Studies on arbuscular mycorrhiza (AM) in the Alentejo (Portugal) using pea mutants resistant to AM fungi as a control tool for field conditions

The utilization and management of arbuscular mycorrhiza (AM) symbiosis may improve production and sustainability of the cropping system. For this purpose, native AM fungi (AMF) were sought and tested for their efficiency to increase plant growth by enhanced P uptake and by alleviation of drought stress. Pot experiments with safflower (Carthamus tinctorius) and pea (Pisum sativum) in five soils (mostly sandy loamy Luvisols) and field experiments with peas were carried out during three years at four different sites. Host plants were grown in heated soils inoculated with AMF or the respective heat sterilized inoculum. In the case of peas, mutants resistant to AMF colonization were used as non-mycorrhizal controls. The mycorrhizal impact on yields and its components, transpiration, and P and N uptake was studied in several experiments, partly under varying P and N levels and water supply. Screening of native AMF by most probable number bioassays was not very meaningful. Soil monoliths were placed in the open to simulate field conditions. Inoculation with a native AMF mix improved grain yield, shoot and leaf growth variables as compared to control. Exposed to drought, higher soil water depletion of mycorrhizal plants resulted in a haying-off effect. The growth response to this inoculum could not be significantly reproduced in a subsequent open air pot experiment at two levels of irrigation and P fertilization, however, safflower grew better at higher P and water supply by multiples. The water use efficiency concerning biomass was improved by the AMF inoculum in the two experiments. Transpiration rates were not significantly affected by AM but as a tendency were higher in non-mycorrhizal safflower. A fundamental methodological problem in mycorrhiza field research is providing an appropriate (negative) control for the experimental factor arbuscular mycorrhiza. Soil sterilization or fungicide treatment have undesirable side effects in field and greenhouse settings. Furthermore, artificial rooting, temperature and light conditions in pot experiments may interfere with the interpretation of mycorrhiza effects. Therefore, the myc- pea mutant P2 was tested as a non-mycorrhizal control in a bioassay to evaluate AMF under field conditions in comparison to the symbiotic isogenetic wild type of var. FRISSON as a new integrative approach. However, mutant P2 is also of nod- phenotype and therefore unable to fix N2. A 3-factorial experiment was carried out in a climate chamber at high NPK fertilization to examine the two isolines under non-symbiotic and symbiotic conditions. P2 achieved the same (or higher) biomass as wild type both under good and poor water supply. However, inoculation with the AMF Glomus manihot did not improve plant growth. Differences of grain and straw yields in field trials were large (up to 80 per cent) between those isogenetic pea lines mainly due to higher P uptake under P and water limited conditions. The lacking N2 fixation in mutants was compensated for by high mineral N supply as indicated by the high N status of the pea mutant plants. This finding was corroborated by the results of a major field experiment at three sites with two levels of N fertilization. The higher N rate did not affect grain or straw yields of the non-fixing mutants. Very efficient AMF were detected in a Ferric Luvisol on pasture land as revealed by yield levels of the evaluation crop and by functional vital staining of highly colonized roots. Generally, levels of grain yield were low, at between 40 and 980 kg ha-1. An additional pot trial was carried out to elucidate the strong mycorrhizal effect in the Ferric Luvisol. A triplication of the plant equivalent field P fertilization was necessary to compensate for the mycorrhizal benefit which was with five times higher grain yield very similar to that found in the field experiment. However, the yield differences between the two isolines were not always plausible as the evaluation variable because they were also found in (small) field test trials with apparently sufficient P and N supply and in a soil of almost no AMF potential. This similarly occurred for pea lines of var. SPARKLE and its non-fixing mycorrhizal (E135) and non-symbiotic (R25) isomutants, which were tested in order to exclude experimentally undesirable benefits by N2 fixation. In contrast to var. FRISSON, SPARKLE was not a suitable variety for Mediterranean field conditions. This raises suspicion putative genetic defects other than symbiotic ones may be effective under field conditions, which would conflict with the concept of an appropriate control. It was concluded that AMF resistant plants may help to overcome fundamental problems of present research on arbuscular mycorrhiza, but may create new ones.

