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Background: Massive high-throughput sequencing of short, hypervariable segments of the 16S ribosomal RNA (rRNA) gene has transformed the methodological landscape describing microbial diversity within and across complex biomes. However, several studies have shown that the methodology rather than the biological variation is responsible for the observed sample composition and distribution. This compromises meta-analyses, although this fact is often disregarded. Results: To facilitate true meta-analysis of microbiome studies, we developed NG-Tax, a pipeline for 16S rRNA gene amplicon sequence analysis that was validated with different mock communities and benchmarked against QIIME as a frequently used pipeline. The microbial composition of 49 independently amplified mock samples was characterized by sequencing two variable 16S rRNA gene regions, V4 and V5-V6, in three separate sequencing runs on Illumina’s HiSeq2000 platform. This allowed for the evaluation of important causes of technical bias in taxonomic classification: 1) run-to-run sequencing variation, 2) PCR–error, and 3) region/primer specific amplification bias. Despite the short read length (~140 nt) and all technical biases, the average specificity of the taxonomic assignment for the phylotypes included in the mock communities was 97.78%. On average 99.95% and 88.43% of the reads could be assigned to at least family or genus level, respectively, while assignment to ‘spurious genera’ represented on average only 0.21% of the reads per sample. Analysis of α- and β-diversity confirmed conclusions guided by biology rather than the aforementioned methodological aspects, which was not achieved with QIIME. Conclusions: Different biological outcomes are commonly observed due to 16S rRNA region-specific performance. NG-Tax demonstrated high robustness against choice of region and other technical biases associated with 16S rRNA gene amplicon sequencing studies, diminishing their impact and providing accurate qualitative and quantitative representation of the true sample composition. This will improve comparability between studies and facilitate efforts towards standardization.

Ancient whole-genome duplications (WGDs)—paleopolyploidy events—are key to solving Darwin’s ‘abominable mystery’ of how flowering plants evolved and radiated into a rich variety of species. The vertebrates also emerged from their invertebrate ancestors via two WGDs, and genomes of diverse gymnosperm trees, unicellular eukaryotes, invertebrates, fishes, amphibians and even a rodent carry evidence of lineage-specific WGDs. Modern polyploidy is common in eukaryotes, and it can be induced, enabling mechanisms and short-term cost-benefit assessments of polyploidy to be studied experimentally. However, the ancient WGDs can be reconstructed only by comparative genomics: these studies are difficult because the DNA duplicates have been through tens or hundreds of millions of years of gene losses, mutations, and chromosomal rearrangements that culminate in resolution of the polyploid genomes back into diploid ones (rediploidisation). Intriguing asymmetries in patterns of post-WGD gene loss and retention between duplicated sets of chromosomes have been discovered recently, and elaborations of signal transduction systems are lasting legacies from several WGDs. The data imply that simpler signalling pathways in the pre-WGD ancestors were converted via WGDs into multi-stranded parallelised networks. Genetic and biochemical studies in plants, yeasts and vertebrates suggest a paradigm in which different combinations of sister paralogues in the post-WGD regulatory networks are co-regulated under different conditions. In principle, such networks can respond to a wide array of environmental, sensory and hormonal stimuli and integrate them to generate phenotypic variety in cell types and behaviours. Patterns are also being discerned in how the post-WGD signalling networks are reconfigured in human cancers and neurological conditions. It is fascinating to unpick how ancient genomic events impact on complexity, variety and disease in modern life.

Both actin and microtubules are major cytoskeletal elements in eukaryotic cells that participate in many cellular processes, including cell division and motility, vesicle and organelle movement, and the maintenance of cell shape. Inside its host cell, the human pathogen Chlamydia trachomatis manipulates the cytoskeleton to promote its survival and enhance its pathogenicity. In particular, Chlamydia induces the drastic rearrangement of both actin and microtubules, which is vital for its entry, inclusion structure and development, and host cell exit. As significant progress in Chlamydia genetics has greatly enhanced our understanding of how this pathogen co-opts the host cytoskeleton, we will discuss the machinery used by Chlamydia to coordinate the reorganization of actin and microtubules.

In the expanding population of immunocompromised patients and those treated in intensive care units, rare fungal infectious agents have emerged as important pathogens, causing invasive infections associated with high morbidity and mortality. These infections may present either as de novo or as breakthrough invasive infections in high-risk patients with hematologic malignancies receiving prophylactic or empirical antifungal therapy or in patients with central venous catheters. Diagnosis and treatment are challenging. Physicians should have a high index of suspicion because early diagnosis is of paramount importance. Conventional diagnostic methods such as cultures and histopathology are still essential, but rapid and more specific molecular techniques for both detection and identification of the infecting pathogens are being developed and hopefully will lead to early targeted treatment. The management of invasive fungal infections is multimodal. Reversal of risk factors, if feasible, should be attempted. Surgical debridement is recommended in localized mold infections. The efficacy of various antifungal drugs is not uniform. Amphotericin B is active against most yeasts, except Trichosporon, as well as against Mucorales, Fusarium, and some species of Paecilomyces and dimorphic fungi. The use of voriconazole is suggested for the treatment of trichosporonosis and scedosporiosis. Combination treatment, though recommended as salvage therapy in some infections, is controversial in most cases. Despite the use of available antifungals, mortality remains high. The optimization of molecular-based techniques, with expansion of reference libraries and the possibility for direct detection of resistance mechanisms, is awaited with great interest in the near future. Further research is necessary, however, in order to find the best ways to confront and destroy these lurking enemies.

