Morphological and molecular identification of new records and new host plants of powdery mildews (Erysiphaceae) from Mexico

Powdery mildew is one of the most severe diseases affecting Cucurbitaceae. We identify the members of Erysiphaceae infecting cucurbits in three producing regions in Mexico. We determined that Golovinomyces ambrosiae, Neoerysiphe sechii, and Podosphaera xanthii infected cultivated and wild cucurbits species or subspecies. Leaf samples showing symptoms and signs of powdery mildew were collected from the northwestern, central western, and eastern regions of Mexico between 2017 and 2020. Species associated with the disease were identified based on morphology and ITS rDNA sequences. All powdery mildew specimens presented only the anamorph. Podosphaera xanthii was the predominant species; it was found in 85.7 % of the samples and in 13 out of 14 species or subspecies of cucurbits. Neoerysiphe sechii was found in 15.23 % of the samples and in only four cucurbit species from the central western and eastern regions of Mexico. Golovinomyces ambrosiae was documented for the first time in a Cucurbitaceae species (Sicyos deppei) from Central West Mexico. Three new hosts were registered for Neoerysiphe sechii (Cucurbita ficifolia, Echinopepon milleflorus, and Sicyos laciniatus) and eight for Podosphaera xanthii (Cucurbita argyrosperma, C. argyrosperma subsp. sororia, C. ficifolia, C. okeechobeensis subsp. martinezii, C. radicans, Sechium edule, Sicyos laciniatus, and S. deppei).

Identification of candidate type 3 effectors that determine host specificity associated with emerging Ralstonia pseudosolanacearum strains

Ralstonia pseudosolanacearum (Rps), previously known as R. solanacearum phylotypes I and III is one of the causal agents of bacterial wilt, a devastating disease that affects more than 250 plant species. Emerging Rps strains were identified infecting new hosts. P824 Rps strain was isolated from blueberry in Florida. Rps strains including PD7123 were isolated from hybrid tea roses in several countries through Europe. P781 is a representative strain of Rps commonly found on mandevilla in Florida. UW757 is a strain isolated from osteospermum plants originating in Guatemala. These strains are phylogenetically closely related and of economic importance on their respective hosts. The objective of this study is to associate the Type 3 Effectors (T3Es) repertoire of these four strains with host specificity. Candidate T3E associated with host specificity to blueberry, tea rose, osteospermum, and mandevilla were identified by sequence homology. Pathogenicity assays on 8 hosts including, blueberry, mandevilla, osteospermum and tea rose with the 4 strains showed that both P824 and PD7123 are pathogenic to blueberry and tea rose. P781 is the only strain pathogenic to mandevilla and P824 is the only strain non-pathogenic to osteospermum. Hypotheses based on correlation of T3E presence/absence and pathogenicity profiles identified 3 candidate virulence and 3 avirulence T3E for host specificity to blueberry and tea rose. Two candidate avirulence T3E were identified for mandevilla, and one candidate virulence for osteospermum. The strategy applied here can be used to reduce the number of host specificity candidate genes in closely related strains with different hosts.

A novel pneumococcal surface protein K of nonencapsulated Streptococcus pneumoniae promotes transmission among littermates in an infant mouse model with influenza A virus co-infection

We established an infant mouse model for colonization and transmission by nonencapsulated Streptococcus pneumoniae (NESp) strains to gain important information about its virulence among children. Invasive pneumococcal diseases have decreased dramatically since the worldwide introduction of pneumococcal capsular polysaccharide vaccines. Increasing prevalence of non-vaccine serotypes including NESp has been highlighted as a challenge in treatment strategy, but the virulence of NESp is not well understood. Protective strategy against NESp colonization and transmission between children require particularly urgent evaluation. NESp lacks capsules, a major virulent factor of pneumococci, but can cause a variety of infections in children and older people. PspK, a specific surface protein of NESp, is a key factor in establishing nasal colonization. In our infant mouse model for colonization and transmission by NESp strains, NESp could establish stable nasal colonization at the same level as encapsulated serotype 6A in infant mice, and could be transmitted between littermates. Transmission was promoted by NESp surface virulence factor PspK and influenza virus co-infection. However, PspK-deletion mutants lost the ability to colonize and transmit to new hosts. Promotion of NESp transmission by influenza was due to increased susceptibility of the new hosts. PspK was a key factor not only in establishment of nasal colonization, but also in transmission to new hosts. PspK may be targeted as a new candidate vaccine for NESp infection in children.

