Correlation analysis of field spore dynamics, disease severity, and major environmental factors for tobacco brown spot

Tobacco brown spot, caused by Alternaria spp., reduces tobacco yield and quality. Monitoring airborne spores is essential for early warning, but the vertical distribution of Alternaria spores within tobacco canopies and its relationship with meteorological factors remain poorly understood. This study aimed to determine the optimal sampling height for Alternaria spores in tobacco fields and to analyze height-dependent correlations with environmental variables. Field experiments were conducted in Fuquan City, Guizhou Province, China, in 2025 using susceptible (Yunyan 87) and moderately resistant (Guiyan 20) tobacco cultivars. Airborne spores were collected using the slide method at three heights (0.5, 1.0, and 1.5 m) and two locations (field center and edge). Meteorological variables, including air temperature, relative humidity, soil temperature, wind speed, total rainfall, and light intensity, were recorded. Relative proportions of spores at each height were calculated, and Pearson and lagged correlation analyses were performed. Spore counts were significantly higher in Yunyan 87 than in Guiyan 20, and higher at the field center than at the edge for both cultivars. The 1.0 m height captured the highest relative proportion of spores (51.8%), followed by 1.5 m (25.3%) and 0.5 m (22.9%). Spore abundance was negatively correlated with air temperature, wind speed, and light intensity, and positively correlated with relative humidity and rainfall. Rainfall showed the strongest positive correlation at a 2-day lag (r = 0.49–0.52). Spores collected at 1.5 m exhibited stronger correlations with wind speed, rainfall, and light intensity than those at lower heights. The middle canopy height (1.0 m) is recommended for spore trap placement at the field center. The height-dependent correlations suggest that meteorological factors exert greater influence on spore dispersal to higher altitudes than on local sporulation. These findings improve understanding of spore spatial distribution and provide a foundation for predictive models of tobacco brown spot. Limitations include the 17-day single-season sampling and the passive slide method; future studies with longer periods and active traps are needed.

Tobacco (Nicotiana tabacum L.) is an important economic crop. In China, the annual planting area is about 1 million hectares, and both production and sales account for about one-third of the world’s total (Sun, 2016). However, tobacco brown spot is an important disease affecting tobacco growth. Initially, small, round, yellowish-brown lesions appear on the leaves. These lesions gradually expand into larger circular spots, ultimately causing a significant reduction in both tobacco yield and quality (Jing et al., 2018). It mainly occurs in the middle and late stages of tobacco development and is characterized by rapid outbreak and fast spread, which damages the quality of tobacco leaves and severely reduces their economic value (Ye, 2010). The disease affects an area of about 100,000 hectares each year, causing economic losses of approximately 100 million yuan (Zhang et al., 2011). To reduce the resulting economic losses, precise detection and continuous prevention and control of tobacco brown spot have become important research directions.

Tobacco brown spot is caused by a fungal pathogen, specifically Alternaria, which thrives at temperatures between 5-38°C, with an optimal range of 25-30°C. Its spores and mycelia can survive for several months when frozen or at -10°C, and can survive for 2 days at 46°C. It can grow in environments with a pH of 4-11, with optimal mycelial growth at pH 6-8 and optimal spore germination at pH 6-7. At temperatures of 24-27°C, over 90% of spores can germinate in a droplet within 5 hours, and germination begins after 12 hours at a relative humidity of 98% (Li et al., 2023; Wang et al., 2023). Tobacco brown spot has a short incubation period and spreads rapidly, causing large-scale crop damage within a short time. It typically occurs during the late growth stage of tobacco leaves, significantly reducing both yield and quality, with yield losses reaching as high as 90% (Zhang et al., 2023). The prevention and control of tobacco brown spot has long been a major challenge in tobacco production. Conventional chemical control methods are associated with several issues, including fungicide resistance and environmental pollution (Zhang et al., 2023), and often fail to achieve satisfactory results. Therefore, effective management strategies for tobacco brown spot remain lacking (Zhang et al., 2023). The life cycle of Alternaria alternata is primarily asexual (anamorphic), though molecular evidence suggests cryptic recombination (Meng et al., 2015). It involves distinct phases of overwintering, germination, infection, colonization, and sporulation. Because these conidia are readily dispersed by wind, monitoring their aerial concentration is essential for disease forecasting, and the simple slide method also known as passive spore trapping, was among the earliest techniques developed for this purpose.

