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July 7, 2019

Persistence of a dominant bovine lineage of group B Streptococcus reveals genomic signatures of host adaptation.

Group B Streptococcus (GBS) is a host-generalist species, most notably causing disease in humans and cattle. However, the differential adaptation of GBS to its two main hosts, and the risk of animal to human infection remain poorly understood. Despite improvements in control measures across Europe, GBS is still one of the main causative agents of bovine mastitis in Portugal. Here, by whole-genome analysis of 150 bovine GBS isolates we discovered that a single CC61 clone is spreading throughout Portuguese herds since at least the early 1990s, having virtually replaced the previous GBS population. Mutations within an iron/manganese transporter were independently acquired by all of the CC61 isolates, underlining a key adaptive strategy to persist in the bovine host. Lateral transfer of bacteriocin production and antibiotic resistance genes also underscored the contribution of the microbial ecology and genetic pool within the bovine udder environment to the success of this clone. Compared to strains of human origin, GBS evolves twice as fast in bovines and undergoes recurrent pseudogenizations of human-adapted traits. Our work provides new insights into the potentially irreversible adaptation of GBS to the bovine environment. © 2016 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

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July 7, 2019

Neuraminidase A-exposed galactose promotes Streptococcus pneumoniae biofilm formation during colonization.

Streptococcus pneumoniae is an opportunistic pathogen that colonizes the nasopharynx. Herein we show that carbon availability is distinct between the nasopharynx and bloodstream of adult humans: glucose is absent from the nasopharynx, whereas galactose is abundant. We demonstrate that pneumococcal neuraminidase A (NanA), which cleaves terminal sialic acid residues from host glycoproteins, exposed galactose on the surface of septal epithelial cells, thereby increasing its availability during colonization. We observed that S. pneumoniae mutants deficient in NanA and ß-galactosidase A (BgaA) failed to form biofilms in vivo despite normal biofilm-forming abilities in vitro Subsequently, we observed that glucose, sucrose, and fructose were inhibitory for biofilm formation, whereas galactose, lactose, and low concentrations of sialic acid were permissive. Together these findings suggested that the genes involved in biofilm formation were under some form of carbon catabolite repression (CCR), a regulatory network in which genes involved in the uptake and metabolism of less-preferred sugars are silenced during growth with preferred sugars. Supporting this notion, we observed that a mutant deficient in pyruvate oxidase, which converts pyruvate to acetyl-phosphate under non-CCR-inducing growth conditions, was unable to form biofilms. Subsequent comparative transcriptome sequencing (RNA-seq) analyses of planktonic and biofilm-grown pneumococci showed that metabolic pathways involving the conversion of pyruvate to acetyl-phosphate and subsequently leading to fatty acid biosynthesis were consistently upregulated during diverse biofilm growth conditions. We conclude that carbon availability in the nasopharynx impacts pneumococcal biofilm formation in vivo Additionally, biofilm formation involves metabolic pathways not previously appreciated to play an important role. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019

Transfer of the methicillin resistance genomic island among staphylococci by conjugation.

Methicillin resistance creates a major obstacle for treatment of Staphylococcus aureus infections. The resistance gene, mecA, is carried on a large (20 kb to?>?60 kb) genomic island, staphylococcal cassette chromosome mec (SCCmec), that excises from and inserts site-specifically into the staphylococcal chromosome. However, although SCCmec has been designated a mobile genetic element, a mechanism for its transfer has not been defined. Here we demonstrate the capture and conjugative transfer of excised SCCmec. SCCmec was captured on pGO400, a mupirocin-resistant derivative of the pGO1/pSK41 staphylococcal conjugative plasmid lineage, and pGO400::SCCmec (pRM27) was transferred by filter-mating into both homologous and heterologous S. aureus recipients representing a range of clonal complexes as well as S. epidermidis. The DNA sequence of pRM27 showed that SCCmec had been transferred in its entirety and that its capture had occurred by recombination between IS257/431 elements present on all SCCmec types and pGO1/pSK41 conjugative plasmids. The captured SCCmec excised from the plasmid and inserted site-specifically into the chromosomal att site of both an isogenic S. aureus and a S. epidermidis recipient. These studies describe a means by which methicillin resistance can be environmentally disseminated and a novel mechanism, IS-mediated recombination, for the capture and conjugative transfer of genomic islands. © 2016 John Wiley & Sons Ltd.

