Genomic evidence of SARS-CoV-2 reinfection cases in southern Brazil.

Cases of reinfection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been reported worldwide. We investigated reinfection cases in a set of more than 30,000 samples, and the SARS-CoV-2 genomes from selected samples from four patients with at least two positive diagnoses with an interval ≥ 45 days between tests were sequenced and analyzed. Comparative genomic and phylogenetic analysis confirmed three reinfection cases and suggested that the fourth one was caused by a virus of the same lineage. Viral sequencing is crucial for understanding the natural course of reinfections and for planning public health strategies for management of COVID-19.

Pangenome inventory of Burkholderia sensu lato, Burkholderia sensu stricto, and the Burkholderia cepacia complex reveals the uniqueness of Burkholderia catarinensis.

Here the pangenome analysis of Burkholderia sensu lato (s.l.) was performed for the first time, together with an updated analysis of the pangenome of Burkholderia sensu stricto, and Burkholderia cepacia complex (Bcc) focusing on the Bcc B. catarinensis specific features of its re-sequenced genome. The pangenome of Burkholderia s.l., Burkholderia s.s., and of the Bcc was open, composed of more than 96% of accessory genes, and more than 62% of unknown genes. Functional annotations showed that secondary metabolism genes belonged to the variable portion of genomes, which might explain their production of several compounds with varied bioactivities. Taken together, this work showed the great variability and uniqueness of these genomes and revealed an underexplored unknown potential in poorly characterized genes. Regarding B. catarinensis 89T, its genome harbors genes related to hydrolases production and plant growth promotion. This draft genome will be valuable for further investigation of its biotechnological potentials.

Genome-based taxonomy of Burkholderia sensu lato: Distinguishing closely related species.

The taxonomy of Burkholderia sensu lato (s.l.) has been revisited using genome-based tools, which have helped differentiate closely related species. Many species from this group are indistinguishable through phenotypic traits and 16S rRNA gene sequence analysis. Furthermore, they also exhibit whole-genome Average Nucleotide Identity (ANI) values in the twilight zone for species circumscription (95-96%), which may impair their correct classification. In this work, we provided an updated Burkholderia s.l. taxonomy focusing on closely related species and give other recommendations for those developing genome-based taxonomy studies. We showed that a combination of ANI and digital DNA-DNA hybridization (dDDH) applying the universal cutoff values of 95% and 70%, respectively, successfully discriminates Burkholderia s.l. species. Using genome metrics with this pragmatic criterion, we demonstrated that i) Paraburkholderia insulsa should be considered a later heterotypic synonym of Paraburkholderia fungorum; ii) Paraburkholderia steynii differs from P. terrae by harboring symbiotic genes; iii) some Paraburkholderia are indeed different species based on dDDH values, albeit sharing ANI values close to 95%; iv) some Burkholderia s.l. indeed represent new species from the genomic viewpoint; iv) some genome sequences should be evaluated with care due to quality concerns.

Genome-based reclassification of Azospirillum brasilense Az39 as the type strain of Azospirillum argentinense sp. nov.

Strain Az39T of Azospirillum is a diazotrophic plant growth-promoting bacterium isolated in 1982 from the roots of wheat plants growing in Marcos Juárez, Córdoba, Argentina. It produces indole-3-acetic acid in the presence of l-tryptophan as a precursor, grows at 20-38 °C (optimal 38 °C), and the cells are curved or spiral-shaped, with diameters ranging from 0.5-0.9 to 1.8-2.2 µm. They contain C16 : 0, C18 : 0 and C18 : 1 ω7c/ω6c as the main fatty acids. Phylogenetic analysis of its 16S rRNA gene sequence confirmed that this strain belongs to the genus Azospirillum, showing a close relationship with Azospirillum baldaniorum Sp245T, Azospirillum brasilense Sp7T and Azospirillum formosense CC-Nfb-7T. Housekeeping gene analysis revealed that Az39T, together with five strains of the genus (Az19, REC3, BR 11975, MTCC4035 and MTCC4036), form a cluster apart from A. baldaniorum Sp245T, A. brasilense Sp7T and A. formosense CC-Nfb-7T. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) between Az39T and the aforementioned type strains revealed values below 96 %, the circumscription limit for the species delineation (ANI: 95.3, 94.1 and 94.0 %; dDDH: 62.9, 56.3 and 55.6 %). Furthermore, a phylogeny evaluation of the core proteome, including 809 common shared proteins, showed an independent grouping of Az39T, Az19, REC3, BR 11975, MTCC4035 and MTCC4036. The G+C content in the genomic DNA of these six strains varied from 68.3 to 68.5 %. Based on the combined phylogenetic, genomic and phenotypic characterization presented here, we consider that strain Az39T, along with strains Az19, REC3, BR 11975, MTCC4035 and MTCC4036, are members of a new Azospirillum species, for which the name Azospirillum argentinense sp. nov. is proposed. The type strain is Az39T (=LBPCV39T=BR 148428T=CCCT 22.01T).