Using the slide method to collect spores is the earliest method for spore sampling. It involves placing a slide coated with a sticky substance (such as Vaseline) horizontally, allowing spores to settle on it by gravity. The slide is then taken back to the laboratory for microscopic examination and counting. Nevertheless, for multi-site, large-scale continuous monitoring, especially for grassroots agricultural plant protection departments, the simple slide capture method is convenient and practical (Cao, 2012). For monitoring purposes, this approach has proven effective, especially in remote areas where the availability of electricity can be problematic. Such devices continue to be used for monitoring plant pathogenic spores—for example, a trap consisting of Vaseline-coated microscope slides fixed at a 45° angle inside a short section of plastic drainpipe has been deployed to monitor coffee rust and sugarcane rust in Mexico (pers. com. Carmen Calderón Ezquerro, UNAM, Mexico). In this design, the drainpipe serves as a rain shield, and the entire unit is mounted on bearings and fitted with a wind vane so that it can freely orient into the wind (West and Kimber, 2015). Similar devices have also been used in the United States to monitor various rust fungi, such as soybean rust (von Qualen and Yang, 2006; Colwell, 2024). The placement height of the slides has a significant impact on capture efficiency. Studies using the slide method to capture rice blast fungus spores in the field have found that the number of spores captured in the air first increases with the elevation of the slide placement and then rapidly decreases (Cao et al., 2004). The interpretation of spore counts obtained by such passive traps, however, is strongly influenced by weather conditions, as temperature, humidity and other environmental variables jointly determine spore production, release and atmospheric transport. Understanding these relationships is therefore essential for translating monitoring data into reliable disease forecasts.

Several studies have emphasized the role of meteorological variables, particularly temperature and humidity, alongside environmental factors such as air pollutants, in shaping the concentration and dispersal of Alternaria spores (Grinn-Gofroń et al., 2019; Picornell et al., 2022; Skjøth et al., 2016). These factors jointly influence sporulation, spore viability, and atmospheric transport. Temperature affects both the growth rate of Alternaria and the timing of spore release, with warmer conditions generally promoting higher spore production (Grewling et al., 2019; Grinn-Gofroń et al., 2019). Humidity, in turn, is critical for spore germination and survival (Attri et al., 2024). Given the complex interactions among these variables, analyzing the correlation between meteorological factors and airborne Alternaria spore abundance is essential for understanding disease dynamics and improving forecasting accuracy.

In current field monitoring practices, spore traps have begun to be widely used to collect spores. France is developing various air sampling methods and continuing to improve spore traps in the field of plant biosecurity. The Coriolis δ air sampler, manufactured by Bertin Technologies (France), is a continuous air sampling device based on patented Coriolis technology that collects liquid samples. This enables a range of rapid detection methods, including PCR and serological analysis (Carvalho et al., 2008). Meanwhile, scientists in Australia have identified several issues related to the use of air samplers and have proposed a series of improvements accordingly (Jackson and Bayliss, 2011). A comprehensive review of air sampling methods, including the applications and limitations of rotating-arm impaction samplers, has also been provided in the ‘It’s a Trap!’ series (Check et al., 2024).

