Pyrenopeziza Brassicae Research Articles

The roles of ascospores and conidia of Pyrenopeziza brassicae in light leaf spot epidemics on winter oilseed rape (Brassica napus) in the UK

SummaryAscospores of Pyrenopeziza brassicae were produced in apothecia (cup‐shaped ascomata) on oilseed rape debris. The conidia, which were morphologically identical to the ascospores, were produced in acervular conidiomata was greater than for lesions caused by ascospores. In June 2000, on the ground under a crop with light on the surface of living oilseed rape tissues. Ascospores were more infective than conidia on oilseed rape leaves. The proportion of lesions caused by conidia located on leaf veins leaf spot, numbers of petioles with apothecia decreased with increasing distance into the crop from the edge of pathways. Air‐borne ascospores of P. brassicae were first collected above debris of oilseed rape affected with light leaf spot on 5 October 1998 and 18 September 1999,12 or 23 days, respectively, after the debris had been exposed outdoors. P. brassicae conidia were first observed on leaves of winter oilseed rape on 6 January 1999 and 15 February 2000, respectively, after plots had been inoculated with debris in November 1998 and October 1999. In 1991/92, numbers of ascospores above a naturally infected crop were small from January to April and increased in June and July. P. brassicae conidia were first observed in February and the percentage plants with leaves, stems or pods with light leaf spot increased greatly in May and June. In 1992/93, in a crop inoculated with debris, numbers of airborne ascospores were small from October to January and increased from April to June. P. brassicae conidia were first observed on leaves in late November and light leaf spot was seen on stems and pods in March and June 1993, respectively.

Effects of temperature and wetness duration on conidial infection, latent period and asexual sporulation ofPyrenopeziza brassicaeon leaves of oilseed rape

Experiments in controlled environments were carried out to determine the effects of temperature and leaf wetness duration on infection of oilseed rape leaves by conidia of the light leaf spot pathogen,Pyrenopeziza brassicae. Visible spore pustules developed on leaves of cv. Bristol inoculated withP. brassicaeconidia at temperatures from 4 to 20°C, but not at 24°C; spore pustules developed when the leaf wetness duration after inoculation was longer than or equal to approximately 6 h at 12–20°C, 10 h at 8°C, 16 h at 6°C or 24 h at 4°C. On leaves of cvs. Capricorn or Cobra, light leaf spot symptoms developed at 8 and 16°C when the leaf wetness duration after inoculation was greater than 3 or 24 h, respectively. The latent period (the time period from inoculation to first spore pustules) ofP. brassicaeon cv. Bristol was, on average, approximately 10 days at 16°C when leaf wetness duration was 24 h, and increased to approximately 12 days as temperature increased to 20°C and to 26 days as temperature decreased to 4°C. At 8°C, an increase in leaf wetness duration from 10 to 72 h decreased the latent period from approximately 25 to 16 days; at 6°C, an increase in leaf wetness duration from 16 to 72 h decreased the latent period from approximately 23 to 17 days. The numbers of conidia produced were greatest at 12–16°C, and decreased as temperature decreased to 8°C or increased to 20°C. At temperatures from 8 to 20°C, an increase in leaf wetness duration from 6 to 24 h increased the production of conidia. There were linear relationships between the number of conidia produced on a leaf and the proportion of the leaf area covered by ‘lesions’ (both log10‐transformed) at different temperatures.

Evidence for a role of cutinase in pathogenicity of Pyrenopeziza brassicae on brassicas

