Brian H Kvitko

University of Georgia | UGA · Department of Plant Pathology

Plant pathogenic bacteria encounter a drastic increase in apoplastic pH during the early stages of plant immunity. The effects of alkalization on pathogen-host interactions have not been comprehensively characterized. Here, we used a global transcriptomic approach to assess the impact of environmental alkalization on Pseudomonas syringae pv. tomato...

Slippery skin of onion caused by Burkholderia gladioli pv. alliicola (Bga) is a common bacterial disease reported from onion growing regions around the world. Despite the increasing attention in recent years, our understanding of the virulence mechanisms of this pathogen remains limited. In this study, we characterized the predicted genetic determi...

Allicin tolerance ( alt ) clusters in phytopathogenic bacteria, which provide resistance to thiosulfinates like allicin, are challenging to find using conventional approaches due to their varied architecture and the paradox of being vertically maintained within genera despite likely being horizontally transferred. This results in significant sequen...

Glacial ice preserves time-sequenced records of preserved microbes, offering access to historic pre-anthropic metagenomes. As proof-of-concept, we three tested methods to recover ancient DNA from a ~250-year ice core from Wyoming's Upper Fremont Glacier for metagenomic sequencing. Direct amplification of filter-concentrated melt water (eDNA) was a...

Pantoea agglomerans is one of four Pantoea species for which strains have been reported in the United States to cause bacterial rot of onion bulbs. However, not all P. agglomerans strains are pathogenic to onion. We characterized onion-associated strains of P. agglomerans to elucidate the genetic and genomic signatures of onion-pathogenic P. agglom...

Members of the bacterial genus Burkholderia are a routine threat to onion production worldwide. In addition to the common onion‐pathogenic species, Burkholderia cepacia , Burkholderia orbicola and Burkholderia gladioli , other Burkholderia species have the potential to cause onion disease. Despite their impacts and long‐known association with onion...

Microbes occur in complex communities within plants as endophytes and establish a network of interactions that can influence plant health positively or negatively. Positive interactions may be synergistic and confer abiotic and biotic stress tolerance. The aim of this study was to identify bacteria and viruses present in storage onion bulbs that we...

Burkholderia gladioli pv. alliicola, B. cepacia, and B. orbicola are common bacterial pathogens of onion. Onions produce organosulfur thiosulfinate defensive compounds after cellular decompartmentalization. Using whole-genome sequencing and in silico analysis, we identified putative thiosulfinate tolerance gene (TTG) clusters in multiple onion-asso...

Burkholderia gladioli pv. alliicola, B.cepacia, and B. orbicola are common bacterial pathogens of onion. Onions produce organosulfur thiosulfinate defensive compounds after cellular decompartmentalization. Using whole genome sequencing and in silico analysis, we identified putative thiosulfinate tolerance gene (TTG) clusters in multiple onion-assoc...

As the name of the genus Pantoea (“of all sorts and sources”) suggests, this genus includes bacteria with a wide range of provenances, including plants, animals, soils, components of the water cycle, and humans. Some members of the genus are pathogenic to plants, and some are suspected to be opportunistic human pathogens; while others are used as m...

Temporospatial regulation of immunity components is essential for properly activating plant defense response. Flagellin-sensing 2 (FLS2) is a surface-localized receptor that recognizes bacterial flagellin. The immune function of FLS2 is compromised in early stages of shoot development. However, the underlying mechanism for the age-dependent FLS2 si...

Phage-derived bacteriocins are highly specific and effective antimicrobial molecules which have successfully been used as prophylactic treatments to prevent phytopathogen infections. Given the specificity of tailocins, a necessary step for broadening the tailocin catalog and for extending applicability across systems and diseases is the screening o...

In the early 1960s, Pseudomonas syringae and other host-specific phytopathogenic proteobacteria were discovered to elicit a rapid, resistance-associated death when infiltrated at high inoculum levels into nonhost tobacco leaves. This hypersensitive reaction (or response; HR) was a useful indicator of basic pathogenic ability. Research over the next...

