Phylogenetic Analysis and Comparative Genomics of Brucella abortus and Brucella melitensis Strains in Egypt.

Brucellosis is a notifiable disease induced by a facultative intracellular Brucella pathogen. In this study, eight Brucella abortus and eighteen Brucella melitensis strains from Egypt were annotated and compared with RB51 and REV1 vaccines respectively. RAST toolkit in the BV-BRC server was used for annotation, revealing genome length of 3,250,377 bp and 3,285,803 bp, 3289 and 3323 CDS, 48 and 49 tRNA genes, the same number of rRNA (3) genes, 583 and 586 hypothetical proteins, 2697 and 2726 functional proteins for B. abortus and B. melitensis respectively. B. abortus strains exhibit a similar number of candidate genes, while B. melitensis strains showed some differences, especially in the SRR19520422 Faiyum strain. Also, B. melitensis clarified differences in antimicrobial resistance genes (KatG, FabL, MtrA, MtrB, OxyR, and VanO-type) in SRR19520319 Faiyum and (Erm C and Tet K) in SRR19520422 Faiyum strain. Additionally, the whole genome phylogeny analysis proved that all B. abortus strains were related to vaccinated animals and all B. melitensis strains of Menoufia clustered together and closely related to Gharbia, Dameitta, and Kafr Elshiek. The Bowtie2 tool identified 338 (eight B. abortus) and 4271 (eighteen B. melitensis) single nucleotide polymorphisms (SNPs) along the genomes. These variants had been annotated according to type and impact. Moreover, thirty candidate genes were predicted and submitted at GenBank (24 in B. abortus) and (6 in B. melitensis). This study contributes significant insights into genetic variation, virulence factors, and vaccine-related associations of Brucella pathogens, enhancing our knowledge of brucellosis epidemiology and evolution in Egypt.

Beyond rational-biosensor-guided isolation of 100 independently evolved bacterial strain variants and comparative analysis of their genomes.

BACKGROUND: In contrast to modern rational metabolic engineering, classical strain development strongly relies on random mutagenesis and screening for the desired production phenotype. Nowadays, with the availability of biosensor-based FACS screening strategies, these random approaches are coming back into fashion. In this study, we employ this technology in combination with comparative genome analyses to identify novel mutations contributing to product formation in the genome of a Corynebacterium glutamicum L-histidine producer. Since all known genetic targets contributing to L-histidine production have been already rationally engineered in this strain, identification of novel beneficial mutations can be regarded as challenging, as they might not be intuitively linkable to L-histidine biosynthesis. RESULTS: In order to identify 100 improved strain variants that had each arisen independently, we performed > 600 chemical mutagenesis experiments, > 200 biosensor-based FACS screenings, isolated > 50,000 variants with increased fluorescence, and characterized > 4500 variants with regard to biomass formation and L-histidine production. Based on comparative genome analyses of these 100 variants accumulating 10-80% more L-histidine, we discovered several beneficial mutations. Combination of selected genetic modifications allowed for the construction of a strain variant characterized by a doubled L-histidine titer (29 mM) and product yield (0.13 C-mol C-mol-1) in comparison to the starting variant. CONCLUSIONS: This study may serve as a blueprint for the identification of novel beneficial mutations in microbial producers in a more systematic manner. This way, also previously unexplored genes or genes with previously unknown contribution to the respective production phenotype can be identified. We believe that this technology has a great potential to push industrial production strains towards maximum performance.

Genome Mining of Novel Targets and Construction of Ladder-shaped melting temperature isothermal amplification Assays for the Identification of Cronobacter sakazakii and Cronobacter malonaticus.

Cronobacter species are potential pathogens that can contaminate powdered infant formula. C. sakazakii and C. malonaticus are the most common species of Cronobacter associated with infections. This study mined new molecular targets for the detection of C. sakazakii and C. malonaticus by using comparative genome approaches. Specific target genes mngB and ompR were obtained and used to detect C. sakazakii and C. malonaticus, respectively. A novel detection method, termed ladder-shape melting temperature isothermal amplification (LMTIA), was developed and evaluated. The detection limit for pure C. sakazakii DNA was 1 pg per reaction and 1 fg per reaction for C. malonaticus. The C. sakazakii, C. malonaticus, and the reference stains were all correctly identified. The amplicons can be successfully visualized and identified by naked eyes when hydroxy naphthol blue dye (HNB dye) was used in the reaction. Therefore, the LMTIA assays developed in this study showed potential application for microorganism identification and detection.

