Morphological Response of Witchweed (Striga asiatica) to In Vitro Culture

Excised sorghum root segments (5–10 mm in length) werecultured for 50 d in four different liquid media containingmineral salts, vitamins, amino acids, glucose, and IAA. Theroots were removed and the remaining medium was solidified withan equal volume of warm 1–6% water agar. Dry unconditionedor conditioned Striga asiatica seeds were transferred to themedium. Some of the seeds germinated and developed into parasitic-typeseedlings. These seedlings had haustoria, tubercles, dense roothairs, branched shoots, and multiple shoot-borne adventitiousroots. The plumule pole developed into a shoot, but the radiclepole displayed only rudimentary development. On the same media,but which had not previously been used to grow sorghum roots,the seedlings displayed a well-developed radicle-derived rootsystem, but the plumule did not grow. Shoots began to appearon the roots only after 35–50 d of culture. These seedlingshad no haustoria, no tubercles, few or scattered root hairs,no shoot-borne adventitious roots and few shoot branches, andappeared to be non-parasitic-type seedlings. Shoots grew ina medium supplemented with IAA and kinetin, but did not in amedium containing NAA plus IBA. On replacement of glucose andIAA with sucrose and 2,4-D, respectively, Striga seeds germinated,and the heart-shaped embryos dedifferentiated into calli. Thecalli have been maintained by subculturing for over 9 months.The results demonstrated that a host signal, in addition tothose for germination and haustorium formation, is requiredfor further development. Moreover, morphogenesis of culturedS. asiatica is influenced by exogenous growth regulators. Key words: Striga asiatica, parasitic weeds, haustoria, Sorghum bicolor, in vitro culture     

A time-course study of early establishment stages of parasitic angiosperm Striga asiatica on susceptible sorghum roots

A time-course study of the early establishment stages of Striga asiatica was carried out on a susceptible sorghum hybrid, CSH 1, using polyethylene bags and whole-root clearing and staining techniques. Preconditioned Striga seeds were applied to different aged segments of primary root but the results did not differ for these different aged segments. Most of the Striga seeds (63%) germinated within 24 h of inoculation on the host roots. The attachment of Striga radicles to host root was rapid and it occurred between 36 and 48 h after inoculation. Only 9% of the germinated Striga seeds attached to the host root but 65% of these attachments successfully penetrated through the epidermis and entered the host cortex within 72 h. Penetration through the cortical cells was difficult; only 17% of attachments were able to reach the endodermis. Penetration took from 12 to 43 h after the first appearance of haustorial cells in the cortex; a total of 84 to 120 h after inoculation on the host root. Penetration through the endodermis and establishment on the host stele was relatively easier, as most of the haustoria reaching the endodermis were able to establish on the host stele. But this is a slow process taking a minimum of 24 h, and a maximum of 60 h after first contact of haustorial cells with the endodermis. The minimum time taken from inoculation of ungerminated Striga seed on the host root to establishment is about 108 h. The results are discussed in relation to published reports on other parasitic species such as Agalinis purpurea.     

Striga gesnerioidesParasitising Cowpea: Development of Infection Structures and Mechanisms of Penetration

Striga gesnerioides(Scrophulariaceae) is an obligate hemiparasiticangiosperm, that infects the roots of cowpea (Vigna unguiculata).The development ofStrigaseedlings was characterized after transferto the surface of cowpea roots using cryo-scanning electronmicroscopy. Contact with the surface of the root caused distinctmorphological changes in theStrigaradicles. Radicle elongationceased 8 h after contact with host roots and radicle hairs developed.These hairs adhered to the surface of the host root. Early stagesof root penetration, i.e. prior to connection to the host vascularsystem were examined by light microscopy and transmission electronmicroscopy to determine the mechanisms by whichStrigainvadesroot tissue. Cowpea roots were penetrated byS. gesnerioidesradiclesafter 48 h. Host cells immediately surrounding the invadingStrigawerecompressed, but other host cells were not affected. Immunocytochemistry,using monoclonal antibodies JIM 5 and JIM 7, showed thatStrigaadvancedintercellularly through the root tissues without extensive removalof pectins at the site of penetration.Copyright 1998 Annalsof Botany Company Striga gesnerioides,Vigna unguiculata, mechanisms of penetration, host-parasite interactions, electron microscopy, immunocytochemistry.     

