Mycelial growth rate and toxin production in the seed pathogen Pyrenophora semeniperda: resource trade‐offs and temporally varying selection

Pyrenophora semeniperda, an important pathogen in Bromus tectorum seed banks in semi‐arid western North America, exhibits >4‐fold variation in mycelial growth rate. Host seeds exhibit seasonal changes in dormancy that affect the risk of pathogen‐caused mortality. The hypothesis tested is that contrasting seed dormancy phenotypes select for contrasting strategies for increasing pathogen fitness, and that increased fitness on nondormant seeds involves a resource trade‐off between toxin production and growth. The strategy for successfully attacking rapidly germinating nondormant seeds at high inoculum loads in autumn involves increased post‐infection aggressiveness to prevent seed escape through germination. An earlier study demonstrated that slow‐growing strains caused higher mortality than faster‐growing strains on nondormant host seeds at high inoculum loads. In this study, production of the toxin cytochalasin B was significantly higher in slower‐growing strains, and was induced only in seeds or in seed‐constituent‐containing media. Its production was reduced in vivo by Bromus tectorum seeds, suggesting direct involvement in pathogenesis on seeds. Fast‐growing strains caused significantly higher mortality than slow‐growing strains at low inoculum loads on dormant seeds, which apparently have resistance that is overcome at high loads or through rapid mycelial proliferation. In a co‐inoculation study, the fast‐growing isolate produced 3 × more stromata than the slow‐growing isolate on dormant seeds, whereas the slow‐growing isolate was twice as successful on nondormant seeds. These results provide evidence that mycelial growth rate variation and associated variation in cytochalasin B production represent a trade‐off maintained through temporally varying selection resulting from seasonal variation in host seed dormancy status.     

Relative contribution of genotype, seed size and environment to secondary seed dormancy potential in Canadian spring oilseed rape (Brassica napus)

Secondary seed dormancy has been linked to seedbank persistence of volunteer oilseed rape (Brassica napus) in western Canada. It has been suggested that there is a genetic component to secondary seed dormancy expression in oilseed rape, but little is known of its importance in relation to non‐genetic factors. In a series of experiments we investigated the relative importance of genotype, seed size, time of windrowing and pre‐ and post‐harvest environment on the expression of secondary seed dormancy. We found that genotype contributed between 44 and 82% to the total variation in secondary seed dormancy. A broad range in secondary seed dormancy expression was observed among 16 genotypes examined. Nevertheless, three‐quarters of the genotypes investigated exhibited relatively high potential for the expression of secondary seed dormancy (back‐transformed mean 71% dormant seeds). Seed size contributed 21% to the total variation, while the influence of seed maturity (harvest regime) on secondary seed dormancy expression was negligible. Despite diverging environmental conditions during the four growing seasons spanning these experiments, the influence of pre‐harvest environment on seed dormancy expression was relatively small and ranged from 0.1% to 4.5%. Secondary seed dormancy potential decreased over time during seed storage. This decrease was greatest when seeds were stored at ambient temperatures and least when seeds were stored at ?70°C.     

Seed dormancy and soil seedbank of the invasive weed Chenopodium hybridum in north‐western China

Seed dormancy and persistence in the soil seedbank play a key role in timing of germination and seedling emergence of weeds; thus, knowledge of these traits is required for effective weed management. We investigated seed dormancy and seed persistence on/in soil of Chenopodium hybridum, an annual invasive weed in north‐western China. Fresh seeds are physiologically dormant. Sulphuric acid scarification, mechanical scarification and cold stratification significantly increased germination percentages, whereas dry storage and treatments with plant growth regulators or nitrate had no effect. Dormancy was alleviated by piercing the seed coat but not the pericarp. Pre‐treatment of seeds collected in 2012 and 2013 with sulphuric acid for 30 min increased germination from 0% to 66% and 62% respectively. Effect of cold stratification on seed germination varied with soil moisture content (MC) and duration of treatment; seeds stratified in soil with 12% MC for 2 months germinated to 39%. Burial duration, burial depth and their interaction had significant effects on seed dormancy and seed viability. Dormancy in fresh seeds was released from October to February, and seeds re‐entered dormancy in April. Seed viability decreased with time for seeds on the soil surface and for those buried at a depth of 5 cm, and 39% and 10%, respectively, were viable after 22 months. Thus, C. hybridum can form at least a short‐lived persistent soil seedbank.     

