Spore Survival Research Articles

Synergistic Interactions among Vacuum, Solar Heating, and UV Irradiation Enhance the Lethality of Interplanetary Space

A Planetary Atmospheric Chamber (PAC) was used to create simulations of interplanetary conditions to test the spore survival of three Bacillus spp. exposed to interacting conditions of vacuum (VAC), simulated solar heating (HEAT), and simulated solar ultraviolet irradiation (UV). Synergism was observed among the experimental factors for all three Bacillus spp. tested that suggested the increased lethality of HEAT and UV when concomitantly exposed to VAC. The most aggressive biocidal effects were observed for assays with VAC + HEAT + UV conditions run simultaneously over short time-steps. The results were used to predict the accumulation of extremely rapid Sterility Assurance Levels (SALs; def., −12 logs of bioburden reduction) measured in a few minutes to a few hours for external surfaces of interplanetary spacecraft. Furthermore, the results were extrapolated to predict that approx. 1 × 104 SAL exposures might be accumulated for external surfaces on the Europa Clipper spacecraft during a 3.5-year transit time between Venus (0.7 AU) and Mars (1.5 AU) during a series of Venus–Earth–Earth gravity assists (VEEGA trajectory) to Jovian space. The results are applicable to external spacecraft surfaces exposed to direct solar heating and UV irradiation during transits though the inner solar system.

Spore germination and lactic acid combined treatment: A new processing strategy for the shelf-life extension of instant wet noodles

Bacillus amyloliquefaciens (BAM) was identified as the predominant spoilage bacteria in instant wet noodles (IWNs). The utilization of industrial acid treatment as a long shelf-life strategy resulted in reduced consumer acceptance due to the acidic taste of the products. This study proposed a processing strategy that integrated spore germination (SG) and lactic acid (LA) treatment to effectively reduce the spore survival rate and extend the shelf life of IWNs. L-histidine, d-glucose, and sodium chloride were highly efficient and safe germinants for BAM spores. In IWNs, compound germinants (1.0 % L-histidine, 0.5 % d-glucose, and 1.0 % sodium chloride) boosted the SG rate by 3.61 times. With synergistic LA treatment, the spore lethality increased by 34.41 % -41.68 %. Under the SG and reduced acid-heat conditions of pH 2.30–2.50, the mortality of spores could reach 92.00 %–93.17 %, which was 14.11 %–15.28 % higher than the industrial acid-heat condition of pH 2.10. DPA, ATP, and membrane potential showed that germinants reduced the spore membrane permeability and promoted the occurrence of spore membrane damage under acid-heat conditions. Moreover, this strategy significantly extended the shelf-life of IWNs by 3.00–5.50 times and controlled the pH ≥ 5.50. Additionally, it improved color, texture, and overall sensory evaluation. Accordingly, this strategy solved the contradiction between the long shelf-life of IWNs and the unacceptable acidification in industrial production.

Effects of Several Fungicides on the Spore Growth Period of Alternaria dauci, a Carrot Black Leaf Blight Fungus, Using a Rezasulin-based Spore Survival Assay

The effects of five fungicides on the spore growth phase of Alternaria dauci, which causes carrot leaf blight, were tested using the spore viability assay (SVA) and agar dilution method (ADM). The average EC50 values for chlorothalonil against seven isolates of A. dauci examined by SVA and ADM were 14.21 μg/ml and more than 100 μg/ml. Dithianon and folpet also had lower EC50 values in SVA than in ADM, while iminoctadine tris-albesilate had lower EC50 value in ADM. For fluazinam, the EC50 values of SVA and ADM were 1.63 and 2.40 μg/ml, respectively. As EC50 values of five fungicides according to the spore growth phase of A. dauci KACC 42997, the efficacy of each fungicide as chlorothalonil, dithianon, and folpet decreased when treated after spore germination rather than when treated with spores before germination. However, iminoctadine tris-albesilate was more effective when treated after spores germinated than when treated before treatment. The excellent effect of fluazinam on the pathogen was maintained until A. dauci KACC 42997 was cultured in potato dextrose broth for 6 hr and the germ tube grew beyond the size of the spore. However, when treated with iminoctadine tris-albesilate and fluazinam after culturing for 12 hr, as the EC50 values of the two fungicides increased to 8.87 and 20.65 μg/ml, their efficacies decreased. The results of this study show that the treatment time of the fungicide should be determined by considering the effect of the fungicide on the spore growth phase of pathogens.