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July 7, 2019

Highlights of the 11th International Bordetella Symposium: from basic biology to vaccine development.

Pertussis is a severe respiratory disease caused by infection with the bacterial pathogen Bordetella pertussis The disease affects individuals of all ages but is particularly severe and sometimes fatal in unvaccinated young infants. Other Bordetella species cause diseases in humans, animals, and birds. Scientific, clinical, public health, vaccine company, and regulatory agency experts on these pathogens and diseases gathered in Buenos Aires, Argentina from 5 to 8 April 2016 for the 11th International Bordetella Symposium to discuss recent advances in our understanding of the biology of these organisms, the diseases they cause, and the development of new vaccines and other strategies to prevent these diseases. Highlights of the meeting included pertussis epidemiology in developing nations, genomic analysis of Bordetella biology and evolution, regulation of virulence factor expression, new model systems to study Bordetella biology and disease, effects of different vaccines on immune responses, maternal immunization as a strategy to prevent newborn disease, and novel vaccine development for pertussis. In addition, the group approved the formation of an International Bordetella Society to promote research and information exchange on bordetellae and to organize future meetings. A new Bordetella.org website will also be developed to facilitate these goals. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019

Co-infection and emergence of rifamycin resistance during a recurrent Clostridium difficile infection.

Clostridium difficile (Peptoclostridium difficile) is a common health care associated infection with a disproportionately high incidence in elderly patients. Disease symptoms range from mild diarrhoea through to life threatening pseudomembranous colitis. Around 20% of patients may suffer recurrent disease which often requires re-hospitalisation of patients.C. difficile was isolated from stool samples from a patient with two recurrent C. difficile infections. PCR-ribotyping, whole genome sequencing and phenotypic assays were used to characterise these isolates.Genotypic and phenotypic screening of C. difficile isolates revealed multiple PCR-ribotypes present, and the emergence of rifamycin resistance during the infection cycle.Understanding both the clinical and bacterial factors that contribute to the course of recurrent infection could inform strategies to reduce recurrence. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019

Expansion of lysine-rich repeats in Plasmodium proteins generates novel localisation sequences that target the periphery of the host erythrocyte.

Repetitive low-complexity sequences, mostly assumed to have no function, are common in proteins that are exported by the malaria parasite into its host erythrocyte. We identify a group of exported proteins containing short lysine-rich tandemly repeated sequences that are sufficient to localise to the erythrocyte periphery where key virulence-related modifications to the plasma membrane and the underlying cytoskeleton are known to occur. Efficiency of targeting is dependent on repeat number, indicating that novel targeting modules could evolve by expansion of short lysine-rich sequences. Indeed, expression GARP fragments from different species shows that two novel targeting sequences have arisen via the process of repeat expansion in this protein. In the protein Hyp12, the targeting function of a lysine-rich sequence is masked by a neighbouring repetitive acidic sequence, further highlighting the importance of repetitive low complexity sequences. We show that sequences capable of targeting the erythrocyte periphery are present in at least nine proteins from Plasmodium falciparum, and one from Plasmodium knowlesi. We find these sequences in proteins known to be involved in erythrocyte rigidification and cytoadhesion, as well as in previously uncharacterised exported proteins. Together, these data suggest that expansion and contraction of lysine-rich repeats could generate targeting sequences de novo as well as modulate protein targeting efficiency and function in response to selective pressure. Copyright © 2016, The American Society for Biochemistry and Molecular Biology.

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July 7, 2019

Assembly and transfer of tripartite integrative and conjugative genetic elements.