Weed seed decay in conventional and diversified cropping systems

Diversified cropping systems can have high soil microbial biomass and thus strong potential to reduce the weed seedbank through seed decay. This study, conducted in Iowa, USA, evaluated the hypothesis that weed seed decay is higher in a diversified 4‐year maize–soyabean–oat/lucerne–lucerne cropping system than in a conventional 2‐year maize–soyabean rotation. Mesh bags filled with either Setaria faberi or Abutilon theophrasti seeds and soil were buried at two depths in the maize phase of the two cropping systems and sampled over a 3‐year period. Setaria faberi seed decay was consistently greater at 2 cm than at 20 cm burial depth and was higher in the more diverse rotation than in the conventional rotation in 1 year. Abutilon theophrasti seeds decayed very little in comparison with seeds of S. faberi. Separate laboratory and field experiments confirmed differences in germination and seed decay among the seed lots evaluated each year. Fusarium, Pythium, Alternaria, Cladosporium and Trichoderma were the most abundant genera colonising seeds of both species. A glasshouse experiment determined a relationship between Pythium ultimum and S. faberi seed decay. Possible differences in seed susceptibility to decay indicate the need to evaluate weed seedbank dynamics in different cropping systems when evaluating overall population dynamics and formulating weed management strategies.     

The roles of light and pericarp on seed dormancy and germination in feral Raphanus sativus (Brassicaceae)

The timing of seed germination may determine the success of a weed species in an agroecosystem, and its expression is modulated by environmental conditions, but also by seed physiology and anatomy. The aims of this study were to investigate the roles of light, pericarp, dry storage and cold stratification on seed dormancy and germination in feral radish, a troublesome agricultural weed in temperate zones of the Americas that reduces crop yields. To this end, we used isolated intact pods and extracted seeds to test germination over time under contrasting temperature, light and storage conditions. Here, we showed that fresh seeds were non‐dormant, but that light and the presence of the pericarp reduced germination, especially under low temperatures. The pericarp reduced the final water content absorbed by seeds inside pods and decreased absorption/dehydration rates. The pericarp showed several small lignified cell layers in the endocarp, and x‐ray images displayed the lack of space between the partially embedded seed and the endocarp. Dry storage and cold stratification were ineffective in breaking the dormancy imposed by the pericarp. The apparent requirement for darkness and the mechanical restriction of the pericarp may have the potential to induce dormancy, spreading the timing of seed germination over a more extended period and hindering the control of feral radish.     

Thermal and hormonal regulation of the dormancy–germination transition in Amaranthus tuberculatus seeds

The transition from seed dormancy to germination is a multi‐step process. However, distinguishing between physiological processes involved in seed dormancy alleviation and those involved in germination has been difficult. We studied the seed dormancy alleviation process in Amaranthus tuberculatus, an important weed species in midwestern USA. Using three A. tuberculatus biotypes that differ in dormancy level, it was determined that stratification reduced seed dormancy from a high to a low level. Temperature alternation alleviated low seed dormancy and triggered germination. Exogenously applied abscisic acid (ABA) and gibberellic acid (GA) had no effect on seeds with high dormancy. However, ABA and paclobutrazol (a GA biosynthesis inhibitor) significantly reduced germination of seeds with low dormancy. Hormones could not replace the effects of stratification or temperature alternation on dormancy alleviation. Based on our results, we propose a seed dormancy–germination transition model in which the dormancy of A. tuberculatus seeds is progressively reduced from a high to a low level; but environmental conditions (i.e. stratification) can accelerate the dormancy alleviation process. Under low dormancy levels, the seed is more sensitive to environmental cues that are responsible for removing dormancy and triggering germination (i.e. temperature alternation). Finally, ABA and GA regulation occurs primarily during the final transition from low dormancy to germination rather than the alleviation of high dormancy.     

Effect of burial depth and soil water regime on the fate of Lithospermum arvense seeds in relation to burial time

Lithospermum arvense is an increasing annual weed in winter crops of the semiarid region of southern Argentina under low impact tillage systems, an agricultural practice that has become popular in recent years. Seed distribution in the soil profile under conventional tillage will change when reduced tillage is implemented, thus affecting the germination microenvironment. The effect of seed burial depth and soil water regime on field germination, enforced dormancy, innate dormancy and seed decay was studied in relation to burial time in a field experiment. In addition, the effect of burial depth on seed germination and seedling emergence was examined under laboratory controlled conditions. Field germination of buried seed ranged from 55% to 65% for shallow (2 cm) and from 5% to 30% for greater depths (20 cm). Enforced dormancy levels were significantly higher among deeper seeds. The amount of innate dormant seeds was reduced to <10% after a year of burial. Lithospermum arvense seedbanks can be classified as short-term persistent. Germination in the laboratory was unaffected by burial depth, while seedling emergence reduction was adequately described by a sigmoidal model. Results indicate that agricultural practices that accumulate L. arvense seeds near the soil surface enhance seedling recruitment.     