Development and Integration of Biocontrol Products in Branched Broomrape (Phelipanche ramosa) management in Tomato

Parasitic weeds of the genera Striga, Orobanche, and Phelipanche pose a severe problem for agriculture because they are difficult to control and are highly destructive to several crops. The present work was carried out during the period October, 2009 to February, 2012 to evaluate the potential of arbuscular mycorrhizal fungi (AMF) to suppress P. ramosa on tomatoes and to investigate the effects of air-dried powder and aqueous extracts from Euphorbia hirta on germination and haustorium initiation in Phelipanche ramosa. The work was divided into three parts: a survey of the indigenous mycorrhizal flora in Sudan, second, laboratory and greenhouse experiments (conducted in Germany and Sudan) to construct a base for the third part, which was a field trial in Sudan. A survey was performed in 2009 in the White Nile state, Sudan to assess AMF spore densities and root colonization in nine fields planted with 13 different important agricultural crops. In addition, an attempt was made to study the relationship between soil physico-chemical properties and AMF spore density, colonization rate, species richness and other diversity indices. The mean percentage of AMF colonization was 34%, ranging from 19-50%. The spore densities (expressed as per 100 g dry soil) retrieved from the rhizosphere of different crops were relatively high, varying from 344 to 1222 with a mean of 798. There was no correlation between spore densities in soil and root colonization percentage. A total of 45 morphologically classifiable species representing ten genera of AMF were detected with no correlation between the number of species found in a soil sample and the spore density. The most abundant genus was Glomus (20 species). The AMF diversity expressed by the Shannon–Weaver index was highest in sorghum (H\= 2.27) and Jews mallow (H\= 2.13) and lowest in alfalfa (H\= 1.4). With respect to crop species, the genera Glomus and Entrophospora were encountered in almost all crops, except for Entrophospora in alfalfa. Kuklospora was found only in sugarcane and sorghum. The genus Ambispora was recovered only in mint and okra, while mint and onion were the only species on which no Acaulospora was found. The hierarchical cluster analysis based on the similarity among AMF communities with respect to crop species overall showed that species compositions were relatively similar with the highest dissimilarity of about 25% separating three of the mango samples and the four sorghum samples from all other samples. Laboratory experiments studied the influence of root and stem exudates of three tomato varieties infected by three different Glomus species on germination of P. ramosa. Root exudates were collected 21or 42 days after transplanting (DAT) and stem exudates 42 DAT and tested for their effects on germination of P. ramosa seeds in vitro. The tomato varieties studied did not have an effect on either mycorrhizal colonization or Phelipanche germination. Germination in response to exudates from 42 day old mycorrhizal plants was significantly reduced in comparison to non-mycorrhizal controls. Germination of P. ramosa in response to root exudates from 21 day old plants was consistently higher than for 42 day-old plants (F=121.6; P<.0001). Stem diffusates from non-mycorrhizal plants invariably elicited higher germination than diffusates from the corresponding mycorrhizal ones and differences were mostly statistically significant. A series of laboratory experiments was undertaken to investigate the effects of aqueous extracts from Euphorbia hirta on germination, radicle elongation, and haustorium initiation in P. ramosa. P. ramosa seeds conditioned in water and subsequently treated with diluted E. hirta extract (10-25% v/v) displayed considerable germination (47-62%). Increasing extract concentration to 50% or more reduced germination in response to the synthetic germination stimulants GR24 and Nijmegen-1 in a concentration dependent manner. P. ramosa germlings treated with diluted Euphorbia extract (10-75 % v/v) displayed haustorium initiation comparable to 2, 