Reduced Odds of Mpox-Associated Hospitalization Among Persons Who Received JYNNEOS Vaccine - California, May 2022-May 2023.

The effectiveness of 1 dose of JYNNEOS vaccine (modified vaccinia Ankara vaccine, Bavarian Nordic) against hospitalization for mpox (caused by Monkeypox virus), has been demonstrated; however, the impact of 2 doses on hospitalization risk, especially among persons infected with HIV, who are at higher risk for severe disease, is an important factor in evaluating vaccine effectiveness against mpox disease severity and Monkeypox virus infection. Surveillance data collected by the California Department of Public Health were used to evaluate whether receipt of 2 doses of JYNNEOS vaccine reduced the odds of hospitalization among persons with mpox. The odds of hospitalization among persons with mpox who had received 1 or 2 JYNNEOS doses were 0.27 (95% CI = 0.08-0.65) and 0.20 (95% CI = 0.01-0.90), respectively, compared with unvaccinated mpox patients. In mpox patients with HIV infection, the odds of hospitalization among those who had received 1 JYNNEOS vaccine dose was 0.28 (95% CI = 0.05-0.91) times that of those who were unvaccinated. No mpox-associated hospitalizations were identified among persons infected with HIV who had received 2 JYNNEOS vaccine doses. To optimize durable immunity, all eligible persons at risk for mpox, especially those infected with HIV, should complete the 2-dose JYNNEOS series.

Diversity and Evolution of Type III Secreted Effectors: A Case Study of Three Families.

A broad range of Gram-negative bacteria employ a type III secretion system (T3SS) to deliver virulence proteins termed type III secreted effectors directly into the cytoplasm of eukaryotic host cells. While effectors can contribute to the colonization of eukaryotic hosts by bacterial symbionts and pathogens, they can also elicit host immune responses that restrict bacterial growth. These opposing selective pressures have shaped the evolution of effector families and may be responsible for their incredible diversity in biochemical function, mechanism of action, and taxonomic distribution. In this chapter, we focus on three distinct effector families whose members are distributed among both plant and animal pathogens. We first discuss the LRR-NEL and YopJ families of effectors. These two effector families possess ubiquitin ligase and acetyltransferase activity, respectively, which in both cases can be directed against host innate immune signal transduction pathways to promote infection. Finally, we discuss the TALE family of transcription activator-like effectors that serve to reprogram host immunity transcriptional responses. This chapter aims to highlight the diversity within these three effector families that results from the strong and dynamic evolutionary forces shaping the interface between host and bacterium.

A High-Throughput, Seedling Screen for Plant Immunity.

An understanding of how biological diversity affects plant-microbe interactions is becoming increasingly important, particularly with respect to components of the pathogen effector arsenal and the plant immune system. Although technological improvements have greatly advanced our ability to examine molecular sequences and interactions, relatively few advances have been made that facilitate high-throughput, in vivo pathology screens. Here, we present a high-throughput, microplate-based, nondestructive seedling pathology assay, and apply it to identify Arabidopsis thaliana effector-triggered immunity (ETI) responses against Pseudomonas syringae type III secreted effectors. The assay was carried out in a 48-well microplate format with spray inoculation, and disease symptoms were quantitatively recorded in a semiautomated manner, thereby greatly reducing both time and costs. The assay requires only slight modifications of common labware and uses no proprietary software. We validated the assay by recapitulating known ETI responses induced by P. syringae in Arabidopsis. We also demonstrated that we can quantitatively differentiate responses from a diversity of plant genotypes grown in the same microplate. Finally, we showed that the results obtained from our assay can be used to perform genome-wide association studies to identify host immunity genes, recapitulating results that have been independently obtained with mature plants.

Identifying Pseudomonas syringae Type III Secreted Effector Function via a Yeast Genomic Screen.

Gram-negative bacterial pathogens inject type III secreted effectors (T3SEs) directly into host cells to promote pathogen fitness by manipulating host cellular processes. Despite their crucial role in promoting virulence, relatively few T3SEs have well-characterized enzymatic activities or host targets. This is in part due to functional redundancy within pathogen T3SE repertoires as well as the promiscuity of individual T3SEs that can have multiple host targets. To overcome these challenges, we generated and characterized a collection of yeast strains stably expressing 75 T3SE constructs from the plant pathogen Pseudomonas syringae This collection is devised to facilitate heterologous genetic screens in yeast, a non-host organism, to identify T3SEs that target conserved eukaryotic processes. Among 75 T3SEs tested, we identified 16 that inhibited yeast growth on rich media and eight that inhibited growth on stress-inducing media. We utilized Pathogenic Genetic Array (PGA) screens to identify potential host targets of P. syringae T3SEs. We focused on the acetyltransferase, HopZ1a, which interacts with plant tubulin and alters microtubule networks. To uncover putative HopZ1a host targets, we identified yeast genes with genetic interaction profiles most similar (i.e., congruent) to the PGA profile of HopZ1a and performed a functional enrichment analysis of these HopZ1a-congruent genes. We compared the congruence analyses above to previously described HopZ physical interaction datasets and identified kinesins as potential HopZ1a targets. Finally, we demonstrated that HopZ1a can target kinesins by acetylating the plant kinesins HINKEL and MKRP1, illustrating the utility of our T3SE-expressing yeast library to characterize T3SE functions.

Expanded type III effector recognition by the ZAR1 NLR protein using ZED1-related kinases.

