Jose C. Huguet-Tapia

Draft Genome Sequence of Streptomyces acidiscabies 84-104, an Emergent Plant Pathogen

A draft genome sequence of the plant pathogen Streptomyces acidiscabies 84-104, an emergent plant pathogen, is presented here. The genome is among the largest of streptomycetes, at more than 11 Mb, and encodes a 100-kb pathogenicity island (PAI) shared with other plant-pathogenic streptomycetes. The presence of this conserved PAI, and the remnants of a conserved integrase/recombinase at its 3 end, supports the hypothesis that S. acidiscabies emerged as a plant pathogen as a result of this acquisition.

Characterization of the Integration and Modular Excision of the Integrative Conjugative Element PAISt in Streptomyces turgidiscabies Car8

PAISt is a large genomic island located in the chromosome of the plant pathogen Streptomyces turgidiscabies Car8. The island carries clustered virulence genes, transfers to other Streptomyces species, and integrates by site-specific recombination at the 8 bp palindrome TTCATGAA. The palindrome is located at the 3′ end of the bacitracin resistance gene (bacA). We demonstrate that PAISt is able to excise in modules by recombination of one internal and two flanking palindromic direct repeats. The gene intSt located at the 3( end of PAISt encodes a tyrosine recombinase. Site-specific recombination activity of intSt was tested and confirmed by heterologous expression in Streptomyces coelicolor. Comparative analysis of PAISt homologues in Streptomyces scabies 87–22 and Streptomyces acidiscabies 84–104 indicates that these islands have been fixed by sequence erosion of intSt and the recombination sites.

Whole genome sequence of the emerging oomycete pathogen Pythium insidiosum strain CDC-B5653 isolated from an infected human in the USA

Pythium insidiosum ATCC 200269 strain CDC-B5653, an isolate from necrotizing lesions on the mouth and eye of a 2-year-old boy in Memphis, Tennessee, USA, was sequenced using a combination of Illumina MiSeq (300 bp paired-end, 14 millions reads) and PacBio (10 Kb fragment library, 356,001 reads). The sequencing data were assembled using SPAdes version 3.1.0, yielding a total genome size of 45.6 Mb contained in 8992 contigs, N50 of 13 Kb, 57% G + C content, and 17,867 putative protein-coding genes. This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession JRHR00000000.

Complete Genome Sequence of the African Strain AXO1947 of Xanthomonas oryzae pv. oryzae

ABSTRACT Xanthomonas oryzae pv. oryzae is the etiological agent of bacterial rice blight. Three distinct clades of X. oryzae pv. oryzae are known. We present the complete annotated genome of the African clade strain AXO194 using long-read single-molecule PacBio sequencing technology. The genome comprises a single chromosome of 4,674,975 bp and encodes for nine transcriptional activator-like (TAL) effectors. The approach and data presented in this announcement provide information for complex bacterial genome organization and the discovery of new virulence effectors, and they facilitate target characterization of TAL effectors.

Genome Content and Phylogenomics Reveal both Ancestral and Lateral Evolutionary Pathways in Plant-Pathogenic Streptomyces Species

ABSTRACT Streptomyces spp. are highly differentiated actinomycetes with large, linear chromosomes that encode an arsenal of biologically active molecules and catabolic enzymes. Members of this genus are well equipped for life in nutrient-limited environments and are common soil saprophytes. Out of the hundreds of species in the genus Streptomyces, a small group has evolved the ability to infect plants. The recent availability of Streptomyces genome sequences, including four genomes of pathogenic species, provided an opportunity to characterize the gene content specific to these pathogens and to study phylogenetic relationships among them. Genome sequencing, comparative genomics, and phylogenetic analysis enabled us to discriminate pathogenic from saprophytic Streptomyces strains; moreover, we calculated that the pathogen-specific genome contains 4,662 orthologs. Phylogenetic reconstruction suggested that Streptomyces scabies and S. ipomoeae share an ancestor but that their biosynthetic clusters encoding the required virulence factor thaxtomin have diverged. In contrast, S. turgidiscabies and S. acidiscabies, two relatively unrelated pathogens, possess highly similar thaxtomin biosynthesis clusters, which suggests that the acquisition of these genes was through lateral gene transfer.

An artificial self-sufficient cytochrome P450 directly nitrates fluorinated tryptophan analogs with a different regio-selectivity

Aromatic nitration is an immensely important industrial process to produce chemicals for a variety of applications, but it often suffers from multiple unsolved challenges. Enzymes as biocatalysts have been increasingly used for organic chemistry synthesis due to their high selectivity and environmental friendliness, but nitration has benefited minimally from the development of biocatalysis. In this work, we aimed to develop TxtE as practical biocatalysts for aromatic nitration. TxtE is a unique class I cytochrome P450 enzyme that nitrates the indole of l-tryptophan. To develop cost-efficient nitration processes, we fused TxtE with the reductase domains of CYP102A1 (P450BM3) and of P450RhF to create class III self-sufficient biocatalysts. The best engineered fusion protein was comparable with wild type TxtE in terms of nitration performance and other key biochemical properties. To demonstrate the application potential of the fusion enzyme, we nitrated 4-F-dl-tryptophan and 5-F-l-tryptophan in large scale enzymatic reactions. Tandem MS/MS and NMR analyses of isolated products revealed altered nitration sites. To our knowledge, these studies represent the first practice in developing biological nitration approaches and lay a solid basis to the use of TxtE-based biocatalysts for the production of valuable nitroaromatics.