Open-source real-time quantitative RT-PCR-based on a RNA standard for the assessment of SARS-CoV-2 viral load.

BACKGROUND: Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) target genes by molecular methods has been chosen as the main approach to identify individuals with Coronavirus disease 2019 (COVID-19) infection. OBJECTIVES: In this study, we developed an open-source RNA standard-based real-time quantitative RT-PCR (RT-qPCR) assay for quantitative diagnostics of SARS-CoV-2 from nasopharynx, oropharynx, saliva and plasma samples. METHODS AND FINDINGS: We evaluated three SARS-CoV-2 target genes and selected the RNA-dependent RNA polymerase (RdRp) gene, given its better performance. To improve the efficiency of the assay, a primer gradient containing 25 primers forward and reverse concentration combinations was performed. The forward and reverse primer pairs with 400 nM and 500 nM concentrations, respectively, showed the highest sensitivity. The LOD95% was ~60 copies per reaction. From the four biological matrices tested, none of them interfered with the viral load measurement. Comparison with the AllplexTM 2019-nCoV assay (Seegene) demonstrated that our test presents 90% sensitivity and 100% specificity. MAIN CONCLUSIONS: We developed an efficient molecular method able to measure absolute SARS-CoV-2 viral load with high replicability, sensitivity and specificity in different clinical samples.

NAD+ biosynthesis in bacteria is controlled by global carbon/nitrogen levels via PII signaling.

NAD+ is a central metabolite participating in core metabolic redox reactions. The prokaryotic NAD synthetase enzyme NadE catalyzes the last step of NAD+ biosynthesis, converting nicotinic acid adenine dinucleotide (NaAD) to NAD+ Some members of the NadE family use l-glutamine as a nitrogen donor and are named NadEGln Previous gene neighborhood analysis has indicated that the bacterial nadE gene is frequently clustered with the gene encoding the regulatory signal transduction protein PII, suggesting a functional relationship between these proteins in response to the nutritional status and the carbon/nitrogen ratio of the bacterial cell. Here, using affinity chromatography, bioinformatics analyses, NAD synthetase activity, and biolayer interferometry assays, we show that PII and NadEGln physically interact in vitro, that this complex relieves NadEGln negative feedback inhibition by NAD+ This mechanism is conserved in distantly related bacteria. Of note, the PII protein allosteric effector and cellular nitrogen level indicator 2-oxoglutarate (2-OG) inhibited the formation of the PII-NadEGln complex within a physiological range. These results indicate an interplay between the levels of ATP, ADP, 2-OG, PII-sensed glutamine, and NAD+, representing a metabolic hub that may balance the levels of core nitrogen and carbon metabolites. Our findings support the notion that PII proteins act as a dissociable regulatory subunit of NadEGln, thereby enabling the control of NAD+ biosynthesis according to the nutritional status of the bacterial cell.

Enterococci from Wild Magellanic Penguins (Spheniscus magellanicus) as an Indicator of Marine Ecosystem Health and Human Impact.

