Pathogen Populations Research Articles

The Mediator complex subunit MoMed15 plays an important role in conferring sensitivity to isoprothiolane by modulating xenobiotic metabolism in M. oryzae.

Rice blast caused by Magnaporthe oryzae is one of the most economically important rice diseases. Fungicides such as isoprothiolane (IPT) have been used extensively for rice blast control, but resistance to IPT in M. oryzae is an emerging threat. In this study, molecular mechanisms of resistance in IPT-resistant mutants were identified. Through whole-genome sequencing and genetic transformation, we identified the gene MoMed15, encoding a transcriptional glutamine-rich co-activator Mediator complex subunit, in which mutations or deletion resulted in moderate IPT resistance. Further research found that MoMed15 physically interacted with the IPT resistance regulatory factor MoIRR to simultaneously regulate both MoIRR expression and the expression of multiple xenobiotic-metabolizing enzymes in response to IPT stress. We hypothesize that some xenobiotic-metabolizing enzymes enhance IPT toxicity by modifying the IPT structure. Variation of MoMed15 affected the recruitment of the transcriptional Mediator complex and decreased the expression of these xenobiotic-metabolizing enzymes, resulting in moderate IPT resistance. We also found that MoPGR1, encoding a protein that activates cytochrome P450 enzymes, was essential to confer IPT sensitivity, and its expression was directly regulated by MoIRR.IMPORTANCEIsoprothiolane (IPT) has been used extensively for the management of rice blast disease and IPT-resistant subpopulations have emerged in Chinese rice fields. The emergence of resistant pathogen populations has led to a steep increase in fungicide use, increasing pesticide risk for the applicator and the environment. The molecular mechanisms of IPT resistance in M. oryzae remain elusive. In this study, we demonstrated that transcriptional co-activator MoMed15 interacts with IPT resistance regulator MoIRR to recruit the Mediator complex, which promotes the expression of xenobiotic-metabolizing enzymes, leading to exacerbated IPT toxicity. The MoMed15 could be used for IPT resistance detection in rice fields.

Hyperspectral imaging analysis for early detection of tomato bacterial leaf spot disease.

Recent advancements in hyperspectral imaging (HSI) for early disease detection have shown promising results, yet there is a lack of validated high-resolution (spatial and spectral) HSI data representing the responses of plants at different stages of leaf disease progression. To address these gaps, we used bacterial leaf spot (Xanthomonas perforans) of tomato as a model system. Hyperspectral images of tomato leaves, validated against in planta pathogen populations for seven consecutive days, were analyzed to reveal differences between infected and healthy leaves. Machine learning models were trained using leaf-level full spectra data, leaf-level Vegetation index (VI) data, and pixel-level full spectra data at four disease progression stages. The results suggest that HSI can detect disease on tomato leaves at pre-symptomatic stages and differentiate bacterial disease spots from abiotic leaf spots. Using VI data as features for machine learning improved overall classification performance by 26-37% compared to the direct use of raw data. Critical wavelength bands and VIs varied across disease progression stages, suggesting that pre-symptomatic disease detection relied more on changes in leaf water content (1400nm) and plant defense hormone-mediated responses (750nm) rather than changes in leaf pigments or internal structure (800-900nm), which may become more crucial during symptomatic stages. In conclusion, this study provides valuable insights into the dynamics of bacterial spot disease, revealing the potential benefits of leaf structure segmentation and VI group pattern analysis in HSI studies for the early detection of leaf diseases.

Wheat Leaf Rust, Caused by Puccinia triticina , and Mitigation Through Host Genetic Resistance

Abstract The obligate fungal pathogen Puccinia triticina causes leaf rust symptoms on wheat worldwide. Epidemics of leaf rust occur during periods of mild weather with high relative humidity. The pathogen infects leaves, causing orange to red pustules, containing millions of urediniospores, reducing the photosynthetic area and causing desiccation and yield loss. Wind-borne urediniospores are produced in continuous asexual cycles on wheat plants. Genetic resistance is the primary control strategy. There are over 80 known leaf rust ( Lr ) resistance genes, and most are race-specific and are active from the seedling stage to maturity. A few Lr genes are nonrace-specific and confer partial resistance to leaf rust and other diseases at the adult plant stage. Combining Lr genes that confer seedling and adult plant resistance has been particularly successful and durable. Pathogen populations are genetically diverse, and races evolve continuously to evade recognition by Lr genes. This results in boom-and-bust cycles when new cultivars with race-specific Lr genes are resistant for some years, but become susceptible, due to evolution of the P. triticina population. Surveillance of P. triticina for the evolution of races helps to develop wheat cultivars with Lr genes that will be resistant to the most prevalent races. Information © The Authors 2024

