Host resistance has become a cornerstone of sustainable production of flue-cured tobacco in regions where root-knot nematodes present a threat to growers. Resistance to races 1 and 3 of M. incognita, historically the most significant root-knot nematode threat to tobacco production, is now widely available in commercially available flue-cured tobacco varieties, and is imparted by the gene Rk1. The same gene also provides resistance to race 1 of M. arenaria. The widespread deployment of this resistance has fostered a shift in root-knot nematode population dynamics, as a result of which M. arenaria race 2 has become the predominant root-knot nematode threat in Virginia. A second resistance gene known to impart resistance to M. javanica, Rk2, has also been incorporated into numerous released cultivars in combination with Rk1. This combination has been demonstrated to impart increased resistance to M. incognita and M. javanica relative to either gene alone. In the present work, eleven greenhouse trials conducted from 2017-2019, as well as two trials conducted in 2018 and 2019 on a cooperating farm, investigated the efficacy of this stacked resistance against M. arenaria race 2 and compared the effect of stacking both resistance genes to the effect of either gene alone relative to a susceptible cultivar. We also evaluated how these forms of resistance compare with resistance possessed by a breeding line with resistance reportedly derived from N. repanda to determine if additional, novel sources of resistance to root-knot nematodes previously identified from other species in the genus Nicotiana could play a role in expanding the genetic diversity of germplasm available for the refinement of host resistance in flue-cured tobacco. Additionally, in light of potential new rule making from the FDA mandating reduced nicotine content of cured tobacco leaf, we investigated the relationship between alkaloid (nicotine) content of flue-cured tobacco and root-knot nematode parasitism, while also evaluating nematode parasitism effects on carbohydrate content. Despite considerable variability in our results, particularly under field conditions, our results demonstrate that stacking Rk1 and Rk2 imparts greater resistance to M. arenaria race 2 than either gene alone, but that an entry possessing resistance reportedly derived from N. repanda exhibited significantly greater resistance to root-knot nematodes than the combination of Rk1 and Rk2 based on root galling, and egg mass and egg production. The alkaloid content of flue-cured tobacco did not appear to have an effect on root-knot nematode parasitism under greenhouse or field conditions, but the presence of the nematode did lead to increased accumulation of nicotine in the roots of plants, while translocation of nicotine to leaves was reduced. Conversely, root-knot nematode parasitism was reduced accumulation of carbohydrates in roots, while having no significant effects on leaf carbohydrate content. / Ph.D. / Root-knot nematodes (Meloidogyne spp.) are microscopic round worms that can cause considerable damage to flue-cured tobacco (Nicotiana tabacum L.), and while not typically responsible for killing plants outright, can reduce the quality of cured tobacco leaf and may predispose plants to a host of other issues, resulting in challenges and economic burdens on growers. Chemicals that effectively control nematodes, which are animals, pose inherent threats to human applicators and may harm the environment in a number of ways, so the use of tobacco varieties that are resistant to root-knot nematodes is increasingly common and essential to sustainable tobacco production. One form of root-knot nematode resistance, called Rk1, has become common and is found in all commercially grown flue-cured tobacco. This form of resistance is effective against 2 ‘races’ of the root-knot nematode M. incognita, which has historically caused tobacco growers the most issues. However, because this resistance is so widely employed, growers have controlled these nematodes, while another species, M. arenaria, has become more prevalent, particularly ‘race’ 2, which is not controlled by Rk1. We know from previous research that another gene, Rk2, provides resistance to some root-knot nematode that Rk1 does not effect, and that combining both genes seems to provide even greater root-knot nematode control than either gene alone. We investigated whether Rk2 is effective at controlling M. arenaria race 2 when it is combined with Rk1 in greenhouse and field experiments. We also investigated how a different, novel type of resistance, which comes from a species of tobacco related to cultivated tobacco, compares with the Rk1/Rk2 resistance in greenhouse trials. Additionally, the FDA has recently suggested that nicotine levels in tobacco leaf should be dramatically reduced to help mitigate adverse human health consequences associated with tobacco consumption. Nicotine may play some role in resistance to root-knot nematode in tobacco, and conversely, root-knot nematodes may impact levels of nicotine, as well as other important chemical constituents of tobacco. We also investigated these questions in greenhouse and field experiments. Our results ultimately demonstrate that combining both Rk1 and Rk2 gives flue-cured tobacco a higher level of resistance to root-knot nematodes than either gene alone, but also suggests that the form of resistance we evaluated from a related Nicotiana species could be even more effective in controlling these nematodes. We observed that the amount of nicotine present in tobacco did not impact nematode parasitism, but that nematode parasitism could lead to lower levels of nicotine in the leaves of plants because the nematodes, which feed on plants roots, cause damage to the plant that interferes in the movement of nicotine from roots to leaves.

Utilizing resistant cultivars is a main control strategy for root-knot nematodes in flue-cured tobacco (Nicotiana tabacum L.). Most commercial cultivars possess the Rk1 gene, providing resistance to races 1 and 3 of Meloidogyne incognita and race 1 of M. arenaria. This initiated a shift in root-knot populations to other species and races, creating a need for resistance to those populations. Numerous cultivars possess a second resistance gene, Rk2. Greenhouse experiments investigated whether possessing both Rk1 and Rk2 increases resistance to a variant of M. incognita race 3 compared to either gene alone, and if high soil temperatures impact their efficacy. Root galling, numbers of egg masses and eggs, and the reproductive index were compared from roots of Coker 371-Gold (susceptible), NC 95 and SC 72 (Rk1Rk1), T-15-1-1 (Rk2Rk2), and STNCB-2-28 and NOD 8 (Rk1Rk1 and Rk2Rk2). The same data were analyzed from plants in open-top root zone cabinet growth chambers set to 25ºC, 30ºC, and 35ºC to examine if resistance is temperature sensitive. Despite variability, Rk1Rk2 entries conferred greater resistance than entries with Rk1 or Rk2 alone. Entries with Rk1 alone reduced galling and reproduction compared to the susceptible control, whereas T-15-1-1 (Rk2) did not, but often suppressed reproduction. An apparent reduction in nematode reproduction was observed at 25ºC and 30ºC on entries possessing Rk1 and Rk1Rk2 compared to the control and Rk2. However, no apparent differences in reproduction occurred on Rk1 and/or Rk2 entries at 35ºC compared to the control, indicating parasitism increased on resistant entries at higher temperatures. / Master of Science in Life Sciences