Bacterial vaginosis is the most prevalent cause of abnormal vaginal discharge in women of childbearing age. It can have a major impact on quality of life and psychological wellbeing if frequently recurrent and strongly symptomatic. The use of molecular techniques to study the vaginal microbiome is increasing our understanding of the dynamic changes in flora that occur in health and disease. It might soon be possible to separate Gardnerella into different pathogenic and non-pathogenic species. Many groups are studying compounds that can disrupt the biofilm which is dominated by Gardnerella and Atopobium vaginae. Several studies in the last decade support the concept of bacterial vaginosis as a sexually transmitted infection.

Klebsiella pneumoniae, a gram-negative bacillus of the Enterobacteriaceae family, is a component of the normal human microbiota and a common cause of community- and healthcare-associated infections. The increasing prevalence of antimicrobial resistance among K. pneumoniae isolates, particularly among those causing healthcare-associated infections, is an important public health concern. Infections caused by these multidrug-resistant organisms, for which safe and effective antimicrobial therapy options are extremely limited, are associated with poor outcomes for patients. The optimal approach to the treatment of infections caused by these multidrug-resistant strains remains undefined, and treatment decisions for an individual patient should be based on a number of organism- (for example, minimum inhibitory concentration) and patient-specific (for example, site of infection) factors. The emergence of pandrug-resistant strains of K. pneumoniae highlights the critical need for consistent implementation of effective strategies for prevention of transmission and infection and for the development of new antimicrobials with activity against these emerging pathogens.

Middle East respiratory syndrome-associated coronavirus (MERS-CoV) has been a significant research focus since its discovery in 2012. Since 2012, 2,040 cases and 712 deaths have been recorded (as of August 11, 2017), representing a strikingly high case fatality rate of 36%. Over the last several years, MERS-CoV research has progressed in several parallel and complementary directions. This review will focus on three particular areas: the origins and evolution of MERS-CoV, the challenges and achievements in the development of MERS-CoV animal models, and our understanding of how novel proteins unique to MERS-CoV counter the host immune response. The origins of MERS-CoV, likely in African bats, are increasingly clear, although important questions remain about the establishment of dromedary camels as a reservoir seeding human outbreaks. Likewise, there have been important advances in the development of animal models, and both non-human primate and mouse models that seem to recapitulate human disease are now available. How MERS-CoV evades and inhibits the host innate immune response remains less clear. Although several studies have identified MERS-CoV proteins as innate immune antagonists, little of this work has been conducted using live virus under conditions of actual infection, but rather with ectopically expressed proteins. Accordingly, considerable space remains for major contributions to understanding unique ways in which MERS-CoV interacts with and modulates the host response. Collectively, these areas have seen significant advances over the last several years but continue to offer exciting opportunities for discovery.

Ancient whole-genome duplications (WGDs)—paleopolyploidy events—are key to solving Darwin’s ‘abominable mystery’ of how flowering plants evolved and radiated into a rich variety of species. The vertebrates also emerged from their invertebrate ancestors via two WGDs, and genomes of diverse gymnosperm trees, unicellular eukaryotes, invertebrates, fishes, amphibians and even a rodent carry evidence of lineage-specific WGDs. Modern polyploidy is common in eukaryotes, and it can be induced, enabling mechanisms and short-term cost-benefit assessments of polyploidy to be studied experimentally. However, the ancient WGDs can be reconstructed only by comparative genomics: these studies are difficult because the DNA duplicates have been through tens or hundreds of millions of years of gene losses, mutations, and chromosomal rearrangements that culminate in resolution of the polyploid genomes back into diploid ones (rediploidisation). Intriguing asymmetries in patterns of post-WGD gene loss and retention between duplicated sets of chromosomes have been discovered recently, and elaborations of signal transduction systems are lasting legacies from several WGDs. The data imply that simpler signalling pathways in the pre-WGD ancestors were converted via WGDs into multi-stranded parallelised networks. Genetic and biochemical studies in plants, yeasts and vertebrates suggest a paradigm in which different combinations of sister paralogues in the post-WGD regulatory networks are co-regulated under different conditions. In principle, such networks can respond to a wide array of environmental, sensory and hormonal stimuli and integrate them to generate phenotypic variety in cell types and behaviours. Patterns are also being discerned in how the post-WGD signalling networks are reconfigured in human cancers and neurological conditions. It is fascinating to unpick how ancient genomic events impact on complexity, variety and disease in modern life.

Neisseria meningitidis is a harmless commensal bacterium finely adapted to humans. Unfortunately, under “privileged” conditions, it adopts a “devious” lifestyle leading to uncontrolled behavior characterized by the unleashing of molecular weapons causing potentially lethal disease such as sepsis and acute meningitis. Indeed, despite the lack of a classic repertoire of virulence genes in N. meningitidis separating commensal from invasive strains, molecular epidemiology and functional genomics studies suggest that carriage and invasive strains belong to genetically distinct populations characterized by an exclusive pathogenic potential. In the last few years, “omics” technologies have helped scientists to unwrap the framework drawn by N. meningitidis during different stages of colonization and disease. However, this scenario is still incomplete and would benefit from the implementation of physiological tissue models for the reproduction of mucosal and systemic interactions in vitro. These emerging technologies supported by recent advances in the world of stem cell biology hold the promise for a further understanding of N. meningitidis pathogenesis.

The bacterial pathogen Listeria monocytogenes (Lm) is the causative agent of listeriosis, a rare but fatal foodborne disease. During infection, Lm can traverse several host barriers and enter the cytosol of a variety of cell types. Thus, consideration of the extracellular and intracellular niches of Lm is critical for understanding the infection process. Here, we review advances in our understanding of Lm infection and highlight how the interactions between the host and the pathogen are context dependent. We discuss discoveries of how Lm senses entry into the host cell cytosol. We present findings concerning how the nature of the various cytoskeleton components subverted by Lm changes depending on both the stage of infection and the subcellular context. We present discoveries of critical components required for Lm traversal of physiological barriers. Interactions between the host gut microbiota and Lm will be briefly discussed. Finally, the importance of Lm biodiversity and post-genomics approaches as a promising way to discover novel virulence factors will be highlighted.

