Next-generation sequencing has become a useful tool for studying transcriptomes. However, these methods typically rely on sequencing short fragments of cDNA, then attempting to assemble the pieces into full-length transcripts. Here, we describe a method that uses PacBio long reads to sequence full-length cDNAs from individual transcriptomes and metatranscriptome samples. We have adapted the PacBio Iso-Seq protocol for use with prokaryotic samples by incorporating RNA polyadenylation and rRNA-depletion steps. In conjunction with SMRT Sequencing, which has average readlengths of 10-15 kb, we are able to sequence entire transcripts, including polycistronic RNAs, in a single read. Here, we show full-length bacterial transcriptomes with the ability to visualize transcription of operons. In the area of metatranscriptomics, long reads reveal unambiguous gene sequences without the need for post-sequencing transcript assembly. We also show full-length bacterial transcripts sequenced after being treated with NEB’s Cappable-Seq, which is an alternative method for depleting rRNA and enriching for full-length transcripts with intact 5’ ends. Combining Cappable-Seq with PacBio long reads allows for the detection of transcription start sites, with the additional benefit of sequencing entire transcripts.
Evolution of a 72-kb cointegrant, conjugative multiresistance plasmid from early community-associated methicillin-resistant Staphylococcus aureus isolates.
Horizontal transfer of plasmids encoding antimicrobial-resistance and virulence determinants has been instrumental in Staphylococcus aureus evolution, including the emergence of community-associated methicillin-resistant S. aureus (CA-MRSA). In the early 1990s the first CA-MRSA isolated in Western Australia (WA), WA-5, encoded cadmium, tetracycline and penicillin-resistance genes on plasmid pWBG753 (~30 kb). WA-5 and pWBG753 appeared only briefly in WA, however, fusidic-acid-resistance plasmids related to pWBG753 were also present in the first European CA-MRSA at the time. Here we characterized a 72-kb conjugative plasmid pWBG731 present in multiresistant WA-5-like clones from the same period. pWBG731 was a cointegrant formed from pWBG753 and a pWBG749-family conjugative plasmid. pWBG731 carried mupirocin, trimethoprim, cadmium and penicillin-resistance genes. The stepwise evolution of pWBG731 likely occurred through the combined actions of IS257, IS257-dependent miniature inverted-repeat transposable elements (MITEs) and the BinL resolution system of the ß-lactamase transposon Tn552 An evolutionary intermediate ~42-kb non-conjugative plasmid pWBG715, possessed the same resistance genes as pWBG731 but retained an integrated copy of the small tetracycline-resistance plasmid pT181. IS257 likely facilitated replacement of pT181 with conjugation genes on pWBG731, thus enabling autonomous transfer. Like conjugative plasmid pWBG749, pWBG731 also mobilized non-conjugative plasmids carrying oriT mimics. It seems likely that pWBG731 represents the product of multiple recombination events between the WA-5 pWBG753 plasmid and other mobile genetic elements present in indigenous CA-MSSA. The molecular evolution of pWBG731 saliently illustrates how diverse mobile genetic elements can together facilitate rapid accrual and horizontal dissemination of multiresistance in S. aureus CA-MRSA.Copyright © 2019 American Society for Microbiology.
Tigecycline is one of the last-resort antibiotics to treat complicated infections caused by both multidrug-resistant Gram-negative and Gram-positive bacteria1. Tigecycline resistance has sporadically occurred in recent years, primarily due to chromosome-encoding mechanisms, such as overexpression of efflux pumps and ribosome protection2,3. Here, we report the emergence of the plasmid-mediated mobile tigecycline resistance mechanism Tet(X4) in Escherichia coli isolates from China, which is capable of degrading all tetracyclines, including tigecycline and the US FDA newly approved eravacycline. The tet(X4)-harbouring IncQ1 plasmid is highly transferable, and can be successfully mobilized and stabilized in recipient clinical and laboratory strains of Enterobacteriaceae bacteria. It is noteworthy that tet(X4)-positive E.?coli strains, including isolates co-harbouring mcr-1, have been widely detected in pigs, chickens, soil and dust samples in China. In vivo murine models demonstrated that the presence of Tet(X4) led to tigecycline treatment failure. Consequently, the emergence of plasmid-mediated Tet(X4) challenges the clinical efficacy of the entire family of tetracycline antibiotics. Importantly, our study raises concern that the plasmid-mediated tigecycline resistance may further spread into various ecological niches and into clinical high-risk pathogens. Collective efforts are in urgent need to preserve the potency of these essential antibiotics.