Sheep anthrax outbreak in Muddaballi Village, Karnataka, India: a comprehensive study of factors

Anthrax, caused by Bacillus anthracis, remains a persistent threat to global public health and livestock industries. This study investigates a sheep anthrax outbreak in Muddaballi Village, Karnataka, India. Muddaballi has a population of 2,450 people and a livestock population of 2,279 animals, including 756 cows, 82 buffaloes, 1,129 sheep, and 312 goats. The village experienced anthrax outbreaks in 2016, 2021, and 2023, with significant sheep mortalities: 4 out of 200 sheep in 2016, 25 out of 150 sheep in 2021, and multiple incidents in 2023 affecting different farmers with mortality rates ranging from 4 to 20 sheep. The ecological characteristics, including an average annual temperature of 27.0°C and annual rainfall of 587 mm, alongside proximity to the Tungabhadra reservoir, played a crucial role in spore survival and transmission. The study highlights socioeconomic factors and Risk Exposure and Mitigation Behaviour (REMB) as critical determinants of anthrax incidence, with farmers reporting strong agreement on the influence of these factors (Likert scale: 4-5). The integration of AI tools for early detection and rapid response, comprehensive disease surveillance, and ensuring vaccine accessibility are essential measures. Training initiatives for local veterinarians and farmers are also crucial to mitigate future outbreaks. This multifaceted approach is vital for effective anthrax prevention and control in Muddaballi and similar regions.

Mapping high probability area for the Bacillus anthracis occurrence in wildlife protected area, South Omo, Ethiopia

Anthrax is a zoonotic disease caused by a spore-forming gram-positive bacterium, Bacillus anthracis. Increased anthropogenic factors inside wildlife-protected areas may worsen the spillover of the disease at the interface. Consequently, environmental suitability prediction for B. anthracis spore survival to locate a high-risk area is urgent. Here, we identified a potentially suitable habitat and a high-risk area for appropriate control measures. Our result revealed that a relatively largest segment of Omo National Park, about 23.7% (1,218 square kilometers) of the total area; 36.6% (711 square kilometers) of Mago National Park, and 29.4% (489 square kilometers) of Tama wildlife Reserve predicted as a high-risk area for the anthrax occurrence in the current situation. Therefore, the findings of this study provide the priority area to focus on and allocate resources for effective surveillance, prevention, and control of anthrax before it causes devastating effects on wildlife.

Encapsulation of Trichoderma reesei and Neurospora crassa in calcium alginate capsules for fungi-mediated self-healing concrete: An explorative study on the protocol, survival of the organisms and CaCO3 biomineralization

Concrete, a predominant building material, is known for its material degradation due to crack formation. The associated durability concerns initiated the use of microorganisms to self-heal concrete cracks by promoting CaCO3 precipitation on their cell walls. Filamentous fungi grow in mycelial networks that provide abundant nucleation sites for the precipitation. As demonstrated by the research on bacteria, protection of the microorganism in the concrete mix is crucial to safeguard its survival. In this work, an encapsulation method based on sodium alginate and calcium lactate was identified in the bacteria-based literature and adapted to the fungi Trichoderma reesei and Neurospora crassa. Results showed spore survival after the encapsulation procedure, however survival was negated when immersed in concrete-relevant conditions. Simultaneously, biomass formation and CaCO3 biomineralization were quantified in liquid media. This study features possibilities to optimize the encapsulation procedure and aids the identification of optimal biomineralization conditions.

The multiplicity of thioredoxin systems meets the specific lifestyles of Clostridia.