Genomic characterization and identification of virulence-related genes in Vibrio nigripulchritudo isolated from white leg shrimp Penaeus vannamei.

Vibrio nigripulchritudo causes vibriosis in penaeid shrimps. Here, we used Illumina and Nanopore sequencing technologies to sequence the genomes of three of its strains (TUMSAT-V. nig1, TUMSAT-V. nig2, and TUMSAT-V. nig3) to explore opportunities for disease management. Putative virulence factors and mobile genetic elements were detected while evaluating the phylogenetic relationship of each isolated strain. The genomes consisted of two circular chromosomes (I and II) plus one or two plasmids. The size of chromosome I ranged from 4.02 to 4.07 Mb with an average GC content of 46%, while the number of predicted CDSs ranged from 3563 to 3644. The size of chromosome II ranged from 2.16 to 2.18 Mb, with an average GC content of 45.5%, and the number of predicted CDSs ranged from 1970 to 1987. Numerous virulence genes were identified related to adherence, antiphagocytosis, chemotaxis, motility, iron uptake, quorum sensing, secretion systems, and toxins in all three genomes. Higher numbers of prophages and genomic islands found in TUMSAT-V. nig1 suggest that the strain has experienced numerous horizontal gene transfer events. The presence of antimicrobial resistance genes suggests that the strains have multidrug resistance. Comparative genomic analysis showed that all three strains belonged to the same clade.

The Complete Chloroplast Genome Sequence of Cicer bijugum, Genome Organization, and Comparison with Related Species.

Background: Chickpea is one of Turkey's most significant legumes, and because of its high nutritional value, it is frequently preferred in human nourishment.Chloroplasts, which have their own genetic material, are organelles responsible for photosynthesis in plant cells and their genome contains non-trivial information about the molecular features and evolutionary process of plants. Objective: Current study aimed at revealing complete chloroplast genome sequence of one of the wild type Cicer species, Cicer bijugum, and comparing its genome with cultivated Cicer species, Cicer arietinum, by using bioinformatics analysis tools. Except for Cicer arietinum, there has been no study on the chloroplast genome sequence of Cicer species.Therefore, we targeted to reveal the complete chloroplast genome sequence of wild type Cicer species, Cicer bijugum, and compare the chloroplast genome of Cicer bijugum with the cultivated one Cicer arietinum. Methods: In this study, we sequenced the whole chloroplast genome of Cicer bijugum, one of the wild types of chickpea species, with the help Next Generation Sequencing platform and compared it with the chloroplast genome of the cultivated chickpea species, Cicer arietinum, by using online bioinformatics analysis tools. Results: We determined the size of the chloroplast genome of C. bijugum as 124,804 bp and found that C. bijugum did not contain an inverted repeat region in its chloroplast genome. Comparative analysis of the C. bijugum chloroplast genome uncovered thirteen hotspot regions (psbA, matK, rpoB, rpoC1, rpoC2, psbI, psbK, accD, rps19, ycf2, ycf1, rps15, and ndhF) and seven of them (matK, accD, rps19, ycf1, ycf2, rps15 and ndhF) could potentially be used as strong molecular markers for species identification. It has been determined that C. bijugum was phylogenetically closer to cultivated chickpea as compared to the other species. Conclusion: It is aimed that the data obtained from this study, which is the first study in which whole chloroplast genomes of wild chickpea species were sequenced, will guide researchers in future molecular, evolutionary, and genetic engineering studies with chickpea species.

Genome analysis of clinical isolate of Campylobacter fetus subspecies fetus MMM01 from India reveals genetic determinants of pathogenesis and adaptation.