Temporal Dynamics of Two Genetically Diverse Downy Mildew Mitochondrial Clades on Cucumber Cultivars in South Carolina and Florida

Downy mildew (DM, Pseudoperonospora cubensis) is an important disease that can significantly reduce yield of field-grown cucumbers in the US. Cucumber cultivars with resistance to DM were widely grown for >30 years. However, resistance broke down in 2004 resulting in severe DM epidemics. Large plot trials were conducted to characterize pathogen virulence and population dynamics on DM tolerant cultivars Peacemaker and Expedition and susceptible SMR-58 in Charleston, SC and Fort Pierce, FL in 2016 and 2017. Rapid DM development on SMR-58 was observed at both locations in 2016 and 2017. Disease progress on Expedition and Peacemaker were slow as compared to SMR-58 in Fort Pierce, FL in 2016. Very low to no DM was observed on Peacemaker or Expedition in Charleston in 2016. In 2017, all cultivars were affected in Charleston, but only SMR-58 had severe DM (95-100%) in Fort Pierce. Pathogen population analysis using DM specific mitochondrial clade (MC1 and MC2) markers showed differences between the P. cubensis populations across location and years on the cucumber cultivars. Both MC1 and MC2 were detected in cucumber plots in SC and FL. The most prevalent DM mitochondrial clade in SC during June 2016 was MC1 primarily on SMR-58 (95-100%). However, in July 2016 that population shifted towards MC2 (45-77%). This suggested that the DM populations can change relatively quickly when suitable environment favors. Mixed populations of MC1 and MC2 continued to be observed in following year in Charleston, SC in 2017. In Fort Pierce, FL a complete shift was observed from MC2 in 2016 to MC1 in 2017. Since both MCs are present in the southeastern United States either at the same time or possibly different times during the production year, cultivars with combined resistance to P. cubensis populations will be needed to reduce reliance on fungicides to manage DM.

Unleashing the synergistic effect of promising fungicides: a breakthrough solution for combating powdery mildew in pea plants.

Pea powdery mildew, caused by Erysiphe pisi, is a major limitation to global pea production. The emergence of fungicide-resistant pathogen populations due to frequent and injudicious pesticide application highlights the importance of exploring the synergistic properties of fungicide combinations. This study investigated the efficacy of difenoconazole, thiophanate-methyl, and sulfur, both individually and in mixtures, against powdery mildew and assessed the interaction types between these fungicides. The results demonstrated that the combination of difenoconazole, thiophanate-methyl, and sulfur was the most effective in reducing, reducing disease severity to 6.10% and minimizing conidial production on foliage. Additionally, this fungicide combination reduced conidial germination by 89.26% in vitro and by 87.50% in a detached leaf assay compared to the control. The treatment also positively impacted leaf chlorophyll content (55.18), green pod yield (22.21 tons ha-1), seed yield (12.29 tons ha-1), and other yield-related parameters. Although statistically significant, this ternary fungicide combination was closely followed by the binary combination of thiophanate-methyl and sulfur, which was the only combination exhibiting synergism in both laboratory and field trials with a synergy factor (SF) > 1. In conclusion, this approach offers improved disease control as part of integrated disease management (IDM) while minimizing the risk of resistant pathogen strains.

