SEED HEALTH OF COMMON BEAN STORED AT CONSTANT MOISTURE AND TEMPERATURE

Fungal incidence in stored common bean (Phaseolus vulgaris L.) is the main concern in order to preserve seed health and viability. The main aim of this study was to analyse these quality parameters in hermetically stored seeds at 10.2, 13.1, 16.2, 18.5% moisture content (MC) and 25, 30, 35, 40°C, through seed germination and health tests. Water activity recorded at 10.2 and 18.5% MC were 0.448 and 0.700, respectively. Low seed moisture content reduced Alternaria spp. incidence at 2530°C. Highest incidence of Fusarium spp. (7.5%) occurred at 16.2% MC and 35-40°C. Highest incidences of Rhizoctonia spp. (8-10%) were recorded at 16.2-18.5% MC and 30-40°C. Penicillium spp. and Aspergillus spp. were predominant throughout the experiment and the highest incidences (80-100%; 20-30%, respectively) were scored at 18.5% MC and 30-35°C and 13.1-18.5% MC at 35°C, respectively. The higher the seed MC the higher the fungi incidence while lower seed MC decreased the incidences by 25%. Storage conditions below 30°C and 13.0% MC appear suitable to preserve common bean seed in relation to viability and health, up to a 8-month period.


INTRODUCTION
Brazil is one of the largest common bean producers in the world, therefore requiring high technology for the maintenance of seed quality parameters as to physical purity, germination and health percentages.Several factors may affect common bean seed conservation, mainly including seed health, moisture content (MC), temperature (T), relative humidity (RH) and the action of fungi and insects.High T and MC accelerate degenerative processes in biological systems, causing gradual, irreversible and accumulative losses in vigour and viability (Delouche & Baskin, 1973).Seeds present a lower respiration rate during storage than external and internal fungi (Lazzari, 1993).
Seeds are the vehicles for transmission of several fungi and frequently introduce new pathogens in exempt areas, so that the integration between seed health and germination tests is recommended to control seed transmitted diseases (Zaumeyer & Thomas, 1957;Singh & Mathur, 1974;Bolkan et al., 1976.;Neergaard, 1977).However, most of the research has Field fungi activity is delayed during storage at low seed MC since they require ≥ 90% air RH for growth (Lazzari, 1993).Fast development and high aggressiveness of these pathogens could kill the seed after sowing due to the action of powerful enzymes and toxins.On the other hand, storage fungi usually develop in seeds in equilibrium at 65-90% air RH (around 12-13% MC) (Loewer et al., 1994).A decrease in field fungi and an increase in storage fungi populations occur after harvest, in an ecological succession.The main objective of this research was to analyse seed germination and fungi incidences in common bean under controlled storage conditions to define an optimum MC, storage period and T required to maintain high seed health and viability.

MATERIAL AND METHODS
Seven kg of common bean seeds cv.IAC-Carioca ETE, were harvested in the 1998-99 season, in Campinas, São Paulo State, Brazil.Seed MC was adjusted at 25°C to 10.2, 13.1, 16.2 and 18.5% MC, from an initial value of 15.1%, either by rehydration over water in a closed plastic box or by dehydration over silica gel, aiming to avoid possible damage to the seeds caused by fast dehydration / rehydration.Seeds were sealed in laminated aluminium-foil packets (polyester structure / aluminium / low-density polyethylene, with a total thickness of 120 mm) and stored in incubators maintained at 25, 30, 35 and 40°C (± 0.5°C).
Seed MC (fresh weight basis) was determined in three 5 g ground seed samples at 130-133°C for 2 h (ISTA, 2004).Water activity (A w ) was determined using three seed samples for each MC in a hygrometer using the dew point technique, at 25°C ± 0.3 (± 0.01A w ).Germination tests were performed at 25°C using 4 × 50 seed replicates for each MC / temperature / sampling date combination, placed in rolled paper towels moistened with deionised water, with initial and final seedling counts recorded at the 5th and 9th days, respectively (ISTA, 2004).
Fungi incidences were determined by the blotter test (Neergaard, 1977), using 200 seeds (20 × 10 replicates) for each MC / temperature / sampling date combination, incubated at 20°C during seven days under 12-h alternating cycles of NUV-light (320-400 nm) and darkness, followed by evaluation under a stereoscopic microscope.Seeds were placed in plastic Petri dishes (9 cm diameter), with three filter papers moistened with sterilised water and previously decontaminated in a 1% sodium hypochlorite solution for 5 minutes (Berjark, 1984;Usberti & Amaral, 1999).Seed fungi incidences were estimated through observations of their structures (Barnett & Hunter, 1972).
Sampling intervals for seed health tests were quite variable due to the different levels of deterioration in relation to MC and storage T (Table 1).Preliminary results revealed no significant differences among initial and sampling time MCs.Fungi incidences for each combination among storage period, T and MC were compared using Fisher's LSD test (p < 0.05).Prior to statistical analyses, germination percentages and fungi incidences were transformed into arcsine √%/100 and √(x+1), respectively.