HowStriga Parasitizes its Host: a TEM and SEM Study

The cellular contact betweenStriga hermonthica andStriga asiaticaand their hosts,Zea mays andSorghum bicolor , was investigatedby light, transmission electron and scanning electron microscopy.The xylem connections between parasites and hosts involve veryspecific, clustered intrusions into the host's water conductingelements, predominantly into the large vessel elements. A singlehaustorial cell can penetrate a host vessel element with morethan one intrusion. All intrusions become covered by an additionalelectron-opaque wall layer. During subsequent differentiation,a dissolution of specific wall parts of the cell intrusionsoccurs so that open, cup- or trunk-like structures result. Thevessel-like host contact can comprise up to five openings withina single intrusion. Concomitantly, the intrusions and the haustorialcells to which they belong lose their protoplasts and transforminto elements which take up water. The walls of the haustorialcells and both wall parts of their appendages become stronglylignified. The water and nutrient absorbing structures insertedinto the host vessel are named ‘oscula’. Withinthe whole haustorial complex of bothStriga species no phloemelements were detected. Translocation of substances from hostto parasite are briefly discussed. Striga hermonthica ; Striga asiatica ; haustorial anatomy; xylem contact; osculum     

The Regulation of the Water Potential Gradient in the Host and Parasite Relationship betweenSorghum bicolorandStriga hermonthica

A glasshouse experiment was carried out to investigate the factorscontrolling the abstraction of xylem fluid from its host bythe parasiteStriga hermonthica(Scrophulariaceae).Strigahad amean daily transpiration rate far exceeding that of its hostsorghum (Sorghum bicolor), with infestation byStrigaalso shownto lower the transpiration rate of the host. Stopping the host'stranspiration was shown to decrease the transpiration rate ofthe parasite. Stopping the parasite's transpiration only gavean initial increase in the host's transpiration rate which wasnot sustained. The parasite had a lower water potential thanits host, values being -0.42 MPa and -0.23 MPa, respectively,and an accompanying higher osmotic pressure of 0.68 MPa against0.51 MPa for sorghum. Modifying the water potential gradientby bagging both partners together showed that the differentialin osmotic pressure and water potential was largely maintainedby the parasite's higher rate of transpiration. A favourablewater potential gradient towards the parasite still existedfollowing the cessation of transpiration, this being generatedby the haustorial resistance to hydraulic conductivity whichwas found to be some 1.5–4.5 times greater than that offeredby the parasite shoot. Both the high rate of transpiration andthe increased resistance across the haustoria would appear tobe necessary means to facilitate the diversion of host resourcesto the parasite.Copyright 1997 Annals of Botany Company Striga hermonthica; sorghum; water relations; haustorium; root parasite     

A Comparative Study of the Haustorial Development of Striga asiatica (L.) Kuntze on Sorghum Cultivars

Ten sorghum cultivars were studied for their mode of Strigaparasitization, and the factors conferring resistance in resistantcultivars most of the Striga haustona failed to penetrate beyondthe endodermis, whereas in susceptible cultivars the haustonapenetrated the endodermis and became established Resistant cultivars showed marked endodermal and pencyclic thickeningand the deposition of silica in their endodermal cells, whichwere lacking in the susceptible cultivars Extra thickening inpencyclic cells as a response to the entrance of haustonum wasobserved in cultivars N-13 and IS-4202 Ten cultivars studied showed differential haustorial reactionsThese reactions included extra thickening in the pericycle inresponse to haustonal infestation, haustonal collapse, tylosts-likeocclusions in the xylem vessels, and the deposition of dark-stainingmaterials in the cortex Although no definite conclusion couldbe drawn regarding the relationship between the degree of mechanicaltissue development and field resistance, there was evidencethat some field-resistant cultivars have strong mechanical tissuesThere could, however, be other factors governing resistanceto Striga in the field Striga asiatica, sorghum, haustorium, anatomy, endodermis, pot test, host resistance mechanism, parasitization, susceptibility     

Tropism and Morphogenesis of Striga Seedlings in the Host Rhizosphere

Using an agar culture technique, studies have been made on thegrowth and development of Striga asiaticaseedlings in the rhizosphereof a host plant(Sorghum Vulgare). The host root exerts a definitepositive chemotropic influence over the growth of the parasiteradicle in the immediate vicinity of the host root, but notfurther. The host root also produces a morphogenic factor whichinduces striking changes in the parasite seedlings. The mechanismof tropism and the significance of the morphogenic factor inthe host rhizosphere are discussed.     