The versatile and ubiquitous Pseudomonas aeruginosa is an opportunistic pathogen causing acute and chronic infections in predisposed human subjects. Here we review recent progress in understanding P. aeruginosa population biology and virulence, its cyclic di-GMP-mediated switches of lifestyle, and its interaction with the mammalian host as well as the role of the type III and type VI secretion systems in P. aeruginosa infection.

Global change is altering oceanic temperature, salinity, pH, and oxygen concentration, directly and indirectly influencing marine microbial food web structure and function. As microbes represent >90% of the ocean’s biomass and are major drivers of biogeochemical cycles, understanding their responses to such changes is fundamental for predicting the consequences of global change on ecosystem functioning. Recent findings indicate that marine archaea and archaeal viruses are active and relevant components of marine microbial assemblages, far more abundant and diverse than was previously thought. Further research is urgently needed to better understand the impacts of global change on virus–archaea dynamics and how archaea and their viruses can interactively influence the ocean’s feedbacks on global change.

A functional comparative genome analysis is essential to understand the mechanisms underlying bacterial evolution and adaptation. Detection of functional orthologs using standard global sequence similarity methods faces several problems; the need for defining arbitrary acceptance thresholds for similarity and alignment length, lateral gene acquisition and the high computational cost for finding bi-directional best matches at a large scale. We investigated the use of protein domain architectures for large scale functional comparative analysis as an alternative method. The performance of both approaches was assessed through functional comparison of 446 bacterial genomes sampled at different taxonomic levels. We show that protein domain architectures provide a fast and efficient alternative to methods based on sequence similarity to identify groups of functionally equivalent proteins within and across taxonomic boundaries, and it is suitable for large scale comparative analysis. Running both methods in parallel pinpoints potential functional adaptations that may add to bacterial fitness.

Multidimensional gradients of inorganic compounds influence microbial activity in diverse pristine and anthropogenically perturbed environments. Here, we suggest that high-throughput cultivation and genetics can be systematically applied to generate quantitative models linking gene function, microbial community activity, and geochemical parameters. Metal resistance determinants represent a uniquely universal set of parameters around which to study and evaluate microbial fitness because they represent a record of the environment in which all microbial life evolved. By cultivating microbial isolates and enrichments in laboratory gradients of inorganic ions, we can generate quantitative predictions of limits on microbial range in the environment, obtain more accurate gene annotations, and identify useful strategies for predicting and engineering the trajectory of natural ecosystems.

With the advent of very rapid and cheap genome analyses and the linkage of these plus microbial metabolomics to potential compound structures came the realization that there was an immense sea of novel agents to be mined and tested. In addition, it is now recognized that there is significant microbial involvement in many natural products isolated from “nominally non-microbial sources”. This short review covers the current screening methods that have evolved and one might even be tempted to say “devolved” in light of the realization that target-based screens had problems when the products entered clinical testing, with off-target effects being the major ones. Modern systems include, but are not limited to, screening in cell lines utilizing very modern techniques (a high content screen) that are designed to show interactions within cells when treated with an “agent”. The underlying principle(s) used in such systems dated back to unpublished attempts in the very early 1980s by the pharmaceutical industry to show toxic interactions within animal cells by using automated light microscopy. Though somewhat successful, the technology was not adequate for any significant commercialization. Somewhat later, mammalian cell lines that were “genetically modified” to alter signal transduction cascades, either up or down, and frequently linked to luciferase readouts, were then employed in a 96-well format. In the case of microbes, specific resistance parameters were induced in isogenic cell lines from approximately the mid-1970s. In the latter two cases, comparisons against parent and sibling cell lines were used in order that a rapid determination of potential natural product “hits” could be made. Obviously, all of these assay systems could also be, and were, used for synthetic molecules. These methods and their results have led to a change in what the term “screening for bioactivity” means. In practice, versions of phenotypic screening are returning, but in a dramatically different scientific environment from the 1970s, as I hope to demonstrate in the short article that follows.

Microbial fermentation has been used historically for the preservation of foods, the health benefits of which have since come to light. Early dairy fermentations depended on the spontaneous activity of the indigenous microbiota of the milk. Modern fermentations rely on defined starter cultures with desirable characteristics to ensure consistency and commercial viability. The selection of defined starters depends on specific phenotypes that benefit the product by guaranteeing shelf life and ensuring safety, texture, and flavour. Lactic acid bacteria can produce a number of bioactive metabolites during fermentation, such as bacteriocins, biogenic amines, exopolysaccharides, and proteolytically released peptides, among others. Prebiotics are added to food fermentations to improve the performance of probiotics. It has also been found that prebiotics fermented in the gut can have benefits that go beyond helping probiotic growth. Studies are now looking at how the fermentation of prebiotics such as fructo-oligosaccharides can help in the prevention of diseases such as osteoporosis, obesity, and colorectal cancer. The potential to prevent or even treat disease through the fermentation of food is a medically and commercially attractive goal and is showing increasing promise. However, the stringent regulation of probiotics is beginning to detrimentally affect the field and limit their application.