Enterococci are commensals that proliferated as animals crawled ashore hundreds of millions of years ago. They are also leading causes of multidrug-resistant hospital-acquired infections. While most studies are driven by clinical interest, comparatively little is known about enterococci in the wild or the effect of human activity on them. Pharmaceutical pollution and runoff from other human activities are encroaching widely into natural habitats. To assess their reach into remote habitats, we investigated the identity, genetic relatedness, and presence of specific traits among 172 enterococcal isolates from wild Magellanic penguins. Four enterococcal species, 18 lineage groups, and different colonization patterns were identified. One Enterococcus faecalis lineage, sequence type 475 (ST475), was isolated from three different penguins, making it of special interest. Its genome was compared to those of other E. faecalis sequence types (ST116 and ST242) recovered from Magellanic penguins, as well as to an existing phylogeny of E. faecalis isolated from diverse origins over the past 100 years. No penguin-derived E. faecalis strains were closely related to dominant clinical lineages. Most possessed intact CRISPR defenses, few mobile elements, and antibiotic resistances limited to those intrinsic to the species and lacked pathogenic features conveyed by mobile elements. Interestingly, plasmids were identified in penguin isolates that also had been reported for other marine mammals. Enterococci isolated from penguins showed limited anthropogenic impact, indicating that they are likely representative of those naturally circulating in the ecosystem inhabited by the penguins. These findings establish an important baseline for detecting the encroachment of human activity into remote planetary environments.IMPORTANCE Enterococci are host-associated microbes that have an unusually broad range, from the built hospital environment to the guts of insects and other animals in remote locations. Despite their occurrence in the guts of animals for hundreds of millions of years, we know little about the properties that confer this range or how anthropogenic activities may be introducing new selective forces. Magellanic penguins live at the periphery of human habitation. It was of interest to examine enterococci from these animals for the presence of antibiotic resistance and other markers reflective of anthropogenic selection. Diverse enterococcal lineages found discount the existence of a single well-adapted intrinsic penguin-specific species. Instead, they appear to be influenced by a carnivorous lifestyle and enterococci present in the coastal sea life consumed. These results indicate that currently, the penguin habitat remains relatively free of pollutants that select for adaptation to human-derived stressors.

Genome-based reclassification of Paenibacillus panacisoli DSM 21345T as Paenibacillus massiliensis subsp. panacisoli subsp. nov. and description of Paenibacillus massiliensis subsp. massiliensis subsp. nov.

Bacteria of the genus Paenibacillus are relevant to humans, animals and plants. The species Paenibacillus massiliensis and Paenibacillus panacisoli are Gram-stain-positive and endospore-forming bacilli isolated from a blood culture of a leukemia patient and from soil of a ginseng field, respectively. Comparative analyses of their 16S rRNA genes revealed that the two Paenibacillus species could be synonyms (99.3% sequence identity). In the present study we performed different genomic analyses in order to evaluate the phylogenetic relationship of these micro-organisms. Paenibacillus massiliensis DSM 16942T and P. panacisoli DSM 21345T presented a difference in their G+C content lower than 1 mol%, overall genome relatedness index values higher than the species circumscription thresholds (average nucleotide identity, 95.57 %; genome-wide ANI, =96.51 %; and orthologous ANI, 96.25 %), and a monophyletic grouping pattern in the phylogenies of the 16S rRNA gene and the proteome core. Considering that these strains present differential biochemical capabilities and that their computed digital DNA-DNA hybridization value is lower than the cut-off for bacterial subspecies circumscription, we suggest that each of them form different subspecies of P. massiliensis, Paenibacillus massiliensis subsp. panacisoli subsp. nov. (type strain DSM 21345T) and Paenibacillus massiliensis subsp. massiliensis subsp. nov. (type strain DSM 16942T).

Systematic review of descriptions of novel bacterial species: evaluation of the twenty-first century taxonomy through text mining.