Nucleotide-binding domain and leucine-rich repeat domain-containing (NLR) proteins are sentinels of plant immunity that monitor host proteins for perturbations induced by pathogenic effector proteins. Here we show that the Arabidopsis ZAR1 NLR protein requires the ZRK3 kinase to recognize the Pseudomonas syringae type III effector (T3E) HopF2a. These results support the hypothesis that ZAR1 associates with an expanded ZRK protein family to broaden its effector recognition spectrum.

The HopF family of Pseudomonas syringae type III secreted effectors.

Pseudomonas syringae is a bacterial phytopathogen that utilizes the type III secretion system to inject effector proteins into plant host cells. Pseudomonas syringae can infect a wide range of plant hosts, including agronomically important crops such as tomatoes and beans. The ability of P. syringae to infect such numerous hosts is caused, in part, by the diversity of effectors employed by this phytopathogen. Over 60 different effector families exist in P. syringae; one such family is HopF, which contains over 100 distinct alleles. Despite this diversity, research has focused on only two members of this family: HopF1 from P. syringae pathovar phaseolicola 1449B and HopF2 from P. syringae pathovar tomato DC3000. In this study, we review the research on HopF family members, including their host targets and molecular mechanisms of immunity suppression, and their enzymatic function. We also provide a phylogenetic analysis of this expanding effector family which provides a basis for a proposed nomenclature to guide future research. The extensive genetic diversity that exists within the HopF family presents a great opportunity to study how functional diversification on an effector family contributes to host specialization.

Image-Based Quantification of Plant Immunity and Disease.

Measuring the extent and severity of disease is a critical component of plant pathology research and crop breeding. Unfortunately, existing visual scoring systems are qualitative, subjective, and the results are difficult to transfer between research groups, while existing quantitative methods can be quite laborious. Here, we present plant immunity and disease image-based quantification (PIDIQ), a quantitative, semi-automated system to rapidly and objectively measure disease symptoms in a biologically relevant context. PIDIQ applies an ImageJ-based macro to plant photos in order to distinguish healthy tissue from tissue that has yellowed due to disease. It can process a directory of images in an automated manner and report the relative ratios of healthy to diseased leaf area, thereby providing a quantitative measure of plant health that can be statistically compared with appropriate controls. We used the Arabidopsis thaliana-Pseudomonas syringae model system to show that PIDIQ is able to identify both enhanced plant health associated with effector-triggered immunity as well as elevated disease symptoms associated with effector-triggered susceptibility. Finally, we show that the quantitative results provided by PIDIQ correspond to those obtained via traditional in planta pathogen growth assays. PIDIQ provides a simple and effective means to nondestructively quantify disease from whole plants and we believe it will be equally effective for monitoring disease on excised leaves and stems.

The rise of the undead: pseudokinases as mediators of effector-triggered immunity.

Pathogens use effector proteins to suppress host immunity and promote infection. However, plants can recognize specific effectors and mount an effector-triggered immune response that suppresses pathogen growth. The YopJ/HopZ family of type III secreted effector proteins is broadly distributed in bacterial pathogens of both animals and plants. These effectors can either suppress host immunity or elicit defense responses depending on the host genotype. In a recent report, we identified an Arabidopsis thaliana pseudokinase ZED1 that is required for the recognition of the Pseudomonas syringae HopZ1a effector. Here we discuss the role of ZED1 in HopZ1a recognition, and present models of effector recognition in plants. We draw parallels between HopZ1a and YopJ effector proteins, and between ZED1 and other immunity-related kinases that can be targeted by pathogen effectors.

The Arabidopsis ZED1 pseudokinase is required for ZAR1-mediated immunity induced by the Pseudomonas syringae type III effector HopZ1a.

Plant and animal pathogenic bacteria can suppress host immunity by injecting type III secreted effector (T3SE) proteins into host cells. However, T3SEs can also elicit host immunity if the host has evolved a means to recognize the presence or activity of specific T3SEs. The diverse YopJ/HopZ/AvrRxv T3SE superfamily, which is found in both animal and plant pathogens, provides examples of T3SEs playing this dual role. The T3SE HopZ1a is an acetyltransferase carried by the phytopathogen Pseudomonas syringae that elicits effector-triggered immunity (ETI) when recognized in Arabidopsis thaliana by the nucleotide-binding leucine-rich repeat (NB-LRR) protein ZAR1. However, recognition of HopZ1a does not require any known ETI-related genes. Using a forward genetics approach, we identify a unique ETI-associated gene that is essential for ZAR1-mediated immunity. The hopZ-ETI-deficient1 (zed1) mutant is specifically impaired in the recognition of HopZ1a, but not the recognition of other unrelated T3SEs or in pattern recognition receptor (PRR)-triggered immunity. ZED1 directly interacts with both HopZ1a and ZAR1 and is acetylated on threonines 125 and 177 by HopZ1a. ZED1 is a nonfunctional kinase that forms part of small genomic cluster of kinases in Arabidopsis. We hypothesize that ZED1 acts as a decoy to lure HopZ1a to the ZAR1-resistance complex, resulting in ETI activation.

A high-throughput forward genetic screen identifies genes required for virulence of Pseudomonas syringae pv. maculicola ES4326 on Arabidopsis.

Successful pathogenesis requires a number of coordinated processes whose genetic bases remain to be fully characterized. We utilized a high-throughput, liquid media-based assay to screen transposon disruptants of the phytopathogen Pseudomonas syringae pv. maculicola ES4326 to identify genes required for virulence on Arabidopsis. Many genes identified through this screen were involved in processes such as type III secretion, periplasmic glucan biosynthesis, flagellar motility, and amino acid biosynthesis. A small set of genes did not fall into any of these functional groups, and their disruption resulted in context-specific effects on in planta bacterial growth.