Several studies have investigated the effect of sampling height on airborne spore collection (Aylor and Taylor, 1983; Khattab and Levetin, 2008; West et al., 2008; Sosnowski et al., 2024; Heger et al., 2026). However, the vertical distribution pattern of Alternaria spp. spores within tobacco canopies and its relationship with meteorological factors remain less explored. This study therefore aimed to determine the optimal sampling height for Alternaria spores in tobacco fields and to analyze the height-dependent correlations with environmental variables. This provides theoretical guidance for related spore-trapping instruments and data support for achieving accurate detection of tobacco brown spot.

Sampling sites and sampling strategy

The experimental site was located in the tobacco commercial fields at the Experimental Farm of the Guizhou Academy of Tobacco Science in Fuquan City (26.74° N, 107.49° E), Guizhou Province. Prior to transplanting, a commercial fertilizer (10:10:25 = N: P 2 O 5: K 2 O) was applied to the soil at a rate of 0.075 kg/m². Four weeks after transplanting, an additional nitrogen fertilizer (containing 20% total nitrogen) was applied at 0.022 kg/m². Tobacco seedlings were raised in 160-cell floating trays and transplanted in mid-April 2025 as single rows with a row spacing of 1.0 m and a plant spacing of 0.6 m (Wang et al., 2016). Two tobacco varieties, Guiyan 20 and Yunyan 87, were selected for the study. At the start of the spore-sampling period (117 days after transplantation), disease severity was assessed visually following the Chinese national standard GB/T 23222—2008 (Grade and investigation method of tobacco diseases and insect pests), which classifies disease severity into six grades (0, 1, 3, 5, 7, 9) based on the proportion of leaf area affected by typical brown spot lesions. Based on these preliminary observations, Yunyan 87 was predominantly rated as grade 5 (severe), whereas Guiyan 20 was predominantly rated as grade 3 (moderate). In each field, two sets of sampling points were established, in which one was in the center of tobcco field and the other was at the edge. Spores were collected at each sampling point using the slide method, with sampling heights set at 0.5 m, 1.0 m, and 1.5 m above the ground. At the time of spore sampling (117 days after transplantation), the plant heights of both cultivars were approximately 1.6–1.8 m, with both at the mature growth stage. Thus, 0.5 m corresponded to the lower canopy near the ground, 1.0 m to the middle canopy where the largest leaf area is concentrated, and 1.5 m to the upper canopy or just above the canopy surface. Microscope slides were mounted vertically on supporting poles at each sampling height. The Vaseline-coated surfaces at the central sampling points faced north, whereas those at the edge sampling points faced west (i.e., towards the interior of the tobacco field). All slides maintained the same orientation throughout the sampling period at each respective location. Although wind direction varied during the experimental period, the prevailing wind was northwesterly. Consequently, both the north-facing (center) and west-facing (edge) surfaces were intermittently exposed to the dominant airflow, albeit with different incident angles. To minimize the potential bias introduced by slide orientation and variable wind direction, we analyzed the data as relative proportions at each height per sampling point per day, rather than using absolute counts directly. Spore collection was conducted from August 11, 2025, to August 27, 2025. Tobacco brown spot is a disease that primarily occurs after topping and during the leaf harvesting and curing stage. Because monitoring airborne spore abundance during the early growth stage is difficult, sampling was initiated after topping when the brown spot epidemic begins to develop. However, leaf harvesting commenced 17 days after the start of sampling, and continued monitoring beyond this point would have substantially compromised data quality due to physical disturbance. Nevertheless, data collected during the early-to-mid epidemic phase can still provide meaningful guidance for disease management. The slides used to capture spores were replaced every 24 hours. The collected slides were covered with cover slips for subsequent counting.

Meteorological data collection

Field environmental meteorological data were provided by a multidimensional environmental meteorological monitoring instrument (Zhejiang Top Cloud Agriculture Technology Co., Ltd., Hangzhou China) deployed near the spore trap. This instrument recorded meteorological factors such as temperature, humidity, and rainfall in real time, and uploaded the data to the crop disease monitoring IoT platform (https://smart.topyn.cn/login/m4J68S/9JN4dN).