Although described bacterial species increased in the twenty-first century, they correspond to a tiny fraction of the actual number of species living on our planet. The volume of textual data of these descriptions constitutes valuable information for revealing trends that in turn could support strategies for improvement of bacterial taxonomy. In this study, a text mining approach was used to generate bibliometric data to verify the state-of-art of bacterial taxonomy. Around 9700 abstracts of bacterial classification containing the expression 'sp. nov.' and published between 2001 and 2018 were downloaded from PubMed and analysed. Most articles were from PR China and the Republic of Korea, and published in the International Journal of Systematic and Evolutionary Microbiology. From about 10 800 species names detected, 93.33 % were considered valid according to the rules of the Bacterial Code, and they corresponded to 82.98 % of the total number of species validated between 2001 and 2018. Streptomyces, Bacillus and Paenibacillus each had more than 200 species described in the period. However, almost 40 % of all species were from the phylum Proteobacteria. Most bacteria were Gram-stain-negative, bacilli and isolated from soil. Thirteen species and one genus homonyms were found. With respect to methodologies of bacterial characterization, the use of terms related to 16S rRNA and polar lipids increased along these years, and terms related to genome metrics only began to appear from 2009 onward, although at a relatively lower frequency. Bacterial taxonomy is known as a conservative discipline, but it gradually changed in terms of players and practices. With the advent of the mandatory use of genomic analyses for species description, we are probably witnessing a turning point in the evolution of bacterial taxonomy.

Genomic metrics analyses indicate that Paenibacillus azotofixans is not a later synonym of Paenibacillus durus.

Paenibacillus durusand Paenibacillus azotofixans, both Gram-stain-positive and endospore-forming bacilli, have been considered to be a single species. However, a preliminary computation of their average nucleotide identity (ANI) values suggested that these species are not synonyms. Given this, the taxonomic attributions of these species were evaluated through genomic and phylogenomic approaches. Although the identity of 16S rRNA gene sequences of P. durus DSM 1735T and P. azotofixans ATCC 35681T are above the circumscription species threshold, genomic metrics analyses indicate otherwise. ANI, gANI and OrthoANI values computed from their genome sequences were around 92 %, below the species limits. Digital DNA-DNA hybridization and MUMi estimations also corroborated these observations. In fact, in all metrics, Paenibacillus zanthoxyli JH29T seemed to be more similar to Paenibacillus azotofixans. ATCC 35681T than P. durus DSM 1735T. Phylogenetic analyses based on concatenated core-proteome and concatenated gyrB, recA, recN and rpoB genes confirmed that P. zanthoxyli is the closest Paenibacillus species to P. azotofixans. A review of the phenotypic profiles from these three species revealed that their biochemical repertoires are very similar, although P. azotofixans ATCC 35681T can be differentiated from P. durus DSM1735T in 13 among more than 90 phenotypic traits. Considering phylogenetic and genomic analyses, Paenibacillus azotofixans should be considered as an independent species, and not as a later synonym of Paenibacillus durus.

Genome-based reclassification of Paenibacillus dauci as a later heterotypic synonym of Paenibacillus shenyangensis.

Paenibacillus shenyangensis and Paenibacillus dauci are Gram-stain-positive, rod-shaped and endospore-forming bacteria originally isolated from soil and carrot samples, respectively, in China. Preliminary comparative genomic analysis showed that these bacteria could constitute a single species. Therefore, in this study, their taxonomic statuses were clarified through distinct genomic metrics and phylogenetic analyses. Paenibacillus shenyangensis A9T and P. dauci H9T presented values of average nucleotide identity (ANI) and its derivative metrics (gANI and OrthoANI) ranging from 97.88 to 98.08 %, and digital DNA-DNA hybridization equal to 89.08 %. Furthermore, the identities of 16S rRNA, gyrB, rpoB, recA and recN genes were all equal or higher than 98.7 %. Phylogenies of these marker genes and the concatenated core proteome were congruent in the sense that P. shenyangensis A9T and P. dauci H9T are the closest type-strains of the genus Paenibacillus. A review of their profiles revealed that these strains do not present pronounced differences at the phenotypic and chemotaxonomic levels. Considering phylogenetic, genomic, phenotypic and chemotaxonomic data, P. dauci should be reclassified as a later heterotypic synonym of P. shenyangensis.

Paenibacillus helianthi sp. nov., a nitrogen fixing species isolated from the rhizosphere of Helianthus annuus L.