Comparative Analysis of Some Morpho-cultural Characters of Venturia inaequalis (Apple Scab Pathogen) under Kashmir Conditions

Apple (Malus × domestica Borkh.) represents one of the most widely cultivated and economically important temperate fruit crops in the world. However, its cultivation faces significant challenges due to various fungal pathogens, particularly apple scab inflicted by Venturia inaequalis, which has a profound impact on the yield and quality of apple, resulting in substantial economic loss. The pervasive nature of V. inaequalis and its ability to damage crops poses a serious threat to sustainable apple production. In this study, the morpho-cultural characteristics of V. inaequalis were systematically analyzed and compared between the Shopian and Baramullah districts of the Kashmir Valley. Ten isolates from each district were grown on Malt Extract Agar (MEA) and Potato Dextrose Agar (PDA) for comparative analysis. The colonies were characterized by a compact hue ranging from greyish-green to blackish and exhibited a densely velvety texture. Mycelial growth was predominantly aerial, with some isolates displaying partial irregularity in colony margins, while others exhibited greater uniformity. The mean colony radii for potato dextrose agar and malt extract agar media was measured as 23.01 mm and 28.50 mm for the Shopian district, and 28.09 mm and 33.33 mm for the Baramullah district, respectively. The mean conidial spore density was recorded at 27.87 spores/cm² for the Baramullah district and 34.67 spores/cm² for the district Shopian, respectively. This variation could be attributed to several factors, including genetic diversity within the pathogen populations, differences in altitude, climate, and other environmental factors, the selection pressure exerted by fungicide applications, the emergence of novel pathogen races, and phenotypic plasticity driven by the differing agricultural practices and microclimatic conditions in the respective regions.

Comparative genomics of Fusarium species causing Fusarium ear rot of maize.

Fusarium ear rot (FER), caused by the fungal pathogen Fusarium verticillioides, stands as one of the most economically burdensome and pervasive diseases affecting maize worldwide. Its impact on food security is particularly pronounced due to the production of fumonisins, toxic secondary metabolites that pose serious health risks, especially for livestock. FER disease severity is complex and polygenic, with few resistance (R) genes being identified for use in breeding resistant varieties. While FER is the subject of several breeding programs, only a few studies have investigated entire populations of F. verticillioides with corresponding virulence data to better understand and characterize the pathogenomics. Here, we sequenced and compared the genomes of 50 Fusarium isolates (43 F. verticillioides and 7 other Fusarium spp.) that were used to inoculate a diverse maize population. Our objectives were to elucidate the genome size and composition of F. verticillioides, explore the variable relationship between fumonisin production and visual disease severity, and shed light on the phylogenetic relationships among the isolates. Additionally, we conducted a comparative analysis of the nucleotide variants (SNPs) and the isolates' effectoromes to uncover potential genetic determinants of pathogenicity. Our findings revealed several promising leads, notably the association of certain gene groups, such as pectate lyase, with disease severity. These genes should be investigated further as putative alleles for breeding resistant maize varieties. We suggest that, beyond validation of the alleles identified in this study, researchers validate each phenotypic dataset on an individual basis, particularly if considering fumonisin concentrations and when using diverse populations. Our study underscores the importance of genomic analysis in tackling FER and offers insights that could inform the development of resilient maize cultivars. By leveraging advances in genomics and incorporating pathogen populations into breeding programs, resistance to FER can be advanced.

Seamless, rapid, and accurate analyses of outbreak genomic data using split k-mer analysis.

Sequence variation observed in populations of pathogens can be used for important public health and evolutionary genomic analyses, especially outbreak analysis and transmission reconstruction. Identifying this variation is typically achieved by aligning sequence reads to a reference genome, but this approach is susceptible to reference biases and requires careful filtering of called genotypes. There is a need for tools that can process this growing volume of bacterial genome data, providing rapid results, but that remain simple so they can be used without highly trained bioinformaticians, expensive data analysis, and long-term storage and processing of large files. Here we describe split k-mer analysis (SKA2), a method that supports both reference-free and reference-based mapping to quickly and accurately genotype populations of bacteria using sequencing reads or genome assemblies. SKA2 is highly accurate for closely related samples, and in outbreak simulations, we show superior variant recall compared with reference-based methods, with no false positives. SKA2 can also accurately map variants to a reference and be used with recombination detection methods to rapidly reconstruct vertical evolutionary history. SKA2 is many times faster than comparable methods and can be used to add new genomes to an existing call set, allowing sequential use without the need to reanalyze entire collections. With an inherent absence of reference bias, high accuracy, and a robust implementation, SKA2 has the potential to become the tool of choice for genotyping bacteria. SKA2 is implemented in Rust and is freely available as open-source software.