Cells are unceasingly confronted by oxidative stresses that oxidize proteins on their cysteines. The thioredoxin (Trx) system, which is a ubiquitous system for thiol and protein repair, is composed of a thioredoxin (TrxA) and a thioredoxin reductase (TrxB). TrxAs reduce disulfide bonds of oxidized proteins and are then usually recycled by a single pleiotropic NAD(P)H-dependent TrxB (NTR). In this work, we first analyzed the composition of Trx systems across Bacteria. Most bacteria have only one NTR, but organisms in some Phyla have several TrxBs. In Firmicutes, multiple TrxBs are observed only in Clostridia, with another peculiarity being the existence of ferredoxin-dependent TrxBs. We used Clostridioides difficile, a pathogenic sporulating anaerobic Firmicutes, as a model to investigate the biological relevance of TrxB multiplicity. Three TrxAs and three TrxBs are present in the 630Δerm strain. We showed that two systems are involved in the response to infection-related stresses, allowing the survival of vegetative cells exposed to oxygen, inflammation-related molecules and bile salts. A fourth TrxB copy present in some strains also contributes to the stress-response arsenal. One of the conserved stress-response Trx system was found to be present both in vegetative cells and in the spores and is under a dual transcriptional control by vegetative cell and sporulation sigma factors. This Trx system contributes to spore survival to hypochlorite and ensure proper germination in the presence of oxygen. Finally, we found that the third Trx system contributes to sporulation through the recycling of the glycine-reductase, a Stickland pathway enzyme that allows the consumption of glycine and contributes to sporulation. Altogether, we showed that Trx systems are produced under the control of various regulatory signals and respond to different regulatory networks. The multiplicity of Trx systems and the diversity of TrxBs most likely meet specific needs of Clostridia in adaptation to strong stress exposure, sporulation and Stickland pathways.

Breeding for Bacterial Wilt Disease Resistance in Brinjal

Brinjal is a warm–season vegetable that belongs to the Solanaceae family. Among the biotic stresses of brinjal, bacterial wilt is a major disease caused by a soil-borne bacterium, Ralstonia solanacearum. Both cultural and chemical disease control methods are not effective due to the wide host range and prolonged spore survival periods in soil. In recent past, the use of resistant varieties and hybrids are recognized as the effective method to control bacterial wilt disease. Information on genetics of resistance to bacterial wilt disease is of prime importance for improvement of varieties and hybrids in brinjal. The bacterial wilt resistance response and resistance mechanisms are strictly based on the environmental conditions and cultivars. In traditional crop improvement methods, the development of varieties and hybrids is expensive and time consuming. The discovery of molecular marker systems increases the speed and precision for developing of varieties and hybrids with desired agronomic traits.

Human in vivo assessment of the survival and germination of Heyndrickxia coagulans SNZ1969 spores delivered via gummy candies

Confectionary products hold promise as unconventional food carriers for probiotic microorganisms. This study explored the delivery of Heyndrickxia coagulans SNZ1969, a spore-forming probiotic, using gummy candies. In this study, we prepared gummy candies containing bacterial spores with a viable count that remained stable during a 24-month shelf-life period, meeting the label claim of at least one billion CFUs per serving (24 g). Then, we carried out an intervention trial involving 24 healthy adults who consumed one serving per day for two weeks followed by an additional two weeks of follow-up. Fecal samples were collected and analyzed with a protocol that allowed the viable counts of SNZ1969, both in spore and vegetative forms. The obtained results revealed that bacterial spores germinated in all volunteers. SNZ1969 persistence in the gut was monitored for two weeks after the end of gummy candy consumption, indicating its potential for prolonged colonization. These findings highlight the potential of unconventional food carriers for probiotic delivery and suggest that spore-forming probiotics can be metabolically active in the human intestine. These findings provide information for the development of food products containing spore-forming probiotics and their potential benefits in promoting gastrointestinal health.