Although many studies have shown that microbes can ectopically stimulate or suppress plant immune responses, the fundamental question of whether the entire preexisting microbiota is indeed required for proper development of plant immune response remains unanswered. Using a recently developed peat-based gnotobiotic plant growth system we found that...

Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of the phosphonate toxin, pantaphos, which is synthesized by putative enzymes encoded by the HiVir (high virulence) gene cluster. The...

Center rot of onion is caused by plant pathogenic Pantoea species which can lead to significant yield losses in the field and during storage. Conventional growers use foliar protectants such as a mixture of copper bactericides and an ethylene-bis-dithiocarbamate (EBDC) fungicide to manage the disease; however, organic growers have limited managemen...

Pantoea ananatis is a member of a Pantoea species complex that causes center rot of bulb onions ( A. cepa ) and also infects other Allium crops like leeks ( Allium porrum ), chives ( Allium schoenoprasum ), bunching onion or Welsh onion ( Allium fistulosum ), and garlic ( Allium sativum ). This pathogen relies on a chromosomal phosphonate biosynthe...

In this work, we report onion pathogenicity determinants in P. stewartii subsp. indologenes and P. allii. We identified two distinct secondary metabolite biosynthetic gene clusters present separately in different strains of onion pathogenic P. stewartii subsp. indologenes. One cluster is similar to the previously described HiVir phosphonate biosynt...

Onion bulb rot can be caused by multiple plant pathogens including bacterial pathogens. During our routine survey of commercial onion farms in 2014, 2020, and 2021, seven putative Rouxiella spp. strains were isolated from symptomatic onion samples in Georgia, United States. Upon fulfilling Koch’s postulates on onion, a genome analysis was conducted...

Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of a phosphonate toxin pantaphos that is catalyzed by putative enzymes encoded by the HiVir gene cluster. The genetic contributions o...

Contaminated fresh produce has been routinely linked to outbreaks of Salmonellosis. Multiple studies have identified Salmonella enterica factors associated with successful colonization of diverse plant niches and tissues. It has also been well documented that S. enterica can benefit from the conditions generated during plant disease by host-compati...

In previous work, we determined the transcriptomic impacts of flg22 pre-induced Pattern Triggered Immunity (PTI) in Arabidopsis thaliana on the pathogen Pseudomonas syringae pv. tomato DC3000 (Pto). During PTI exposure we observed expression patterns in Pto reminiscent of those previously observed in a Pto algU mutant. AlgU is a conserved extracyto...

Pseudomonas syringae is a diverse phytopathogenic species complex, and includes strains that can cause disease across a wide variety of plant species. Much previous research into the molecular basis of immunity and infection has focused on pathogen and plant responses in a handful of model strains and hosts, and with a tacit assumption that early s...

Onion center rot is caused by at least four species of Pantoea ( P. ananatis, P. agglomerans, P. allii, and P. stewartii subsp. indologenes ). Critical onion pathogenicity determinants for P. ananatis were recently described but whether those determinants are common among other onion-pathogenic Pantoea species remains unknown. In this work, we repo...

Integrative Conjugative Elements (ICEs) are replicons that can insert and excise from chromosomal locations in a site-specific manner, can conjugate across strains, and which often carry a variety of genes useful for bacterial growth and survival under specific conditions. Although ICEs have been identified and vetted within certain clades of the a...

Species of Pantoea represent a group of plant pathogenic bacteria that infect a variety of agro-economically important plant species. Among these, a complex of P. ananatis, P. allii, P. agglomerans, and P. stewartii subsp. indologenes cause Center rot in onion, resulting in significant economic losses. As species of Pantoea are phenotypically close...

In previous work, we determined the transcriptomic impacts of flg22 pre-induced Pattern Triggered Immunity (PTI) in Arabidopsis thaliana on the pathogen Pseudomonas syringae pv. tomato DC3000 (Pto). During PTI exposure we observed expression patterns in Pto reminiscent of those previously observed in a Pto algU mutant. AlgU is a conserved extracyto...