In this study we report the whole genome sequencing (WGS) based analysis of blood-borne Campylobacter fetus subsp. fetus MMM01 isolated from a diabetic patient to obtain deeper insights in to the virulence and host adaptability. The sequenced genome of C. fetus subsp. fetus MMM01 along with reference genomes retrieved from NCBI was subjected to various in-silico analysis including JSpecies, MLST server, PATRIC server, VFanalyzer, CARD, PHASTER to understand their phylogenetic relation, virulence and antimicrobial resistance profile. The genome had a size of 1,788,790 bp, with a GC content of 33.09%, nearly identical to the reference strain C. fetus subsp. fetus 82-40. The MLST based phylogenetic tree constructed revealed the polyphyletic branching and MMM01 (ST25) was found to be closely related to ST11, both belong to the sap-A serotype which are more common in human infections. VFanalyzer identified 88 protein-coding genes coding for several virulence factors including Campylobacter adhesion to fibronectin, flagellar apparatus, cytolethal distending toxin operons and Campylobacter invasion antigen proteins which enhance the virulence of bacteria along with resistance genes against antibiotics including fluoroquinolone, chloramphenicol, tetracycline, and aminoglycoside in MMM01, which points to enhanced survival and pathogenicity of this zoonotic pathogen. It was interesting to find that MMM01 lacked FGI-II island found in most of the clinical isolates, which encoded CRISPR Cas and prophage II regions. More details about the complexity and evolution of this zoonotic pathogen could be learned from future studies that concentrate on comparative genome analysis using larger genome datasets.

Assembly and Annotation of Red Spruce (Picea rubens) Chloroplast Genome, Identification of Simple Sequence Repeats, and Phylogenetic Analysis in Picea.

We have sequenced the chloroplast genome of red spruce (Picea rubens) for the first time using the single-end, short-reads (44 bp) Illumina sequences, assembled and functionally annotated it, and identified simple sequence repeats (SSRs). The contigs were assembled using SOAPdenovo2 following the retrieval of chloroplast genome sequences using the black spruce (Picea mariana) chloroplast genome as the reference. The assembled genome length was 122,115 bp (gaps included). Comparatively, the P. rubens chloroplast genome reported here may be considered a near-complete draft. Global genome alignment and phylogenetic analysis based on the whole chloroplast genome sequences of Picea rubens and 10 other Picea species revealed high sequence synteny and conservation among 11 Picea species and phylogenetic relationships consistent with their known classical interrelationships and published molecular phylogeny. The P. rubens chloroplast genome sequence showed the highest similarity with that of P. mariana and the lowest with that of P. sitchensis. We have annotated 107 genes including 69 protein-coding genes, 28 tRNAs, 4 rRNAs, few pseudogenes, identified 42 SSRs, and successfully designed primers for 26 SSRs. Mononucleotide A/T repeats were the most common followed by dinucleotide AT repeats. A similar pattern of microsatellite repeats occurrence was found in the chloroplast genomes of 11 Picea species.

Roles of Species-Specific Legumains in Pathogenicity of the Pinewood Nematode Bursaphelenchus xylophilus.

Peptidases are very important to parasites, which have central roles in parasite biology and pathogenesis. In this study, by comparative genome analysis, genome-wide peptidase diversities among plant-parasitic nematodes are estimated. We find that genes encoding cysteine peptidases in family C13 (legumain) are significantly abundant in pine wood nematodes Bursaphelenchus genomes, compared to those in other plant-parasitic nematodes. By phylogenetic analysis, a clade of B. xylophilus-specific legumain is identified. RT-qPCR detection shows that these genes are highly expressed at early stage during the nematode infection process. Utilizing transgene technology, cDNAs of three species-specific legumain were introduced into the Arabidopsis γvpe mutant. Functional complementation assay shows that these B. xylophilus legumains can fully complement the activity of Arabidopsis γVPE to mediate plant cell death triggered by the fungal toxin FB1. Secretory activities of these legumains are experimentally validated. By comparative transcriptome analysis, genes involved in plant cell death mediated by legumains are identified, which enrich in GO terms related to ubiquitin protein transferase activity in category molecular function, and response to stimuli in category biological process. Our results suggest that B. xylophilu-specific legumains have potential as effectors to be involved in nematode-plant interaction and can be related to host cell death.

Telomere-to-telomere genome assembly of asparaginase-producing Trichoderma simmonsii.