Supernatants from apple cutin-induced cultures of the light leaf spot pathogen Pyrenopeziza brassicae were shown to contain a single esterase with a pI of approx. 4.4 and a MW of 21kDa. Analysis of the hydrolysis products derived from enzymatic cleavage of3H labelled cutin by TLC demonstrated that these supernatants had cutinolytic activity. Treatment with the serine esterase inhibitors ebelactones A and B resulted in 50% inhibition of theP. brassicae esterase activity at concentrations of 4 and 0.4μ gml−1, respectively. It is therefore concluded that following induction in vitro with apple cutin, P. brassicae produces a single extracellular esterase with cutinolytic activity.A putative cutinase gene was cloned by heterologous Southern hybridization of P. brassicae genomic DNA using the Botrytis cinerea cutinase gene as a probe. The ORF (706bp) is predicted to encode a polypeptide of 203 amino acids with a molecular weight of 20.379kDa and a pI of 4.38. This polypeptide shows high levels of homology with the cutinases of B. cinerea and of other fungi, and contains the conserved histidine, aspartic acid and serine catalytic triad. Expression of the putative cutinase gene was shown to correlate with induction of esterase activity in vitro.SEM analysis of Brassica napus leaves infected withP. brassicae confirmed that direct penetration of the host surface occurs with no evidence of appressorial formation. The ebelactones A (100μ gml−1) and B (10μ gml−1) had no effect on germination of P. brassicae conidia in vitro; however, a marked decrease in pathogenicity of P. brassicae was observed on B. napus plants treated with 100μ gml−1ebelactone A or 10μ gml−1ebelactone B. In these plants, failed penetration was observed and the number of acervular conidiomata was greatly reduced. These results suggest a functional role for fungal cutinolytic activity in pathogenicity of P. brassicae on B. napus.

Effects of stem canker (Leptosphaeria maculans) and light leaf spot (Pyrenopeziza brassicae) on yield of winter oilseed rape (Brassica napus) in southern England

The relationships between yield loss and incidence or severity of stem canker and light leaf spot in winter oilseed rape were analysed by correlation and regression analyses, using data from experiments at Rothamsted, England in 1992/93, 1994/95 and 1995/96. Growth stages (GS) 6,3/6,4 and 4,0/4,5 were identified as the critical points for relating percentage yield loss to stem canker and light leaf spot (on stems), respectively. Critical point (CP) and area under disease progress curve (AUDPC) models relating percentage yield loss to combined incidence or severity of stem canker and light leaf spot (stems) in each experiment were constructed by linear regression. There were no differences in the CP models for incidence between 1992/93, 1994/95 and 1995/96 experiments, or in the AUDPC models for incidence between 1992/93 and 1994/95 experiments. Therefore, a general CP model relating percentage yield loss (ΔY) to combined incidence of stem canker (Si) at GS 6,3/6,4 and light leaf spot (stems) (Li) at GS 4,0/4,5 was constructed using data from the three experiments: ΔY = 0·85 + 0·079Si + 0·065Li(R2 = 43·7%,P < 0·001, 92 df). A general AUDPC model relating ΔYto the AUDPC of combined incidence of stem canker (Sia) from GS 5·7 to GS 6·5 and light leaf spot (stems) (Lia) from GS 4·0 to GS 6·3 was constructed using data from the 1992/93 and 1994/95 experiments: ΔY = 0·07 + 0·00096Sia + 0·0026Lia(R2 = 43·6%,P < 0·001, 68 df). These two general yield‐loss models were tested with data from Rothamsted in 1993/94 and Boxworth in 1992/93. The predictive accuracy of the CP model based on combined incidence of stem canker and light leaf spot (stems) was better than that of the AUDPC model. Yield losses predicted by summing the estimates from individual models for incidence of stem canker alone (GS 6,3/6,4) and light leaf spot alone (on leaves at GS 3,3) were greater than observed yield losses in experiments at Rothamsted in 1992/93, 1993/94, 1994/95 and 1995/96 and at Boxworth in 1992/93.

Cloning and expression studies during vegetative and sexual development of Pbs1, a septin gene homologue from Pyrenopeziza brassicae

A septin gene homologue designated Pyrenopeziza brassicae septin 1 ( Pbs1) has been identified and cloned from the plant pathogenic fungus Pyrenopeziza brassicae and its expression analysed. Pbs1 is present in both mating types and in a single copy within each genome and is transcribed in proportionate levels during both vegetative and sexual growth.

Epidemiology in relation to methods for forecasting light leaf spot (Pyrenopeziza brassicae) severity on winter oilseed rape (Brassica napus) in the UK