Viability of Bacillus coagulans spores with ofloxacin & ornidazole: Implications for gut health

The balance of gut microbiota significantly impacts host health. Disruption of the natural gut flora, often caused by infections or the use of broad-spectrum antimicrobial drugs, can lead to dysbiosis, causing gastrointestinal disorders, such as diarrhea. Probiotics show promising outcomes in restoring gut health, but concerns remain about their interaction with antimicrobials and the viability of spores in the intended gut location. To address these uncertainties, the current study was devised to evaluate how well spores-forming bacteria endure and develop in an environment where antimicrobial agents are present.: The study investigated the survival and growth of spores under the influence of broad-spectrum antimicrobial agents, Ofloxacin and Ornidazole. To cultivate the spores in the presence of these antimicrobials, a mixture of MRS broth and PNY agar media was used. The number of colonies that developed were measured to assess the extent of spore survival and germination.In a simulated environment resembling human intestinal pH, spores exhibited viability. Starting with an initial count of 1.38 billion CFU, the spores multiplied to 8.75 billion CFU at 24 hours and further reached to 86.25 billion CFU at 72 hours in the presence of Ofloxacin and Ornidazole. On the other hand, the viable count reached to the level of 88 billion CFU in the absence of antimicrobial agents.This study offers evidence that spores are able to remain viable and germinate when co-administered with Ofloxacin and Ornidazole.

Impact of Protein Aggregates on Sporulation and Germination of Bacillus subtilis.

In order to improve our general understanding of protein aggregate (PA) management and impact in bacteria, different model systems and processes need to be investigated. As such, we developed an inducible synthetic PA model system to investigate PA dynamics in the Gram-positive model organism Bacillus subtilis. This confirmed previous observations that PA segregation in this organism seems to follow the Escherichia coli paradigm of nucleoid occlusion governing polar localization and asymmetric segregation during vegetative growth. However, our findings also revealed that PAs can readily persist throughout the entire sporulation process after encapsulation in the forespore during sporulation. Moreover, no deleterious effects of PA presence on sporulation, germination and spore survival against heat or UV stress could be observed. Our findings therefore indicate that the sporulation process is remarkably robust against perturbations by PAs and misfolded proteins.

Crystal structure and induced stability of trimeric BxpB: implications for the assembly of BxpB-BclA complexes in the exosporium of Bacillus anthracis.

The outermost exosporium layer of Bacillus anthracis spores, the causative agents of anthrax, is comprised of a basal layer and an external hair-like nap. The nap includes filaments composed of trimers of the collagen-like glycoprotein BclA. Essentially all BclA trimers are attached to the spore in a process in which part of the 38-residue amino-terminal domain (NTD) of BclA forms an extremely stable interaction with the basal layer protein BxpB. Evidence indicates that the BclA-BxpB interaction is direct and requires trimeric BxpB. To further investigate the nature of the BclA-BxpB interaction, we determined the crystal structure of BxpB. The structure was trimeric with each monomer consisting of 11 β strands with connecting loops. The structure did not include apparently disordered amino acids 1-19, which contain the only two cysteine residues of the 167-residue BxpB. The orientation of the structure reveals regions of BxpB that could be involved in interacting with the BclA NTD and with adjacent cysteine-rich proteins in the basal layer. Furthermore, the BxpB structure closely resembles that of the 134-residue carboxyl-terminal domain of BclA, which forms trimers that are highly resistant to heat and detergent. We demonstrated that BxpB trimers do not share this resistance. However, when BxpB trimers are mixed with a peptide containing residues 20-38 of BclA, they form a complex that is as stable as BclA-BxpB complexes extracted from spores. Together, our results provide new insights into the mechanism of BclA-BxpB attachment and incorporation into the exosporium. IMPORTANCE The B. anthracis exosporium plays major roles in spore survival and infectivity, but the complex mechanism of its assembly is poorly understood. Key steps in this process are the stable attachment of collagen-like BclA filaments to the major basal layer structural protein BxpB and the insertion of BxpB into an underlying basal layer scaffold. The goal of this study is to further elucidate these interactions thereby advancing our understanding of exosporium assembly, a process shared by many spore-forming bacteria including important human pathogens.