Integrative Conjugative Elements (ICEs) are replicons that can insert and excise from chromosomal locations in a site specific manner, can conjugate across strains, and which often carry a variety of genes useful for bacterial growth and survival under specific conditions. Although ICEs have been identified and vetted within certain clades of the a...

This study provides a taxonomic characterization of three bacterial strains isolated from onion seedlings in Georgia USA. Yellow-colored colonies were isolated, and a diffusible fluorescent pigment was visible under ultraviolet light on King’s medium B. Preliminary analysis of the basic phenotype tests and 16S rRNA gene sequence analysis indicated...

Three phytopathogenic bacterial strains (Pc19-1 T , Pc19-2 and Pc19-3) were isolated from seedlings displaying water-soaked, dark brown-to-black, necrotic lesions on pepper ( Capsicum annuum ) leaves in Georgia, USA. Upon isolation on King’s medium B, light cream-coloured colonies were observed and a diffusible fluorescent pigment was visible under...

Pantoea ananatis, a gram negative and facultative anaerobic bacterium is a member of a Pantoea spp. complex that causes center rot of onion, which significantly affects onion yield and quality. This pathogen does not have typical virulence factors like type II or type III secretion systems but appears to require a biosynthetic gene-cluster, HiVir/P...

Bacterial phytopathogens deliver effector proteins into host cells as key virulence weapons to cause disease. Extensive studies revealed diverse functions and biochemical properties of different effector proteins from pathogens. In this study, we show that the Pseudomonas syringae effector AvrE, the founding member of a broadly conserved and pathol...

Cotton bacterial blight (CBB), caused by Xanthomonas citri pv. malvacearum (Xcm), was a major disease of cotton in the United States in the early part of the 20 th century. The recent reemergence of CBB revealed many gaps in our understanding of this important disease. In this study, we employed a wild type (WT) field isolate of Xcm from Georgia US...

Pantoea ananatis is a gram-negative bacterium and the primary causal agent of center rot of onions in Georgia. Previous genomic studies identified two virulence gene clusters, HiVir and alt, associated with center rot. The HiVir gene cluster is required to induce necrosis on onion tissues via synthesis of pantaphos, (2-hydroxy[phosphono-methyl)male...

Pantoea stewartii subsp. indologenes is a causative agent of leafspot of foxtail millet and pearl millet; however, novel strains were recently identified that are pathogenic on onion. We phenotypically and genotypically characterized seventeen Pantoea stewartii subsp. indologenes strains from onion and other sources (pearl millet, foxtail millet, g...

Cotton bacterial blight (CBB) was a major disease of cotton in the United States in the early part of the 20th century. The recent reemergence of CBB, caused by Xanthomonas citri pv. malvacearum ( Xcm ) revealed many gaps in our understanding of this important disease. In this study, we employed a field isolate of Xcm from Georgia USA (WT) to gener...

Pantoea ananatis is a gram-negative bacterium and the primary causal agent of center rot of onions in Georgia. Previous genomic studies identified two virulence gene clusters, HiVir and alt, associated with center rot. The HiVir gene cluster is required to induce necrosis on onion tissues via synthesis of a predicted small molecule toxin. The alt g...

Pseudomonas syringae can rapidly deploy specialized functions to deal with abiotic and biotic stresses. Host niches pose specific sets of environmental challenges driven, in part, by immune defenses. Bacteria use a “just-in-time” strategy of gene regulation, meaning that they only produce the functions necessary for survival as needed. Extracytopla...

With an increase in foodborne illnesses associated with the consumption of fresh produce, it is important to understand the interactions between human bacterial enteric pathogens and plants. It was previously established that diseased plants can create a permissive environment for opportunistic endophytic colonization of enteric pathogens. However,...

Background: Pantoea ananatis is a member of a Pantoea spp. complex that causes center rot of onion, which significantly affects onion yield and quality. This pathogen does not have typical virulence factors like type II or type III secretion systems but appears to require a biosynthetic gene-cluster, HiVir/PASVIL (located chromosomally), for a phos...