Pyrenopeziza brassicae, cause of light leaf spot of oilseed rape, has a complex polycyclic life cycle. It can be difficult to control light leaf spot in winter oilseed rape in the UK since it is not easy to optimise fungicide application timing. Early autumn infections are usually symptomless and recognisable lesions do not develop until the epidemic has progressed further by the spring. Light leaf spot often has a patchy distribution in winter oilseed rape crops and estimation of disease incidence can be difficult. There is evidence that epidemics are initiated primarily by ascospores produced from apothecia that survive the summer inter-crop period on infected debris. Subsequent development of the epidemic during the winter and spring is maintained by rain-splashed conidia that spread light leaf spot from initial foci. Understanding the relative roles of ascospores and conidia in the light leaf spot life cycle is crucial for forecasting epidemic severity and developing control strategies. The current web-based regional forecast system provides an autumn forecast of the incidence of light leaf spot that can be expected the following spring. This is based on survey data which assesses the occurrence of disease the previous July, and weather factors, such as deviations from summer mean temperature and winter rainfall. The forecast can be updated throughout the autumn and winter and includes crop-specific elements so that growers can adjust risks by inputting information about cultivar, sowing date and fungicide use. Crop-specific forecasts can be confirmed by assessing the incidence of light leaf spot. Such assessments will become easier when immunodiagnostic methods for detection of the disease become available. Incorporation of information on spore biology (e.g. apothecial maturation, ascospore release and infection conditions) is considered as a component of the interactive, continuously updated, crop-specific, web-based forecasts which are needed in the future.

Winter oilseed rape with high levels of resistance to Pyrenopeziza brassicae derived from wild Brassica species

Oilseed rape cultivars possess inadequate levels of resistance to light leaf spot disease, caused by the ascomycete Pyrenopeziza brassicae Sutton & Rawlinson. High levels of resistance to this disease were found within wild accessions of Brassica oleracea and B. rapa. This resistance was introgressed into agronomically acceptable winter oilseed rape breeding lines. Seedling resistance was determined by two genes. One of these, derived from B. rapa and positioned on linkage group N1, resulted in no apparent symptoms following infection, while the other, derived from B. oleracea and positioned on N16, resulted in black necrotic flecks and a reduced amount of sporulation compared with standard cultivars. Several agronomically acceptable double haploid lines were developed which expressed very high levels of adult plant resistance.

Development of PCR based diagnostic techniques for the two mating types of Pyrenopeziza brassicae (light leaf spot) on winter oilseed rape (Brassica napus ssp. oleifera)

Two sets of PCR primers were designed from DNA sequence which flanked the 3′ end of thePyrenopeziza brassicae mating type idiomorphs (MAT-1 and MAT-2). The first set of primers facilitated the amplification of a 750 bp product from genomic DNA of 50 isolates of P. brassicae , which included both mating types. The 750 bp PCR product was also amplified from DNA extracted from Brassica napus tissue infected with P. brassicae . The 750 bp PCR product was not produced when these diagnostic primers were used in PCR reactions containing DNA from a number of other fungal species, including fungal pathogens of oilseed rape and Tapesia yallundae , which is closely related to P. brassicae . The second set of primers were designed to amplify different sized products from the two mating types of P. brassicae . These mating type specific primers facilitated the amplification of a 440 bp product from all MAT-1 isolates tested and a 535 bp product from 30% of the MAT-2 isolates tested. The value of molecular diagnostic techniques for the identification of P. brassicae in oilseed rape crops, for distinguishing between mating types of the fungus and in disease forecasting is discussed.

Cloning of the mating type loci from Pyrenopeziza brassicae reveals the presence of a novel mating type gene within a discomycete MAT 1‐2 locus encoding a putative metallothionein‐like protein

This article was published in Mol Microbiol (1998) 30: 799–806. In order to move the fungal mating type field towards a standard nomenclature, as suggested by Arie et al. (1997), the mating type designations for Pyrenopeziza brassicae have been reversed. The MAT 1-1 locus described by Singh and Ashby (1998), which was based on the genetic nomenclature proposed by Courtice and Ingram (1987), becomes MAT-2 (EMBL AJ006072). As a consequence the PHB1 gene, encoding the HMG protein, becomes PHB2 and vice versa. The MAT 1-2 locus described by Singh and Ashby (1998) becomes MAT-1 (EMBL AJ006073) and encodes three genes designated PAD1, PMT1 and PHB1.