Synergistic interactions of biocontrol agents and chemical fungicides enhance the disease resistance in Vigna radiata (L.) against Macrophomina phaseolina-associated with dry root rot

Macrophomina phaseolina, a necrotrophic fungus responsible for dry root rot disease in Vigna radiata L. (greengram) causes extensive yield losses. Biocontrol agents are reported to suppress phytopathogens, mitigate biotic stress, and improve growth and yield of plants. Here, Bacillus subtilis PSB-5 and Trichoderma sp. TD-6 along with chemical fungicides; mancozeb (MZB) and carbendazim (CBZM) were evaluated alone/or in combination for their effectiveness against M. phaseolina-induced root rot disease in greengram. B. subtilis and Trichoderma sp. suppressed phyto-fungal growth by 72 and 78 %, respectively. Both bioagents synthesized indole-3-acetic acid, ammonia, and solubilized insoluble phosphate, released antimicrobial substances including siderophore, hydrogen cyanide, and various extracellular lytic enzymes. Further, efficacy of bioagents and fungicides (at 50 and 100 ppm) were tested (single and combined) against M. phaseolina-infected greengram cultivated in pot soils. While evaluating, bioagents and fungicides increased plant height, biomass, and vigor indices of infected greengram in order: PSB-5 + TD-6 > PSB-5 > TD-6 > MZB + CBZM (100 ppm) > CBZM (100 ppm) > MZB (100 ppm). Further, among treatment, dual inoculation of bioagents (B. subtilis + Trichoderma), maximally increased total chlorophyll (59.4 %), carotenoid (68 %), root phosphorous (69.6 %), root nitrogen (65.7 %), total soluble protein (80 %), soluble sugar (76 %), pod yield (82.3 %) and seed protein (68 %) over infected plants. Moreover, biotic stress-induced oxidative stress biomarkers (proline, malondialdehyde, electrolyte leakage) and reactive oxygen species were significantly (p ≤ 0.05) lowered in fungal-infected greengram following fungicides and bioagents application. Furthermore, combined inoculation of bioagents significantly (p ≤ 0.05) reduced percent disease index (PDI) and increased % protection in biotic-stressed greengram. Besides, bioagents potentially increased disease resistance in M. phaseolina-infected greengram plants by modulating defense responses such as activities of β-1, 3 glucanase, catalase, phenylalanine ammonia lyase, polyphenol oxidase, peroxidase, and superoxide dismutase which significantly protected plants from fungal pathogen. Soil populations of pathogen and bioagents declined and considerably increased, respectively, at harvest. Application of antagonistic bacterial and fungal strains enhances greengram resistance to M. phaseolina while concurrently offering agriculture an environmentally friendly choice. Also, these strains can be applied as biocontrol agents in disease suppression over chemical fungicides.

Confirmation of oat crown rust disease in Taiwan.

Oat is a minor forage crop grown in Taiwan. Only a few historical records of oat rust disease have been reported in the country. Therefore, the pathogen population remains poorly characterized. A rust-like disease outbreak was detected at the Experimental Farm of National Taiwan University in 2019, which caused significant damage to field experiments. To determine the identity of the pathogen responsible for this disease outbreak, we collected infected foliar material. Disease signs suggested infection by the oat crown rust fungus. Hence, common procedures in rust pathology were applied to confirm the identity of the pathogen with phenotypic and molecular diagnostic techniques. A total of 50 field pathogen samples from infected oat cultivars were collected in 2019 and five single pustule rust isolates were obtained in 2020 and 2021. These isolates were initially identified as Puccinia coronata var. avenae f. sp. avenae (Pca) based on the phylogenetic analysis of nrITS sequence data. This identification was subsequently confirmed through whole-genome phylogeny, which showed that the representative Taiwanese isolate NTU1 clustered with other Pca representative strains in Basidiomycota. Phenotyping assays across 36 oat differential lines demonstrated that Taiwanese isolates are phenotypically similar with relatively low virulence. This study presents the first molecular confirmation of Pca in Taiwan and reports the virulence profiles of Taiwanese Pca population.