Pneumocystis jirovecii is an unusual ascomycetous fungus that can be detected in the lungs of healthy individuals. Transmission from human to human is one of its main characteristics in comparison with other fungi responsible for invasive infections. P. jirovecii is transmitted through the air between healthy individuals, who are considered to be the natural reservoir, at least transiently. In immunocompromised patients, P. jirovecii multiplies, leading to subacute infections and acute life-threatening pneumonia, called Pneumocystis pneumonia [PCP]. PCP is caused by genotypically distinct mixtures of organisms in more than 90% of cases, reinforcing the hypothesis that there is constant inhalation of P. jirovecii from different contacts over time, although reactivation of latent organisms from previous exposures may be possible. Detection of P. jirovecii DNA without any symptoms or related radiological signs has been called “colonization”. This situation could be considered as the result of recent exposure to P. jirovecii that could evolve towards PCP, raising the issue of cotrimoxazole prophylaxis for at-risk quantitative polymerase chain reaction (qPCR)-positive immunocompromised patients. The more accurate way to diagnose PCP is the use of real-time quantitative PCR, which prevents amplicon contamination and allows determination of the fungal load that is mandatory to interpret the qPCR results and manage the patient appropriately. The detection of P. jirovecii in respiratory samples of immunocompromised patients should be considered for potential risk of developing PCP. Many challenges still need to be addressed, including a better description of transmission, characterization of organisms present at low level, and prevention of environmental exposure during immunodepression.

The discovery and adaption of bacterial clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated (Cas) systems has revolutionized the way researchers edit genomes. Engineering of catalytically inactivated Cas variants (nuclease-deficient or nuclease-deactivated [dCas]) combined with transcriptional repressors, activators, or epigenetic modifiers enable sequence-specific regulation of gene expression and chromatin state. These CRISPR–Cas-based technologies have contributed to the rapid development of disease models and functional genomics screening approaches, which can facilitate genetic target identification and drug discovery. In this short review, we will cover recent advances of CRISPR–dCas9 systems and their use for transcriptional repression and activation, epigenome editing, and engineered synthetic circuits for complex control of the mammalian genome.

Despite their inherent toxicity and the global spread of bacterial resistance, aminoglycosides (AGs), an old class of microbial drugs, remain a valuable component of the antibiotic arsenal. Recent studies have continued to reveal the fascinating biochemistry of AG biosynthesis and the rich potential in their pathway engineering. In particular, parallel pathways have been shown to be common and widespread in AG biosynthesis, highlighting nature’s ingenuity in accessing diverse natural products from a limited set of genes. In this review, we discuss the parallel biosynthetic pathways of three representative AG antibiotics—kanamycin, gentamicin, and apramycin—as well as future directions towards the discovery and development of novel AGs.

The human immunodeficiency virus (HIV) evolves rapidly owing to the combined activity of error-prone reverse transcriptase, recombination, and short generation times, leading to extensive viral diversity both within and between hosts. This diversity is a major contributing factor in the failure of the immune system to eradicate the virus and has important implications for the development of suitable drugs and vaccines to combat infection. This review will discuss the recent technological advances that have shed light on HIV evolution and will summarise emerging concepts in this field.

A single extra-intestinal pathogenic Escherichia coli (ExPEC) clone, named sequence type (ST) 131, is responsible for millions of global antimicrobial-resistant (AMR) infections annually. Population genetics indicate that ST131 consists of different clades (i.e. A, B, and C); however, clade C is the most dominant globally. A ST131 subclade, named C1-M27, is emerging in Japan and has been responsible for the recent increase in AMR ExPEC in that country. The sequential acquisition of several virulence and AMR genes associated with mobile genetic elements during the 1960s to 1980s primed clade C (and its subclades C1 and C2) for success in the 1990s to 2000s. IncF plasmids with F1:A2:B20 and F2:A1:B replicons have shaped the evolution of the C1 and C2 subclades. It is possible that ST131 is a host generalist with different accessory gene profiles. Compensatory mutations within the core genome of this clone have counterbalanced the fitness cost associated with IncF plasmids. ST131 clade C had dramatically changed the population structure of ExPEC, but it still remains unclear which features of this clade resulted in one of the most unprecedented AMR successes of the 2000s.

Invasive aspergillosis (IA) has been increasingly reported in populations other than the historical hematology patients and there are new questions about the performance of microbiological tools. Microscopy and culture have been completed by biomarkers, either antigens or DNA, and in blood or respiratory specimens or both. First studied in hematology, the antigen galactomannan performance in serum is low in other patient populations where the pathophysiology of the infection can be different and the prevalence of IA is much lower. DNA detection with polymerase chain reaction (PCR) in blood or serum (or both) has reached a certain level of acceptance thanks to consensus methods based on real-time quantitative PCR (qPCR). When used on respiratory specimens, galactomannan and qPCR depend on standardization of the sampling and the diverse mycological procedures. Thus, culture remains the main diagnostic criterion in critically ill patients. The current trend toward more effective anti-mold prophylaxis in hematology hampers the yield of a screening strategy, as is usually performed in hematology. Therefore, circulating biomarkers as confirmatory tests should be considered and their performance should be reappraised in each new setting. The use of azole prophylaxis also raises the issue of selecting azole-resistance Aspergillus fumigatus isolates. Ideally, the biomarkers will be more efficient when individual genetic risks of IA are defined. Culture, though not standardized, remains a key element for the diagnosis of IA and has the advantage to easily detect molds other than A. fumigatus. It is still unclear whether next-generation sequencing will replace culture in the future.

Congenital cytomegalovirus (CMV) remains a leading cause of disability in children. Understanding the pathogenesis of infection from the mother via the placenta to the neonate is crucial if we are to produce new interventions and provide supportive mechanisms to improve the outcome of congenitally infected children. In recent years, some major goals have been achieved, including the diagnosis of primary maternal CMV infection in pregnant women by using the anti-CMV IgG avidity test and the diagnosis and prognosis of foetal CMV infection by using polymerase chain reaction real-time tests to detect and quantify the virus in amniotic fluid. This review summarises recent advances in our understanding and highlights where challenges remain, especially in vaccine development and anti-viral therapy of the pregnant woman and the neonate. Currently, no therapeutic options during pregnancy are available except those undergoing clinical trials, whereas valganciclovir treatment is recommended for congenitally infected neonates with moderately to severely symptomatic disease.