Pantoea ananatis is a member of a Pantoea spp. complex that causes center rot of onion, which significantly affects onion yield and quality. This pathogen does not have typical virulence factors like type II or type III secretion systems but appears to require a biosynthetic gene-cluster, HiVir/PASVIL (located chromosomally), for a phosphonate seco...

Unlike most characterized bacterial plant pathogens, the broad-host-range plant pathogen Pantoea ananatis lacks both the virulence-associated type III and type II secretion systems. In the absence of these typical pathogenicity factors, P. ananatis induces necrotic symptoms and extensive cell death in onion tissue dependent on the HiVir proposed se...

Onion ( Allium. cepa L), garlic ( A. sativum L.), and other members of the Allium genus produce volatile antimicrobial thiosulfinates upon cellular damage. Allicin has been known since the 1950s as the primary antimicrobial thiosulfinate compound and odorant produced by garlic. However, the roles of endogenous thiosulfinate production in host-bacte...

A hallmark of multicellular organisms is their ability to maintain physiological homeostasis by communicating among cells, tissues, and organs. In plants, intercellular communication is largely dependent on plasmodesmata (PD), which are membrane-lined channels connecting adjacent plant cells. Upon immune stimulation, plants close PD as part of thei...

Successful pathogens must efficiently defeat or delay host immune responses, including those triggered by release or exposure of microbe-associated molecular patterns (MAMPs). Knowledge of the molecular details leading to this phenomenon in genuine plant-pathogen interactions is still scarce. We took advantage of the well-established Arabidopsis th...

An error was observed in Step 2 of section 3.3. Direct cAMP Assay of this chapter. The same was replaced with the correction below.

Significance Plants have evolved a powerful innate immune system to defend against microbial pathogens. Despite extensive studies, how plant immunity ultimately inhibits bacterial pathogen growth is largely unknown, due to difficulties in profiling bacterial responses in planta . In this study, we established two methods for in planta bacterial tra...

Pseudomonas syringae is one of the best-studied plant pathogens and serves as a model for understanding host–microorganism interactions, bacterial virulence mechanisms and host adaptation of pathogens as well as microbial evolution, ecology and epidemiology. Comparative genomic studies have identified key genomic features that contribute to P. syri...

Pattern triggered Immunity (PTI) can confer broad defense against diverse microbes and pathogens with disparate lifestyles through the detection of microbial extracellular signatures by surface exposed pattern recognition receptors. However, unlike recognition of pathogen effectors by cytosolic resistance proteins, PTI is typically not associated w...

Pseudomonas syringae pv. tomato DC3000 is an important model plant pathogen, with a fully annotated genome and multiple compatible plant hosts. Very few studies have examined the regulation of DC3000 gene expression in vivo. We developed a quantitative reverse transcription-polymerase chain reaction assay to monitor transcriptional changes in DC300...

Pantoea ananatis is a member of the family Enterobacteriaceae and an enigmatic plant pathogen with a broad host range. Although P. ananatis strains can be aggressive on onion causing foliar necrosis and onion center rot, previous genomic analysis has shown that P. ananatis lacks the primary virulence secretion systems associated with other plant pa...

Large Pantoea Plasmid LPP-1 alignment of proto-type strain PNA 97-1 with AJ13355 pEA320 (Accession CP020943; NC_017533.1) Alignment generated with Geneious MAUVE plugin.

Venn diagram of genes shared among the pathogenic isolate PNA 97-1R; nonpathogenic isolates PANS 99-36, and genomic type strain LMG 20103 (Accession CP001875.2). The number of coding sequences are derived from the gene_presence_absense.csv output of the ROARY pan-genome analysis using default parameters (Supplementary File 2).

wgMLST cladogram of Pantoea strains generated through PGAdb-builder platform. Outgroups P. agglomerans P10C and P. stewartii subsp. indologens LMG 2632 (Accession NZ_LIME00000000.1 and JPKO00000000.1).