Germination rates of weedy radish populations (Raphanus spp.) altered by crop‐wild hybridisation,not human‐mediated changes to soil moisture

Cultivated plants are known to readily hybridise with their wild relatives, sometimes forming populations with weedier life‐history strategies than their progenitors. Due to altered precipitation patterns from human‐induced global climate change, crop‐wild hybrid populations may have new and unpredictable environmental tolerances relative to parental populations, which would further challenge farming and land‐management weed control strategies. To recognise the role of seed dormancy variation in weed invasion, we compared seedbank dynamics of two cross‐type populations (wild radish, Raphanus raphanistrum, and crop‐wild hybrid radish, R. raphanistrum × R. sativus) across a soil moisture gradient. In a seed‐burial experiment, we assessed relative rates of seed germination, dormancy and seed mortality over two years across cross types (crop‐wild hybrid or wild) and watering treatments (where water was withheld, equal to annual rainfall, or double annual rainfall). Weekly population censuses in 2012 and 2013 assessed the frequency and timing of seedling emergence within a growing season. Generally, germination rates were two times higher and seed dormancy was 58% lower in hybrid versus wild populations. Surprisingly, experimental soil moisture conditions did not determine seedbank dynamics over time. Yet, seed bank dynamics changed between years, potentially related to different amounts of annual rainfall. Thus, variation in seedbank dynamics may be driven by crop‐wild hybridisation rates and, potentially, annual variation in soil moisture conditions.     

Behaviour of buried and surface-sown seeds of Parthenium hysterophorus

Parthenium hysterophorus L. seeds were buried at a depth of 5 cm for periods of 2–24 months to determine their longevity. The majority (73.7%) of these seeds were still viable after 24 months of burial. The remainder could not be recovered (18.0%) or were no longer viable (8.3%). There was a log-linear decline in persistence of germinable seeds over time, which indicated a constant rate of loss and a half-life of about 6 years. Seedling emergence from surface-sown seeds was also studied. Although there was considerable rainfall (31 mm), seedlings did not emerge during the first month of this experiment. In the succeeding 3 months, there was substantial seedling emergence after rainfall, and 51.4% of seeds had germinated by the end of the fourth month. After 5 months had passed, further seedling emergence was not detected, and intact seeds could not be located. These findings suggest that seed incorporation into the soil is important to the long-term persistence of P . hysterophorus seeds. In an initial test of germination, unburied seeds from the same seed lot exhibited a degree of innate dormancy, and this may explain the delayed germination observed in the surface-sown seeds. In the seed burial and recovery experiment, innate dormancy was lost after 2 months of burial in the field, although in situ germination of buried seed remained low for at least 24 months. Therefore, it appears that more than one dormancy mechanism may contribute to the persistence of P. hysterophorus seeds.     

Assessing wind and mammals as seed dispersal vectors in an invasive legume

While some plants have modified seed structures to facilitate dispersal, many lack such specialised adaptations, making their mode of dispersal unclear. This can be particularly problematic for predicting shifts in species ranges or tracking the spread of invasive plants. As an example, the seed size and shape of the invasive legume, Lespedeza cuneata, suggest that wind and attachment to animals are not important for dispersal, yet populations can spread surprising distances within a few years. Using a series of experiments conducted in the laboratory and field, we tested wind and mammal fur as mediators of seed dispersal. To test wind dispersal, seed traps were arranged radially around a patch of L. cuneata and seeds were collected following dispersal. Attachment to mammal fur was tested by fitting pelts of a deer, coyote and raccoon to artificial torsos and determining seed retention in both the field and the laboratory. Laboratory trials also examined the influence of wet versus dry conditions. Our results showed that wind direction strongly influenced dispersal distance and seeds were readily dispersed by mammal fur. The number of seeds retained was species specific, depending on fur depth and mammal size, with seed retention increasing under wet conditions. Together, these results suggest that both wind and mammal fur contribute to the movement of L. cuneata across grasslands. Consequently, both dispersal vectors should be considered when designing and implementing control strategies.     