In the past century, advances in antibiotics and vaccination have dramatically altered the incidence and clinical outcomes of bacterial meningitis. We review the shifting epidemiology of meningitis in children, including after the implementation of vaccines that target common meningitic pathogens and the introduction of intrapartum antibiotic prophylaxis offered to mothers colonized with Streptococcus agalactiae. We also discuss what is currently known about the pathogenesis of meningitis. Recent studies of the human microbiome have illustrated dynamic relationships of bacterial and viral populations with the host, which may potentiate the risk of bacterial meningitis.

Antibiotics have been widely used for a number of decades for human therapy and farming production. Since a high percentage of antibiotics are discharged from the human or animal body without degradation, this means that different habitats, from the human body to river water or soils, are polluted with antibiotics. In this situation, it is expected that the variable concentration of this type of microbial inhibitor present in different ecosystems may affect the structure and the productivity of the microbiota colonizing such habitats. This effect can occur at different levels, including changes in the overall structure of the population, selection of resistant organisms, or alterations in bacterial physiology. In this review, I discuss the available information on how the presence of antibiotics may alter the microbiota and the consequences of such alterations for human health and for the activity of microbiota from different habitats.

Filamentous fungi have proven to be a better-suited model system than unicellular yeasts in analyses of cellular processes such as polarized growth, exocytosis, endocytosis, and cytoskeleton-based organelle traffic. For example, the filamentous fungus Neurospora crassa develops a variety of cellular forms. Studying the molecular basis of these forms has led to a better, yet incipient, understanding of polarized growth. Polarity factors as well as Rho GTPases, septins, and a localized delivery of vesicles are the central elements described so far that participate in the shift from isotropic to polarized growth. The growth of the cell wall by apical biosynthesis and remodeling of polysaccharide components is a key process in hyphal morphogenesis. The coordinated action of motor proteins and Rab GTPases mediates the vesicular journey along the hyphae toward the apex, where the exocyst mediates vesicle fusion with the plasma membrane. Cytoplasmic microtubules and actin microfilaments serve as tracks for the transport of vesicular carriers as well as organelles in the tubular cell, contributing to polarization. In addition to exocytosis, endocytosis is required to set and maintain the apical polarity of the cell. Here, we summarize some of the most recent breakthroughs in hyphal morphogenesis and apical growth in N. crassa and the emerging questions that we believe should be addressed.

About 2,500 papers dated 2014–2016 were recovered by searching the PubMed database for Streptomyces, which are the richest known source of antibiotics. This review integrates around 100 of these papers in sections dealing with evolution, ecology, pathogenicity, growth and development, stress responses and secondary metabolism, gene expression, and technical advances. Genomic approaches have greatly accelerated progress. For example, it has been definitively shown that interspecies recombination of conserved genes has occurred during evolution, in addition to exchanges of some of the tens of thousands of non-conserved accessory genes. The closeness of the association of Streptomyces with plants, fungi, and insects has become clear and is reflected in the importance of regulators of cellulose and chitin utilisation in overall Streptomyces biology. Interestingly, endogenous cellulose-like glycans are also proving important in hyphal growth and in the clumping that affects industrial fermentations. Nucleotide secondary messengers, including cyclic di-GMP, have been shown to provide key input into developmental processes such as germination and reproductive growth, while late morphological changes during sporulation involve control by phosphorylation. The discovery that nitric oxide is produced endogenously puts a new face on speculative models in which regulatory Wbl proteins (peculiar to actinobacteria) respond to nitric oxide produced in stressful physiological transitions. Some dramatic insights have come from a new model system for Streptomyces developmental biology, Streptomyces venezuelae, including molecular evidence of very close interplay in each of two pairs of regulatory proteins. An extra dimension has been added to the many complexities of the regulation of secondary metabolism by findings of regulatory crosstalk within and between pathways, and even between species, mediated by end products. Among many outcomes from the application of chromosome immunoprecipitation sequencing (ChIP-seq) analysis and other methods based on “next-generation sequencing” has been the finding that 21% of Streptomyces mRNA species lack leader sequences and conventional ribosome binding sites. Further technical advances now emerging should lead to continued acceleration of knowledge, and more effective exploitation, of these astonishing and critically important organisms.

Mitochondria are double membrane organelles of endosymbiotic origin, best known for constituting the centre of energetics of a eukaryotic cell. They contain their own mitochondrial genome, which as a consequence of gradual reduction during evolution typically contains less than two dozens of genes. In this review, we highlight the extremely diverse architecture of mitochondrial genomes and mechanisms of gene expression between the three sister groups constituting the phylum Euglenozoa - Euglenida, Diplonemea and Kinetoplastea. The earliest diverging euglenids possess a simplified mitochondrial genome and a conventional gene expression, whereas both are highly complex in the two other groups. The expression of their mitochondrial-encoded proteins requires extensive post-transcriptional modifications guided by complex protein machineries and multiple small RNA molecules. Moreover, the least studied diplonemids, which have been recently discovered as a highly abundant component of the world ocean plankton, possess one of the most complicated mitochondrial genome organisations known to date.

High-throughput sequencing of PCR-amplified taxonomic markers (like the 16S rRNA gene) has enabled a new level of analysis of complex bacterial communities known as microbiomes. Many tools exist to quantify and compare abundance levels or OTU composition of communities in different conditions. The sequencing reads have to be denoised and assigned to the closest taxa from a reference database. Common approaches use a notion of 97% similarity and normalize the data by subsampling to equalize library sizes. In this paper, we show that statistical models allow more accurate abundance estimates. By providing a complete workflow in R, we enable the user to do sophisticated downstream statistical analyses, whether parametric or nonparametric. We provide examples of using the R packages dada2, phyloseq, DESeq2, ggplot2 and vegan to filter, visualize and test microbiome data. We also provide examples of supervised analyses using random forests and nonparametric testing using community networks and the ggnetwork package.

