SEGUNDO MÉTODO: PRECIPITACIÓN CONTROLADA

For Penicillium, a different effect was observed in both experiments. Penicillium CFU numbers were consistently higher in the onion residue treatment than in the other treatments (up to 20 times in the second experiment) and the second hypothesis Ho’’ was also rejected. This finding suggests that Penicillium could be promoted by the onion residue. This promotion is not simply due to the addition of OM in the soil because in the oat treatment levels were not as high despite a larger incorporation of OM. It is likely that the decomposition of the onion residue encouraged the proliferation of Penicillium colonies in the soil. To the best of the author’s knowledge, no research has been done on the impact of dry onion scale residues and Penicillium

as several Penicillium species such as P. albocoremium Frisvad (Frisvad), P. allii

Vincent & Pitt and P. digitatum (Pers.) Sacc. are responsible for post-harvest storage rots of onion (Overy et al., 2005; Raju & Naik, 2006) and P. glabrum which was isolated in this study has also been reported as an onion storage pathogen (Samson

et al., 2002). The results obtained in these two experiments suggest that, the dry onion scales residues left behind after the harvest of onion could promote Penicillium

growth. This is of critical importance for onion growers as Penicillium strains responsible for “storage rots” come from the field. The onion bulbs are contaminated when they are left to dry on the ground just before harvest. This can cause large economical loss to onion growers. Therefore, continuous onion rotation systems could lead to the build up of a large Penicillium inoculum and findings from this research support the concept of rotation for the onion cropping system as it will help to maintain low Penicillium inoculum levels in the field.

A regression analysis also showed that when Penicillium CFU numbers were high,

Trichoderma CFU numbers were reduced. This analysis confirms the results obtained in the first pot trial and is in accordance with previous research (Eastburn & Butler, 1988a; Eastburn & Butler, 1988b) and therefore, the thrid hypothesis ho’’’ was rejected. These authors looked at different abiotic and biotic factors affecting the population density and distribution of indigenous T. harzianum in an alfalfa (Medicago sativa L.) field at the microsite level. They looked at the effect of several fungal genera including Aspergillus, Fusarium, and Penicillium. They found that

Penicillium miczynskii Zaleski was negatively associated with the presence of T. harzianum in the soil. They did not investigate the effect of other Penicillium species on other species of Trichoderma but it is reasonable to think that similar negative associations were taking place in the onion treatment of both experiments. This could be explained either by direct suppression of Trichoderma growth by the release of inhibitory molecules during the decomposition of the onion scales residues, by indirect competition through the promotion of Penicillium growth or by amensalism where Penicillium is restricting Trichoderma growth in the rhizosphere soil or possibly on the TSM-LU plates when Penicillium CFU numbers are very high. Penicillium

species have long been known for their ability to release inhibitory secondary metabolites such as antibiotics to suppress the growth of organisms in their vicinity

(Nicoletti et al., 2007). The mode of action resulting in Trichoderma suppression is investigated further in the next chapter.

3.4.3. Trichoderma species diversity

The diversity results in both experiments did not differ from the long term field experiment (Chapter Two) as the same five Trichoderma species were recovered in this experiment. The overall Trichoderma species proportions in the rhizosphere of onion plant were variable between treatments and at different sampling times however, in the first experiment three species T. hamatum (39%), T. harzianum

(27%) T. koningii (25%) dominated the Trichoderma population representing 91% of the total population. However, in the second experiment, only T. hamatum and T. koningii were the dominant species (79%) while the remaining species T. atroviride,

T. asperellum and T. harzianum were in relatively equal proportions. Overall, the results obtained in this study were in accordance with what was previously observed in the long term field experiment and supports the idea that these five species could coexist in the rhizosphere and competition between them might be very limited since proportions seem to be relatively stable. However, they may occupy different niches along the root, which was not investigated in the present study as the entire root system was combined. This aspect is investigated further in Chapter Five. Interestingly, the onion residue treatment in the second experiment was mainly dominated by two species (T. hamatum 70% and T. koningii 20%) and had the lowest diversity with only three species present.

In addition, a treatment effect was detected in the first experiment for the presence frequency of T. harzianum but not for the other four species so the fourth hypothesis Ho’’’’ was only rejected for T, harzianum. From the presence frequency data for T. harzianum there was some evidence that onion residues could inhibit its presence in the soil. The lower occurrence of T. harzianum in the onion residue treatments of both experiments could be caused by several factors including the release of inhibitory compounds from the decomposition of onion scales, competition from the high Penicillium population and competition for the same nutrients or the release by

Penicillium CFU of secondary metabolites such as antibiotics that would inhibit this species in particular. These possible factors are investigated in the next chapter.

Comparison in species diversity between the rhizosphere and the rhizoplane soil in the first experiment revealed that diversity was lower in the rhizoplane soil as only four species were recovered instead of five as with the rhizosphere soil. Trichoderma atroviride was never recovered in the rhizoplane soil indicating that T. atroviride

might favour the rhizosphere soil where competition from soil microorganisms such as rhizobacteria would be less intense. Interestingly, the proportion of T. asperellum

in the rhizoplane soil was the second largest behind T. hamatum. Therefore, T. asperellum might behave in the opposite way to T. atroviride by favouring the rhizoplane soil. This observation could indicate that onion could exert a selective pressure on the colonisation location of Trichoderma species. This would be in accordance with Kurakov and Kostina (2001) study which reported that colonisation of plant root surface by Trichoderma species was species-specific. This requires further research to establish if Trichoderma species favour the colonisation of the onion root surface. If active selection does take place, the success of the application of Trichoderma in situ as a biological control agent could be increased by selecting species that the onion plant will encourage.