This study describes the characterization of type 2 IncC plasmids pC-Ec20-KPC and pC-Ec2-KPC, carrying blaKPC-2 gene, from two multiresistant E. coli recovered in the University Hospital of Larissa, in 2018.Escherichia coli, Ec-2Lar and Ec-20Lar, were recovered from rectal swabs from two patients, during the monthly surveillance cultures. Transfer experiments by conjugation were carried out with E. coli recipients. blaKPC-carrying plasmids were characterized by S1 profiling. Isolates were typed by MLST. Whole bacterial genome was sequenced using the Sequel platform.Both E. coli isolates, belonging to ST648, transferred the blaKPC-2 to E. coli A15 laboratory strain by conjugation. Plasmid analysis revealed that the transconjugants harbored blaKPC-positive plasmids of different sizes. Analysis of plasmid sequences showed that, in both isolates, blaKPC-2 gene was carried on type 2 IncC plasmids pC-Ec20-KPC and pC-Ec2-KPC. Both plasmids carried the ARI-B resistance island, which consisted of several resistance genes, intact and truncated copies of several mobile elements, and a 25,571-bp segment harboring coding sequences for an iron transporter. The blaKPC-2 gene was part of the transposon Tn4401a, which was bounded by direct repeats of 5 bp (TCCTT) suggesting its transposition into the IncC plasmids.To our knowledge, this is the first report on complete nucleotide sequences of type 2 IncC plasmids. These findings, which hypothesize the acquisition of KPC-2-encoding transposon Tn4401a by an IncC replicon, indicate the ongoing need for molecular surveillance studies of MDR pathogens. Additionally, they underline the increasing clinical importance of the IncC plasmid family.Copyright © 2019. Published by Elsevier Ltd.
Detection of transferable oxazolidinone resistance determinants in Enterococcus faecalis and Enterococcus faecium of swine origin in Sichuan Province, China.
The aim of this study was to detect the transferable oxazolidinone resistance determinants (cfr, optrA and poxtA) in E. faecalis and E. faecium of swine origin in Sichuan Province, China.A total of 158 enterococci strains (93 E. faecalis and 65 E. faecium) isolated from 25 large-scale swine farms were screened for the presence of cfr, optrA and poxtA by PCR. The genetic environments of cfr, optrA and poxtA were characterized by whole genome sequencing. Transfer of oxazolidinone resistance determinants was determined by conjugation or electrotransformation experiments.The transferable oxazolidinone resistance determinants, cfr, optrA and poxtA, were detected in zero, six, and one enterococci strains, respectively. The poxtA in one E. faecalis strain was located on a 37,990 bp plasmid, which co-harbored fexB, cat, tet(L) and tet(M), and could be conjugated to E. faecalis JH2-2. One E. faecalis strain harbored two different OptrA variants, including one variant with a single substitution, Q219H, which has not been reported previously. Two optrA-carrying plasmids, pC25-1, with a size of 45,581 bp, and pC54, with a size of 64,500 bp, shared a 40,494 bp identical region that contained genetic context IS1216E-fexA-optrA-erm(A)-IS1216E, which could be electrotransformed into Staphylococcus aureus. Four different chromosomal optrA gene clusters were found in five strains, in which optrA was associated with Tn554 or Tn558 that were inserted into the radC gene.Our study highlights the fact that mobile genetic elements, such as plasmids, IS1216E, Tn554 and Tn558, may facilitate the horizontal transmission of optrA or poxtA.Copyright © 2019. Published by Elsevier Ltd.
Molecular Characterization of a Multidrug-Resistant Klebsiella pneumoniae Strain R46 Isolated from a Rabbit
To investigate the mechanisms of multiple resistance and the horizontal transfer of resistance genes in animal pathogens, we characterized the molecular structures of the resistance gene-related sequences in a multidrug-resistant Klebsiella pneumoniae strain R46 isolated from a rabbit. Molecular cloning was performed to clone the resistance genes, and minimum inhibitory concentrations (MICs) were measured to determine the resistance characteristics of the cloned genes and related strains. A conjugation experiment was conducted to assess the transferability of the resistance plasmids. Sequencing and comparative genomic methods were used to analyze the structures of the resistance gene-related sequences. The K. pneumoniae R46 genome consisted of a chromosome and three resistance plasmids named pR46-27, pR46-42, and pR46-270, respectively. The whole genome encoded 34 antibiotic resistance genes including a newly identified chromosome-encoded florfenicol resistance gene named mdfA2. pR46-270, besides encoding 26 antibiotic resistance genes, carried four clusters of heavy metal resistance genes and several virulence-related genes or gene clusters. The plasmid-encoded resistance genes were mostly associated with mobile genetic elements. The plasmid with the most similarity to the floR gene-harboring plasmid pR46-27 was pCTXM-2271, a plasmid from Escherichia coli. The results of this work demonstrated that the plasmids with multidrug resistance genes were present in animal-derived bacteria and more florfenicol resistance genes such as mdfA2 could be present in bacterial populations. The resistance genes encoded on the plasmids may spread between the bacteria of different species or genera and cause the resistance dissemination.