BACKGROUND: Trichoderma is a genus of fungi in the family Hypocreaceae and includes species known to produce enzymes with commercial use. They are largely found in soil and terrestrial plants. Recently, Trichoderma simmonsii isolated from decaying bark and decorticated wood was newly identified in the Harzianum clade of Trichoderma. Due to a wide range of applications in agriculture and other industries, genomes of at least 12 Trichoderma spp. have been studied. Moreover, antifungal and enzymatic activities have been extensively characterized in Trichoderma spp. However, the genomic information and bioactivities of T. simmonsii from a particular marine-derived isolate remain largely unknown. While we screened for asparaginase-producing fungi, we observed that T. simmonsii GH-Sj1 strain isolated from edible kelp produced asparaginase. In this study, we report a draft genome of T. simmonsii GH-Sj1 using Illumina and Oxford Nanopore technologies. Furthermore, to facilitate biotechnological applications of this species, RNA-sequencing was performed to elucidate the transcriptional profile of T. simmonsii GH-Sj1 in response to asparaginase-rich conditions. RESULTS: We generated ~ 14 Gb of sequencing data assembled in a ~ 40 Mb genome. The T. simmonsii GH-Sj1 genome consisted of seven telomere-to-telomere scaffolds with no sequencing gaps, where the N50 length was 6.4 Mb. The total number of protein-coding genes was 13,120, constituting ~ 99% of the genome. The genome harbored 176 tRNAs, which encode a full set of 20 amino acids. In addition, it had an rRNA repeat region consisting of seven repeats of the 18S-ITS1-5.8S-ITS2-26S cluster. The T. simmonsii genome also harbored 7 putative asparaginase-encoding genes with potential medical applications. Using RNA-sequencing analysis, we found that 3 genes among the 7 putative genes were significantly upregulated under asparaginase-rich conditions. CONCLUSIONS: The genome and transcriptome of T. simmonsii GH-Sj1 established in the current work represent valuable resources for future comparative studies on fungal genomes and asparaginase production.

Comparative analysis using the draft genome sequence of California poppy (Eschscholzia californica) for exploring the candidate genes involved in benzylisoquinoline alkaloid biosynthesis.

Genome characterization of California poppy (Eschscholzia californica cv. "Hitoezaki"), which produces pharmaceutically important benzylisoquinoline alkaloids (BIAs), was carried out using the draft genome sequence. The numbers of tRNA and rRNA genes were close to those of the other plant species tested, whereas the frequency of repetitive sequences was distinct from those species. Comparison of the predicted genes with those of Amborella trichopoda, Nelumbo nucifera, Solanum lycopersicum, and Arabidopsis thaliana, and analyses of gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway indicated that the enzyme genes involved in BIA biosynthesis were highly enriched in the California poppy genome. Further comparative analysis using the genome information of Papaver somniferum and Aquilegia coerulea, both BIA-producing plants, revealed that many genes encoding BIA biosynthetic enzymes, transcription factors, transporters, and candidate proteins, possibly related to BIA biosynthesis, were specifically distributed in these plant species.

Biodegradation of phenol by a highly tolerant strain Rhodococcus ruber C1: Biochemical characterization and comparative genome analysis.

A novel phenol-degrading strain was isolated and identified as Rhodococcus ruber C1. The degradation analysis shows that 1806 mg/L of phenol can be completely degraded by strain C1 within 38 h, and the maximum specific growth rate (µmax=1.527 h-1) and maximum specific phenol degradation rate (qmax=3.674 h-1) indicate its excellent phenol metabolism capability. More importantly, phenol can be degraded by strain C1 in the temperature range of 20-45 °C within 72 h, and with longer degradation time, phenol can be completely degraded even at 10, 15 and 50 °C. The whole genome of strain C1 was sequenced, and a comparative genome analysis of strain C1 with 36 other genomes of Rhodococcus was performed. A remarkable gene family expansion occurred during the evolution of Rhodococcus, and a comprehensive evolutionary picture of Rhodococcus at genomic level was presented. Moreover, the copy number of genes involved in phenol metabolism was compared among genus Rhodococcus, and the results demonstrate high phenol degradation capability of strain C1 at genomic level. These findings suggest that Rhodococcus ruber C1 is a bacterium capable of degrading phenol efficiently in the temperature range of 10-50 °C.

Genome-wide analysis of proline-rich extension-like receptor protein kinase (PERK) in Brassica rapa and its association with the pollen development.