Impact of cultivar on survival of volunteer oilseed rape populations in fields is more important than field management

A joint assessment of two separate approaches investigated the occurrence of volunteer oilseed rape (Brassica napus L.; OSR ) as affected by cultivar, field history and environment. Approach I comprised surveys for volunteers on >100 farmers’ fields in Germany in the years 2009 and 2010. Volunteers were assigned to the cultivars grown in previous years by inter‐simple sequence repeat‐PCR and cluster analysis. High‐dormancy cultivars resulted in 0–7 and low‐dormancy cultivars in 0–1.3 volunteers m^?2. Highest numbers originated from the most recent harvests 2007 and 2006. Approach II was a meta‐analysis based on 116 data sets from field trials and farmers’ fields in Germany to evaluate and to rank the impact of management factors on the soil seedbank and volunteers in following crops. Varietal disposition to seed dormancy turned out to be the significantly most relevant factor. The contribution of varietal dormancy to variation in the soil seedbank and of volunteers in the 1st and 2nd following crop was 1.2–2.3 times as great as the contribution of post‐harvest tillage. Up to 45% of the variation in the observations originated from factors that can be controlled by human actions on a given location in a given year. The overall analysis confirmed the results from independent short‐term trials and showed that both agronomists and breeders need to contribute to reducing OSR volunteers.     

Using panicle dry weight to estimate seed production in Echinochloa crus‐galli

A better understanding of weed seed production is a key element for any long‐term management allowing some weeds to shed seeds. The challenge with measuring seed production in weeds is the large effort required in terms of time and labour. For the weed species Echinochloa crus‐galli, it was tested whether the number of seeds per panicle dry weight or per panicle length can be used to estimate seed production. Experiments were conducted in three maize fields in north‐eastern Germany. The effect of factors that could influence this relationship, such as the time of seedling emergence, the density of E. crus‐galli, the control intensity of other weeds, seed predation and field, was included. A few days before maize harvest, all panicles were removed and weighed, panicle length was measured, and for a subsample of 178 panicles, the number of seeds was counted manually. Panicle dry weight predicted the number of seeds per panicle better (R² = 0.92) than did panicle length (R² = 0.69). The other factors except for ‘field’ and ‘seed predation’ had no effect on these relationships. The relationships between seed number and panicle dry weight found in this study closely resembled those reported in an earlier study. Based on our results, we conclude that both plant traits are appropriate for the estimatation of seed production, depending on required level of precision and availablilty of resources for the evaluation of sustainable weed management strategies.     

Efficacy of UV‐C radiation to reduce seedborne anthracnose (Colletotrichum acutatum) from Andean lupin (Lupinus mutabilis)

The potential of UV‐C radiation of Andean lupin (Lupinus mutabilis) seeds to eradicate seedborne infections of anthracnose caused by Colletotrichum acutatum was investigated. UV‐C doses from 0 to 691.2 kJ m^?2 (resulting from 0 to 96 h of exposure time) on disease incidence reduction and germination on artificially and naturally infected seed were evaluated. The degree of incidence reduction and seed germination was dependent on the dose of UV‐C. The UV‐C doses of 86.4 kJ m^?2 and higher reduced incidence from 6% to 7% to undetectable levels, but these UV‐C doses also reduced seed germination. UV‐C can deleteriously affect physiological processes and overall growth. To assess its impact, L. mutabilis seeds irradiated with UV‐C doses of 57.6 and 86.4 kJ m^?2 were grown. Seedlings grown from noninfected seed and UV‐C treated seed showed an increased concentration of chlorophyll and protein contents, as well as an increase in the activation of defence enzymes peroxidase and catalase, in comparison with plants grown from infected seed. UV‐C doses resulted in seed emergence and seedling dry weight rates that were similar to the noninfected control or better than the fungicide control. Moreover, 57.6 kJ m^?2 reduced transmission of the pathogen from seed to the plantlets by 80%, while 86.4 kJ m^?2 apparently eradicated the pathogen, under greenhouse conditions. The use of UV‐C, first reported here, is advantageous for controlling anthracnose in lupin.     