The ADEP Biosynthetic Gene Cluster in Streptomyces hawaiiensis NRRL 15010 Reveals an Accessory clpP Gene as a Novel Antibiotic Resistance Factor.
The increasing threat posed by multiresistant bacterial pathogens necessitates the discovery of novel antibacterials with unprecedented modes of action. ADEP1, a natural compound produced by Streptomyces hawaiiensis NRRL 15010, is the prototype for a new class of acyldepsipeptide (ADEP) antibiotics. ADEP antibiotics deregulate the proteolytic core ClpP of the bacterial caseinolytic protease, thereby exhibiting potent antibacterial activity against Gram-positive bacteria, including multiresistant pathogens. ADEP1 and derivatives, here collectively called ADEP, have been previously investigated for their antibiotic potency against different species, structure-activity relationship, and mechanism of action; however, knowledge on the biosynthesis of the natural compound and producer self-resistance have remained elusive. In this study, we identified and analyzed the ADEP biosynthetic gene cluster in S. hawaiiensis NRRL 15010, which comprises two NRPSs, genes necessary for the biosynthesis of (4S,2R)-4-methylproline, and a type II polyketide synthase (PKS) for the assembly of highly reduced polyenes. While no resistance factor could be identified within the gene cluster itself, we discovered an additional clpP homologous gene (named clpPADEP) located further downstream of the biosynthetic genes, separated from the biosynthetic gene cluster by several transposable elements. Heterologous expression of ClpPADEP in three ADEP-sensitive Streptomyces species proved its role in conferring ADEP resistance, thereby revealing a novel type of antibiotic resistance determinant.IMPORTANCE Antibiotic acyldepsipeptides (ADEPs) represent a promising new class of potent antibiotics and, at the same time, are valuable tools to study the molecular functioning of their target, ClpP, the proteolytic core of the bacterial caseinolytic protease. Here, we present a straightforward purification procedure for ADEP1 that yields substantial amounts of the pure compound in a time- and cost-efficient manner, which is a prerequisite to conveniently study the antimicrobial effects of ADEP and the operating mode of bacterial ClpP machineries in diverse bacteria. Identification and characterization of the ADEP biosynthetic gene cluster in Streptomyces hawaiiensis NRRL 15010 enables future bioinformatics screenings for similar gene clusters and/or subclusters to find novel natural compounds with specific substructures. Most strikingly, we identified a cluster-associated clpP homolog (named clpPADEP) as an ADEP resistance gene. ClpPADEP constitutes a novel bacterial resistance factor that alone is necessary and sufficient to confer high-level ADEP resistance to Streptomyces across species.Copyright © 2019 American Society for Microbiology.
Increased prevalence of Escherichia coli strains from food carrying blaNDM and mcr-1-bearing plasmids that structurally resemble those of clinical strains, China, 2015 to 2017.
Introduction: Emergence of resistance determinants of blaNDM and mcr-1 has undermined the antimicrobial effectiveness of the last line drugs carbapenems and colistin. Aim: This work aimed to assess the prevalence of blaNDM and mcr-1 in E. coli strains collected from food in Shenzhen, China, during the period 2015 to 2017. Methods: Multidrug-resistant E. coli strains were isolated from food samples. Plasmids encoding mcr-1 or blaNDM genes were characterised and compared with plasmids found in clinical isolates.ResultsAmong 1,166 non-repeated cephalosporin-resistant E. coli strains isolated from 2,147 food samples, 390 and 42, respectively, were resistant to colistin and meropenem, with five strains being resistant to both agents. The rate of resistance to colistin increased significantly (p?0.01) from 26% in 2015 to 46% in 2017, and that of meropenem resistance also increased sharply from 0.3% in 2015 to 17% in 2017 (p?0.01). All meropenem-resistant strains carried a plasmid-borne blaNDM gene. Among the colistin-resistant strains, three types of mcr-1-bearing plasmids were determined. Plasmid sequencing indicated that these mcr-1 and blaNDM-bearing plasmids were structurally similar to those commonly recovered from clinical isolates. Interestingly, both mcr-1-bearing and blaNDM-bearing plasmids were transferrable to E. coli strain J53 under selection by meropenem, yet only mcr-1-bearing plasmids were transferrable under colistin selection. Conclusion: These findings might suggest that mobile elements harbouring mcr-1 and blaNDM have been acquired by animal strains and transmitted to our food products, highlighting a need to prevent a spike in the rate of drug resistant food-borne infections.