A multilocus perspective on the evolutionary dynamics of multistrain pathogens

Many human pathogens, including malaria, dengue, influenza, Streptococcus pneumoniae , and cytomegalovirus, coexist as multiple genetically distinct strains. Understanding how these multistrain pathogens evolve is of critical importance for forecasting epidemics and predicting the consequences of vaccination. One factor believed to play an important role is naturally acquired immunity. Consequently, a large body of research has sought to predict how acquired immunity molds the genomics of pathogen populations (i.e., what shapes pathogen strain structure). The diversity of existing models has resulted in conflicting evolutionary predictions and has sparked an ongoing debate about which predictions are most broadly applicable. Here, we adopt a multilocus population genetics perspective that unifies the predictions of existing models. We identify three key factors that determine the role of naturally acquired immunity in the evolution of pathogen strain structure: i) the strength and specificity of immune protections, ii) the dynamic immunological landscape, and iii) the number of loci coding for the antigens of the pathogen. Isolating and discussing these three factors clarifies the relationship among previous models of multistrain dynamics, and establishes a solid theoretical foundation for the study of the evolutionary epidemiology of multistrain pathogens.

Single-Cell Transcriptomics Reveals a Pivotal Role of DOCK2 in Sjögren Disease.

Sjögren disease (SjD) is an autoimmune condition characterized by the dysfunction of the salivary and lacrimal glands. The study aimed to decipher the pathogenic cell populations and their immunologic pathways in the salivary glands. We further determined the therapeutic effect of inhibiting dedicator of cytokinesis 2 (DOCK2) shared by novel clusters of CD8+ T cells in an SjD mouse model. This study employed single-cell RNA sequencing to examine the composition and dynamics of immune cells in the salivary glands of SjD mice. By analyzing the transcriptomic data and employing clustering analysis, a specific target was identified, leading to the treatment of mice with a targeted inhibitor. The results showed diverse immune cell types, including B cells, CD4+ T cells, CD8+ T cells, macrophages, and natural killer cells. We identified specific clusters possessing phenotypic characteristics of immune cell subpopulations, thereby showing specific genes/pathways associated with the disease. The most striking finding was the elevated expression of DOCK2 in CD8+ T cells in the SjD model. This discovery is significant because subsequent treatment with a DOCK2 inhibitor 4-[3-(2-Chlorophenyl)-2-propen-1-ylidene]-1-phenyl-3,5-pyrazolidinedione (CPYPP) led to a marked amelioration of SjD signs. The effectiveness of DOCK2 inhibition in alleviating SjD signs highlights the potential of DOCK2 as a therapeutic target, opening new avenues for treatment strategies that could modulate the immune response more effectively in SjD.

Commercial bacteriophage preparations for the control of Listeria monocytogenes and Shiga toxin-producing Escherichia coli in raw and pasteurized milk

Listeria monocytogenes was the etiologic agent in nearly all recent outbreaks in North America attributed to pasteurized dairy products, whereas Escherichia coli O157 infections were responsible for most of the rare, yet serious complications from outbreaks involving unpasteurized dairy. This study determined the susceptibility of selected strains of L. monocytogenes and Shiga toxin-producing E. coli (STEC) to commercial phage preparations and their ability to control these pathogens in pasteurized and raw milk during 7-day storage at 7 °C. Both phage products demonstrated high lytic efficiency against 17 strains of L. monocytogenes whereas the efficiency of E. coli phages was more variable against 11 strains of O157 and non-O157 STEC. Broth microdilution assays identified effective endpoint multiplicities of infection (MOI) ranging from log 2.53 to 5.13, which differed between strains of L. monocytogenes and phage products. Mean log MOIs of E. coli phages against STEC also varied within and between products from 0.43 to 7.05. Despite these observations, the change in counts over time of three L. monocytogenes strains exposed to phage in pasteurized milk (log MOI 6) was similar with counts ∼ 4 log CFU/mL lower than control at day 7. Results for STEC O157 varied by strain but counts were lower than control in all cases by 72 h thorough day 7. Titers on isolates of both pathogens isolated from pasteurized milk indicated that the surviving populations were less susceptible to phage. The addition of a phage preparation to raw milk did not reduce populations of either pathogen or affect the change in counts of any strain over time when compared to control. Reduced efficacy in raw milk may be attributed to reduced phage binding as titers in raw milk decreased steadily (∼2 log PFU/mL) during storage. Commercial phage products may be a promising pathogen control intervention for pasteurized dairy products, warranting further investigation.