Integrative and conjugative elements (ICEs) are ubiquitous mobile genetic elements present as “genomic islands” within bacterial chromosomes. Symbiosis islands are ICEs that convert nonsymbiotic mesorhizobia into symbionts of legumes. Here we report the discovery of symbiosis ICEs that exist as three separate chromosomal regions when integrated in their hosts, but through recombination assemble as a single circular ICE for conjugative transfer. Whole-genome comparisons revealed exconjugants derived from nonsymbiotic mesorhizobia received three separate chromosomal regions from the donor Mesorhizobium ciceri WSM1271. The three regions were each bordered by two nonhomologous integrase attachment (att) sites, which together comprised three homologous pairs of attL and attR sites. Sequential recombination between each attL and attR pair produced corresponding attP and attB sites and joined the three fragments to produce a single circular ICE, ICEMcSym(1271) A plasmid carrying the three attP sites was used to recreate the process of tripartite ICE integration and to confirm the role of integrase genes intS, intM, and intG in this process. Nine additional tripartite ICEs were identified in diverse mesorhizobia and transfer was demonstrated for three of them. The transfer of tripartite ICEs to nonsymbiotic mesorhizobia explains the evolution of competitive but suboptimal N2-fixing strains found in Western Australian soils. The unheralded existence of tripartite ICEs raises the possibility that multipartite elements reside in other organisms, but have been overlooked because of their unusual biology. These discoveries reveal mechanisms by which integrases dramatically manipulate bacterial genomes to allow cotransfer of disparate chromosomal regions.

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July 7, 2019

Selvamicin, an atypical antifungal polyene from two alternative genomic contexts.

The bacteria harbored by fungus-growing ants produce a variety of small molecules that help maintain a complex multilateral symbiosis. In a survey of antifungal compounds from these bacteria, we discovered selvamicin, an unusual antifungal polyene macrolide, in bacterial isolates from two neighboring ant nests. Selvamicin resembles the clinically important antifungals nystatin A1 and amphotericin B, but it has several distinctive structural features: a noncationic 6-deoxymannose sugar at the canonical glycosylation site and a second sugar, an unusual 4-O-methyldigitoxose, at the opposite end of selvamicin’s shortened polyene macrolide. It also lacks some of the pharmacokinetic liabilities of the clinical agents and appears to have a different target. Whole genome sequencing revealed the putative type I polyketide gene cluster responsible for selvamicin’s biosynthesis including a subcluster of genes consistent with selvamicin’s 4-O-methyldigitoxose sugar. Although the selvamicin biosynthetic cluster is virtually identical in both bacterial producers, in one it is on the chromosome, in the other it is on a plasmid. These alternative genomic contexts illustrate the biosynthetic gene cluster mobility that underlies the diversity and distribution of chemical defenses by the specialized bacteria in this multilateral symbiosis.

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July 7, 2019

Characterization and comparative overview of complete sequences of the first plasmids of Pandoraea across clinical and non-clinical strains.

To date, information on plasmid analysis in Pandoraea spp. is scarce. To address the gap of knowledge on this, the complete sequences of eight plasmids from Pandoraea spp. namely Pandoraea faecigallinarum DSM 23572(T) (pPF72-1, pPF72-2), Pandoraea oxalativorans DSM 23570(T) (pPO70-1, pPO70-2, pPO70-3, pPO70-4), Pandoraea vervacti NS15 (pPV15) and Pandoraea apista DSM 16535(T) (pPA35) were studied for the first time in this study. The information on plasmid sequences in Pandoraea spp. is useful as the sequences did not match any known plasmid sequence deposited in public databases. Replication genes were not identified in some plasmids, a situation that has led to the possibility of host interaction involvement. Some plasmids were also void of par genes and intriguingly, repA gene was also not discovered in these plasmids. This further leads to the hypothesis of host-plasmid interaction. Plasmid stabilization/stability protein-encoding genes were observed in some plasmids but were not established for participating in plasmid segregation. Toxin-antitoxin systems MazEF, VapBC, RelBE, YgiT-MqsR, HigBA, and ParDE were identified across the plasmids and their presence would improve plasmid maintenance. Conjugation genes were identified portraying the conjugation ability amongst Pandoraea plasmids. Additionally, we found a shared region amongst some of the plasmids that consists of conjugation genes. The identification of genes involved in replication, segregation, toxin-antitoxin systems and conjugation, would aid the design of drugs to prevent the survival or transmission of plasmids carrying pathogenic properties. Additionally, genes conferring virulence and antibiotic resistance were identified amongst the plasmids. The observed features in the plasmids shed light on the Pandoraea spp. as opportunistic pathogens.