Advantage of the F2:A1:B- IncF Pandemic Plasmid over IncC Plasmids in In Vitro Acquisition and Evolution of blaCTX-M Gene-Bearing Plasmids in Escherichia coli.
Despite a fitness cost imposed on bacterial hosts, large conjugative plasmids play a key role in the diffusion of resistance determinants, such as CTX-M extended-spectrum ß-lactamases. Among the large conjugative plasmids, IncF plasmids are the most predominant group, and an F2:A1:B- IncF-type plasmid encoding a CTX-M-15 variant was recently described as being strongly associated with the emerging worldwide Escherichia coli sequence type 131 (ST131)-O25b:H4 H30Rx/C2 sublineage. In this context, we investigated the fitness cost of narrow-range F-type plasmids, including the F2:A1:B- IncF-type CTX-M-15 plasmid, and of broad-range C-type plasmids in the K-12-like J53-2 E. coli strain. Although all plasmids imposed a significant fitness cost to the bacterial host immediately after conjugation, we show, using an experimental-evolution approach, that a negative impact on the fitness of the host strain was maintained throughout 1,120 generations with the IncC-IncR plasmid, regardless of the presence or absence of cefotaxime, in contrast to the F2:A1:B- IncF plasmid, whose cost was alleviated. Many chromosomal and plasmid rearrangements were detected after conjugation in transconjugants carrying the IncC plasmids but not in transconjugants carrying the F2:A1:B- IncF plasmid, except for insertion sequence (IS) mobilization from the fliM gene leading to the restoration of motility of the recipient strains. Only a few mutations occurred on the chromosome of each transconjugant throughout the experimental-evolution assay. Our findings indicate that the F2:A1:B- IncF CTX-M-15 plasmid is well adapted to the E. coli strain studied, contrary to the IncC-IncR CTX-M-15 plasmid, and that such plasmid-host adaptation could participate in the evolutionary success of the CTX-M-15-producing pandemic E. coli ST131-O25b:H4 lineage.Copyright © 2019 Mahérault et al.
Salmonella Genomic Island 3 Is an Integrative and Conjugative Element and Contributes to Copper and Arsenic Tolerance of Salmonella enterica.
Salmonella genomic island 3 (SGI3) was first described as a chromosomal island in Salmonella 4,,12:i:-, a monophasic variant of Salmonella enterica subsp. enterica serovar Typhimurium. The SGI3 DNA sequence detected from Salmonella 4,,12:i:- isolated in Japan was identical to that of a previously reported one across entire length of 81?kb. SGI3 consists of 86 open reading frames, including a copper homeostasis and silver resistance island (CHASRI) and an arsenic tolerance operon, in addition to genes related to conjugative transfer and DNA replication or partitioning, suggesting that the island is a mobile genetic element. We successfully selected transconjugants that acquired SGI3 after filter-mating experiments using the S. enterica serovars Typhimurium, Heidelberg, Hadar, Newport, Cerro, and Thompson as recipients. Southern blot analysis using I-CeuI-digested genomic DNA demonstrated that SGI3 was integrated into a chromosomal fragment of the transconjugants. PCR and sequencing analysis demonstrated that SGI3 was inserted into the 3′ end of the tRNA genes pheV or pheR The length of the target site was 52 or 55?bp, and a 55-bp attI sequence indicating generation of the circular form of SGI3 was also detected. The transconjugants had a higher MIC against CuSO4 compared to the recipient strains under anaerobic conditions. Tolerance was defined by the cus gene cluster in the CHASRI. The transconjugants also had distinctly higher MICs against Na2HAsO4 compared to recipient strains under aerobic conditions. These findings clearly demonstrate that SGI3 is an integrative and conjugative element and contributes to the copper and arsenic tolerance of S. enterica.Copyright © 2019 American Society for Microbiology.
