Histological and Molecular Developmental Mechanism of Striga Weed (Striga hermonthica) Parasitizing Maize (Zea mays)

Abstract

Smallholder farmers in sub-Saharan Africa (SSA) have for decades battled with Striga hermonthica, a root parasitic angiosperm that constrains maize production leading to yield losses of up to 100%. The most sustainable strategies in control of S. hermonthica can only arise from a clear understaning of the parasite molecular mechanism during parasitism. Currently there is no clear understanding of mechanisms involved in haustoria development and the resistance mechanisms in some existing maize genotypes. Such an understanding could lead to several management approaches via RNA interference and gene overexpression techniques. This potential for developing resistance against S. hermonthica is also further constrained by the paucity of candidate genes to target and lack of efficient high throughput gene screening protocols. Viral induced gene silencing (VIGS) provides an easy and effective strategy in screening for putative candidate genes for targeting through knockdown techniques but has not been optimised in S. hermonthica. This studys' specific objectives were: (i) To Advance our understanding of the resistance mechanisms and biology of S. hermonthica parasitism on a susceptible (Namba nane) and tolerant (KSTP’94) open pollinated varieties of maize in Kenya, (ii) To identify some of the genes involved in S. hermonthica haustoriagenesis, (iii) To develop a VIGS protocol for functional genomics in the parasitic plant S. hermonthica. The two maize varieties were planted in S. hermonthica infested soil collected from Alupe followed by data collection on the number of germinated S. hermonthica plants and haustoria, in the different varieties. Further histological analysis were done by cross-sectioning formalin acetic alcohol (FAA) fixed S. hermonthica haustoria attached to maize root of the different varieties. Using a predictive literature search on other well studied parasitic plants like Orobanchea and triphysaria, gene specific primers were generated and used to clone followed by sanger sequencing of the fragments. For the VIGS protocol development, the tobacco rattle virus (TRV2 and TRV1) in agrobacterium GV3101 designed to silence phytoene desaturase (PDS) gene were introduced into S. hermonthica using agro-drench and agro-infiltration methods. The results show that the maize varieties response to S. hermonthica was different in the sense that there was delayed parasite emergence on the KSTP’94 (90 days after planting) compared to Namba nane where the parasite started emerging at 42 days after planting. Additionally, there was an average of 5±0.5 Striga plants with only 10.7±0.7% infestation rate while Namba nane had an average of 26±0.7 S. hermonthica plants per variety with 50.3±0.9% infestation rate. The average number of haustoria in the KSTP’94 was low (18±2.6) compared to 43±2.3 in Namba nane variety at 112 days after planting. Histologically, KSTP’94 variety exhibited resistance to haustoria penetration at the host cortex and endodermis and only in few cases did the parasite haustoria make connections to the vasculature. In Namba nane variety, the haustoria penetrated easily through the cortex and endodermis and made connections to the vasculature. Five key haustoria formation genes were found to be expressed in S. hermonthica haustoria through RT-PCR; Expansin, Cysteine protease, Mannose 6-phosphate reductase, Tvpirin and Quinone oxidoreductase. The effectiveness of agrodrench and agroinfiltration guided VIGS was determined in Striga via photobleaching phenotypes on leaves at 14 and 7 days post infection, respectively. The photobleaching however cleared at 28 days after planting. The transformation efficiency for VIGS protocol was 60±2.9% in agro-infiltration and 10.3±1.5% in agrodrench. Summarily, the results provide baseline information on the possible responses to local maize germplasm to S. hermonthica and the candidate genes involved in haustoria formation. The study developed a VIGS protocol that could be used for genomic studies in S. hermonthica. Further the genes identified in S. hermonthica could be used in parasite management via RNAi constructs.