Dormancy,germination and emergence of Urochloa panicoides regulated by temperature

Urochloa panicoides is an annual weed of summer crops. In Argentina, in subhumid areas with monsoon rainfall, it germinates and establishes in a single flush. To (i) identify the environmental factors that modify its seed dormancy level and germination and (ii) quantify the parameters describing the thermal behaviour of the germination and emergence dynamics of this weed under non‐limiting water conditions, we established a set of germination experiments performed (i) under controlled conditions using seeds after ripened for 3 or 6 months in different thermal and hydric conditions and (ii) under field conditions, where the soil temperature was modified by applying different shading levels. Seed dormancy level remained high with 3 months after ripening in all treatments. After 6 months, seeds stored at 4°C in dry conditions did not germinate at any temperature, while seeds stored at 25°C in dry conditions and in situ germinated c. 20% and 60% respectively. Germination percentage was higher in seeds harvested before their natural dispersal. The base, optimum and maximum temperatures for seed germination were 6, 35 and 45°C respectively. Shading reduced the number of emerged seedlings, possibly by reducing the soil thermal amplitude. The results explained the dormancy‐breaking mechanism of U. panicoides that allows a high germination rate in the field when rainfall occurs.     

How much of seed dormancy in weeds can be related to seed traits?

Seed dormancy contributes to species persistence in unpredictable environments and is a key process to be taken into account in weed dynamics models. As the level of seed dormancy, photosensitivity and the dates of dormancy induction and release are difficult to measure, our objective was to relate weed seed dormancy with morphological, chemical or physiological seed traits and with expert knowledge. Dormancy of four species was studied experimentally during a 2‐year seed burial experiment. Experiments were supplemented with data from the literature to increase the number of species analysed, resulting in a data set of 29 species. Proportions of non–dormant seeds were higher for elongated than spherical seeds, even when accounting for phylogenetic relatedness between species. Elongated seeds, which tend to remain on the soil surface in undisturbed habitats, may have been selected for lack of dormancy and immediate germination to limit mortality due to predation. Dormancy increased with seed coat thickness, which can act as a chemical and physical barrier to germination, while no relation was found with seed lipid or protein content. No correlation was found between photosensitivity parameters and any of the species traits analysed. Variations in dormancy dates (induction and release) were highly correlated with average field emergence season estimated from expert knowledge. The observed correlations suggest that the level of dormancy results both from direct and from indirect effects of traits being involved in trade‐offs together with seed mortality.     

Plant–herbivore asynchrony necessitates augmentative releases of the exotic beetle,Zygogramma bicolorata,to enhance the biological control of P arthenium hysterophorus

Systematic information on the quantitative impact of Z ygogramma bicolorata on the biology of P arthenium hysterophorus is crucial as the seeds of this weed continue to germinate from the accumulated soil seed bank throughout the year in the form of different germinating flushes, while the activity of the beetle ceases during winter as it enters diapause. Therefore, plant–herbivore interactions need to be explored to develop predictions of the overall impact of the introduced beetle on the weed. The findings revealed that defoliation by Z . bicolorata had a significant impact on the plant height, density and flower production in flushes F ₃, F ₄ and F ₅, but not in F ₁ and F ₂ that exhibited longer periodicity, profuse branching, a longer flowering period and maximum flower production and contributed mostly to the existing seed soil bank. Therefore, total depletion of the existing soil seed bank was not possible. Consequently, the effect of augmentative field releases of laboratory‐reared beetles was explored on F ₁ and F ₂ in February for three consecutive years (2011–2013). Before initiating the trial, random soil samples were taken from the plots that were assigned to the paired treatments (i.e. with the beetle and without the beetle [insecticide‐treated]) and it was found that the seed bank in those samples did not differ. The single release of Z . bicolorata adults at five per plant at the six‐leaf stage significantly reduced the soil seed bank, compared to without the biocontrol agent, irrespective of the flushes at the end of the season.     

Modelling Chloris virgata germination and emergence under different temperature and light quality conditions