To assess the representativeness of the meteorological conditions during the sampling period (August 11–27, 2025), we compared the recorded data with the 10-year (2015–2024) historical climate records for Fuquan City for the same 17-day interval (August 11–27). During the 2025 sampling window, the mean air temperature was 27.2°C, the mean relative humidity was 82.0%, and the total rainfall was 117.7 mm. These values were generally consistent with the 10-year averages (2015–2024) for the same 17-day interval, which were 27.1°C, 80.0%, and 143.3 mm, respectively. Although the 2025 season received somewhat less rainfall than the historical average, the overall meteorological conditions during the sampling period were broadly representative of the typical August climate in this region.

Spore count

Spores of Alternaria spp. collected on glass slides were identified under a light microscope. No stain or mounting medium was applied; the slides were directly covered with coverslips for observation. Spores of Alternaria spp. collected on glass slides were identified under a light microscope based on morphological characteristics, including conidial size, septation, and shape. The small-spored, muriform Alternaria conidia observed in this study, characterized by their solitary occurrence, ovoid to obclavate shape, transverse and longitudinal septa, and variable presence or absence of a short beak, were assigned to Alternaria sect. Alternaria (Woudenberg et al., 2015). For each sampling height at each location, one microscope slide was exposed daily and replaced every 24 h over the 17-day sampling period, yielding 17 replicate slides per height per location (n = 17). Each slide was covered with two coverslips for observation, and each coverslip defined an observation area of 572 mm² (i.e., a total of two areas, each 572 mm²). The number of collected spores was counted daily for subsequent analysis.

Data analysis

All experimental data were recorded and processed using Microsoft Excel (2024, Microsoft Corp., Redmond, WA, USA). Statistical analysis were performed using SPSS (version 27.0, IBM Corp., Armonk, NY, USA). To compare spore capture efficiency among the three heights, we first calculated, for each sampling point on each day, the total number of spores collected across the three heights. For days where the total was greater than zero, we calculated the percentage of spores at each height relative to that daily total. These percentages were then averaged across all sampling points and all sampling days for each height. This relative proportion approach avoids overweighting data from locations with exceptionally high absolute spore counts. Pearson correlation coefficients were calculated to assess the relationships between the number of Alternaria spp. collected at different heights and various meteorological factors.

To assess delayed effects, lagged Pearson correlation analyses were performed for rainfall and relative humidity at lags 1, 2, and 3 days (i.e., spore counts correlated with meteorological values from 1, 2, and 3 days prior). All analyses were conducted in R version 4.5.2 (R Core Team, 2025).

Monitoring findings from mild and severe disease plots

Monitored using the passive slide method were airborne Alternaria spp. spores (Figure 1A). In the selected Yunyan 87 and Guiyan 20 tobacco fields, the number of Alternaria spp. spores collected at both the center and edge of Yunyan 87 was higher than that of Guiyan 20. (Figure 1B). For both cultivars, more spores were collected at the field center than at the edge. (Figure 1C). Recorded 117 days after seedling transplantation was the highest spore count for both cultivars. Specifically, for Yunyan 87, collected at the field edge were 2 spores mm - ², whereas at the field center, 8.72 spores mm - ² were collected. For Guiyan 20, 0.13 spores mm - ² were collected at the edge and 0.47 spores mm - ² at the center. Occurring 117 days after transplantation was the largest rainfall event during the entire spore collection period, suggesting a close relationship between rainfall and the release of Alternaria spp. spores.

Spore collection at three heights

To compare spore capture efficiency across heights while minimizing bias from location-specific differences in absolute spore abundance, we analyzed the relative proportion of spores at each height per sampling point per day. Across all valid sampling events, the mean relative proportion of spores was highest at 1.0 m (51.8%), followed by 1.5 m (25.3%) and 0.5 m (22.9) (Figure 2). This pattern indicates that the middle canopy level (1.0 m) consistently captured the largest share of airborne spores, regardless of the substantial differences in absolute spore counts among cultivars and field locations. Therefore, 1.0 m above ground is recommended as the optimal vertical layer for spore collector placement within the tobacco canopy under the conditions of this study.