SARS-CoV-2 Variants of Concern Infect the Respiratory Tract and Induce Inflammatory Response in Wild-Type Laboratory Mice

The emergence of new severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants of concern pose a major threat to public health, due to possible enhanced virulence, transmissibility and immune escape. These variants may also adapt to new hosts, in part through mutations in the spike protein. In this study, we evaluated the infectivity and pathogenicity of SARS-CoV-2 variants of concern in wild-type C57BL/6 mice. Six-week-old mice were inoculated intranasally with a representative virus from the original B.1 lineage, or the emerging B.1.1.7 and B.1.351 lineages. We also infected a group of mice with a mouse-adapted SARS-CoV-2 (MA10). Viral load and mRNA levels of multiple cytokines and chemokines were analyzed in the lung tissues on day 3 after infection. Our data show that unlike the B.1 virus, the B.1.1.7 and B.1.351 viruses are capable of infecting C57BL/6 mice and replicating at high concentrations in the lungs. The B.1.351 virus replicated to higher titers in the lungs compared with the B.1.1.7 and MA10 viruses. The levels of cytokines (IL-6, TNF-α, IL-1β) and chemokine (CCL2) were upregulated in response to the B.1.1.7 and B.1.351 infection in the lungs. In addition, robust expression of viral nucleocapsid protein and histopathological changes were detected in the lungs of B.1.351-infected mice. Overall, these data indicate a greater potential for infectivity and adaptation to new hosts by emerging SARS-CoV-2 variants.

Biology and engineering of integrative and conjugative elements: Construction and analyses of hybrid ICEs reveal element functions that affect species-specific efficiencies

Integrative and conjugative elements (ICEs) are mobile genetic elements that reside in a bacterial host chromosome and are prominent drivers of bacterial evolution. They are also powerful tools for genetic analyses and engineering. Transfer of an ICE to a new host involves many steps, including excision from the chromosome, DNA processing and replication, transfer across the envelope of the donor and recipient, processing of the DNA, and eventual integration into the chromosome of the new host (now a stable transconjugant). Interactions between an ICE and its hosts throughout the life cycle likely influence the efficiencies of acquisition by new hosts. Here, we investigated how different functional modules of two ICEs, Tn916 and ICEBs1, affect the transfer efficiencies into different host bacteria. We constructed hybrid elements that utilize the high-efficiency regulatory and excision modules of ICEBs1 and the conjugation genes of Tn916. These elements produced more transconjugants than Tn916, likely due to increased excision frequencies. We also found that several Tn916 and ICEBs1 components can substitute for one other. Using B. subtilis donors and three Enterococcus species as recipients, we found that different hybrid elements were more readily acquired by some species than others, demonstrating species-specific interactions in steps of the ICE life cycle. This work demonstrates that hybrid elements utilizing the efficient regulatory functions of ICEBs1 can be built to enable efficient transfer into and engineering of a variety of other species.

Establishment and new hosts of the non-native seed beetle Stator limbatus (Coleoptera, Chrysomelidae, Bruchinae) on acacias in Europe