Specific aspects of bacteriophage regulation: international practices and future developments

INTRODUCTION. The increasing prevalence of multidrug-resistant strains of pathogens determines the need for fundamentally new antibacterial agents, including bacteriophage preparations. The consistent implementation of phage therapy is hindered by the lack of generally accepted standardised regulatory documents governing the legal and methodological aspects of the production and preclinical and clinical studies of bacteriophage preparations.AIM. This study aimed to analyse the international experience with the production and lifecycle management of bacteriophage preparations, as well as the main regulatory requirements for the control of their quality, safety, and efficacy.DISCUSSION. It is difficult to develop virulent bacteriophage preparations in accordance with the existing requirements for other medicinal products because of the biological characteristics of bacteriophages, the wide variety of bacteriophage strains, and the potential for rapid changes both in the bacteriophage population and in the pathogen population. Therefore, it is reasonable to develop streamlined marketing authorisation routes for phage therapies and methods for the assessment of their safety and efficacy. As part of these efforts, it is necessary to assess the adverse events specific to this group of medicinal products, such as the risks of lysogeny, resistance to bacteriophages, and antibiotic resistance gene transfer between bacterial strains. The pharmaceutical development of bacteriophage preparations can be based on several approaches. Many countries worldwide, including the United States, are implementing the concept of Quality by Design, considering approaches based on the Biological Master File, and conducting Expanded Access programmes. The Active Substance Master File procedure allows the submission of a separate document package covering only part of the registration dossier for regulatory approval. Expanded Access programmes provide individual patients with access to innovative medicinal products without approved treatment protocols. In the Russian Federation, the commercial production of bacteriophage medicinal products is conducted in accordance with the quality standards specified in the State Pharmacopoeia of the Russian Federation.CONCLUSIONS. There are fundamental differences in the approaches to phage therapy and its regulation around the world and in the Russian Federation. It is reasonable to supplement the current national guidelines for the safety and efficacy evaluation of bacteriophage preparations, in particular, to specify the requirements for conducting preclinical studies.

Exploring pathogen population density as a metric for understanding post-COVID infectious disease surges.

After the easing of COVID-19 restrictions, peaks of common infectious diseases surpassed pre-pandemic levels, raising questions about causes and ways to monitor these changes. A proposed measure, the Pathogen Population Density (PPD) score, could help track these shifts. PPD refers to the concentration of infectious agents within a population at a given time and location, serving as a potential indicator of infection levels in susceptible individuals at the population level. It is likely that PPD remains relatively stable within a specific community, as an equilibrium forms between infections and susceptibility. During the pandemic, nonpharmaceutical interventions (NPIs) led to a reduction in infectious diseases, possibly lowering population immunity and decreasing the PPD score. Once NPIs were lifted, the PPD score likely increased sharply due to a larger pool of susceptible individuals, causing more primary infections and stronger recurrent infections, faster transmission, and more severe pathogenic outcomes at the individual level. Monitoring the PPD score over time could help predict when infection peaks will occur. PPD is influenced by factors such as public health strategies, vaccination programs, and the behavior of high-risk individuals. As a quantitative measure, PPD has the potential to serve as a valuable predictive and monitoring tool, helping public health officials anticipate and track changes in infectious disease dynamics. It could be an effective tool for managing future outbreaks or pandemics and serve as a communication tool between scientists and the public to understand the emergence of new disease peaks.

Field-scale gene flow of fungicide resistance in Pyrenophora teres f. teres and the effect of selection pressure on the population structure.