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July 7, 2019

Genomic analyses of multidrug resistant Pseudomonas aeruginosa PA1 resequenced by single-molecule real-time sequencing.

As a third-generation sequencing (TGS) method, single-molecule real-time (SMRT) technology provides long read length, and it is well suited for resequencing projects and de novo assembly. In the present study, Pseudomonas aeruginosa PA1 was characterized and resequenced using SMRT technology. PA1 was also subjected to genomic, comparative and pan-genomic analyses. The multidrug resistant strain PA1 possesses a 6,498,072 bp genome and a sequence type of ST-782. The genome of PA1 was also visualized, and the results revealed the details of general genome annotations, virulence factors, regulatory proteins (RPs), secretion system proteins, type II toxin-antitoxin (T-A) pairs and genomic islands. Whole genome comparison analysis suggested that PA1 exhibits similarity to other P. aeruginosa strains but differs in terms of horizontal gene transfer (HGT) regions, such as prophages and genomic islands. Phylogenetic analyses based on 16S rRNA sequences demonstrated that PA1 is closely related to PAO1, and P. aeruginosa strains can be divided into two main groups. The pan-genome of P. aeruginosa consists of a core genome of approximately 4,000 genes and an accessory genome of at least 6,600 genes. The present study presented a detailed, visualized and comparative analysis of the PA1 genome, to enhance our understanding of this notorious pathogen. © 2016 The Author(s).

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July 7, 2019

Genome sequencing and comparative genomics analysis revealed pathogenic potential in Penicillium capsulatum as a novel fungal pathogen belonging to Eurotiales.

Penicillium capsulatum is a rare Penicillium species used in paper manufacturing, but recently it has been reported to cause invasive infection. To research the pathogenicity of the clinical Penicillium strain, we sequenced the genomes and transcriptomes of the clinical and environmental strains of P. capsulatum. Comparative analyses of these two P. capsulatum strains and close related strains belonging to Eurotiales were performed. The assembled genome sizes of P. capsulatum are approximately 34.4 Mbp in length and encode 11,080 predicted genes. The different isolates of P. capsulatum are highly similar, with the exception of several unique genes, INDELs or SNPs in the genes coding for glycosyl hydrolases, amino acid transporters and circumsporozoite protein. A phylogenomic analysis was performed based on the whole genome data of 38 strains belonging to Eurotiales. By comparing the whole genome sequences and the virulence-related genes from 20 important related species, including fungal pathogens and non-human pathogens belonging to Eurotiales, we found meaningful pathogenicity characteristics between P. capsulatum and its closely related species. Our research indicated that P. capsulatum may be a neglected opportunistic pathogen. This study is beneficial for mycologists, geneticists and epidemiologists to achieve a deeper understanding of the genetic basis of the role of P. capsulatum as a newly reported fungal pathogen.

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July 7, 2019

Variant exported blood-stage proteins encoded by Plasmodium multigene families are expressed in liver stages where they are exported into the parasitophorous vacuole.

Many variant proteins encoded by Plasmodium-specific multigene families are exported into red blood cells (RBC). P. falciparum-specific variant proteins encoded by the var, stevor and rifin multigene families are exported onto the surface of infected red blood cells (iRBC) and mediate interactions between iRBC and host cells resulting in tissue sequestration and rosetting. However, the precise function of most other Plasmodium multigene families encoding exported proteins is unknown. To understand the role of RBC-exported proteins of rodent malaria parasites (RMP) we analysed the expression and cellular location by fluorescent-tagging of members of the pir, fam-a and fam-b multigene families. Furthermore, we performed phylogenetic analyses of the fam-a and fam-b multigene families, which indicate that both families have a history of functional differentiation unique to RMP. We demonstrate for all three families that expression of family members in iRBC is not mutually exclusive. Most tagged proteins were transported into the iRBC cytoplasm but not onto the iRBC plasma membrane, indicating that they are unlikely to play a direct role in iRBC-host cell interactions. Unexpectedly, most family members are also expressed during the liver stage, where they are transported into the parasitophorous vacuole. This suggests that these protein families promote parasite development in both the liver and blood, either by supporting parasite development within hepatocytes and erythrocytes and/or by manipulating the host immune response. Indeed, in the case of Fam-A, which have a steroidogenic acute regulatory-related lipid transfer (START) domain, we found that several family members can transfer phosphatidylcholine in vitro. These observations indicate that these proteins may transport (host) phosphatidylcholine for membrane synthesis. This is the first demonstration of a biological function of any exported variant protein family of rodent malaria parasites.