Chloris virgata is a problematic weed around the world. Prediction of weed germination rates could be a useful strategy to optimise timing of weed control actions. We studied the germination and emergence of C. virgata collected seeds under different after-ripening treatments and different exhumation dates after seed dispersal, to estimate seed dormancy level and predict weed emergence dynamics under field conditions. Three experiments were conducted under controlled conditions to determine base, optimum and maximum germination temperatures (T_b, T_o and T_m respectively) and comprised: (a) exposure of seeds to gradually increasing and decreasing temperatures between 5 and 35°C; (b) exposure of seeds to different constant temperatures; and (c) exposure of seeds to different light quality conditions (red – far red ratio) and temperature regimes (constant and alternating temperatures). To explore genuine environmental conditions, a field experiment was performed to determine weed emergence under different shading levels. Finally, with the data obtained, a thermal time model for dormancy release was used to predict C. virgata seedling emergence in the Argentine Pampas region. Seeds after-ripened in cold and wet conditions and constant 25°C showed the highest germination percentages. The values of T_b (7°C), T_o (28°C) and T_m (40°C) remained constant at all exhumation dates. Neither light quality nor thermal regime modified the final germination percentages. However, shading delayed seedling emergence under field conditions, even when it was adjusted by thermal time. These results may allow predicting C. virgata emergence in temperate regions and help to improve weed control in integrated weed management strategies.     

Survival of seeds from perennial biomass species during commercial‐scale anaerobic digestion

Tall perennial grass species can be utilised as bioenergy feedstocks, but some are considered invasive species. Using biomass from such species as feedstocks for anaerobic digestion (AD) may introduce the risk of disseminating viable seeds onto agricultural lands during digestate application. To evaluate this risk, we investigated the survival rates of perennial grass seeds obtained from biomass species during AD. After removal from the digester, seeds were germinated and stained with tetrazolium chloride to determine viability. During three experimental runs, batches of 100 seeds from four species were exposed to 0, 2, 6, 12, 24, 48, 72 and 168 h of mesophilic (38°C) AD within a commercial‐scale digester. Seed viability of Phalaris arundinacea, Phragmites australis, Panicum virgatum and Solanum lycopersicum was reduced by 95% (LT₉₅) after 29, 52, 98 and 105 h of AD respectively. Commercial digesters that utilise perennial grasses as a feedstock typically have retention times ranging from 240 to 1480 h, which greatly exceeds the LT₉₅ values found in this study. Anaerobic digestion resulted in the rapid death of seeds in all species tested, suggesting unwanted dissemination of perennial grass species via digestate application to agricultural land is unlikely.     

A review of the relationship between primary and secondary dormancy,with reference to the volunteer crop weed oilseed rape (Brassica napus)

Fresh seeds of oilseed rape (Brassica napus) are reported to be nondormant and nonphotoblastic. However, a portion of the seeds can be induced into a light‐requiring state (secondary dormancy) for germination and also exhibit dormancy cycling. Thus, if seeds become buried in the soil they can form a persistent seedbank and become a serious volunteer weed in succeeding crops. The capacity of nondormant seeds of B. napus to be induced into secondary dormancy is contrary to results of studies on fresh nondormant seeds of some other species. A reanalysis of published and unpublished data shows that fresh seeds of this species have some degree of primary dormancy and that there is a significant relationship between primary dormancy and the capacity to enter secondary dormancy. However, most germination tests on B. napus have not been done in enough detail to detect primary dormancy (or not) in fresh seeds of this species. The usefulness of information on the relationship between primary dormancy and the capacity of the seeds to enter secondary dormancy is discussed in relation to management of weedy volunteers of this species.     

Long‐term survival of Acidovorax citrulli in citron melon (Citrullus lanatus var. citroides) seeds

Long‐term survival of Acidovorax citrulli in citron melon (Citrullus lanatus var. citroides) seeds was investigated. Citron melon seed lots infected with A. citrulli were generated in the field by inoculating either the pistils (stigma) or pericarps (ovary wall) of the female blossoms. Seventeen A. citrulli isolates from 14 different haplotypes belonging to two different groups (group I and II) were used for inoculation. After confirming that 100% of seed lots were infected, they were stored at 4°C and 50% RH for 7 years. After storage, the viability of A. citrulli cells from individual lots was determined by plating macerated seeds on semiselective medium as well as growing seeds for 14 days and scoring for bacterial fruit blotch symptoms. The type of A. citrulli isolate (group I or group II) used did not significantly influence bacterial survival. However, A. citrulli survival was significantly greater in seed lots generated via pistil inoculation (52·9 and 29·4%) than via pericarp inoculation (23·5 and 17·6%). Repetitive extragenic palindrome (rep)‐PCR on A. citrulli isolated from citron melon seed lots after storage displayed similar fingerprinting patterns to those of the reference strains originally used for blossom inoculation, indicating that cross‐contamination did not occur. The results indicate that A. citrulli may survive/overwinter in citron melon seeds for at least 7 years and bacterial survival in seed was influenced more by method of blossom inoculation than by the type of bacterial isolate.