The effectiveness of fungicides to control foliar fungal crop diseases is being diminished by the increasing spread of resistances to fungicides. One approach that may help to maintain efficacy is remediation of resistant populations by sensitive ones. However, the success of such approaches can be compromised by re-incursion of resistance through aerial spore dispersal; although, knowledge of localized gene flow is lacking. Here, we report on a replicated mark-release-recapture field experiment with several treatments set up to study spore-dispersal-mediated gene flow of a mutated allele that confers demethylase inhibitor resistance in Pyrenophora teres f. teres (Ptt). Artificial inoculation of the host, barley (Hordeum vulgare), was successful across the 12-ha trial, where the introduced sensitive- and resistant-populations were, respectively, 6- and 13-fold the DNA concentration of the native Ptt population. Subsequent disease pressure remained low which hampered spread of the epidemic to such extent that gene flow was not detected at, or beyond 2.5 m from source points. In the absence of gene flow, plots were assessed for treatment effects; fungicide applied to populations that contained 14.3% of allele mutation increased in frequency to 24.5%, whereas sensitive populations had no change in structure. Untreated controls of native Ptt population remained genetically stable, yet untreated controls that were inoculated with sensitive Ptt had half the resistance frequency of the native population structure. The trial demonstrates the potential for management to remediate fungicide resistant pathogen populations, where localized gene flow is minimal; to safeguard chemical crop protection into the future.

The effect of Plantago major seed mucilage fortified with cell-free supernatant (CFS) of probiotic Lactococcus lactis NJ414 on extending shelf life and ensuring the safety of lamb meat slices stored at 4 °C

The present study was conducted to create a bioactive edible coating using Plantago major mucilage (PMM) and the cell-free supernatant (CFS) of probiotic Lactococcus lactis NJ414 to extend the shelf life and ensure the safety of lamb meat slices stored at 4 °C. The PMM+2%CFS coated slices showed a significant decrease in total viable count (8.14–5.48 log CFU/g), and psychrotrophic bacteria count (5.24–3.54 log CFU/g). Additionally, the pH value decreased from 5.93 to 5.71, the thiobarbituric acid (TBA) reduced by 65%, and the peroxide value (PV) decreased by 60%. Moreover, PMM+2%CFS coated slices had higher hardness (68.87 N vs. 62.10 N), moisture content (67.57% vs. 64.35%), and overall acceptance (6.40 vs. 3.00) compared to the control. Adaptive neuro-fuzzy inference systems (ANFIS) and gaussian process regression (GPR) models were used to predict the population of microbial pathogens, physicochemical and sensory characteristics of coated lamb products based on three inputs including PMM (0% and 2%), CFS (0%, 1% and 2%), time (1, 3, 6 and 9 days). The result showed that artificial intelligence-based models, especially ANFIS (R2 > 0.99) are able to estimate the population of microbial pathogens, physiochemical and sensory characteristics.

Sustaining Soil Health in High Tunnels: A Paradigm Shift toward Soil-centered Management

This review was conducted to synthesize current knowledge, learn producer and Extension specialist perspectives, and identify gaps in understanding of the role of soil health in sustaining production in high tunnel (HT) systems. This synthesis includes findings from scholarly resources related to soil health in HTs, including research and Extension-based literature, perspectives from experienced HT producers and technical assistance providers, and the direct observations of a broad network of university research and Extension personnel working with HTs. Findings are intended to identify knowledge gaps and additional research and Extension resource needs of greatest priority to the HT producer community and technical assistance providers that support them at the time of publication. A review of 68 research articles and 58 Extension resources was conducted. Focus group interviews were conducted with small groups of experienced HT farmers in four regions of the eastern half of the United States, with in-depth farm case studies conducted in individual farmers in three of these regions. Growers across regions identified soil fertility management, soilborne diseases, soil compaction, and lack of consistency of soil analyses specific to HTs as the greatest soil-related challenges to HT production. Research and resources for technical assistance providers on mitigation strategies to remediate yield-limiting HT soil conditions, such as excessive soil salinity and high pathogen populations, were also lacking. As such, process-based research on techniques such as leaching, soil steaming, solarization, and anaerobic soil disinfestation in tunnels that consider short- and long-term costs, benefits, and effects on soil and plant productivity should be prioritized in the future when considering the impact of HT production on soil health. Interviews also indicated a need for networking opportunities for technical assistance providers across agencies (e.g., Natural Resources Conservation Service, Extension, nongovernmental organizations). Despite a high and increasing rate of adoption, there is currently a lack of information about maintaining HT systems. Given that HTs play a critical and growing economic role for specialty crop growers throughout the eastern United States, comprehensive intervention across the research–Extension spectrum to sustain productivity in HT systems is recommended.