Three facultatively anaerobic endospore-forming bacteria were isolated from the rhizosphere of sunflowers grown in fields of Rio Grande do Sul State, Brazil. The designated type strain P26ET was previously identified as a sunflower growth promoting bacterium and is able to fix nitrogen and to excrete ammonia. According to analyses of 16S rRNA gene sequences, P26ET presented similarity values above 98.8% in relation to Paenibacillus azotifigens NF2-4-5T, Paenibacillus graminis RSA19T, Paenibacillus jilunlii Be17T, Paenibacillus salinicaeni LAM0A28T, and Paenibacillus sonchi X19-5T. Phylogenetic reconstructions based on 16S rRNA gene and core proteome data showed that the strains P26ET, P3E and P32E form a distinct clade, which did not include any type strain of the currently described Paenibacillus species. Also, genomic comparisons using average nucleotide identity (ANI), Orthologous ANI and in silico DNA-DNA hybridization revealed similarity ranges below the recommended thresholds when the three isolates from sunflower were compared to their close relatives. The DNA G + C content of strain P26ET was determined to be 49.4 mol%. The major cellular fatty acids are anteiso-C15:0 and iso-C15:0, representing about 58 and 14% of the total fatty acids in P26ET, respectively. Based on different taxonomic genomic metrics, phylogeny, and phenotypic data, we propose that strain P26ET (= DSM 102269 = BR10509) represents a novel species within the genus Paenibacillus, for which the name Paenibacillus helianthi sp. nov. is proposed.

The genomes of three Bradyrhizobium sp. isolated from root nodules of Lupinus albescens grown in extremely poor soils display important genes for resistance to environmental stress.

Lupinus albescens is a resistant cover plant that establishes symbiotic relationships with bacteria belonging to the Bradyrhizobium genus. This symbiosis helps the development of these plants in adverse environmental conditions, such as the ones found in arenized areas of Southern Brazil. This work studied three Bradyrhizobium sp. (AS23, NAS80 and NAS96) isolated from L. albescens plants that grow in extremely poor soils (arenized areas and adjacent grasslands). The genomes of these three strains were sequenced in the Ion Torrent platform using the IonXpress library preparation kit, and presented a total number of bases of 1,230,460,823 for AS23, 1,320,104,022 for NAS80, and 1,236,105,093 for NAS96. The genome comparison with closest strains Bradyrhizobium japonicum USDA6 and Bradyrhizobium diazoefficiens USDA110 showed important variable regions (with less than 80% of similarity). Genes encoding for factors for resistance/tolerance to heavy metal, flagellar motility, response to osmotic and oxidative stresses, heat shock proteins (present only in the three sequenced genomes) could be responsible for the ability of these microorganisms to survive in inhospitable environments. Knowledge about these genomes will provide a foundation for future development of an inoculant bioproduct that should optimize the recovery of degraded soils using cover crops.

Characteristics of serogroup 20 S.pneumoniae isolates from Brazil.

BACKGROUND: Although serogroup 20 is not part of any conjugate pneumococcal vaccine, its serotype 20A, but not 20B, belongs to the polysaccharide 23-valent formula. Little is known about its clinical, laboratorial and epidemiological characteristics. METHODS: The purpose of this study was to evaluate the bacterial genotypes (by PFGE and MLST), clinical characteristics of patients (from review of medical records) and antimicrobial susceptibility of serogroup 20 isolates which were recovered from patients with invasive pneumococcal disease (IPD) from 2007 to 2012. Subtyping to determine 20A and 20B types was also performed by sequencing the genes of the cps locus. RESULTS: Sixteen isolates were genotyped and were highly related. All pneumococci were resistant to tetracycline and 31 % were non-susceptible to trimethoprim/sulfamethoxazole. Penicillin MIC ranged from 0.004 to 1 µg/mL and non-susceptibility (MIC ≥ 0.12 µg/mL) was observed in 5/16 isolates (31 %). All isolates belonged to subtype 20B. Most patients were male with a median age of 62 years and presented at least one underlying disease (mostly respiratory conditions). All isolates belonged to ST8889 and to a unique PFGE clone. CONCLUSIONS: A high clonal occurrence of serotype 20B pneumococci recovered from patients with IPD in Brazil was observed. As a non-PCV10 serotype, selective pressure may be responsible for this unusual occurrence of serogroup 20. However, temporal variation effect should not be underestimated; therefore it is an issue that warrants continued monitoring.

