Diversity of Strains in the Pseudomonas syringae Complex Causing Bacterial Stem Blight of Alfalfa (Medicago sativa) in the United States.

Alfalfa growers in the Intermountain West of the United States have recently seen an increased incidence in bacterial stem blight (BSB), which can result in significant herbage yield losses from the first harvest. BSB has been attributed to Pseudomonas syringae pv. syringae and P. viridiflava; however, little is known about the genetic diversity and pathogenicity of these bacteria or their interaction with alfalfa plants. Here, we present a comprehensive phylogenetic and phenotypic analysis of P. syringae and P. viridiflava strains causing BSB on alfalfa. A multilocus sequence analysis found that they grouped exclusively with P. syringae PG2b and P. viridiflava PG7a. Alfalfa symptoms caused by both bacterial groups were indistinguishable, although there was a large range in mean disease scores for individual strains. Overall, PG2b strains incited significantly greater disease scores than those caused by PG7a strains. Inoculated plants showed browning in the xylem and collapse of epidermal and pith parenchyma cells. Inoculation with a mixture of PG2b and PG7a strains did not result in synergistic activity. The populations of PG2b and PG7a strains were genetically diverse within their clades and did not group by location or haplotype. The PG2b strains had genes for production of the phytotoxin coronatine, which is unusual in PG2b strains. The results indicate that both pathogens are well established on alfalfa across a wide geographic range and that a recent introduction or evolution of more aggressive strains as the basis for emergence of the disease is unlikely.

Pseudomonas viridiflava: An internal outsider of the Pseudomonas syringae species complex.

Pseudomonas viridiflava is a gram-negative pseudomonad that is phylogenetically placed within the Pseudomonas syringae species complex. P. viridiflava has a wide host range and causes a variety of symptoms in different plant parts, including stems, leaves, and blossoms. Outside of its role as a pathogen, P. viridiflava also exists as an endophyte, epiphyte, and saprophyte. Increased reports of P. viridiflava causing disease on new hosts in recent years coincide with increased research on its genetic variability, virulence, phylogenetics, and phenotypes. There is high variation in its core genome, virulence factors, and phenotypic characteristics. The main virulence factors of this pathogen include the enzyme pectate lyase and virulence genes encoded within one or two pathogenicity islands. The delineation of P. viridiflava in the P. syringae complex has been investigated using several molecular approaches. P. viridiflava comprises its own species, within the complex. While seemingly an outsider to the complex as a whole due to differences in the core genome and virulence genes, low average nucleotide identity to other of P. syringae complex members, and some phenotypic traits, it remains as part of the complex. Defining phylogenetic, phenotypic, and genomic characteristics of P. viridiflava in comparison to other P. syringae members is important to understanding this pathogen and for the development of disease resistance and management practices. TAXONOMY: Kingdom Bacteria; Phylum Proteobacteria; Class Gammaproteobacteria; Family Pseudomonadaceae; Genus Pseudomonas; Species Pseudomonas syringae species complex, Genomospecies 6, Phylogroup 7 and 8. MICROBIOLOGICAL PROPERTIES: Gram-negative, fluorescent, aerobic, motile, rod-shaped, oxidase negative, arginine dihydrolase negative, levan production negative (or positive), potato rot positive (or negative), tobacco hypersensitivity positive. GENOME: There are two complete genomes, five chromosome-level genomes, and 1,540 genomes composed of multiple scaffolds of P. viridiflava available in the National Center for Biotechnology Information Genome database. The median total length of these assemblies is 5,975,050 bp, the median number of protein coding genes is 5,208, and the median G + C content is 59.3%. DISEASE SYMPTOMS: P. viridiflava causes a variety of disease symptoms, including spots, streaks, necrosis, rots, and more in above- and below-ground plant parts on at least 50 hosts. EPIDEMIOLOGY: There have been several significant disease outbreaks on field and horticultural crops caused by P. viridiflava since the turn of the century. P. viridiflava has been reported as a pathogen, epiphyte, endophyte, and saprophyte. This species has been isolated from a variety of environmental sources, including asymptomatic wild plants, snow, epilithic biofilms, and icepacks.

There is no evidence that Podoctidae carry eggs of their own species: Reply to Machado and Wolff (2017).

In our recent publication (Sharma et al., 2017), we tested the hypothesis that eggs attached to the legs of male Podoctidae (Opiliones, Laniatores) constituted a case of paternal care, using molecular sequence data in tandem with multiple sequence alignments to test the prediction that sequences of the eggs and the adults that carried them would indicate conspecific identity. We discovered that the sequences of the eggs belonged to spiders, and thus rejected the paternal care hypothesis for these species. Machado and Wolff (2017) recently critiqued our work, which they regarded as a non-critical interpretation and over-reliance on molecular sequence data, and defended the traditional argument that the eggs attached to podoctids are in fact harvestman eggs. Here we show that additional molecular sequence data also refute the identity of the eggs as conspecific harvestman eggs, using molecular cloning techniques to rule out contamination. We show that individual gene trees consistently and reliably place the egg and adult sequences in disparate parts of the tree topology. Phylogenetic methods consistently place all egg sequences within the order Araneae (spiders). We submit that evidence for the paternal care hypothesis based on behavioral, morphological, and natural history approaches is either absent or insufficient for concluding that the eggs of podoctids are conspecific.

A multilocus phylogeny of Podoctidae (Arachnida, Opiliones, Laniatores) and parametric shape analysis reveal the disutility of subfamilial nomenclature in armored harvestman systematics.

The taxonomy and systematics of the armored harvestmen (suborder Laniatores) are based on various sets of morphological characters pertaining to shape, armature, pedipalpal setation, and the number of articles of the walking leg tarsi. Few studies have tested the validity of these historical character systems in a comprehensive way, with reference to an independent data class, i.e., molecular sequence data. We examined as a test case the systematics of Podoctidae, a family distributed throughout the Indo-Pacific. We tested the validity of the three subfamilies of Podoctidae using a five-locus phylogeny, and examined the evolution of dorsal shape as a proxy for taxonomic utility, using parametric shape analysis. Here we show that two of the three subfamilies, Ibaloniinae and Podoctinae, are non-monophyletic, with the third subfamily, Erecananinae, recovered as non-monophyletic in a subset of analyses. Various genera were also recovered as non-monophyletic. As first steps toward revision of Podoctidae, the subfamilies Erecananinae Roewer, 1912 and Ibaloniinae Roewer, 1912 are synonymized with Podoctinae Roewer, 1912 new synonymies, thereby abolishing unsubstantiated subfamilial divisions within Podoctidae. We once again synonymize the genus Paralomanius Goodnight & Goodnight, 1948 with Lomanius Roewer, 1923 revalidated. We additionally show that eggs carried on the legs of male Podoctidae are not conspecific to the males, falsifying the hypothesis of paternal care in this group.