Changes in patterns of protein synthesis during haustorial development of Striga hermonthica (Del.) Benth. seedlings

This study focuses upon the developmental transition of theparasitic plant Striga hermonthica from its freeliving state(germinated seedling) to its parasitic state after developmentof an infection organ: the haustorium. A new method has beendeveloped that allows the production of gram quantities of germinatedand haustorially-induced Striga seedlings, thereby facilitatingbiochemical and molecular analysis of haustorial induction.Water-soluble proteins have been extracted from germinated seeds(stage A) and seedlings treated with 2,6-dimethoxy-p-benzoquinone(2,6-DMBQ) to induce haustorium (stage B). Samples were analysedby two-dimensional polyacrylamide gel electrophoresis and quantitativeas well as qualitative differences could be observed. In particulara group of four highly abundant acidic proteins (molecular weight39 kDa, pl 5.1, 5.3, 5.3, 5.6) and three other proteins (molecularweight 12 kDa, pl 6.9; 17 kDa, pl 4.4; 17 kDa, pl 4.45) wereseen in stage A while at least four proteins (molecular weight21.5 kDa, pl 6.4; 21.5 kDa, pl 6.3; 31 kDa, pl 5.1; 34 kDa,pl 6.2) were present in greater abundance in stage B. In orderto compare watersoluble protein with newly synthesized proteinpatterns, mRNAs from the two stages of development were isolatedand cell-free translation products analysed by 2-D PAGE. Two-Dgels of cell-free translation products showed the appearanceof six proteins in stage B (molecular weight ranging from 10to 35 kDa) and the presence of three acidic proteins in stageA with one protein (molecular weight 40 kDa) very similar insize to the triplet of proteins in the water-soluble protein2-D gels. Key words: Striga hermonthica (Del.) Benth., haustorium, 2-D PAGE, 2,6-DMBQ, translation in vitro     

Uptake and Partitioning of Nitrogen by Maize Infected withStriga hermonthica

The uptake and partitioning of nitrogen (N) by maize infectedwith the parasitic angiosperm,Striga hermonthicawas investigatedin sand culture in a glasshouse. The purpose was to determinethe effect ofStrigaon N uptake and partitioning in maize. Maizewas grown at 22, 66 and 133 mg N per plant and sampled fivetimes. There was no significantStrigaxN interaction in any measuredresponse. Leaf dry matter ofStriga-infected maize, averagedover all N treatments, was 92% that of uninfected maize at thefour-leaf stage but by the 18-leaf stage it had decreased to58%. Similarly, stem dry matter of infected maize which was91% that of uninfected maize at the four-leaf stage was only42% at the 18-leaf stage. Root dry matter was similar for infectedand uninfected maize. N concentration in the leaf, stem androot declined asymptotically from the first to the last samplingdate for both infected and uninfected maize. The asymptoticvalue of N concentration inStriga-infected maize was 16% greaterin the leaf, 55% in the stem, and 21% in the root than in uninfectedmaize. The concentration of N inStrigawas higher than in maizeat the 16- and 18-leaf stages. Uptake of N was similar for infectedand uninfected plants at the four–eight leaf stage butat the eight–12 leaf stage, N uptake by infected maizewas 52% that of uninfected maize. However, the proportion oftotal plant nitrogen partitioned to the root was greater (P<0.001)forStriga-infected maize. These results showed that the extentto whichS. hermonthicareduced maize growth and N uptake, butincreased the proportion of N partitioned to the roots, didnot depend on the rate of N fertilizer applied.Copyright 1998Annals of Botany Company Maize; nitrogen; partitioning;Striga hermonthica; uptake.     

