28.4). Moreover, the population patterns of single nucleotide variants from symptomatic and asymptomatic carriers obtained with a self-sampling scheme confirmed the presence of several fixed variants, and differences in allelic frequencies among localities. We identified the mutation N:S194L of the nucleocapsid protein associated with symptomatic patients. Phylogenetically, this mutation is frequent in Mexican sub-clades. Our results highlight the dual and complementary role of spike and nucleocapsid proteins in adaptive evolution of SARS-CoV-2 to their hosts and provide a baseline for specific follow-up of mutations of concern during the vaccination stage.

The American cranberry ( Vaccinium macrocarpon Ait.) is an iconic North American fruit crop of great cultural and economic importance. Cranberry can be considered a fruit crop model due to its unique fruit nutrient composition, overlapping generations, recent domestication, both sexual and asexual reproduction modes, and the existence of cross-compatible wild species. Development of cranberry molecular resources started very recently; however, further genetic studies are now being limited by the lack of a high-quality genome assembly. Here, we report the first chromosome-scale genome assembly of cranberry, cultivar Stevens, and a draft genome of its close wild relative species Vaccinium microcarpum . More than 92% of the estimated cranberry genome size (492 Mb) was assembled into 12 chromosomes, which enabled gene model prediction and chromosome-level comparative genomics. Our analysis revealed two polyploidization events, the ancient γ-triplication, and a more recent whole genome duplication shared with other members of the Ericaeae, Theaceae and Actinidiaceae families approximately 61 Mya. Furthermore, comparative genomics within the Vaccinium genus suggested cranberry- V. microcarpum divergence occurred 4.5 Mya, following their divergence from blueberry 10.4 Mya, which agrees with morphological differences between these species and previously identified duplication events. Finally, we identified a cluster of subgroup-6 R2R3 MYB transcription factors within a genomic region spanning a large QTL for anthocyanin variation in cranberry fruit. Phylogenetic analysis suggested these genes likely act as anthocyanin biosynthesis regulators in cranberry. Undoubtedly, these new cranberry genomic resources will facilitate the dissection of the genetic mechanisms governing agronomic traits and further breeding efforts at the molecular level.