Methods for assessment of light leaf spot (Pyrenopeziza brassicae) on winter oilseed rape (Brassica napus) in the UK

Summary.Light leaf spot (Pyrenopeziza brassicae) was assessed as % plants with light leaf spot, % leaves with light leaf spot or % leaf area with light leaf spot in winter oilseed rape field experiments done at different sites (Rothamsted, Hertfordshire; Boxworth, Cambridgeshire; near Aberdeen, Scotland), with different cultivars (e.g. Bristol and Capitol), different fungicide treatments, on plants sampled at different dates. Regression analyses on data from these experiments showed that there were consistently good relationships between % leaves with light leaf spot and % plants with light leaf spot for plants sampled during the autumn and winter, until the % plants with light leaf spot approached 100%. The slopes and positions of regression lines were sometimes affected by cultivar, fungicide treatment or sampling date, but not by site. The relationship between % leaf area with light leaf spot (square root‐transformed) and % leaves with light leaf spot was less consistent than that between % leaves with light leaf spot and % plants with light leaf spot and was sometimes affected by cultivar, fungicide treatment or sampling date but not by site. The relationship between % leaf area with light leaf spot (square root‐transformed) and % plants with light leaf spot was also inconsistent and was sometimes affected by cultivar, fungicide treatment, sampling date and site.

Diagnosis of light leaf spot (Pyrenopeziza brassicae) on winter oilseed rape (Brassica napus) in the UK

Summary.Light leaf spot lesions were generally first observed as light green areas on leaves of UK winter oilseed rape crops in January or February and later became brittle and bleached. Elongated lesions, which were brown with indistinct edges, developed on stems in the spring and summer, when lesions were also observed on flower buds, pedicels and pods. Development of diagnostic white pustules (spore masses of Pyrenopeziza brassicae, which erupt through surfaces of infected tissues) for confirmation of light leaf spot infection on symptomless plants or plants with indistinct or ambiguous symptoms in the autumn, winter or spring was enhanced by incubating plants in polyethylene bags. In experiments with artificially inoculated plants, glasshouse‐grown plants exposed in infected crops and plants sampled from crops, white pustules developed at all incubation temperatures from 2oC to 20oC on infected leaves of different cultivars. The period of incubation required before the appearance of pustules decreased as the time that had already elapsed since the initial infection increased. The longest periods of incubation were required at the lowest temperatures (2oC or 5oC) but leaves senesced and abscised from plants most quickly at the highest temperatures (15oC or 20oC), suggesting that the optimal incubation temperature was between 10oC and 15oC.

Identification of QTL involved in field resistance to light leaf spot (Pyrenopeziza brassicae) and blackleg resistance (Leptosphaeria maculans) in winter rapeseed (Brassica napus L.)

Quantitative trait loci (QTL), involved in the polygenic field resistance of rapeseed (Brassica napus L.) to light leaf spot disease, were mapped using 288 DNA markers on 152 doubled-haploid (DH) lines derived from the cross ‘Darmor-bzh’בYudal’. Over two years (1995 and 1996), the DH population was evaluated for light leaf spot resistance on leaves (L) and stems (S), and for blackleg disease resistance in same field trials. For the L resistance criterion, a total of five and seven QTL were detected in 1995 and in 1996 respectively, accounting for 53% and 57% of the genotypic variation. For the S criterion, three and five QTL were identified in 1995 and in 1996 respectively, explaining 29% and 43% of the genotypic variation. The locations of the QTL detected were quite consistent over the two years (4- and 2-year common QTL for L and S, respectively). Three genomic regions, located on the DY5, DY10 and DY11 groups, were common to the resistance on leaves and stems. In comparison with the QTL for blackleg resistance described by Pilet et al. (1998), two regions on the DY6 and DY10 groups, were associated with the two disease resistances. These ‘multiple disease resistance’ (‘MDR’) QTL may correspond to genes involved in common resistance mechanisms towards the two pathogens or else to clusters of resistance genes.

Comparison of cytokinin and cytokinin-O-glucoside cleaving β-glucosidase productionin vitrobyVenturia inaequalisand other phytopathogenic fungi with differing modes of nutritionin planta