BACKGROUND: Proline-rich extension-like receptor protein kinases (PERKs) are an important class of receptor kinases located in the plasma membrane, most of which play a vital role in pollen development. RESULTS: Our study identified 25 putative PERK genes from the whole Brassica rapa genome (AA). Phylogenetic analysis of PERK protein sequences from 16 Brassicaceae species divided them into four subfamilies. The biophysical properties of the BrPERKs were investigated. Gene duplication and synteny analyses and the calculation of Ka/Ks values suggested that all 80 orthologous/paralogous gene pairs between B. rapa and A. thaliana, B. nigra and B. oleracea have experienced strong purifying selection. RNA-Seq data and qRT-PCR analyses showed that several BrPERK genes were expressed in different tissues, while some BrPERKs exhibited high expression levels only in buds. Furthermore, comparative transcriptome analyses from six male-sterile lines of B. rapa indicated that 7 BrPERK genes were downregulated in all six male-sterile lines. Meanwhile, the interaction networks of the BrPERK genes were constructed and 13 PERK coexpressed genes were identified, most of which were downregulated in the male sterile buds. CONCLUSION: Combined with interaction networks, coexpression and qRT-PCR analyses, these results demonstrated that two BrPERK genes, Bra001723.1 and Bra037558.1 (the orthologs of AtPERK6 (AT3G18810)), were downregulated beginning in the meiosis II period of male sterile lines and involved in anther development. Overall, this comprehensive analysis of some BrPERK genes elucidated their roles in male sterility.

Genome-based reclassification of Lactobacillus casei: emended classification and description of the species Lactobacillus zeae.

Taxonomic relationships between Lactobacillus casei, Lactobacillus paracasei and Lactobacillus zeae have long been debated. Results of previous analyses have shown that overall genome relatedness indices (such as average nucleotide identity and core nucleotide identity) between the type strains L. casei ATCC 393T and L. zeae ATCC 15820T were 94.6 and 95.3 %, respectively, which are borderline for species definition. However, the digital DNA‒DNA hybridization value was 57.3 %, which was clearly lower than the species delineation threshold of 70 %, and hence raised the possibility that L. casei could be reclassified into two species. To re-evaluate the taxonomic relationship of these taxa, multilocus sequence analysis (MLSA) based on the concatenated five housekeeping gene (dnaJ, dnaK, mutL, pheS and yycH) sequences, phylogenomic and core genome multilocus sequence typing analyses, gene presence and absence profiles using pan-genome analysis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) profiling analysis, cellular fatty acid compositions, and phenotype analysis were carried out. The results of phenotypic characterization, MLSA, whole-genome sequence-based analyses and MALDI-TOF MS profiling justified an independent species designation for the L. zeae strains, and supported an emended the description of the name of Lactobacillus zeae (ex Kuznetsov 1956) Dicks et al. 1996, with ATCC 15820T (=DSM 20178T=BCRC 17942T) as the type strain.

Comparative genomic analysis of Pseudomonas amygdali pv. lachrymans NM002: Insights into its potential virulence genes and putative invasion determinants.

Pseudomonas amygdali pv. lachrymans is currently of important plant pathogenic bacteria that causes cucumber angular leaf spot worldwide. The pathogen has been studied for its roles in pathogenicity and plant inheritance resistance. To further delineate traits critical to virulence, invasion and survival in the phyllosphere, we reported the first complete genome of P. amygdali pv. lachrymans NM002. Analysis of the whole genome in comparison with three closely-related representative pathovars of P. syringae identified the conservation of virulence genes, including flagella and chemotaxis, quorum-sensing systems, two-component systems, and lipopolysaccharide and antiphagocytosis. It also revealed differences of invasion determinants, such as type III effectors, phytotoxin (coronatine, syringomycin and phaseolotoxin) and cell wall-degrading enzyme, which may contribute to infectivity. The aim of this study was to derive genomic information that would reveal the probable molecular mechanisms underlying the virulence, infectivity and provide a better understanding of the pathogenesis of the P. syringae pathovars.

Construction of the first high-density genetic linkage map and identification of seed yield-related QTLs and candidate genes in Elymus sibiricus, an important forage grass in Qinghai-Tibet Plateau.