Identification and Differentiation of the Fusarium graminearum NX-2 Chemotype Using High-Resolution Melting (HRM).

Fusarium head blight causes significant yield losses in wheat and other cereals and contaminates grain products with trichothecene mycotoxins. F. graminearum isolates are classified into different chemotypes depending on the type of mycotoxin produced, including the type B trichothecenes 3-acetyl deoxynivalenol (3-ADON), 15-acetyl deoxynivalenol (15-ADON), nivalenol (NIV), and the recently identified type A trichothecene NX-2. Molecular tools to differentiate NX-2 producers from other chemotypes have remained relatively laborious and time consuming. In this study, we developed and validated a high-resolution melting (HRM) assay that can identify NX-2 producers quickly and cost-effectively. By analyzing TRI1 coding sequences from 183 geographically diverse isolates representing all four F. graminearum chemotypes, we selected a 75-base pair region containing four non-synonymous single nucleotide polymorphisms (SNPs) that are specific to the NX-2 genotypes. The amplicon generated two HRM profiles, one of which was specific for only NX-2. We confirmed that the assay is robust across qPCR platforms and unambiguously differentiates NX-2 from other chemotypes using a panel of 72 diverse isolates previously collected from North America. The HRM assay was also successful in identifying NX-2 producers directly from DNA extracted from infected wheat spikes with varying levels of disease severity and fungal DNA. The assay can detect as little as 0.01 ng of fungal DNA in a background of 50 ng of plant DNA. This new diagnostic assay can be used for high-throughput molecular detection of the NX-2 chemotype of F. graminearum from infected plant samples and culture collections, thus making it a valuable tool for surveys of contemporary and historical FHB pathogen populations.

Beijing Central Asian Outbreak strain cluster of Mycobacterium tuberculosis detected in Irkutsk region, Russia

Background. Irkutsk region maintains a high incidence rate of multidrug-resistant (MDR) tuberculosis (TB). Detection of MDR-associated Mycobacterium tuberculosis strains in Irkutsk region requires dynamic assessment of the TB pathogen population, taking into account the emergence of a new resistant variant of Beijing Central Asian Outbreak (CAO).The aim of the study. To assess changes in the genotypic structure of M. tuberculosis strains circulating in the Irkutsk region over a ten-year period.Materials and methods. A total of 732 M. tuberculosis strains (196 strains for 2021– 2022, 536 strains for 2011–2015) were studied using MIRU-VNTR and SNP typing.Results. The MDR level increased to 67.4 % with an increase of pre-extensive drug resistance (pre-XDR) (33.2 %) (p < 0.001). In the modern sample, the dominance of the Beijing genotype increased (83.7 %) due to an increase in the proportions of the B0/W148 (38.8 %) and CAO (12.8 %) subtypes with a stable overall level of Central Asian Russian (36.8 %) and other Beijing strains (8.2 %). Strains other than the Beijing genotype belonged mainly to the Euro-American lineage (Lineage 4): LAM (8.9 %), Ural (2.7 %), Haarlem (2.0 %), S (0.5 %) and L4-unclassified (5.3 %); 25 isolates were not classified. In the 2011–2015 sample, LAM was more common than in the modern sample (10.8 % vs. 3.6 %; p < 0.01). The increase in MDR and preXDR was statistically significant among Beijing B0/W148 strains (93.4 % vs. 66.1 %; p < 0.001).Conclusion. Unfavorable trends of significant spread of MDR and pre-XDR of the Beijing genotype strains were revealed. Among the Beijing strains, not only B0/W148 but also the Beijing CAO subtype, which was previously rare in Siberia, are the most successful; they have the highest levels of MDR and pre-XDR and a tendency to widespread distribution in all groups of TB patients.