5-Dimethoxy-p-benzoquinon (DMBQ) at 20 µM. Euphorbia extract applied during conditioning reduced haustorium initiation in a concentration dependent manner. E. hirta extract or air-dried powder, applied to soil, induced considerable P. ramosa germination. Pot experiments were undertaken in a glasshouse at the University of Kassel, Germany, to investigate the effects of P. ramosa seed bank on tomato growth parameters. Different Phelipanche seed banks were established by mixing the parasite seeds (0 - 32 mg) with the potting medium in each pot. P. ramosa reduced all tomato growth parameters measured and the reduction progressively increased with seed bank. Root and total dry matter accumulation per tomato plant were most affected. P. ramosa emergence, number of tubercles, and tubercle dry weight increased with the seed bank and were, invariably, maximal with the highest seed bank. Another objective was to determine if different AM fungi differ in their effects on the colonization of tomatoes with P. ramosa and the performance of P. ramosa after colonization. Three AMF species viz. GIomus intraradices, Glomus mosseae and Glomus Sprint® were used in this study. For the infection, P. ramosa seeds (8 mg) were mixed with the top 5 cm soil in each pot. No mycorrhizal colonization was detected in un-inoculated control plants. P. ramosa infested, mycorrhiza inoculated tomato plants had significantly lower AMF colonization compared to plants not infested with P. ramosa. Inoculation with G. intraradices, G. mosseae and Glomus Sprint® reduced the number of emerged P. ramosa plants by 29.3, 45.3 and 62.7% and the number of tubercles by 22.2, 42 and 56.8%, respectively. Mycorrhizal root colonization was positively correlated with number of branches and total dry matter of tomatoes. Field experiments on tomato undertaken in 2010/12 were only partially successful because of insect infestations which resulted in the complete destruction of the second run of the experiment. The effects of the inoculation with AMF, the addition of 10 t ha-1 filter mud (FM), an organic residues from sugar processing and 36 or 72 kg N ha-1 on the infestation of tomatoes with P. ramosa were assessed. In un-inoculated control plants, AMF colonization ranged between 13.4 to 22.1% with no significant differences among FM and N treatments. Adding AMF or FM resulted in a significant increase of branching in the tomato plants with no additive effects. Dry weights were slightly increased through FM application when no N was applied and significantly at 36 kg N ha-1. There was no effect of FM on the time until the first Phelipanche emerged while AMF and N application interacted. Especially AMF inoculation resulted in a tendency to delayed P. ramosa emergence. The marketable yield was extremely low due to the strong fruit infestation with insects mainly whitefly Bemisia tabaci and tomato leaf miner (Tuta absoluta). Tomatoes inoculated with varied mycorrhiza species displayed different response to the insect infestation, as G. intraradices significantly reduced the infestation, while G. mosseae elicited higher insect infestation. The results of the present thesis indicate that there may be a potential of developing management strategies for P. ramosa targeting the pre-attachment stage namely germination and haustorial initiation using plant extracts. However, ways of practical use need to be developed. If such treatments can be combined with AMF inoculation also needs to be investigated. Overall, it will require a systematic approach to develop management tools that are easily applicable and affordable to Sudanese farmers. It is well-known that proper agronomical practices such as the design of an optimum crop rotation in cropping systems, reduced tillage, promotion of cover crops, the introduction of multi-microbial inoculants, and maintenance of proper phosphorus levels are advantageous if the mycorrhiza protection method is exploited against Phelipanche ramosa infestation. Without the knowledge about the biology of the parasitic weeds by the farmers and basic preventive measures such as hygiene and seed quality control no control strategy will be successful, however.