Microbial Protocols for Spacecraft: 3. Spore Monolayer Preparation Methods for Ultraviolet Irradiation Exposures.

Developing robust microbial survival models for interplanetary and planetary spacecraft requires precise inactivation kinetics for vehicle bioburdens. To generate such data, reliable protocols are required for preparing, testing, and assaying microbial cells or spores on simulated spacecraft materials. New data are presented on the utility of the liquid droplet protocol for applying Bacillus subtilis spores to aluminum coupons. Results indicate that low-density spore monolayers should be created between 2 and 5 × 106 spores per cm2 on individual coupons to prevent the formation of aggregates or multilayers of spores. Such aggregation or multilayers will interfere with the precision of characterizing the effects of UV irradiation on spore survival. Optimum spore monolayers are defined as spore monolayers without overlapping or clustered cells and in which all spores will receive UV photons during assays. The best spore monolayers were created with sterile deionized water (SDIW) on uncoated aluminum coupons, or with SDIW + Triton X-100 (at 0.5 × of the critical micellar concentration) on either uncoated Al-coupons or on Chemfilm Class 1A-coated coupons. The Triton X-100 surfactant improved the uniformity of the monolayers without affecting the sensitivity of the spores to UV irradiation. Furthermore, spore layers created at either 2 × 107 or 2 × 108 spores/cm2 created multi-stacking effects that clearly reduced the precision of the UV irradiation assays. A set of standardized protocols is suggested for spacecraft processing and planetary protection communities to permit directly comparing results from divergent labs.

Addition of sodium alginate capsules containing Lysinibacillus sphaericus for self-healing of cracks in mortars

The biomineralization technique of crack remediation in construction mortar can be achieved through the incorporation of calcifying bacteria within the mortar matrix. However, the thermal and autogenous shrinkage associated with the hydration of cement can be detrimental to the survival of the bacterial spores. Encapsulation of the spores in a matrix such as superabsorbent hydrogels, specifically sodium alginate, can provide protection from such detrimental conditions while also serving as a water reservoir for the metabolic activity of the bacterial cells. This paper investigated the addition of sodium alginate capsules prepared through spherification in construction mortars, aiming to preserve and optimize rheological, physical and mechanical properties, and to release bacteria when cracks occur. Mortar formulations with capsules containing Lysinibacillus sphaericus were calculated in order to maintain the granulometric distribution of standard formulation, thus obtaining products with excellent rheological, physical and mechanical properties, and allowing the addition of up to 1% capsules. Multivariate analysis of variance was applied at each age, showing that formulations with capsules kept the same properties as the original formula. After cracking, it was possible to observe the action of the released bacteria, through the formation of crystalline structures at the cracks. The capsules were prepared through a simple and cheap process, and it was proved that not only they do not affect concrete properties, but also are effective in both protecting the bacteria during mixing and hardening and releasing it after cracking.

Antifungal Activity of Perillaldehyde on Fusarium solani and Its Control Effect on Postharvest Decay of Sweet Potatoes.

Root rot caused by Fusarium solani is one of the major postharvest diseases limiting sweet potato production. Here, antifungal activity and the action mode of perillaldehyde (PAE) against F. solani were investigated. A PAE concentration of 0.15 mL/L in air (mL/L air) markedly inhibited the mycelial growth, spore reproduction and spore viability of F. solani. A PAE vapor of 0.25 mL/L in air could control the F. solani development in sweet potatoes during storage for 9 days at 28 °C. Moreover, the results of a flow cytometer demonstrated that PAE drove an increase in cell membrane permeability, reduction of mitochondrial membrane potential (MMP) and accumulation of reactive oxygen species (ROS) in F. solani spores. Subsequently, a fluorescence microscopy assay demonstrated that PAE caused serious damage to the cell nuclei in F. solani by inducing chromatin condensation. Further, the spread plate method showed that the spore survival rate was negatively correlated with the level of ROS and nuclear damage, of which the results indicated that PAE-driven ROS accumulation plays a critical role in contributing to cell death in F. solani. In all, the results revealed a specific antifungal mechanism of PAE against F. solani, and suggest that PAE could be a useful fumigant for controlling the postharvest diseases of sweet potatoes.