BACKGROUND: Elymus sibiricus is an ecologically and economically important perennial, self-pollinated, and allotetraploid (StStHH) grass, widely used for forage production and animal husbandry in Western and Northern China. However, it has low seed yield mainly caused by seed shattering, which makes seed production difficult for this species. The goals of this study were to construct the high-density genetic linkage map, and to identify QTLs and candidate genes for seed-yield related traits. RESULTS: An F2 mapping population of 200 individuals was developed from a cross between single genotype from "Y1005" and "ZhN06". Specific-locus amplified fragment sequencing (SLAF-seq) was applied to construct the first genetic linkage map. The final genetic map included 1971 markers on the 14 linkage groups (LGs) and was 1866.35 cM in total. The length of each linkage group varied from 87.67 cM (LG7) to 183.45 cM (LG1), with an average distance of 1.66 cM between adjacent markers. The marker sequences of E. sibiricus were compared to two grass genomes and showed 1556 (79%) markers mapped to wheat, 1380 (70%) to barley. Phenotypic data of eight seed-related traits (2016-2018) were used for QTL identification. A total of 29 QTLs were detected for eight seed-related traits on 14 linkage groups, of which 16 QTLs could be consistently detected for two or three years. A total of 6 QTLs were associated with seed shattering. Based on annotation with wheat and barley genome and transcriptome data of abscission zone in E. sibiricus, we identified 30 candidate genes for seed shattering, of which 15, 7, 6 and 2 genes were involved in plant hormone signal transcription, transcription factor, hydrolase activity and lignin biosynthetic pathway, respectively. CONCLUSION: This study constructed the first high-density genetic linkage map and identified QTLs and candidate genes for seed-related traits in E. sibiricus. Results of this study will not only serve as genome-wide resources for gene/QTL fine mapping, but also provide a genetic framework for anchoring sequence scaffolds on chromosomes in future genome sequence assembly of E. sibiricus.

Genome sequence of the opportunistic human pathogen Magnusiomyces capitatus.

The yeast Magnusiomyces capitatus is an opportunistic human pathogen causing rare yet severe infections, especially in patients with hematological malignancies. Here, we report the 20.2 megabase genome sequence of an environmental strain of this species as well as the genome sequences of eight additional isolates from human and animal sources providing an insight into intraspecies variation. The distribution of single-nucleotide variants is indicative of genetic recombination events, supporting evidence for sexual reproduction in this heterothallic yeast. Using RNAseq-aided annotation, we identified genes for 6518 proteins including several expanded families such as kexin proteases and Hsp70 molecular chaperones. Several of these families are potentially associated with the ability of M. capitatus to infect and colonize humans. For the purpose of comparative analysis, we also determined the genome sequence of a closely related yeast, Magnusiomyces ingens. The genome sequences of M. capitatus and M. ingens exhibit many distinct features and represent a basis for further comparative and functional studies.

Comparative genome analysis reveals key genetic factors associated with probiotic property in Enterococcus faecium strains.

BACKGROUND: Enterococcus faecium though commensal in the human gut, few strains provide a beneficial effect to humans as probiotics while few are responsible for the nosocomial infection. Comparative genomics of E. faecium can decipher the genomic differences responsible for probiotic, pathogenic and non-pathogenic properties. In this study, we compared E. faecium strain 17OM39 with a marketed probiotic, non-pathogenic non-probiotic (NPNP) and pathogenic strains. RESULTS: E. faecium 17OM39 was found to be closely related with marketed probiotic strain T110 based on core genome analysis. Strain 17OM39 was devoid of known vancomycin, tetracycline resistance and functional virulence genes. Moreover, E. faecium 17OM39 genome was found to be more stable due to the absence of frequently found transposable elements. Genes imparting beneficial functional properties were observed to be present in marketed probiotic T110 and 17OM39 strains. Genes associated with colonization and survival within gastrointestinal tract was also detected across all the strains. CONCLUSIONS: Beyond shared genetic features; this study particularly identified genes that are responsible for imparting probiotic, non-pathogenic and pathogenic features to the strains of E. faecium. Higher genomic stability, absence of known virulence factors and antibiotic resistance genes and close genomic relatedness with marketed probiotics makes E. faecium 17OM39 a potential probiotic candidate. The work presented here demonstrates that comparative genome analyses can be applied to large numbers of genomes, to find potential probiotic candidates.

Elucidation of the anti-hyperammonemic mechanism of Lactobacillus amylovorus JBD401 by comparative genomic analysis.