Early symptoms of apple scab caused byVenturia inaequalisinclude leaf curl and green island formation, both of which are indicative of an imbalance of plant growth regulators. To test whetherV. inaequalishas the potential to contribute to this phenomenon, cytokinin productionin vitrowas analysed quantitatively by high performance liquid chromatography followed by enzyme linked immunosorbant assay. Both zeatin riboside and isopentenyl adenosine type cytokinins were detected in extracts from mycelia and culture filtrates. The highest amounts of cytokinins were recovered from mycelium grown in Murashige and Skoog glucose vitamins medium (MSGV). These were five-fold greater than those recovered from malt extract salts medium (MES).To ascertain whether cleavage of biologically inactive cytokinin glucosides into free cytokinins by fungal β-glucosidases contributed to this phenomenon, the production of cytokinin-glucoside cleaving β-glucosidasesin vitrobyV. inaequaliswas compared with a range of other phytopathogenic fungi, with differing modes of nutritionin planta. All of the fungi tested apart fromBotrytis cinerea, produced β-glucosidase which specifically cleaved the substrate, Zeatin-O-glucoside ((OG)Z). The levels of (OG)Z cleaving β-glucosidase from mycelia of the hemibiotrophsV. inaequalisandLeptosphaeria maculans, grown in MSGV were 12-fold and seven-fold higher, respectively than when grown in MES. In comparison, mycelia (OG)Z cleaving β-glucosidase activity in mycelium of the biotrophCladosporium fulvumgrown in MSGV was over four-fold higher than MES. Spore washes from the obligate biotrophErysiphe graminisshowed no detectable (OG)Z cleaving β-glucosidase activity. No significant difference in (OG)Z cleaving β-glucosidase activity was seen between mycelium grown in MES and MSGV for the hemibiotroph,Pyrenopeziza brassicae. For the necrotrophic fungi,Penicillium expansum, Monilinia fructicola, B. cinerea, Alternaria brassicaeandAlternaria brassicicolaeither no (OG)Z activity was detectable or no significant difference in levels of (OG)Z cleaving β-glucosidase activity were found between the nutrient rich and growth limiting media. These results are discussed in relation to factors that may contribute to the biotrophic nutrition of fungal phytopathogens.

Effects of light leaf spot (Pyrenopeziza brassicae) on yield of winter oilseed rape (Brassica napus)

SummaryThe relationship between development of light leaf spot and yield loss in winter oilseed rape was analysed, initially using data from three experiments at sites near Aberdeen in Scotland in the seasons 1991/92, 1992/93 and 1993/94, respectively. Over the three seasons, single‐point models relating yield to light leaf spot incidence (% plants with leaves with light leaf spot) at GS 3.3 (flower buds visible) generally accounted for more of the variance than single‐point models at earlier or later growth stages. Only in 1992/93, when a severe light leaf spot epidemic developed on leaves early in the season, did the single‐point model for disease severity on leaves at GS 3.5/4.0 account for more of the variance than that for disease incidence at GS 3.3. In 1991/92 and 1992/3, when reasonably severe epidemics developed on stems, the single‐point model for light leaf spot incidence (stems) at GS 6.3 accounted for as much of the variance. Two‐point (disease severity at GS 3.3 and GS 4.0) and AUDPC models (disease incidence/severity) accounted for more of the variance than the single‐point model based on disease incidence at GS 3.3 in 1992/93 but not in the other two seasons. Therefore, a simple model using the light leaf spot incidence at GS 3.3 (x) as the explanatory variable was selected as a predictive model to estimate % yield loss (yr): yr= 0.32x– 0.57. This model fitted all three data sets from Scotland, When data sets from Rothamsted, Rosemaund and Thurloxton in England were used to test it, this single‐point predictive model generally fitted the data well, except when yield loss was clearly not related to occurrence of light leaf spot. However, the regression lines relating observed yield loss to light leaf spot incidence at GS 3.3 often had smaller slopes than the line produce, by the model based on Scottish data.

Genetic variation among field isolates of Pyrenopeziza brassicae

Amplified fragment length polymorphism (AFLP) markers were used to determine genetic diversity and population structure of Pyrenopeziza brassicae, the causal agent of light leaf spot of Brassica spp. Fungal isolates were sampled from six regions in the UK, one region in Germany and one region in France. A high level of genetic diversity was found (HT = 58%), with most variation attributed to within regions (HS = 43%), which suggests that sexual reproduction is frequent. FST values suggested significant population differentiation between England and the continent, but not between Scotland and England and Scotland and the continent. Overall, a moderate but significant level of regional differentiation was found (FST = 16 ± 4.0). There was no correlation between FST values and distance, indicating that long‐distance dispersal by natural factors does not occur at high frequencies. However, the lack of differentiation among populations from Aberdeen, Winchester and Cambridge suggests that seed transmission or other artificial methods of dispersal may be important.