Stator limbatus is a phytophagous beetle native to warm regions of North and Central America, feeding on Fabaceae seeds and one of the most polyphagous species within the subfamily Bruchinae, here reported for the first time in Europe and on new hosts. Adult beetles emerged from Acacia spp. seeds collected in the islands of Corsica (France), and Sardinia (Italy). The wide presence in Sardinia and Corsica supports the hypothesis that this alien species was introduced several years ago. In both islands, S. limbatus emerged from Acacia mearnsii seeds, with infestation rates of up to 74.2 and 90.8% in 2019 and 2020, respectively. This seed beetle also emerged from two previously unreported host species, Acacia saligna and A. pycnantha, showing highest infestation rates of 4.0 and 95.1%, respectively. Both Acacia species are reported as new host associations with S. limbatus. Overall, seed infestation rates recorded in 2019 and 2020 indicate that S. limbatus is well established and that Mediterranean bioclimatic conditions are suitable for its population increase in size. This study lays the foundations for further research on known and potential host species and the spread and distribution of S. limbatus in Europe.

qnrA gene diversity in Shewanella spp.

Members of Shewanella are ubiquitous in aquatic environments, some of which have been implicated in human infections. The progenitors of antibiotic resistance genes with clinical relevance, such as qnrA genes, have been identified in Shewanella. qnrA code for a pentapeptide repeat protein that protects type II topoisomerases, decreasing susceptibility to quinolones and fluoroquinolones. In this study, 248 genomes of 49 Shewanella species were analysed as well as 33 environmental isolates belonging to 10 Shewanella species. The presence of the qnrA gene was detected in 22.9% of the genomes and 15.2% of the isolates. The gene was more often detected in Shewanella algae , but was also detected in Shewanella carassii , Shewanella chilikensis , Shewanella haliotis and Shewanella indica . The identified genes encoded the previously described variants QnrA3 (in 22 genomes of one species), QnrA2 (eight genomes and three species), QnrA1 (six genomes and two species), QnrA7 (five genomes and two species), QnrA10 (two genomes of one species) and QnrA4 (one genome). In addition, 11 novel variants with 3 to 7 amino acid substitutions were identified (in 13 genomes and one environmental isolate). The presence of this gene appears to be species-specific although within some species several variants were detected. The study presents a previously unknown diversity of qnrA in Shewanella , highlighting the role of this genus as progenitor and reservoir of these genes. Further studies are needed to determine the phenotypes conferred by the new variants and the mechanisms that may mediate the transfer of these genes to new hosts.

Preliminary Characterization of Phage-like Particles from the Male-Killing Mollicute Spiroplasma poulsonii (an Endosymbiont of Drosophila)

Viruses are vastly abundant and influential in all ecosystems, and are generally regarded as pathogens. Viruses of prokaryotes (themselves highly diverse and abundant) are known as bacteriophages or phages. Phages engage in diverse associations with their hosts, and contribute to regulation of biogeochemical processes, horizontal movement of genes, and control of bacterial populations. Recent studies have revealed the influential role of phage in the association of arthropods and their heritable endosymbiotic bacteria (e.g. the Proteobacteria genera Wolbachia and Hamiltonella). Despite prior studies (~30 years ago) documenting presence of phage in the mollicute Spiroplasma infecting Drosophila, genomic sequences of such phage are lacking, and their effects on the Spiroplasma-Drosophila interaction have not been comprehensively characterized. The present work isolated phage-like particles from the male-killing Spiroplasma poulsonii (strains NSRO and MSRO-Br) harbored by Drosophila melanogaster. Isolated particles were subjected to DNA sequencing, assembly, and annotation. Our results recovered three ~19 kb phage-like contigs (two in NSRO and one in MSRO-Br), and two smaller non-phage-like contigs encoding a known Spiroplasma toxin and an insertion element. Whole or parts of the particle-derived contigs were found in the genome assemblies of members of the Spiroplasma poulsonii clade. Although our results do not allow us to distinguish whether the contigs obtained represent infective phage-like particles capable of transmitting their DNA to new hosts, their encoding of several typical phage genes suggests that they are at least remnants of functional phage. We discuss potential implications of our findings and suggest future directions.

Staphylococcal Phages Adapt to New Hosts by Extensive Attachment Site Variability

A growing number of humans are being infected by antibiotic resistant Staphylococcus aureus originating from livestock. The preference of S. aureus for humans or animals is in part determined by factors encoded by viruses (phages) that reside in the bacterial genome.