A functional comparative genome analysis is essential to understand the mechanisms underlying bacterial evolution and adaptation. Detection of functional orthologs using standard global sequence similarity methods faces several problems; the need for defining arbitrary acceptance thresholds for similarity and alignment length, lateral gene acquisition and the high computational cost for finding bi-directional best matches at a large scale. We investigated the use of protein domain architectures for large scale functional comparative analysis as an alternative method. The performance of both approaches was assessed through functional comparison of 446 bacterial genomes sampled at different taxonomic levels. We show that protein domain architectures provide a fast and efficient alternative to methods based on sequence similarity to identify groups of functionally equivalent proteins within and across taxonomic boundaries. As the computational cost scales linearly, and not quadratically with the number of genomes, it is suitable for large scale comparative analysis. Running both methods in parallel pinpoints potential functional adaptations that may add to bacterial fitness.

Probiotics are “live microorganisms which, when consumed in adequate amounts, confer a health benefit to the host”. A number of attributes are highly sought after among these microorganisms, including immunomodulation, epithelial barrier maintenance, competitive exclusion, production of short-chain fatty acids, and bile salt metabolism. Bacteriocin production is also generally regarded as a probiotic trait, but it can be argued that, in contrast to other traits, it is often considered a feature that is desirable, rather than a key probiotic trait. As such, the true potential of these antimicrobials has yet to be realised.

Candida albicans is an important human fungal pathogen, in terms of both its clinical significance and its use as an experimental model for scientific investigation. Although this opportunistic pathogen is a natural component of the human flora, it can cause life-threatening infections in immunosuppressed patients. There are currently a limited number of antifungal molecules and drug targets, and increasing resistance to the front-line therapeutics, demonstrating a clear need for new antifungal drugs. Understanding the biology of this pathogen is an important prerequisite for identifying new drug targets for antifungal therapeutics. In this review, we highlight some recent developments that help us to understand how virulence traits are regulated at the molecular level, in addition to technical advances that improve the ability of genome editing in C. albicans.

In recent years, substantial progress has been made in understanding the molecular and cell biology of the human parasite Entamoeba histolytica, an important pathogen with significant global impact. This review outlines some recent advances in the Entamoeba field in the last five years, focusing on areas that have not recently been discussed in detail: (i) molecular mechanisms regulating parasite gene expression, (ii) new efforts at drug discovery using high-throughput drug screens, and (iii) the effect of gut microbiota on amoebiasis.

Modern society is fueled by fossil energy produced millions of years ago by photosynthetic organisms. Cultivating contemporary photosynthetic producers to generate energy and capture carbon from the atmosphere is one potential approach to sustaining society without disrupting the climate. Algae, photosynthetic aquatic microorganisms, are the fastest growing primary producers in the world and can therefore produce more energy with less land, water, and nutrients than terrestrial plant crops. We review recent progress and challenges in developing bioenergy technology based on algae. A variety of high-value products in addition to biofuels can be harvested from algal biomass, and these may be key to developing algal biotechnology and realizing the commercial potential of these organisms. Aspects of algal biology that differentiate them from plants demand an integrative approach based on genetics, cell biology, ecology, and evolution. We call for a systems approach to research on algal biotechnology rooted in understanding their biology, from the level of genes to ecosystems, and integrating perspectives from physical, chemical, and social sciences to solve one of the most critical outstanding technological problems.

Leishmania has a plastic genome, and drug pressure can select for gene copy number variation (CNV). CNVs can apply either to whole chromosomes, leading to aneuploidy, or to specific genomic regions. For the latter, the amplification of chromosomal regions occurs at the level of homologous direct or inverted repeated sequences leading to extrachromosomal circular or linear amplified DNAs. This ability of Leishmania to respond to drug pressure by CNVs has led to the development of genomic screens such as Cos-Seq, which has the potential of expediting the discovery of drug targets for novel promising drug candidates.

How and when eukaryotic cells make the irrevocable commitment to divide remain central questions in the cell-cycle field. Parallel studies in yeast and mammalian cells seemed to suggest analogous control mechanisms operating during the G1 phase—at Start or the restriction (R) point, respectively—to integrate nutritional and developmental signals and decide between distinct cell fates: cell-cycle arrest or exit versus irreversible commitment to a round of division. Recent work has revealed molecular mechanisms underlying this decision-making process in both yeast and mammalian cells but also cast doubt on the nature and timing of cell-cycle commitment in multicellular organisms. These studies suggest an expanded temporal window of mitogen sensing under certain growth conditions, illuminate unexpected obstacles and exit ramps on the path to full cell-cycle commitment, and raise new questions regarding the functions of cyclin-dependent kinases (CDKs) that drive G1 progression and S-phase entry.

In this study, we interpreted RNA-seq time-course data of three developmental stages of Plasmodium species by clustering genes based on similarities in their expression profile without prior knowledge of the gene function. Functional enrichment of clusters of upregulated genes at specific time-points reveals potential targetable biological processes with information on their timings. We identified common consensus sequences that these clusters shared as potential points of coordinated transcriptional control. Five cluster groups showed upregulated profile patterns of biological interest. This included two clusters from the Intraerythrocytic Developmental Cycle (cluster 4 = 16 genes, and cluster 9 = 32 genes), one from the sexual development stage (cluster 2 = 851 genes), and two from the gamete-fertilization stage in the mosquito host (cluster 4 = 153 genes, and cluster 9 = 258 genes). The IDC expressed the least numbers of genes with only 1448 genes showing any significant activity of the 5020 genes (~29%) in the experiment. Gene ontology (GO) enrichment analysis of these clusters revealed a total of 671 uncharacterized genes implicated in 14 biological processes and components associated with these stages, some of which are currently being investigated as drug targets in on-going research. Five putative transcription regulatory binding motifs shared by members of each cluster were also identified, one of which was also identified in a previous study by separate researchers. Our study shows stage-specific genes and biological processes that may be important in antimalarial drug research efforts. In addition, timed-coordinated control of separate processes may explain the paucity of factors in parasites.