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July 7, 2019

Emerging infectious disease implications of invasive mammalian species: the greater white-toothed shrew (Crocidura russula) is associated with a novel serovar of pathogenic Leptospira in Ireland.

The greater white-toothed shrew (Crocidura russula) is an invasive mammalian species that was first recorded in Ireland in 2007. It currently occupies an area of approximately 7,600 km2 on the island. C. russula is normally distributed in Northern Africa and Western Europe, and was previously absent from the British Isles. Whilst invasive species can have dramatic and rapid impacts on faunal and floral communities, they may also be carriers of pathogens facilitating disease transmission in potentially naive populations. Pathogenic leptospires are endemic in Ireland and a significant cause of human and animal disease. From 18 trapped C. russula, 3 isolates of Leptospira were cultured. However, typing of these isolates by standard serological reference methods was negative, and suggested an, as yet, unidentified serovar. Sequence analysis of 16S ribosomal RNA and secY indicated that these novel isolates belong to Leptospira alstonii, a unique pathogenic species of which only 7 isolates have been described to date. Earlier isolations were limited geographically to China, Japan and Malaysia, and this leptospiral species had not previously been cultured from mammals. Restriction enzyme analysis (REA) further confirms the novelty of these strains since no similar patterns were observed with a reference database of leptospires. As with other pathogenic Leptospira species, these isolates contain lipL32 and do not grow in the presence of 8-azagunaine; however no evidence of disease was apparent after experimental infection of hamsters. These isolates are genetically related to L. alstonii but have a novel REA pattern; they represent a new serovar which we designate as serovar Room22. This study demonstrates that invasive mammalian species act as bridge vectors of novel zoonotic pathogens such as Leptospira.

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July 7, 2019

Decay of sexual trait genes in an asexual parasitoid wasp.

Trait loss is a widespread phenomenon with pervasive consequences for a species’ evolutionary potential. The genetic changes underlying trait loss have only been clarified in a small number of cases. None of these studies can identify whether the loss of the trait under study was a result of neutral mutation accumulation or negative selection. This distinction is relatively clear-cut in the loss of sexual traits in asexual organisms. Male-specific sexual traits are not expressed and can only decay through neutral mutations, whereas female-specific traits are expressed and subject to negative selection. We present the genome of an asexual parasitoid wasp and compare it to that of a sexual lineage of the same species. We identify a short-list of 16 genes for which the asexual lineage carries deleterious SNP or indel variants, whereas the sexual lineage does not. Using tissue-specific expression data from other insects, we show that fifteen of these are expressed in male-specific reproductive tissues. Only one deleterious variant was found that is expressed in the female-specific spermathecae, a trait that is heavily degraded and thought to be under negative selection in L. clavipes. Although the phenotypic decay of male-specific sexual traits in asexuals is generally slow compared with the decay of female-specific sexual traits, we show that male-specific traits do indeed accumulate deleterious mutations as expected by theory. Our results provide an excellent starting point for detailed study of the genomics of neutral and selected trait decay.

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July 7, 2019

Genomic insights into Campylobacter jejuni virulence and population genetics

Campylobacter jejuni has long been recognized as a main food-borne pathogen in many parts of the world. Natural reservoirs include a wide variety of domestic and wild birds and mammals, whose intestines offer a suitable biological niche for the survival and dissemination of the organism. Understanding the genetic basis of the biology and pathogenicity of C. jejuni is vital to prevent and control Campylobacter-associated infections. The recent progress in sequencing techniques has allowed for a rapid increase in our knowledge of the molecular biology and the genetic structures of Campylobacter. Single-molecule realtime (SMRT) sequencing, which goes beyond four-base sequencing, revealed the role of DNA methylation in modulating the biology and virulence of C. jejuni at the level of epigenetics. In this review, we will provide an up-to-date review on recent advances in understanding C. jejuni genomics, including structural features of genomes, genetic traits of virulence, population genetics, and epigenetics.

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