Analysis of the correlation between spore populations and meteorological factors

The collected Alternaria spp. counts showed a negative correlation with air temperature, wind speed, and light intensity, and a positive correlation with air humidity and total rainfall, with a weaker correlation with soil temperature. Among them, the Alternaria spp. counts collected at 1.5 m showed a stronger correlation with wind speed, total rainfall, and light intensity than those at 1.0 m and 0.5 m (Table 1). Specifically, at 1.5 m, the correlation coefficient with wind speed was -0.136, compared to -0.089 at 0.5 m and -0.073 at 1.0 m. Also observed at 1.5 m was a higher correlation with total rainfall than that at 0.5 m or 1.0 m. Likewise, the negative correlation with light intensity was more pronounced at 1.5 m than at 0.5 m and 1.0 m. These findings suggested that spore populations at greater heights were more sensitive to meteorological fluctuations.

Lagged correlations with meteorological factors

Lagged Pearson correlation analysis was performed for all meteorological variables at lags 0 to 5 days (Table 2). Overall, among the five meteorological factors examined, rainfall exhibited the most pronounced delayed positive effect on airborne Alternaria spore counts, whereas relative humidity showed only weak contemporaneous correlations, and both wind speed and light intensity were negatively associated across all lags without significance.

Specifically, rainfall presented the strongest positive correlation with spore abundance at a lag of 2 days, with correlation coefficients ranging from 0.49 to 0.52 across the three sampling heights, suggesting a 48h delay between rain events and peak spore release (Figure 3). For lags 3 to 5 days, these correlations gradually declined. Regarding relative humidity, only weak contemporaneous correlations were observed, and no clear lag pattern was detected. As for wind speed and light intensity, consistently negative correlations with spore counts were found across all lags, albeit not statistically significant (|r| < 0.25, p > 0.40). These negative associations were consistent with physical dilution of spores by wind and light-inhibited sporulation.

As the pathogen of tobacco brown spot, the presence and quantity of Alternaria spp. spores are important indicators of the occurrence and severity of the disease (Cheng et al., 2011). Achieving accurate monitoring of the number of Alternaria spp. spores in the field environment can also enable precise prediction of the occurrence and development of tobacco brown spot. Therefore, accurately collecting spore data in the field is a very important condition. This study aimed to elucidate the spatial distribution pattern of Alternaria spp. spores in tobacco fields affected by brown spot. Specifically, it was investigated whether spore abundance was greater in the middle of the field compared to the field edges, and it was determined which of three vertical heights (0.5 m, 1.0 m, and 1.5 m) allowed for the collection of a higher number of spores. It should be noted that the 17-day sampling duration in this study, although covering the critical early-to-mid epidemic window after topping, represents a relatively limited dataset. It should also be noted that the 17-day sampling period represents a single season. Although the meteorological conditions during this period were broadly consistent with the 10-year averages for temperature and humidity, the total rainfall (117.7 mm) was somewhat lower than the 10-year average (143.3 mm). Interannual variability in weather patterns, particularly rainfall, may influence spore dispersal dynamics and the timing of peak spore release. Multi-year monitoring across multiple seasons would be valuable to validate the generalizability of our findings. For more robust and generalizable conclusions, future studies could extend the sampling period to encompass a longer portion of the epidemic cycle, thereby capturing a wider range of environmental conditions and spore release events.