The field of herpes simplex virus (HSV) latency and reactivation has been marked by controversy, which is not unexpected considering the complexities of the biology involved. While controversy is an important tool for digging to the bottom of difficult issues, we propose that unproductive conflict in the field arises in part from poorly defined terminology and the need for a collective framework. The uses of advanced global molecular and next-generation sequencing approaches and an increasing array of in vitro model systems have provided new molecular-level insights into HSV latency and reactivation, with the promise of expanding our concepts of these processes. However, our current framework and language are inadequate to effectively integrate new data streams into the established theories. In this brief perspective, we look back into the past to examine when and how the lexicon of HSV latency and reactivation arose in the literature and its evolution. We propose to open a dialogue among investigators for the purpose of updating and clearly defining terms used to describe these processes and to build a collective integrated framework to move our field forward.

A functional comparative genome analysis is essential to understand the mechanisms underlying bacterial evolution and adaptation. Detection of functional orthologs using standard global sequence similarity methods faces several problems; the need for defining arbitrary acceptance thresholds for similarity and alignment length, lateral gene acquisition and the high computational cost for finding bi-directional best matches at a large scale. We investigated the use of protein domain architectures for large scale functional comparative analysis as an alternative method. The performance of both approaches was assessed through functional comparison of 446 bacterial genomes sampled at different taxonomic levels. We show that protein domain architectures provide a fast and efficient alternative to methods based on sequence similarity to identify groups of functionally equivalent proteins within and across taxonomic bounderies. As the computational cost scales linearly, and not quadratically with the number of genomes, it is suitable for large scale comparative analysis. Running both methods in parallel pinpoints potential functional adaptations that may add to bacterial fitness.

The ability of the pathogenic fungus Cryptococcus neoformans to cause life-threatening meningoencephalitis in immunocompromised individuals is due in large part to elaboration of a capsule consisting of polysaccharide fibers. The size of the cell-associated capsule is remarkably responsive to a variety of environmental and host conditions, but the mechanistic details of the regulation, synthesis, trafficking, and attachment of the polysaccharides are poorly understood. Recent studies reveal a complex network of transcription factors that influence capsule elaboration in response to several different signals of relevance to disease (e.g., iron deprivation). The emerging complexity of the network is consistent with the diversity of conditions that influence the capsule and illustrates the responsiveness of the fungus to both the environment and mammalian hosts.

The wide spread of gene exchange and loss in the prokaryotic world has prompted the concept of ‘lateral genomics’ to the point of an outright denial of the relevance of phylogenetic trees for evolution. However, the pronounced coherence congruence of the topologies of numerous gene trees, particularly those for (nearly) universal genes, translates into the notion of a statistical tree of life (STOL), which reflects a central trend of vertical evolution. The STOL can be employed as a framework for reconstruction of the evolutionary processes in the prokaryotic world. Quantitatively, however, horizontal gene transfer (HGT) dominates microbial evolution, with the rate of gene gain and loss being comparable to the rate of point mutations and much greater than the duplication rate. Theoretical models of evolution suggest that HGT is essential for the survival of microbial populations that otherwise deteriorate due to the Muller’s ratchet effect. Apparently, at least some bacteria and archaea evolved dedicated vehicles for gene transfer that evolved from selfish elements such as plasmids and viruses. Recent phylogenomic analyses suggest that episodes of massive HGT were pivotal for the emergence of major groups of organisms such as multiple archaeal phyla as well as eukaryotes. Similar analyses appear to indicate that, in addition to donating hundreds of genes to the emerging eukaryotic lineage, mitochondrial endosymbiosis severely curtailed HGT. These results shed new light on the routes of evolutionary transitions, but caution is due given the inherent uncertainty of deep phylogenies.

High-throughput sequencing of PCR-amplified taxonomic markers (like the 16S rRNA gene) has enabled a new level of analysis of complex bacterial communities known as microbiomes. Many tools exist to quantify and compare abundance levels or microbial composition of communities in different conditions. The sequencing reads have to be denoised and assigned to the closest taxa from a reference database. Common approaches use a notion of 97% similarity and normalize the data by subsampling to equalize library sizes. In this paper, we show that statistical models allow more accurate abundance estimates. By providing a complete workflow in R, we enable the user to do sophisticated downstream statistical analyses, including both parameteric and nonparametric methods. We provide examples of using the R packages dada2, phyloseq, DESeq2, ggplot2 and vegan to filter, visualize and test microbiome data. We also provide examples of supervised analyses using random forests, partial least squares and linear models as well as nonparametric testing using community networks and the ggnetwork package.

Intracellular single-celled parasites belonging to the large phylum Apicomplexa are amongst the most prevalent and morbidity-causing pathogens worldwide. In this review, we highlight a few of the many recent advances in the field that helped to clarify some important aspects of their fascinating biology and interaction with their hosts. Plasmodium falciparum causes malaria, and thus the recent emergence of resistance against the currently used drug combinations based on artemisinin has been of major interest for the scientific community. It resulted in great advances in understanding the resistance mechanisms that can hopefully be translated into altered future drug regimens. Apicomplexa are also experts in host cell manipulation and immune evasion. Toxoplasma gondii and Theileria sp., besides Plasmodium sp., are species that secrete effector molecules into the host cell to reach this aim. The underlying molecular mechanisms for how these proteins are trafficked to the host cytosol (T. gondii and Plasmodium) and how a secreted protein can immortalize the host cell (Theileria sp.) have been illuminated recently. Moreover, how such secreted proteins affect the host innate immune responses against T. gondii and the liver stages of Plasmodium has also been unraveled at the genetic and molecular level, leading to unexpected insights. Methodological advances in metabolomics and molecular biology have been instrumental to solving some fundamental puzzles of mitochondrial carbon metabolism in Apicomplexa. Also, for the first time, the generation of stably transfected Cryptosporidium parasites was achieved, which opens up a wide variety of experimental possibilities for this understudied, important apicomplexan pathogen.

