The combined top growth of all six root stock seedlings was increased with the addition of VAN fungi and no phosphorus. However, the combined growth with VAN and phosphorus was slightly more. The root stock seedlings decreased in dependency on VAN in the following order: sour orange, Cleopatra mandarin, sweet orange, Rough lemon, Rangpur lime and Carrizo citrange. Root-shoot ratios indicate that root stock mycorrhizal dependency decreases as their capacity for root production increases. Emphasizing the importance of previous work, Timmer and Leyden state the interactions of copper and phosphorus in the fumigation-mycorrhizal syndrome are important. Copper deficiency has been frequently observed in citrus seedlings following the application of phosphate fertilizer. According to Timmer and Leyden, the application of phosphorus induces copper deficiency by stimulating growth of non-mycorrhizal seedlings until copper becomes limiting nutritionally. However, phosphorus-induced copper deficiency appears to be due to phosphorus inhibition of mycorrhizal development of seedlings inoculated with Glomus fasciculatus. Rhodes stresses the fact that mycorrhizae are recognized as being significantly beneficial to host-plant relationships, particularly where root systems are restricted and nutrient systems are low—although he makes no special reference to citrus. Mehraveran also discusses the mycorrhizal dependency of six citrus cultivars .Plants with mycorrhizae and a given phosphorus level are healthier than non-mycorrhizal plants with an equal phosphorus level. The type of mycorrhizae is important; the soil type, area of origin, effects on nematodes, Phytophthora, photosynthetic activity, and hydraulics are all factors to consider. Thus, Menge et al. found that in greenhouse experiments the addition of Glomus fasciculatus significantly increased the growth of Troyer citrange seedlings in 20 of 26 methyl bromide-fumigated soils from Southern California.

Of the six soils in which the mycorrhizal fungus provided no growth increase, two were greenhouse soils ,growing blueberries in pots three were nursery soils, and one was a field soil. Presence of the fungus increased foliar phosphorus, potassium, and copper and decreased foliar magnesium and sodium concentrations in the leaves of the Troyer citrange in the majority of the citrus soils. They present an interesting table on the mycorrhizal dependency of Troyer on Glomus fasciculatus on the 26 soils. While much of the work has been done with sandy soils, the whole soil-mycorrhizal complex is extremely important. Obviously there are different species of the mycorrhizal fungus and one wonders if they are equally effective and if their response varies in a different soil environment. To determine this, Graham, Linderman and Menge tested six VAN isolates. They found that isolates of Glomus fasciculatus was most efficient, and Glomus macrocarpum the least effective. Growth enhancement was significantly greater for Glomus isolates from California than from Florida. It is also stressed that growth enhancement for VAN fungi may vary with the soil type. Johnson reports on the effects of phosphorus nutrition on mycorrhizal colonization, photosynthesis, growth, and nutrient composition in sour orange seedlings. The sour orange seedlings were inoculated with the VAN fungus Glomus intraradices and fertilized with weekly applications of phosphorus. The photosynthetic rates correlated with a high phosphorus content in the leaf tissue of the central plants, but Johnson could find no correlation for the VAN-infected seedlings. He suggests that factors in addition to improved phosphorus nutrition influence the photosynthetic rate of VAN plants. The conclusions of Edriss, Davis and Burger were somewhat similar. Using sour orange seedlings and inoculating them with the mycorrhizal fungus Gigaspora heterogama, they found that the cytokinin production was greater than that of the check non-mycorrhizal plants despite the fact that there were similar dry weights and phosphorus concentrations in the leaves. The enhancement of the cytokinin production seemed to be associated with the mycorrhizal infection rather than increased phosphorus uptake.

The effect of VAN inoculations on nematode populations was first investigated by O’Bannon et al. in greenhouse studies in Florida. They found that when Rough lemon seedlings were inoculated with the citrus nematode, Tylenchulus semipenetrans and then transplanted into soil infected with VAN Glomus mosseae, that the presence of the fungus increased seedling growth. The seedling suppression by the citrus nematode alone was greater than the checks or the VAN inoculated. They made no studies on other nematodes such as the burrowing nematode Radopholus similis. Similarly, Hussey and Roncadori report that nematode suppression of vegetative growth or yield are partly offset by the presence of a VAN fungus. Using Rough lemon seedlings they found that the presence of the mycorrhizal fungus lessened the nematodes’ attraction to the citrus roots, hindered penetration, and the subsequent development and reproduction of the citrus nematode was suppressed. They do not specify why, except that a healthier plant is a more resistant plant. To some extent, this same hypothesis exists with the VAN fungus relationship with Phytophthora. Davis and Menge working with Pineapple sweet orange seedlings, present evidence that suggested that there was some tolerance to Phytophthora parasitica in seedlings infected with Glomus fasciculatus. They also felt this effect was caused by the ability of mycorrhizal roots to absorb more phosphorus and possibly other nutrients than non-mycorrhizal roots, as evidenced by root health and greater phosphorus uptake. Again, a healthier plant is a more resistant plant. They did not report on any studies with Phytophthora citrophthora. Davis and Menge again suggest that VAN fungi have a variable influence on the tolerance of seedlings of Pineapple sweet orange and Troyer citrange to Phytophthora parasitica. There are a number of papers on the effects of VAN fungi and water relationships of host citrus seedlings. The first report perhaps was that of Levy and Krikun . These two Israeli researchers working with Rough lemon seedlings studied recovery from water stress on similar-sized VAN infected seedlings and non-mycorrhizal seedlings. They found the VAN-infected seedlings affected stomatal conductance, photosynthesis and proline accumulation but not leaf water potential.