RAST functional categories for predicted protein coding sequences of Pantoea ananatis strains.

Phaster predicted phage regions in PNA 97-1R. Green indicated intact phage regions whereas red indicates incomplete phage regions.

Sequencing and assembly statistics for Illumina draft-genomes of Pantoea ananatis strains.

Primers and conditions used for amplification and sequencing.

IslandPath DIMOB genomic islands identified in WGS P. ananatis strains.

ROARY 10 strain WGS Analysis. The presence of each CDS in the respective genome is listed below each labeled strain column. If a cell is blank the respective strain does not have the gene. Logic equations were used to highlight cells, resulting in genes present in pathogenic strains and absent in non-pathogenic strains.

ROARY 3 strain WGS Analysis. The presence of each CDS in the respective genome is listed below each labeled strain column. If a cell is blank the respective strain does not have the gene. Summation equations were used to calculate the numeric values for the Venn Diagram in Supplementary Figure 3.

Dendrogram derived by cluster analysis of similarities between Pantoea ananatis strains based on repetitive extragenic palindromic polymerase chain reaction (REP-PCR) DNA genomic profiles. Analysis was performed using Dice's (1945) coefficient in BioNumerics software package (Applied Math, Kortrijk, Belgium).

BRIG analysis of P. ananatis strains and the OVR A-D loci to the reference strain PNA 97-1R. BRIG is a locally run analysis that generates figures based on blastn comparisons of query genomes to a reference strain.

Center rot of onion is an economically important disease caused by three Pantoea spp.: Pantoea ananatis, P. agglomerans, and P. allii. Symptoms caused by these three species are similar and include white streaking and necrosis of foliage; and, in some cases, the bacterium may enter the bulb, causing liquefaction and rot of bulb scales. Two bacteria...

Pseudomonas syringae pv. tomato DC3000 (DC3000) is an important model plant pathogen, with a fully annotated genome and multiple compatible plant hosts. Very few studies have examined the regulation of DC3000 gene expression in vivo . We developed a RT-qPCR assay to monitor transcriptional changes in DC3000 inoculated into Arabidopsis thaliana leav...

An accurate and complete roster of the Type III effector (T3E) proteins translocated by the P. syringae Type III secretion system (T3SS) into host cells is critical to understanding the pathogen’s interactions with plants. The adenylate cyclase (Cya) reporter offers a highly sensitive and robust assay for monitoring the translocation of T3Es. T3Es...

The plant cytoskeleton underpins the function of a multitude of cellular mechanisms, including those associated with developmental- and stress-associated signaling processes. In recent years, the actin cytoskeleton has been demonstrated to play a key role in plant immune signaling, including a recent demonstration that pathogens target actin filame...

Background The combination of imaging technologies and luciferase-based bioluminescent bacterial reporter strains provide a sensitive and simple non-invasive detection method (photonic bioimaging) for the study of diverse biological processes, as well as efficacy of therapeutic interventions, in live animal models of disease. The engineering of bio...

Whole genome sequencing revealed the presence of a genomic anomaly in the region of 4.7 to 4.9 Mb of the Pseudomonas syringae pv. tomato (Pst) DC3000 genome. The average read depth coverage of Pst DC3000 whole genome sequencing results suggested that a 165 kb segment of the chromosome had doubled in copy number. Further analysis confirmed the 165 k...

We have made significant improvements to a broad-host-range system for the cloning and manipulation of large bacterial genomic regions based on site-specific recombination between directly repeated oriT sites during conjugation. Using two suicide capture vectors carrying flanking homology regions, oriT sites are recombined on either side of the tar...

Pseudomonas syringae pv. tomato DC3000 produces the phytotoxin coronatine, a major determinant of the leaf chlorosis associated with DC3000 pathogenesis. The DC3000 PSPTO4723 (cmaL) gene is located in a genomic region encoding type III effectors; however, it promotes chlorosis in the model plant Nicotiana benthamiana in a manner independent of type...