The persistence of time: the lifespan of Bacillus anthracis spores in environmental reservoirs

Anthrax is a lethal bacterial zoonosis primarily affecting herbivorous wildlife and livestock. Upon host death Bacillus anthracis vegetative cells form spores capable of surviving for years in soil. Anthrax transmission requires host exposure to large spore doses. Thus, conditions that facilitate higher spore concentrations or promote spore survival will increase the probability that a pathogen reservoir infects future hosts. We investigated abiotic and pathogen genomic variation in relation to spore concentrations in surface soils (0–1 cm depth) at 40 plains zebra (Equus quagga) anthrax carcass sites in Namibia. Specifically, how initial spore concentrations and spore survival were affected by seasonality associated with the timing of host mortality, local soil characteristics, and pathogen genomic variation. Zebras dying of anthrax in wet seasons—the peak season for anthrax in Etosha National Park—had soil spore concentrations 1.36 orders of magnitude higher than those that died in dry seasons. No other variables considered affected spore concentrations, and spore survival rates did not differ among sites. Surface soils at these pathogen reservoirs remained culture positive for a range of 3.8–10.4 years after host death. Future research could evaluate if seasonal patterns in spore concentrations are driven by differences in sporulation success or levels of terminal bacteremia.

Dry/wet cycling reduces spore germination and viability in six peatland bryophytes.

Dry/wet cycling driven by water level fluctuation in wetlands may strongly influence the destiny of seeds. However, how dry/wet cycling affects spore survival and germinability in peatland bryophytes is poorly understood. Six peatland bryophytes, three hummock- and three hollow-dwelling Sphagnum species, were chosen as study species. We tested the effects of dry (60% air RH)/wet (waterlogging) cycle frequency (once per 12, 8 or 4 days for low, medium or high, respectively) and ratio (3:1, 1:1 or 1:3 dry:wet time per cycle) on spore germinability, viability, dormancy percentage and protonema development. Dry/wet cycling significantly reduced spore germination percentage and viability and slowed protonema development in all Sphagnum species, being more pronounced with higher dry/wet cycling frequencies. The hummock species S. capillifolium and S. fuscum had higher spore germination percentage after the continuous dry treatment, while the hollow species S. angustifolium, S. squarrosum and S. subsecundum showed the opposite response, compared to the continuously wet treatment. Except for S. squarrosum, spore viability was higher after the dry than after the wet treatment. Spore viability and dormancy percentage were higher after a dry/wet ratio of 1:3 than after ratios of 3:1 and 1:1. Our study shows that both germinability and viability of bryophyte spores are reduced by dry/wet cycling (especially when frequent) in peatlands. This emphasizes the need to ensure constant water levels and low frequencies of water level fluctuation, which are relevant in connection with wetland restoration, to promote Sphagnum spore survival and establishment in peatlands after disturbances.

Survival of Clostridioides difficile spores in thermal and chemo-thermal laundering processes and influence of the exosporium on their adherence to cotton bed sheets.

Clostridioides difficile spores were previously demonstrated to survive industrial laundering. Understanding interactions between heat, disinfectants and soiling (e.g. bodily fluids) affecting C. difficile spore survival could inform the optimization of healthcare laundry processes. Reducing spore attachment to linen could also enhance laundering efficacy. This study aimed to compare the sensitivity of C. difficile spores to heat and detergent, with and without soiling and to investigate adherence to cotton. Survival of C. difficile spores exposed to industrial laundering temperatures (71-90°C), reference detergent and industrial detergent was quantified with and without soiling. The adherence to cotton after 0 and 24 h air drying was determined with the exosporium of C. difficile spores partially or fully removed. Clostridioides difficile spores were stable at 71°C for 20 min (≤0·37 log10 reduction) while 90°C was sporicidal (3 log10 reduction); soiling exerted a protective effect. Industrial detergent was more effective at 71°C compared to 25°C (2·81 vs 0·84 log10 reductions), however, specifications for sporicidal activity (>3 log10 reduction) were not met. Clostridioides difficile spores increasingly adhered to cotton over time, with 49% adherence after 24 h. Removal of the exosporium increased adherence by 19-23% compared to untreated spores. Further understanding of the role of the exosporium in attachment to cotton could enhance spore removal and aid decontamination of linen.