They suggest that most of the effect of mycorrhizal association is on stomatal regulation rather than on root resistance. Syvertsen studied the hydraulic conductivity of four commercial citrus root stocks. The hydraulic conductivity was estimated using a special pressure chamber technique. He found that Carrizo citrange and Rough lemon had the highest root conductivity, whereas Cleopatra mandarin and sour orange had the least. Further hydraulic studies were conducted by Graham and Syvertsen . They used seedlings of Carrizo citrange and sour orange grown in a low phosphorus sandy soil and either inoculated with Glomus intraradices or fertilized with phosphorus. The mycorrhizal-infected seedlings had sufficient levels of leaf phosphorus, but the non-mycorrhizal seedlings were phosphorus deficient. The root-shoot ratio of both root stocks was reduced by the mycorrhizal colonization, but root hydraulic conductivity per unit root length of mycorrhizal Carrizo and sour orange was more than twice that of non-mycorrhizal seedlings. The mycorrhizal plants had higher transpiration rates, apparently increased bythe conductivity of the roots. The authors felt the response was due to the mycorrhizal enhancement of phosphorus nutrition. In further studies, Graham and Syvertsen grew seedlings of five citrus root stocks in a low phosphorus sandy soil. The seedlings were incorporated into three treatments: inoculated with Glomus intraradices, non-inoculated, but fertilized with phosphorus, and non-inoculated and no phosphorus added. The order of the mycorrhizal dependency of the five root-stocks is as follows: sour orange = Cleopatra mandarin > Swingle citrumelo > Carrizo citrange > trifoliate orange. The less dependent root stocks, i.e., trifoliate orange and Carrizo citrange, had greater leaf phosphorus, finer roots, and slower growth rates than sour orange and Cleopatra mandarin. Rootstocks with a lower mycorrhizal dependency also generally had greater hydraulic conductivity of the roots,frambuesa cultivo greater transpiration and carbon dioxide assimilation rates. In additional but similar studies, Syvertsen and Graham amplify their work again with seedlings of Carrizo citrange, trifoliate orange, sour orange, Swingle citrumelo, and Cleopatra mandarin. Whole plant transpiration and maximum rates of net gas exchange or carbon dioxide and water vapor from single leaves were positively correlated with the hydraulic conductivity of the seedlings. Leaf nitrogen and phosphorus content and shoot-root ratio were also positively correlated with root conductivity. The differences in soil water depletion and plant water relations of trifoliate orange and Carrizo citrange during drought and recovery cycle is related to their root conductivity. They stress that the capability of root systems to conduct water and mineral elements is a very important factor in plant growth and physiological activity. The possibilities of seed inoculation or field inoculation was first mentioned by Newcomb . Hattingh and Gerdemann inoculated sour orange seed, successfully developing a special technique. They coated the seed with a mycorrhizal inoculum in a 1% solution of methyl cellulose. Menge, Lembright and Johnson indicated that commercial production of mycorrhizal inoculum for use in fumigated or sterilized soil was being attempted in several locations in the United States.

They felt the only current way to produce suitable quantities of mycorrhizal inoculum was on roots of susceptible host plants. Contamination by other pathogenic organisms can be a problem. Maybe better methods are available. The most common method for inoculating citrus in the field and in the greenhouse has been to mix the mycorrhizal inoculum with the soil prior to planting or transplanting. Menge et al. felt banding, layering and root inoculation were more efficient than seed inoculation. Root fragments can also be an important source of inoculation. Graham and Fardelmann found that root pieces stored up to one year under moist conditions did not lose the colonization potential with Glomus epigaeum. However, drying reduced this potential to nearly zero after nine months. Glomus intraradices was found sporulating in citrus roots found in orchard soil. They propose that dead root fragments account for a high percentage of the propagules in the citrus soil. The propagation of mycorrhizae cannot be too difficult. For a number of years a citrus nurseryman in California offered mycorrhizae for sale for inoculative purposes. This service is no longer available either because of propagation problems or little demand by the citrus industry. The knowledge of the necessity of the mycorrhizae in the seed bed or the transplant stage is the important thing. The mycorrhizae will gradually re-infect a fumigated soil. The nurseryman could leave the fumigated site fallow for a year or plant host cover crops of cereals and grasses or legumes prior to planting a seedbed or transplanting. Furthermore, field grown nursery trees are gradually diminishing. Container growing is so much more efficient and economical, and container trees can be grown in a shorter period of time. There is also less transplant shock. When the author visited South Africa in 1982, there were only three field grown nurseries left and they ceased with that planting. South Africa is essentially 100% in container growing. Spain and Australia are now heavily into container growing, with more citrus producing areas following suit. If the container planting mix is properly planned and prepared, there will no longer be this problem. For further information on this problem, a nice review of the total VAN potential benefits and interactions is presented by Graham . While the article is a review, it nicely presents the views and facts in pathology, horticulture, and physiology. A book on the subject is currently being written by J. A. Menge of the Department of Plant Pathology, Citrus Research Center, Riverside. Differences in cellular structure have infrequently but effectively been used to distinguish a limited number of stock species. Thus, Penzig found a striking difference in the cellular structure of the pith of twigs of trifoliate orange and sour orange, and this may probably also be true of the roots. Swingle suggested this method of distinguishing between these two species when used in Satsuma production in the U.S. Gulf states. Longitudinal sections of the pith of young stems of the trifoliate orange exhibit an irregular arrangement of thin-walled cells [labeled as] Fig. 66A, while similar sections of sour orange stems show only uniform thin-walled cells arranged in regular series and an entire absence of the crossplates of thick-walled cells . Wolf extended this method to distinguish Yuzu, which was found to have only thin-walled pith cells similar to those of the sour orange, but irregularly arranged . Thus, it differs from the sour orange, in which the cells are arranged in regular series or chains, and from the trifoliate, in which there are crossplates of thick-walled cells .