Incipient Parallel Evolution of SARS-CoV-2 Deltacron Variant in South Brazil.

With the coexistence of multiple lineages and increased international travel, recombination and gene flow are likely to become increasingly important in the adaptive evolution of SARS-CoV-2. These processes could result in genetic introgression and the incipient parallel evolution of multiple recombinant lineages. However, identifying recombinant lineages is challenging, and the true extent of recombinant evolution in SARS-CoV-2 may be underestimated. This study describes the first SARS-CoV-2 Deltacron recombinant case identified in Brazil. We demonstrate that the recombination breakpoint is at the beginning of the Spike gene. The 5' genome portion (circa 22 kb) resembles the AY.101 (Delta), and the 3' genome portion (circa 8 kb nucleotides) is most similar to the BA.1.1 (Omicron). Furthermore, evolutionary genomic analyses indicate that the new strain emerged after a single recombination event between lineages of diverse geographical locations in December 2021 in South Brazil. This Deltacron, AYBA-RS, is one of the dozens of recombinants described in 2022. The submission of only four sequences in the GISAID database suggests that this lineage had a minor epidemiological impact. However, the recent emergence of this and other Deltacron recombinant lineages (XD, XF, and XS) suggests that gene flow and recombination may play an increasingly important role in the COVID-19 pandemic. We explain the evolutionary and population genetic theory that supports this assertion, concluding that this stresses the need for continued genomic surveillance. This monitoring is vital for countries where multiple variants are present, as well as for countries that receive significant inbound international travel.

Viral load is associated with mitochondrial dysfunction and altered monocyte phenotype in acute severe SARS-CoV-2 infection.

Monocytes play a major role in the initial innate immune response to SARS-CoV-2. Although viral load may correlate with several clinical outcomes in COVID-19, much less is known regarding their impact on innate immune phenotype. We evaluated the monocyte phenotype and mitochondrial function in severe COVID-19 patients (n = 22) with different viral burden (determined by the median of viral load of the patients) at hospital admission. Severe COVID-19 patients presented lower frequency of CD14 + CD16- classical monocytes and CD39 expression on CD14 + monocytes, and higher frequency of CD14 + CD16 + intermediate and CD14-CD16 + nonclassical monocytes as compared to healthy controls independently of viral load. COVID-19 patients with high viral load exhibited increased GM-CSF, PGE-2 and lower IFN-α as compared to severe COVID-19 patients with low viral load (p < 0.05). CD14 + monocytes of COVID-19 patients with high viral load presented higher expression of PD-1 but lower HLA-DR on the cell surface than severe COVID-19 patients with low viral load. All COVID-19 patients presented decreased monocyte mitochondria membrane polarization, but high SARS-CoV-2 viral load was associated with increased mitochondrial reactive oxygen species. In this sense, higher viral load induces mitochondrial reactive oxygen species generation associated with exhaustion profile in CD14 + monocytes of severe COVID-19 patients. Altogether, these data shed light on new pathological mechanisms involving SARS-CoV-2 viral load on monocyte activation and mitochondrial function, which were associated with COVID-19 severity.

SARS-CoV-2 introduction and lineage dynamics across three epidemic peaks in Southern Brazil: massive spread of P.1.