Conjugal Transfer, Whole-Genome Sequencing, and Plasmid Analysis of Four mcr-1-Bearing Isolates from U.S. Patients.
Four Enterobacteriaceae clinical isolates bearing mcr-1 gene-harboring plasmids were characterized. All isolates demonstrated the ability to transfer colistin resistance to Escherichia coli; plasmids were stable in conjugants after multiple passages on nonselective media. mcr-1 was located on an IncX4 (n?=?3) or IncN (n?=?1) plasmid. The IncN plasmid harbored 13 additional antimicrobial resistance genes. Results indicate that the mcr-1-bearing plasmids in this study were highly transferable in vitro and stable in the recipients.This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.
Genomic Analysis of Emerging Florfenicol-Resistant Campylobacter coli Isolated from the Cecal Contents of Cattle in the United States.
Genomic analyses were performed on florfenicol-resistant (FFNr) Campylobacter coli isolates recovered from cattle, and the cfr(C) gene-associated multidrug resistance (MDR) plasmid was characterized. Sixteen FFNrC. coli isolates recovered between 2013 and 2018 from beef cattle were sequenced using MiSeq. Genomes and plasmids were found to be closed for three of the isolates using the PacBio system. Single nucleotide polymorphisms (SNPs) across the genome and the structures of MDR plasmids were investigated. Conjugation experiments were performed to determine the transferability of cfr(C)-associated MDR plasmids. The spectrum of resistance encoded by the cfr(C) gene was further investigated by agar dilution antimicrobial susceptibility testing. All 16 FFNr isolates were MDR and exhibited coresistance to ciprofloxacin, nalidixic acid, clindamycin, and tetracycline. All isolates shared the same resistance genotype, carrying aph (3′)-III, hph, ?aadE (truncated), blaOXA-61, cfr(C), and tet(O) genes plus a mutation of GyrA (T86I). The cfr(C), aph (3′)-III, hph, ?aadE, and tet(O) genes were colocated on transferable MDR plasmids ranging in size from 48 to 50?kb. These plasmids showed high sequence homology with the pTet plasmid and carried several Campylobacter virulence genes, including virB2, virB4, virB5, VirB6, virB7, virB8, virb9, virB10, virB11, and virD4 The cfr(C) gene conferred resistance to florfenicol (8 to 32?µg/ml), clindamycin (512 to 1,024?µg/ml), linezolid (128 to 512?µg/ml), and tiamulin (1,024?µg/ml). Phylogenetic analysis showed SNP differences ranging from 11 to 2,248 SNPs among the 16 isolates. The results showed that the cfr(C) gene located in the conjugative pTet MDR/virulence plasmid is present in diverse strains, where it confers high levels of resistance to several antimicrobials, including linezolid, a critical drug for treating infections by Gram-positive bacteria in humans. This report highlights the power of genomic antimicrobial resistance surveillance to uncover the intricacies of transmissible coresistance and provides information that is needed for accurate risk assessment and mitigation strategies.IMPORTANCECampylobacter is a leading cause of foodborne diarrheal illness worldwide, with more than one million cases each year in the United States alone. The global emergence of antimicrobial resistance in this pathogen has become a growing public health concern. Florfenicol-resistant (FFNr) Campylobacter has been very rare in the United States. In this study, we employed whole-genome sequencing to characterize 16 multidrug-resistant Campylobacter coli isolates recovered from cattle in the United States. A gene [cfr(C)] was found to be responsible for resistance not only to florfenicol but also to several other antimicrobials, including linezolid, a critical drug for treating infections by Gram-positive bacteria in humans. The results showed that cfr(C) is located in a conjugative pTet MDR/virulence plasmid. This report highlights the power of antimicrobial resistance surveillance to uncover the intricacies of transmissible coresistance and provides information that is needed for accurate risk assessment and mitigation strategies.
Diverse Commensal Escherichia coli Clones and Plasmids Disseminate Antimicrobial Resistance Genes in Domestic Animals and Children in a Semirural Community in Ecuador.
