Expression of a crown gall biological control phenotype in an avirulent strain of Agrobacterium vitis by addition of the trifolitoxin production and resistance genes

BACKGROUND: Agrobacterium vitis is a causal agent of crown-gall disease. Trifolitoxin (TFX) is a peptide antibiotic active only against members of a specific group of α-proteobacteria that includes Agrobacterium and its close relatives. The ability of TFX production by an avirulent strain of Agrobacterium to reduce crown gall disease is examined here. RESULTS: TFX was shown to be inhibitory in vitro against several A. vitis strains. TFX production, expressed from the stable plasmid pT2TFXK, conferred biological control activity to an avirulent strain of A. vitis. F2/5, against three virulent, TFX-sensitive strains of A. vitis tested on Nicotiana glauca. F2/5(pT2TFXK) is significantly reduces number and size of galls when co-inoculated with tumorigenic strain CG78 at a 10:1 ratio, but is ineffective at 1:1 or 1:10 ratios. F2/5(pT2TFXK) is effective when co-inoculated with tumorigenic strain CG435 at 10:1 and 1:1 ratios, but not at a 1:10 ratio. When F2/5(pT2TFXK) is co-inoculated with CG49 at a 10:1 ratio, the incidence of gall formation does not decline but gall size decreases by more than 70%. A 24 h pre-inoculation with F2/5(pT2TFXK) does not improve biological control at the 1:10 ratio. CONCLUSIONS: TFX production by an avirulent strain of Agrobacterium does confer in that strain the ability to control crown gall disease on Nicotiana glauca. This is the first demonstration that the production of a ribosomally synthesized, post-translationally modified peptide antibiotic can confer reduction in plant disease incidence from a bacterial pathogen

Identification of the Genes Required for the Culture of Liberibacter crescens, the Closest Cultured Relative of the Liberibacter Plant Pathogens

Here Tn5 random transposon mutagenesis was used to identify the essential elements for culturing Liberibacter crescens BT-1 that can serve as antimicrobial targets for the closely related pathogens of citrus, Candidatus Liberibacter asiaticus (Las) and tomato and potato, Candidatus Liberibacter solanacearum (Lso). In order to gain insight on the virulence, metabolism, and culturability of the pathogens within the genus Liberibacter, a mini-Tn5 transposon derivative system consisting of a gene specifying resistance to kanamycin, flanked by a 19-base-pair terminal repeat sequence of Tn5, was used for the genome-wide mutagenesis of L. crescens BT-1 and created an insertion mutant library. By analyzing the location of insertions using Sanger and Illumina Mi-Seq sequencing, 314 genes are proposed as essential for the culture of L. crescens BT-1 on BM-7 medium. Of those genes, 76 are not present in the uncultured Liberibacter pathogens and, as a result, suggest molecules necessary for the culturing these pathogens. Those molecules include the aromatic amino acids, several vitamins, histidine, cysteine, lipopolysaccharides, and fatty acids. In addition, the 238 essential genes of L. crescens in common with L. asiaticus are potential targets for the development of therapeutics against the disease

Complete genome sequence of Liberibacter crescens BT-1

Liberibacter crescens BT-1, a Gram-negative, rod-shaped bacterial isolate, was previously recovered from mountain papaya to gain insight on Huanglongbing (HLB) and Zebra Chip (ZC) diseases. The genome of BT-1 was sequenced at the Interdisciplinary Center for Biotechnology Research (ICBR) at the University of Florida. A finished assembly and annotation yielded one chromosome with a length of 1,504,659 bp and a G+C content of 35.4%. Comparison to other species in the Liberibacter genus, L. crescens has many more genes in thiamine and essential amino acid biosynthesis. This likely explains why L. crescens BT-1 is culturable while the known Liberibacter strains have not yet been cultured. Similar to Candidatus L. asiaticus psy62, the L. crescens BT-1 genome contains two prophage regions

Drivers of archaeal ammonia-oxidizing communities in soil

Soil ammonia-oxidizing archaea (AOA) are highly abundant and play an important role in the nitrogen cycle. In addition, AOA have a significant impact on soil quality. Nitrite produced by AOA and further oxidized to nitrate can cause nitrogen loss from soils, surface and groundwater contamination, and water eutrophication. The AOA discovered to date are classified in the phylum Thaumarchaeota. Only a few archaeal genomes are available in databases. As a result, AOA genes are not well annotated, and it is difficult to mine and identify archaeal genes within metagenomic libraries. Nevertheless, 16S rRNA and comparative analysis of ammonia monooxygenase sequences show that soils can vary greatly in the relative abundance of AOA. In some soils, AOA can comprise more than 10% of the total prokaryotic community. In other soils, AOA comprise less than 0.5% of the community. Many approaches have been used to measure the abundance and diversity of this group including DGGE, T-RFLP, q-PCR, and DNA sequencing. AOA have been studied across different soil types and various ecosystems from the Antarctic dry valleys to the tropical forests of South America to the soils near Mount Everest. Different studies have identified multiple soil factors that trigger the abundance of AOA. These factors include pH, concentration of available ammonia, organic matter content, moisture content, nitrogen content, clay content, as well as other triggers. Land use management appears to have a major effect on the abundance of AOA in soil, which may be the result of nitrogen fertilizer used in agricultural soils. This review summarizes the published results on this topic and suggests future work that will increase our understanding of how soil management and edaphoclimatic factors influence AOA

Important denominator between autoimmune comorbidities: a review of class II HLA, autoimmune disease, and the gut

Human leukocyte antigen (HLA) genes are associated with more diseases than any other region of the genome. Highly polymorphic HLA genes produce variable haplotypes that are specifically correlated with pathogenically different autoimmunities. Despite differing etiologies, however, many autoimmune disorders share the same risk-associated HLA haplotypes often resulting in comorbidity. This shared risk remains an unanswered question in the field. Yet, several groups have revealed links between gut microbial community composition and autoimmune diseases. Autoimmunity is frequently associated with dysbiosis, resulting in loss of barrier function and permeability of tight junctions, which increases HLA class II expression levels and thus further influences the composition of the gut microbiome. However, autoimmune-risk-associated HLA haplotypes are connected to gut dysbiosis long before autoimmunity even begins. This review evaluates current research on the HLA-microbiome-autoimmunity triplex and proposes that pre-autoimmune bacterial dysbiosis in the gut is an important determinant between autoimmune comorbidities with systemic inflammation as a common denominator

Influence of Fecal Sample Storage on Bacterial Community Diversity

Previous studies have identified a correlation, either positive or negative, between specific stool bacteria strains and certain autoimmune diseases. These conflicting data may relate to sample collection. The aim of this work was to evaluate the influence of the collection parameters of time and temperature on bacterial community composition. Samples were taken from healthy children and immediately divided in 5 sub-samples. One sample was frozen immediately at -80°C, while the other aliquots were frozen 12, 24, 48, and 72h later DNA extracted from each sample was used to amplify the 16S rRNA with barcoded primers. The amplified products were pooled and partial 16S rRNA sequences were obtained by pyrosequencing. Person-to-person variability in community diversity was high. A list of those taxa that comprise at least 1% of the community was made for each individual. None of these were present in high numbers in all individuals. The Bacteroides were present in the highest abundance in three of four subjects. A total of 23,701 16S rRNA sequences were obtained with an average of 1,185 reads per sample with an average length of 200 bases. Although pyrosequencing of amplified 16S rRNA identified changes in community composition over time (~10%), little diversity change was observed at 12 hours (3.06%) with gradual changes occurring after 24 (8.61%), 48 (9.72%), and 72 h (10.14%), post collection