Successful rescue and field establishment of native banana varieties severely affected by rhizome rot

Rhizome rot disease caused by Erwinia spp. is emerging as a major problem in banana nurseries and young plantations worldwide. Management of the disease is possible only in the initial stages of development. Currently no method is available for rescuing plant material already infected with this pathogen. A total of 95 Nanjanagud Rasabale and 212 Elakki Bale suckers were collected from different growing regions of Karnataka, India. During nursery maintenance of these lines, severe Erwinia infection was noticed. We present a method to rescue infected plants and establish them under field conditions. Differences were noticed in infection severity amongst the varieties and their accessions. Field data revealed good establishment and growth of most rescued plants under field conditions. The discussed rescue protocol coupled with good field management practices resulted in 89.19 and 82.59 percent field establishment of previously infected var. Nanjanagud Rasabale and var. Elakki Bale plants, respectively.

Simultaneous Drying and Separation of Composite Agricultural Material for Hay Processing

In composite agricultural materials such as grass, tee, medicinal plants; leaves and stems have a different drying time. By this behavior, after leaving the dryer, the stems may have greater moisture content than desired, while the leaves one minor, which can cause either the appearance of fungi or the collapse of the over-dried material. Taking into account that a lot of grass is dehydrated in forced air dryers, especially rotary drum dryers, this research was developed in order to establish conditions enabling to make a separation of the components during the drying process in order to provide a homogeneous product at the end. For this, a rotary dryer consisting of three concentric cylinders and a circular sieve aligned with the more internal cylinder was proposed; so that, once material enters into the dryer in the area of the inner cylinder, stems pass through sieve to the middle and then continue towards the external cylinder, while the leaves continue by the inner cylinder. For this project, a mixture of Ryegrass and White Clover was used. The characteristics of the components of a mixture were: Drying Rate in thin layer and in rotation, Bulk density, Projected Area, Terminal velocity, weight/Area Ratio, Flux through Rotary sieve. Three drying temperatures; 40°C, 60° C and 80° C, and three rotation speeds; 10 rpm, 20 rpm and 40 rpm were evaluated. It was found that the differences in drying time are the less at 80 °C when the dryer rotates at 40 rpm. Above this speed, the material adheres to the walls of the dryer or sieve and does not flow. According to the measurements of terminal velocity of stems and leaves of the components of the mixture, the speed of the air should be less than 1.5 m s-1 in the inner drum for the leaves and less than 4.5 m s-1 in middle and outer drums for stems, in such way that only the rotational movement of the dryer moves the material and achieves a greater residence time. In other hand, the best rotary sieve separation efficiencies were achieved when the material is dry, but the results are good in all the moisture contents. The best rotary speed of sieve is within the critical rotational speed, i.e. 20 rpm. However, the rotational speed of the dryer, including the sieve in line with the inner cylinder should be 10 rpm or less in order to achieve the greatest residence times of the material inside the dryer and the best agitation through the use of lifting flights. With a finite element analysis of a dryer prototype, using an air flow allowing speeds of air already stated, I was found that the best performance occurs when, through a cover, air enters the dryer front of the Middle cylinder and when the inner cylinder is formed in its entirety through a sieve. This way, air flows in almost equal amounts by both the middle and external cylinders, while part of the air in the Middle cylinder passes through the sieve towards the inner cylinder. With this, leaves do not adhere to the sieve and flow along drier, thanks to the rotating movement of the drums and the showering caused by the lifting flights. In these conditions, the differences in drying time are reduced to 60 minutes, but the residence time is higher for the stems than for leaves, therefore the components of the mixture of grass run out of the dryer with the same desired moisture content.

A survey of brassica vegetable smallholder farmers in the Gauteng and Limpopo provinces of South Africa

A study was taken to investigate the types of brassica vegetables mostly grown by smallholder farmers in two provinces of South Africa. Thirty-one smallholder vegetable farmers in the Gauteng province and Waterberg district in the Limpopo province were surveyed. In addition, the study also sought to establish the common diseases, the management strategies used and problems encountered by the farmers. Farmers were interviewed using a questionnaire with closed and open–ended questions. The results indicated that the smallholder farmers mostly grew cabbage (93.6%) as their main brassica crop followed by rape (41.2%). Thirty percent of farmers could not identify or name the predominant disease/s encountered in their fields. Major diseases encountered by farmers surveyed were an unknown disease/s (33.3%), black rot (26.7%), Alternaria leaf spot (6.7%) and white rust (6.7%). Smallholder farmers have inadequate technical information available especially relating to crop diseases, their identification and control. Farmers encountered challenges with black rot disease especially on cabbage, rape and kale and the disease was a problem during winter and summer. Generally, the smallholder farmers used crop rotation (74.2%) as a major practice to manage the diseases experienced. They rotated their brassica vegetables with other crops/vegetables like tomatoes, onions, beetroots and maize. Most of the farmers interviewed (61.3%) did not use chemicals to control diseases, whereas 38.7% of them used chemicals. This was mostly because they lacked information and knowledge, high costs associated with use of chemical fungicides and some were shifting towards organic farming. From the study it was noted that there was a need for technical support to improve farmers’ knowledge on disease identification and control within the surveyed areas.