It is important to clarify the valid comparisons that can be made from the spore count data presented in this study. The relative proportion analysis presented in Figure 2, which was calculated as the percentage of spores at each height relative to the daily total per sampling point, is not affected by slide orientation and therefore can be reliably used to evaluate spore capture efficiency across different heights. In contrast, the absolute spore count data used in Figures 1B, C, 3, and Tables 1, 2 require more cautious interpretation. Because the passive slides at the center and edge positions faced different orientations (north-facing at the center vs. west-facing at the edge), direct comparisons between the center and edge locations within each plot may be confounded by slide orientation and wind direction, rather than reflecting true differences in airborne spore concentration. However, comparisons between the two cultivars at the same location—i.e., Yunyan 87 center vs. Guiyan 20 center (both north-facing), and Yunyan 87 edge vs. Guiyan 20 edge (both west-facing)—remain valid, as the slide orientation was consistent within each location. These comparisons support the conclusion that spore abundance is positively associated with cultivar susceptibility. Comparison of the two selected tobacco cultivars, Yunyan 87 and Guiyan 20, revealed marked differences in disease susceptibility. Consistent with its higher susceptibility to tobacco brown spot, Yunyan 87 exhibited significantly greater spore counts than Guiyan 20 at both field center (8.72 vs. 0.47 spores mm - ²) and edge (2.00 vs. 0.13 spores mm - ²). This substantial difference indicates that cultivars with more severe disease support greater pathogen reproduction and release more airborne spores. The positive correlation between airborne Alternaria spp. spore load and field disease severity is therefore strongly supported. From a management perspective, monitoring spore concentrations could serve as a reliable indicator of local epidemic risk, enabling early intervention in susceptible cultivars. These findings highlight the value of cultivar-specific spore surveillance for precision disease forecasting.

The optimal sampler location and height depend primarily on the specific information required and should account for several key factors. These include the source characteristics and aerodynamic properties of the target bioaerosols, the surrounding vegetation or physical structures, the operational constraints of the sampler, and the meteorological conditions governing the release, dispersal, and deposition of fungal spores across vertical and horizontal gradients (Khattab and Levetin, 2008). The higher spore counts in the field center are explained by greater inoculum production from concentrated lesions, the attenuation of spore density along a distance-dependent dispersal gradient toward the edges, and microclimatic conditions characterized by reduced wind speed and increased canopy shading that facilitate spore retention centrally.

Fixed installations at ground level (0.5 - 1.5 m) are most commonly applied for fungal spore sampling, providing representative measurements of airborne spore concentrations from localised sources and supporting the determination of dispersal gradients (Lacey and Venette, 1995). To determine the optimal height for collecting Alternaria spores in tobacco fields, this study evaluated three sampling heights (0.5 m, 1.0 m, and 1.5 m). The relative proportion analysis identified 1.0 m as the most effective sampling height within the tobacco canopy, with an average of 52.5% of spores captured at this height across all sampling positions and dates. This finding is consistent with the vertical distribution pattern expected for spores released from foliar lesions within the canopy, where the middle height approximates the primary source layer and thus yields the highest collection efficiency.

The observed correlations between airborne Alternaria spp. spore abundance and meteorological factors are consistent with both the biological characteristics of the pathogen and the physical processes underlying spore dispersal. The positive associations with relative humidity and rainfall are supported by previous studies (Chávez et al., 2025), which reported that Alternaria spore concentrations correlate positively with humidity and with rainfall accumulated over the preceding three days. These patterns are reinforced by field observations indicating that extended dew periods and humid conditions favor spore germination and infection, with disease severity increasing under frequent wet-dry cycles. In contrast, the negative correlation with light intensity aligns with recent evidence showing that continuous darkness significantly enhances sporulation in Alternaria alternata (Sun et al., 2025; Deng et al., 2024; Li et al., 2025). With respect to temperature, although optimal mycelial growth occurs between 25 °C and 30 °C (Hatzipapas et al., 2002), more recent large-scale analysis suggest that higher air temperatures are associated with increased spore concentrations across most sites, with temperature exerting a stronger influence than land use (Plaza et al., 2025). The negative correlation with wind speed observed in this study can be explained by established aerobiological principles, whereby prevailing wind direction and speed govern spore transport and local concentration; higher wind velocities facilitate dispersal away from source areas, thereby reducing local collection efficiency (Apangu et al., 2023; Gregory, 1945; Rodríguez-Fernández et al., 2023). The two-day lagged positive correlation between rainfall and spore abundance aligns with the time required for conidial maturation and release under sustained high humidity. Timmer et al. (1998) showed that Alternaria conidia are produced abundantly when infected tissue remains near 100% RH for 24 h, and that rainfall events trigger massive conidial release. In tobacco, moisture is the dominant factor driving brown spot development, supporting our observation of a 48h window of increased airborne spores. Although our gravity slide method and short sampling window are limitations, recent machine-learning studies on potato (Meno Fariñas et al., 2022) demonstrated that integrating lagged meteorological predictors with spore data substantially improves forecasting accuracy. Collectively, these findings underscore the critical role of microclimatic conditions in shaping airborne spore dynamics and highlight the value of integrating meteorological data into predictive models for tobacco brown spot.