Genomic surveillance of SARS-CoV-2 is paramount for understanding viral dynamics, contributing to disease control. This study analyzed SARS-CoV-2 genomic diversity in Rio Grande do Sul (RS), Brazil, including the first reported case in each Regional Health Coordination and cases from three epidemic peaks. Ninety SARS-CoV-2 genomes from RS were sequenced and analyzed through comparison with SARS-CoV-2 datasets available in GISAID for phylogenetic inference and mutation analysis. Among the first reported cases, we found the following lineages: B.1 (33.3%), B.1.1.28 (26.7%), B.1.1 (13.3%), B.1.1.33 (10.0%), and A (6.7%), evidencing SARS-CoV-2 introduction by both international origin and community-driven transmission. We found predominance of B.1.1.33 (50.0%) and B.1.1.28 (35.0%) during the first epidemic peak (July-August 2020), emergence of P.2 (55.6%) in the second peak (November-December 2020), and massive spread of P.1 and related sequences (78.4%), such as P.1-like-II, P.1.1 and P.1.2 in the third peak (February-April, 2021). Eighteen novel mutation combinations were found among P.1 genomes, and 22 different spike mutations and/or deletions among P.1 and related sequences. This study shows the dispersion of SARS-CoV-2 lineages in Southern Brazil and describes SARS-CoV-2 diversity during three epidemic peaks, highlighting the spread of P.1 and the high genetic diversity of currently circulating lineages. Genomic monitoring of SARS-CoV-2 is essential to guide health authorities' decisions to control COVID-19 in Brazil.

Genomic Metrics Applied to Rhizobiales (Hyphomicrobiales): Species Reclassification, Identification of Unauthentic Genomes and False Type Strains.

Taxonomic decisions within the order Rhizobiales have relied heavily on the interpretations of highly conserved 16S rRNA sequences and DNA-DNA hybridizations (DDH). Currently, bacterial species are defined as including strains that present 95-96% of average nucleotide identity (ANI) and 70% of digital DDH (dDDH). Thus, ANI values from 520 genome sequences of type strains from species of Rhizobiales order were computed. From the resulting 270,400 comparisons, a ≥95% cut-off was used to extract high identity genome clusters through enumerating maximal cliques. Coupling this graph-based approach with dDDH from clusters of interest, it was found that: (i) there are synonymy between Aminobacter lissarensis and Aminobacter carboxidus, Aurantimonas manganoxydans and Aurantimonas coralicida, "Bartonella mastomydis," and Bartonella elizabethae, Chelativorans oligotrophicus, and Chelativorans multitrophicus, Rhizobium azibense, and Rhizobium gallicum, Rhizobium fabae, and Rhizobium pisi, and Rhodoplanes piscinae and Rhodoplanes serenus; (ii) Chelatobacter heintzii is not a synonym of Aminobacter aminovorans; (iii) "Bartonella vinsonii" subsp. arupensis and "B. vinsonii" subsp. berkhoffii represent members of different species; (iv) the genome accessions GCF_003024615.1 ("Mesorhizobium loti LMG 6,125T"), GCF_003024595.1 ("Mesorhizobium plurifarium LMG 11,892T"), GCF_003096615.1 ("Methylobacterium organophilum DSM 760T"), and GCF_000373025.1 ("R. gallicum R-602 spT") are not from the genuine type strains used for the respective species descriptions; and v) "Xanthobacter autotrophicus" Py2 and "Aminobacter aminovorans" KCTC 2,477T represent cases of misuse of the term "type strain". Aminobacter heintzii comb. nov. and the reclassification of Aminobacter ciceronei as A. heintzii is also proposed. To facilitate the downstream analysis of large ANI matrices, we introduce here ProKlust ("Prokaryotic Clusters"), an R package that uses a graph-based approach to obtain, filter, and visualize clusters on identity/similarity matrices, with settable cut-off points and the possibility of multiple matrices entries.

Emergence of the novel SARS-CoV-2 lineage VUI-NP13L and massive spread of P.2 in South Brazil.

In this study, we analyzed 340 whole genomes of SARS-CoV-2, which were sampled between April and November 2020 in 33 cities of Rio Grande do Sul, South Brazil. We demonstrated the circulation of two novel emergent lineages, VUI-NP13L and VUI-NP13L-like, and five major lineages that had already been assigned (B.1.1.33, B.1.1.28, P.2, B.1.91, B.1.195). P.2 and VUI-NP13L demonstrated a massive spread in October 2020. Constant and consistent genomic surveillance is crucial to identify newly emerging SARS-CoV-2 lineages in Brazil and to guide decision making in the Brazilian Public Healthcare System.