Pacbio sequencing of copper-tolerant Xanthomonas citri reveals presence of a chimeric plasmid structure and provides insights into reassortment and shuffling of transcription activator-like effectors among X. citri strains

BackgroundXanthomonas citri, a causal agent of citrus canker, has been a well-studied model system due to recent availability of whole genome sequences of multiple strains from different geographical regions. Major limitations in our understanding of the evolution of pathogenicity factors in X. citri strains sequenced by short-read sequencing methods have been tracking plasmid reshuffling among strains due to inability to accurately assign reads to plasmids, and analyzing repeat regions among strains. X. citri harbors major pathogenicity determinants, including variable DNA-binding repeat region containing Transcription Activator-like Effectors (TALEs) on plasmids. The long-read sequencing method, PacBio, has allowed the ability to obtain complete and accurate sequences of TALEs in xanthomonads. We recently sequenced Xanthomonas citri str. Xc-03-1638-1-1, a copper tolerant A group strain isolated from grapefruit in 2003 from Argentina using PacBio RS II chemistry. We analyzed plasmid profiles, copy number and location of TALEs in complete genome sequences of X. citri strains.ResultsWe utilized the power of long reads obtained by PacBio sequencing to enable assembly of a complete genome sequence of strain Xc-03-1638-1-1, including sequences of two plasmids, 249xa0kb (plasmid harboring copper resistance genes) and 99xa0kb (pathogenicity plasmid containing TALEs). The pathogenicity plasmid in this strain is a hybrid plasmid containing four TALEs. Due to the intriguing nature of this pathogenicity plasmid with Tn3-like transposon association, repetitive elements and multiple putative sites for origins of replication, we might expect alternative structures of this plasmid in nature, illustrating the strong adaptive potential of X. citri strains. Analysis of the pathogenicity plasmid among completely sequenced X. citri strains, coupled with Southern hybridization of the pathogenicity plasmids, revealed clues to rearrangements of plasmids and resulting reshuffling of TALEs among strains.ConclusionsWe demonstrate in this study the importance of long-read sequencing for obtaining intact sequences of TALEs and plasmids, as well as for identifying rearrangement events including plasmid reshuffling. Rearrangement events, such as the hybrid plasmid in this case, could be a frequent phenomenon in the evolution of X. citri strains, although so far it is undetected due to the inability to obtain complete plasmid sequences with short-read sequencing methods.

Whole-Genome Sequence of Ralstonia solanacearum P673, a Strain Capable of Infecting Tomato Plants at Low Temperatures

ABSTRACT Ralstonia solanacearum is the causal agent of bacterial wilt, one of the most destructive bacterial plant diseases. We present the whole-genome sequence of the strain P673 (phylotype IIB, sequevar 4). This strain is capable of producing disease in tomato plants at low temperatures. P673 has 311 unique genes.

Feather mites play a role in cleaning host feathers: New insights from DNA metabarcoding and microscopy

Parasites and other symbionts are crucial components of ecosystems, regulating host populations and supporting food webs. However, most symbiont systems, especially those involving commensals and mutualists, are relatively poorly understood. In this study, we have investigated the nature of the symbiotic relationship between birds and their most abundant and diverse ectosymbionts: the vane‐dwelling feather mites. For this purpose, we studied the diet of feather mites using two complementary methods. First, we used light microscopy to examine the gut contents of 1,300 individual feather mites representing 100 mite genera (18 families) from 190 bird species belonging to 72 families and 19 orders. Second, we used high‐throughput sequencing (HTS) and DNA metabarcoding to determine gut contents from 1,833 individual mites of 18 species inhabiting 18 bird species. Results showed fungi and potentially bacteria as the main food resources for feather mites (apart from potential bird uropygial gland oil). Diatoms and plant matter appeared as rare food resources for feather mites. Importantly, we did not find any evidence of feather mites feeding upon bird resources (e.g., blood, skin) other than potentially uropygial gland oil. In addition, we found a high prevalence of both keratinophilic and pathogenic fungal taxa in the feather mite species examined. Altogether, our results shed light on the long‐standing question of the nature of the relationship between birds and their vane‐dwelling feather mites, supporting previous evidence for a commensalistic–mutualistic role of feather mites, which are revealed as likely fungivore–microbivore–detritivore symbionts of bird feathers.

Phylogenomic analysis supports multiple instances of polyphyly in the oomycete peronosporalean lineage

The study of biological diversification of oomycetes has been a difficult task for more than a century. Pioneer researchers used morphological characters to describe this heterogeneous group, and physiological and genetic tools expanded knowledge of these microorganisms. However, research on oomycete diversification is limited by conflicting phylogenies. Using whole genomic data from 17 oomycete taxa, we obtained a dataset of 277 core orthologous genes shared among these genomes. Analyses of this dataset resulted in highly congruent and strongly supported estimates of oomycete phylogeny when we used concatenated maximum likelihood and coalescent-based methods; the one important exception was the position of Albugo. Our results supported the position of Phytopythium vexans (formerly in Pythium clade K) as a sister clade to the Phytophthora-Hyaloperonospora clade. The remaining clades comprising Pythium sensu lato formed two monophyletic groups. One group was composed of three taxa that correspond to Pythium clades A, B and C, and the other group contained taxa representing clades F, G and I, in agreement with previous Pythium phylogenies. However, the group containing Pythium clades F, G and I was placed as sister to the Phytophthora-Hyaloperonospora-Phytopythium clade, thus confirming the lack of monophyly of Pythium sensu lato. Multispecies coalescent methods revealed that the white blister rust, Albugo laibachii, could not be placed with a high degree of confidence. Our analyses show that genomic data can resolve the oomycete phylogeny and provide a phylogenetic framework to study the evolution of oomycete lifestyles.