Furthermore, a distinct spatial distribution pattern was observed across both cultivars, with spore collections from the middle of the fields being significantly higher than those from the edges. However, the practical application of this finding is constrained by agricultural practices. The relatively large footprint of current spore-trapping equipment poses a risk of interfering with routine farming operations, particularly in mechanized production systems. Given the significantly reduced collection efficiency at the field edge, the development of more compact and less intrusive spore-trapping devices represents a critical avenue for future research to enable in-field monitoring without disrupting standard agronomic practices.

Alternaria spp. is one of the most abundant fungal spores in the air (Apangu et al., 2023), widely spread during the tobacco harvest season, experiencing several short disease cycles each season. During the disease cycles, the extent of conidial dispersal varies over time and space, and it is considered a ‘medium-high’ risk pathogen (Wang et al., 2016). Accurate monitoring of the airborne dispersal patterns of Alternaria spp. spores would enable tobacco growers to implement timely and targeted interventions, thereby mitigating the incidence and severity of tobacco brown spot and significantly reducing associated economic losses. Furthermore, a comprehensive understanding of the spatiotemporal dynamics of spore dispersal is essential for optimizing the design of monitoring instruments and enhancing the predictive accuracy of disease models. Addressing these knowledge gaps would alleviate a critical concern for tobacco production.

In this study, we investigated the spatial distribution of airborne Alternaria spp. spores in tobacco fields and their relationships with key meteorological factors using the simple and low-cost slide method. The results demonstrate that spore abundance is positively correlated with disease severity, with the susceptible cultivar Yunyan 87 showing significantly higher spore counts than the medium-resistant cultivar Guiyan 20. Spore numbers were consistently greater at the field center than at the edge, and under our experimental conditions, 1.0 m above ground was identified as the optimal sampling height, though this may require adjustment for different cultivars or growth stages.

Meteorological analysis revealed that spore abundance was negatively correlated with air temperature, wind speed, and light intensity, and positively correlated with humidity and rainfall. Notably, rainfall exhibited a two-day lagged positive effect on spore counts, suggesting a 48-h window between rain events and peak spore release. Spores collected at 1.5 m height showed stronger correlations with wind, rainfall, and light than those at lower heights, indicating height-dependent microclimatic influences.

These findings provide practical guidance for field monitoring of tobacco brown spot: a spore trap placed at 1.0 m in the field center is recommended for optimal collec-tion efficiency. The observed relationships between spore dynamics and meteorological variables lay a foundation for developing predictive models for disease forecasting.

Given the 17-day sampling duration and the use of a passive gravity slide method, the results should be considered as an exploratory rather than definitive characterization of spore dispersal. Future studies with longer sampling periods, active trapping devices, and integration of molecular detection methods are needed to validate and extend these conclusions. Nonetheless, this study demonstrates that even low-cost, low-technology approaches can yield valuable insights into the epidemiology of tobacco brown spot and support practical disease management decisions.