The increased prevalence of antimicrobial resistance (AMR) among Enterobacteriaceae has had major clinical and economic impacts on human medicine. Many of the multidrug-resistant (multiresistant) Enterobacteriaceae found in humans are community acquired, and some of them are possibly linked to food animals (i.e., livestock raised for meat and dairy products). In this study, we examined whether numerically dominant commensal Escherichia coli strains from humans (n?=?63 isolates) and domestic animals (n?=?174 isolates) in the same community and with matching phenotypic AMR patterns were clonally related or shared the same plasmids. We identified 25 multiresistant isolates (i.e., isolates resistant to more than one antimicrobial) that shared identical phenotypic resistance patterns. We then investigated the diversity of E. coli clones, AMR genes, and plasmids carrying the AMR genes using conjugation, replicon typing, and whole-genome sequencing. All of the multiresistant E. coli isolates (from children and domestic animals) analyzed had at least 90 or more whole-genome SNP differences between one another, suggesting that none of the strains was recently transferred. While the majority of isolates shared the same antimicrobial resistance genes and replicons, DNA sequencing indicated that these genes and replicons were found on different plasmid structures. We did not find evidence of the clonal spread of AMR in this community: instead, AMR genes were carried on diverse clones and plasmids. This presents a significant challenge for understanding the movement of AMR in a community.IMPORTANCE Even though Escherichia coli strains may share nearly identical phenotypic AMR profiles and AMR genes and overlap in space and time, the diversity of clones and plasmids challenges research that aims to identify sources of AMR. Horizontal gene transfer appears to play a more significant role than clonal expansion in the spread of AMR in this community.Copyright © 2019 Salinas et al.
Spreading Patterns of NDM-Producing Enterobacteriaceae in Clinical and Environmental Settings in Yangon, Myanmar.
The spread of carbapenemase-producing Enterobacteriaceae (CPE), contributing to widespread carbapenem resistance, has become a global concern. However, the specific dissemination patterns of carbapenemase genes have not been intensively investigated in developing countries, including Myanmar, where NDM-type carbapenemases are spreading in clinical settings. In the present study, we phenotypically and genetically characterized 91 CPE isolates obtained from clinical (n = 77) and environmental (n = 14) samples in Yangon, Myanmar. We determined the dissemination of plasmids harboring genes encoding NDM-1 and its variants using whole-genome sequencing and plasmid analysis. IncFII plasmids harboring blaNDM-5 and IncX3 plasmids harboring blaNDM-4 or blaNDM-7 were the most prevalent plasmid types identified among the isolates. The IncFII plasmids were predominantly carried by clinical isolates of Escherichia coli, and their clonal expansion was observed within the same ward of a hospital. In contrast, the IncX3 plasmids were found in phylogenetically divergent isolates from clinical and environmental samples classified into nine species, suggesting widespread dissemination of plasmids via horizontal transfer. Half of the environmental isolates were found to possess IncX3 plasmids, and this type of plasmid was confirmed to transfer more effectively to recipient organisms at a relatively low temperature (25°C) compared to the IncFII plasmid. Moreover, various other plasmid types were identified harboring blaNDM-1, including IncFIB, IncFII, IncL/M, and IncA/C2, among clinical isolates of Klebsiella pneumoniae or Enterobacter cloacae complex. Overall, our results highlight three distinct patterns of the dissemination of blaNDM-harboring plasmids among CPE isolates in Myanmar, contributing to a better understanding of their molecular epidemiology and dissemination in a setting of endemicity.Copyright © 2019 American Society for Microbiology.
Supernumerary B chromosomes (Bs) are extra karyotype units in addition to A chromosomes, and are found in some fungi and thousands of animals and plant species. Bs are uniquely characterized due to their non-Mendelian inheritance, and represent one of the best examples of genomic conflict. Over the last decades, their genetic composition, function and evolution have remained an unresolved query, although a few successful attempts have been made to address these phenomena. A classical concept based on cytogenetics and genetics is that Bs are selfish and abundant with DNA repeats and transposons, and in most cases, they do not carry any function. However, recently, the modern quantum development of high scale multi-omics techniques has shifted B research towards a new-born field that we call “B-omics”. We review the recent literature and add novel perspectives to the B research, discussing the role of new technologies to understand the mechanistic perspectives of the molecular evolution and function of Bs. The modern view states that B chromosomes are enriched with genes for many significant biological functions, including but not limited to the interesting set of genes related to cell cycle and chromosome structure. Furthermore, the presence of B chromosomes could favor genomic rearrangements and influence the nuclear environment affecting the function of other chromatin regions. We hypothesize that B chromosomes might play a key function in driving their transmission and maintenance inside the cell, as well as offer an extra genomic compartment for evolution.