Water activity
A w is the quotient of seed vapour pressure over pure water vapour pressure at the same temperature and is an important parameter in storage studies since it is closely related to rate and intensity of common bean seed deterioration (Sartori, 1996).The A w values recorded on each seed MC (45-70% RH) were 0.448,

Statistical analyses
Statistical analyses of fungi incidences on common bean seeds are presented in Table 2, for each com-bination of storage period, T and MC.No statistical interaction were recorded among fungi incidences, storage T and seed MC.Penicillium spp.and Aspergillus spp.revealed the highest incidences among fungi throughout the experiment (Figures 1 and 2).Regression lines of fungi incidences were observed in shorter Storage period (days)    storage periods according to increases on seed MC and storage T.
The determination coefficients estimated for storage fungi incidences (Figure 1), ranging from 0.08 to 1, revealed a decreasing tendency, according to increases on MC and storage T.However, the values recorded for field fungi incidences (Figure 2) were quite variable, ranging from zero to 1, without showing a specific tendency.

Storage fungi incidences
Penicillium spp.and Aspergillus spp.are the main storage fungi in common bean and usually invade the seeds during and after maturation, causing damage as soon as they find appropriate conditions.The primary coloniser is Aspergillus spp., which subsequently allows the development of Penicillium spp.(Faiad et al., 1996).Penicillium spp.and Aspergillus spp.incidences scored in common bean seeds stored at 25, 30, 35, 40°C and 10.2, 13.1, 16.2, 18.5% MC are presented in Figure 1.
Penicillium spp.-Penicillium spp.had the highest incidences in common bean seeds, mainly at 18.5% MC and 30-40°C (80-100%) while still higher values (around 60%) were recorded at 25°C and 16.2-18.5%MC.The higher the storage T, the higher the fungus invasion during early storage periods.Lowest seed MC (10.2-13.1%)reduced fungus incidences; however, at 30-35°C, the values reached 10%, while at 40°C some values were higher than 20%.The best T range for fungus invasion was 30-35°C and MC higher than 16%.
Highest incidences of Penicillium spp.were observed at 18.5% MC at all T, while Aspergillus spp.contamination was only pronounced at 35°C; so, the higher the seed MC, the higher the fungal incidences at early storage periods.Additional high T effects on fungi incidences could also be noted.Penicillium spp.and Aspergillus spp.presented the highest incidences in common bean seeds during hermetic storage.These results agree with Terveit (1945); Wilcox et al. (1974); Bolkan et al. (1976); Dhingra & Sinclair (1978) and Hernandez et al. (1994).

Seed germination
Increases in MC reduced seed viability and this effect was more pronounced for highest seed MC (16.2, 18.5%), unrelated to storage T (Figure 3).The determination coefficients calculated for germination percentages were quite high, ranging from 0.49 to 0.94, without showing a specific tendency according to MC and storage T.
Some germination percentage reductions observed throughout the storage period might be influenced by previous high incidences of Penicillium spp.(60.5% at 130-day; 36.5% at 132-day) (Figure 1).Moreover, some early high incidences of Aspergillus spp. at 40 to 60-day storage could play an additional role in the deterioration process.
Similar results were reported by Chisholm & Coates (1997), evaluating germination percentages and fungi incidences in three leguminous seeds during storage with subsequent germination reduction and increase in fungi incidences mainly at 28°C.Stored beans (10.3-14.2%MC at 30°C) with high initial seed germination percentage and MC around 11.5% might maintain viability for eight months (Aguirre & Peske, 1991).Sanhewe & Ellis (1996) have also reported that bean seed quality was higher at cold temperatures during development and maturation.

CONCLUSIONS
Storage conditions with moisture content and temperature lower than 13.1% and 30°C, respectively, appear to be adequate for maintenance of seed viability and healthiness up to 8-months storage.Such storage conditions, which might be easily reached by seed sun drying and open storage, suggest a closing remark as to potential benefits and offer seed producers with a strategy for maintaining seed viability.
in open-stored common bean, not taking into account controlled environments.