Transport of Glutamate and Glutamine in the Light Leaf Spot Pathogen Pyrenopeziza brassicae

AbstractThe kinetics of glutamic acid and glutamine uptake in the light leaf spot fungus, Pyrenopeziza brassicae, are biphasic. At low and high concentrations, glutamic acid and glutamine share a high‐affinity and a low‐affinity carrier, respectively, with Kms of 4.0 and 4.4 μm for uptake of glutamic acid and glutamine, respectively, by the high‐affinity system, and Kms of 580 and 560 μm for uptake of glutamic acid and glutamine by the low‐affinity system. The data suggest that glutamic acid and glutamine are taken up by a different system to that responsible for the uptake of ornithine, arginine, lysine and asparagine, and may represent system IV described in Neurospora crassa for the uptake of acidic amino acids.

Sugar transport in the light leaf spot pathogen Pyrenopeziza brassicae

In the light leaf spot fungus Pyrenopeziza brassicae, the kinetics of uptake of sucrose, glucose and fructose are all biphasic. At low and high concentrations, glucose and fructose share a high-affinity and a low-affinity hexose carrier respectively, with K m s of 3.5 and 4.6 μM for uptake of glucose and fructose respectively by the high-affinity system, and K s s of 690 and 750 uM for uptake of glucose and fructose by the low-affinity system. The data also suggest the existence of separate high-affinity and low-affinity uptake systems for sucrose. P. brassicae possesses both soluble and paniculate invertase activity, with pH optima of 5.0 and 4.0 respectively. Activity of the particulate invertase is considerably in excess of the highest rates of sucrose uptake.

Sugar transport in the light leaf spot pathogen Pyrenopeziza brassicae

In the light leaf spot fungus Pyrenopeziza brassicae, the kinetics of uptake of sucrose, glucose and fructose are all biphasic. At low and high concentrations, glucose and fructose share a high-affinity and a low-affinity hexose carrier respectively, with Kms of 3.5 and 4.6 μM for uptake of glucose and fructose respectively by the high-affinity system, and Kss of 690 and 750 uM for uptake of glucose and fructose by the low-affinity system. The data also suggest the existence of separate high-affinity and low-affinity uptake systems for sucrose. P. brassicae possesses both soluble and paniculate invertase activity, with pH optima of 5.0 and 4.0 respectively. Activity of the particulate invertase is considerably in excess of the highest rates of sucrose uptake.

The use of AFLP fingerprinting for the detection of genetic variation in fungi

A new PCR-based technique for the detection of inter- and intraspecific genetic variation has been tested on isolates of the fungal phytopathogens Cladosporium fulvum and Pyrenopeziza brassicae . The method is based on the selective PCR amplification of restriction fragments from digests of genomic DNA. We show that the technique is very efficient at detecting polymorphisms, even in species where very little variation could previously be found by RFLP analysis. 21 primer combinations were used on four isolates of P. brassicae , detecting a total of 162 polymorphisms (mean = 4·1 polymorphisms per primer combination per pair of isolates). Four primer combinations were used on eight isolates of C. fulvum , detecting a total of 32 polymorphisms (mean = 3·3 polymorphisms per primer combination per pair of isolates). Primer combinations varied in their ability to detect variation, ranging from 0 to 24 polymorphisms between P. brassicae isolates and 0 to 10 polymorphisms between C. fulvum isolates. AFLP fingerprints were highly reproducible and have great potential as a tool for evaluating genetic diversity of fungal pathogens.

The incorporation of pathogen spores into rain‐splash droplets: a modelling approach

Simple, theoretical, physical principles and existing experimental data were used to derive an analytical model to describe the incorporation of plant pathogen spores into splash droplets. Data were obtained from experiments on splash dispersal of spores of Pseudocercosporella herpotrichoides (cereal eyespot), Pyrenopeziza brassicae (oilseed rape light leaf spot) and Septoria nodorum (wheat glume blotch). In these experiments, incident drops of diameter 4–5 mm were allowed to fall onto spore suspensions 0.5 mm deep with 1.2 × 105 to 6.5 × 105 spores/mL. The analytical model was constructed as the product of three functions of droplet diameter which described, respectively, the frequency distribution of droplet diameters, the proportion of droplets carrying spores and the mean number of spores in spore‐carrying droplets in each diameter category. The frequency distribution of droplet sizes was described by a log‐normal distribution, the proportion of droplets carrying spores was described by an exponential function and the adimensional spore concentration in spore‐carrying droplets was described by a power law. The cumulative proportions of spores in droplets in diameter categories of increasing diameter were calculated to compare observed and fitted data.