Salmonella enterica serovar Typhimurium is a foodborne pathogen causing inflammatory disease in the intestine following diarrhea and is responsible for thousands of deaths worldwide. Many in vitro investigations using cell culture models are available, but these do not represent the real natural environment present in the intestine of infected hosts. Several in vivo animal models have been used to study the host-pathogen interaction and to unravel the immune responses and cellular processes occurring during infection. An animal model for Salmonella-induced intestinal inflammation relies on the pretreatment of mice with streptomycin. This model is of great importance but still shows limitations to investigate the host-pathogen interaction in the small intestine in vivo. Here, we review the use of mouse models for Salmonella infections and focus on a new small animal model using 1-day-old neonate mice. The neonate model enables researchers to observe infection of both the small and large intestine, thereby offering perspectives for new experimental approaches, as well as to analyze the Salmonella-enterocyte interaction in the small intestine in vivo.

In the 1990s several biocontrol agents on that contained Burkholderia strains were registered by the United States Environmental Protection Agency (EPA). After risk assessment these products were withdrawn from the market and a moratorium was placed on the registration of Burkholderia-containing products, as these strains may pose a risk to human health. However, over the past few years the number of novel Burkholderia species that exhibit plant-beneficial properties and are normally not isolated from infected patients has increased tremendously. In this commentary we wish to summarize recent efforts that aim at discerning pathogenic from beneficial Burkholderia strains.

Mitochondria are double membrane organelles of endosymbiotic origin, best known for constituting the centre of energetics of a eukaryotic cell. They contain their own mitochondrial genome, which as a consequence of gradual reduction during evolution typically contains less than two dozens of genes. In this review, we highlight the extremely diverse architecture of mitochondrial genomes and mechanisms of gene expression between the three sister groups constituting the phylum Euglenozoa - Euglenida, Diplonemea and Kinetoplastea. The earliest diverging euglenids possess a simplified mitochondrial genome and a conventional gene expression, whereas both are highly complex in the two other groups. The expression of their mitochondrial-encoded proteins requires extensive post-transcriptional modifications guided by complex protein machineries and multiple small RNA molecules. Moreover, the least studied diplonemids, which have been recently discovered as a highly abundant component of the world ocean plankton, possess one of the most complicated mitochondrial genome organisations known to date.

Biotechnology has almost unlimited potential to change our lives in very exciting ways. Many of the chemical reactions that produce these products can be fully optimized by performing them at extremes of temperature, pressure, salinity, and pH for efficient and cost-effective outcomes. Fortunately, there are many organisms (extremophiles) that thrive in extreme environments found in nature and offer an excellent source of replacement enzymes in lieu of mesophilic ones currently used in these processes. In this review, I discuss the current uses and some potential new applications of extremophiles and their products, including enzymes, in biotechnology.

In this perspective, we highlight recent examples and trends in metabolic engineering and synthetic biology that demonstrate the synthetic potential of enzyme and pathway engineering for natural product discovery. In doing so, we introduce natural paradigms of secondary metabolism whereby simple carbon substrates are combined into complex molecules through “scaffold diversification”, and subsequent “derivatization” of these scaffolds is used to synthesize distinct complex natural products. We provide examples in which modern pathway engineering efforts including combinatorial biosynthesis and biological retrosynthesis can be coupled to directed enzyme evolution and rational enzyme engineering to allow access to the “privileged” chemical space of natural products in industry-proven microbes. Finally, we forecast the potential to produce natural product-like discovery platforms in biological systems that are amenable to single-step discovery, validation, and synthesis for streamlined discovery and production of biologically active agents.

Plastids and mitochondria derive from prokaryotic symbionts that lost most of their genes after the establishment of endosymbiosis. In consequence, relatively few of the thousands of different proteins in these organelles are actually encoded there. Most are now specified by nuclear genes. The most direct way to reconstruct the evolutionary history of plastids and mitochondria is to sequence and analyze their relatively small genomes. However, understanding the functional diversification of these organelles requires the identification of their complete protein repertoires – which is the ultimate goal of organellar proteomics. In the meantime, judicious combination of proteomics-based data with analyses of nuclear genes that include interspecies comparisons and/or predictions of subcellular location is the method of choice. Such genome-wide approaches can now make use of the entire sequences of plant nuclear genomes that have emerged since 2000. Here I review the results of these attempts to reconstruct the evolution and functions of plant DNA-containing organelles, focusing in particular on data from nuclear genomes. In addition, I discuss proteomic approaches to the direct identification of organellar proteins and briefly refer to ongoing research on non-coding nuclear DNAs of organellar origin (specifically, nuclear mitochondrial DNA and nuclear plastid DNA).

Microbes are now known to participate in an extensive repertoire of cooperative behaviors such as biofilm formation, production of extracellular public-goods, group motility, and higher-ordered multicellular structures. A fundamental question is how these cooperative tasks are maintained in the face of non-cooperating defector cells. Recently, a number of molecular mechanisms including facultative participation, spatial sorting, and policing have been discovered to stabilize cooperation. Often these different mechanisms work in concert to reinforce cooperation. In this review, we describe bacterial cooperation and the current understanding of the molecular mechanisms that maintain it.