BACKGROUND: Recent experimental evidence showed that lactobacilli could be used as potential therapeutic agents for hyperammonemia. However, lack of understanding on how lactobacilli reduce blood ammonia levels limits application of lactobacilli to treat hyperammonemia. RESULTS: We report the finished and annotated genome sequence of L. amylovorus JBD401 (GenBank accession no. CP012389). L. amylovorus JBD401 reducing blood ammonia levels dramatically was identified by high-throughput screening of several thousand probiotic strains both within and across Lactobacillus species in vitro. Administration of L. amylovorus JBD401 to hyperammonemia-induced mice reduced the blood ammonia levels of the mice to the normal range. Genome sequencing showed that L. amylovorus JBD401 had a circular chromosome of 1,946,267 bp with an average GC content of 38.13%. Comparative analysis of the L. amylovorus JBD401 genome with L. acidophilus and L. amylovorus strains showed that L. amylovorus JBD401 possessed genes for ammonia assimilation into various amino acids and polyamines Interestingly, the genome of L. amylovorus JBD401 contained unusually large number of various pseudogenes suggesting an active stage of evolution. CONCLUSIONS: L. amylovorus JBD401 has genes for assimilation of free ammonia into various amino acids and polyamines which results in removal of free ammonia in intestinal lumen to reduce the blood ammonia levels in the host. This work explains the mechanism of how probiotics reduce blood ammonia levels.

Comparative genomics analysis to differentiate metabolic and virulence gene potential in gastric versus enterohepatic Helicobacter species.

BACKGROUND: The genus Helicobacter are gram-negative, microaerobic, flagellated, mucus-inhabiting bacteria associated with gastrointestinal inflammation and classified as gastric or enterohepatic Helicobacter species (EHS) according to host species and colonization niche. While there are over 30 official species, little is known about the physiology and pathogenic mechanisms of EHS, which account for most in the genus, as well as what genetic factors differentiate gastric versus EHS, given they inhabit different hosts and colonization niches. The objective of this study was to perform a whole-genus comparative analysis of over 100 gastric versus EHS genomes in order to identify genetic determinants that distinguish these Helicobacter species and provide insights about their evolution/adaptation to different hosts, colonization niches, and mechanisms of virulence. RESULTS: Whole-genome phylogeny organized Helicobacter species according to their presumed gastric or EHS classification. Analysis of orthologs revealed substantial heterogeneity in physiological and virulence-related genes between gastric and EHS genomes. Metabolic reconstruction predicted that unlike gastric species, EHS appear asaccharolytic and dependent on amino/organic acids to fuel metabolism. Additionally, gastric species lack de novo biosynthetic pathways for several amino acids and purines found in EHS and instead rely on environmental uptake/salvage pathways. Comparison of virulence factor genes between gastric and EHS genomes identified overlapping yet distinct profiles and included canonical cytotoxins, outer membrane proteins, secretion systems, and survival factors. CONCLUSIONS: The major differences in predicted metabolic function suggest gastric species and EHS may have evolved for survival in the nutrient-rich stomach versus the nutrient-devoid environments, respectively. Contrasting virulence factor gene profiles indicate gastric species and EHS may utilize different pathogenic mechanisms to chronically infect hosts and cause inflammation and tissue damage. The findings from this study provide new insights into the genetic differences underlying gastric versus EHS and support the need for future experimental studies to characterize these pathogens.

Pectin methylesterase inhibitor (PMEI) family can be related to male sterility in Chinese cabbage (Brassica rapa ssp. pekinensis).

Pectin-related genes play significant roles in pollen development and pollen tube growth, and their allelic variations are one of the major reasons for the abnormal development of male gametophyte. Currently, little is known about the role of the PMEI family in male sterility of plants. In this study, 97 putative PMEI genes were identified in Brassica rapa genome. By a phylogenetic analysis, the PMEI family was divided into 10 clades with highly conserved structural characteristics. The publically available RNA-seq data on different tissues of B. rapa accession Chiifu-401-42 revealed that 23 PMEI isoforms were flower-specific genes. We created a recessive genic male sterile mutant (ftms) in Chinese cabbage. This mutant was a doubled haploid line with stable inheritance, derived from Chinese cabbage 'FT' generated through a combination of radiation mutagenesis and isolated microspore culture. The transcriptome profiles of the floral buds of ftms and its wild-type line 'FT' were determined using RNA-seq. A total of 17 PMEI genes were found to be differentially expressed; all of them were down-regulated in ftms compared to their levels in 'FT'. Consistent with the transcriptome data, all these genes were observed to be highly expressed in the floral buds of 'FT' using qRT-PCR analysis. Of these, eight genes were specifically expressed in the floral buds of 'FT'; three of these (Bra019903, Bra014099, and Bra032239) were stamen-specific genes. The results contribute to further elucidation of the regulatory mechanisms underlying male sterility in Chinese cabbage.