Antimicrobial peptide CB-M exhibits direct antifungal activity against Botrytis cinerea and induces disease resistance to gray mold in cherry tomato fruit

Gray mold is a common postharvest disease caused by Botrytis cinerea that causes fruit decay and reduces fruit storage quality. In this study, the antifungal activity of antimicrobial peptide CB-M against B. cinerea and its mechanism of action, as well as the ability of CB-M to induce disease resistance in cherry tomato fruit, were investigated. CB-M had a strong inhibitory effect on spore survival and mycelial growth of B. cinerea, and these effects were enhanced with increased CB-M concentration. Moreover, significant leakages of cellular constituents and fluorescence microscopy assay confirmed that CB-M treatment could destroy the cell membrane integrity of B. cinerea. Scanning electron microscopy observations further demonstrated obvious distortion and shriveling of B. cinerea hyphae. Gel retardation and SDS-PAGE assay showed that CB-M could likely bind to cellular DNA and degrade/inhibit protein synthesis in B. cinerea. Furthermore, preincubation with CB-M before pathogen inoculation significantly decreased lesion diameter and incidence of gray mold caused by B. cinerea in tomato, cherry tomato, and grape fruit throughout whole storage time. Cherry tomato was used to explore the ability of CB-M to induce gray mold resistance, and the results showed that CB-M preincubation enhanced the activities of superoxide dismutase, peroxidase, and catalase, accelerated the accumulation of pathogenesis-related proteins, flavonoids, and total phenolics, and increased the activities of β-1,3-glucanase, chitinase, and phenylalanine ammonia-lyase during storage. These results indicated that CB-M is a good substitute for fungicides and has excellent prospects for controlling postharvest gray mold in horticultural crops.

Disease in regenerating pine forests linked to temperature and pathogen spillover from the canopy

Abstract Previous investigations of the role of pathogens as drivers of community dynamics in forests have mostly focused on interspecific interactions between canopy trees and seedlings via soil feedbacks. However, feedbacks can also occur directly between canopy trees and seedlings when spores of foliar and stem pathogens fall onto seedlings regenerating underneath the canopy. We studied pathogen spillover between canopy trees and conspecific and heterospecific regeneration in the Pinus nigra–Diplodia sapinea pathosystem. We sampled 70 pine stands distributed across a temperature gradient of 9.5–13.7°C (mean annual temperature). In each stand, we linked spore load and spore survival in the canopy with pathogen biomass and disease severity in regenerating seedlings. The density of regenerating seedlings and the health status of conspecific and heterospecific seedlings were also measured. The strength of canopy–understorey interactions was correlated with both climatic and stand variables. The most severe symptoms of disease in regenerating seedlings were found in the warmest stands. Structural equation models suggested that disease severity in seedlings was mediated by an increased spore spillover from the canopy. Temperature also increased the pathogen survival rate within seedlings, further contributing to disease severity. Interestingly, disease severity in the canopy did not correlate with spore load in the canopy, suggesting that adult trees were spilling pathogens onto the understorey regardless of their health status. Pathogen spillover increased in more open stands. Greater disease severity in seedlings correlated with stands with a higher density of oak seedlings. Synthesis. These results suggest an understudied mechanism of the Janzen–Connell theory, according to which, canopy–understorey negative feedbacks could be driven by canopy pathogens. Our data also suggest that temperature shapes canopy–understorey interactions, which implies that under a warming climate scenario, an acceleration of conspecific negative feedbacks could be expected, with implications for forest regeneration dynamics.