Structure and Development of the Primary Haustorium in Alectra vogelii Benth. (Scrophulariaceae)

Anatomical observations were made on the structure and developmentof the primary haustorium of Alectra vogelii. Its developmentinvolves a mutual aggressive growth of both the host and parasitetissues resulting in the formation of a very large and complextuberous organ. One of the host tissues whose growth is stimulatedby parasite infection is the pericycle whose cells divide repeatedlyand grow around and within the parasite haustorial cortex. Fromvarious points of the proliferating host pericycle, roots becomeinitiated and eventually the entire surface of the haustoriumbecomes covered with these roots. We have referred to them as‘haustorial roots’, a term which we have re-examinedand redefined. True xylary connections are established not onlybetween the parasite and the host root but also between theparasite and these ‘haustorial roots’. The uniquedevelopment of primary haustorium and ‘haustorial roots’in A. vogelii is discussed in relation to the development ofprimary haustoria in other root parasites.     

Haustorial Development and Growth Benefit to Seedlings of the Root Hemiparasitic Tree Nuytsia floribunda(Labill.) R.Br. in Association with Various Hosts

Dry matter gains and haustorial production of pot-cultured seedlingsof Nuytsia floribunda were assessed after a 12 month periodof association singly with each of a range of potential woodyhost species. One species,Adenanthos cygnorum , of similar sizeto most parasitized hosts, served as measure of response ofNuytsia in a non-benefiting situation. Rated on this basis,all 23 parasitized hosts elicited greater mean dry weights ofNuytsia than when on Adenanthos, and seven of these instanceswere highly significant. Numbers and weights of penetratingand presumably functional haustoria formed on a host were broadlycorrelated with growth benefit to Nuytsia, but there were notableinstances of unusually poor or great benefit from a host relativeto the complement of haustoria involved. Experiments in whichhaustoria-bearing associations of Nuytsia partnered with nodulatedAcacia hosts (Acacia acuminata and A. cyclops) were fed¹⁵N₂showedsignificant transfer of¹⁵N to the parasite, but failed to determinewhether the label had been acquired through haustoria or directlyby Nuytsia roots following turnover of nodule and root residuesof the host in the rooting medium. A parallel study using theunusual non-protein amino acid, djenkolic acid, as a markerof benefit from the djenkolic acid-containing host A. cyclops,showed appearance and progressive build-up of the compound infoliage of Nuytsia over a 6 month period after partnering thespecies in pot culture. Presence of the compound at final harvestin xylem sap of both partners but not in soil solution of thecultures strongly indicated xylem transfer via haustoria asthe principal avenue for N benefit to the parasite. Resultsare discussed in relation to a recent evaluation of haustorialstructure and functioning of N. floribunda. Copyright 2000 Annalsof Botany Company Root hemiparasite, Nuytsia, Loranthaceae, growth benefit, haustorial production, nitrogen transfer from hosts     

In Vitro Haustorium Development in Roots and Root Cultures of the Hemiparasitic Plant Triphysaria Versicolor

Parasitic plants in the Orobanchaceae invade host plant roots through root organs called haustoria. Parasite roots initiate haustorium development when exposed to specific secondary metabolites that are released into the rhizosphere by host plant roots. While molecular approaches are increasingly being taken to understand the genetic mechanism underlying these events, a limitation has been the lack of a transformation system for parasitic plants. Since the haustorium development occurs in roots of Orobanchaceae, root cultures may be suitable material for transient or stable transformation experiments. To this end, root cultures were obtained from explants, and subsequently calluses, from the hemiparasitic plant Triphysaria versicolor. The cultured roots retained their competence to form haustoria when exposed to host roots, host root exudates, or purified haustorium-inducing factors. The root culture haustoria invaded host roots and initiated a vascular continuity between the parasite and host roots. The ontogeny of haustoria development on root cultures was indistinguishable from that on seedlings roots. Root cultures should provide useful material for molecular studies of haustorium development.     

Haustorial Structure and Functioning of the Root Hemiparastic Tree Nuytsia floribunda(Labill.) R.Br. and Water Relationships with its Hosts

Observations on the origin and mature structure of the haustoriumof the Western Australian Christmas tree (Nuytsia floribunda)corroborate and extend the findings of earlier workers. We showthat the previously described sclerenchymatous ‘horn’or ‘prong’ formed within the haustorium acts asa sickle-like cutting device which transversely severs the hostroot and then becomes lodged in haustorial collar tissue directlyopposite to that where it originated. The cutting process isdeduced to be rapid and the gland-like fluid filled structurein the haustorium is suggested to generate a hydrostatic forcedriving the device through the host root. The haustorial parenchymacells at the tight junction between the endophytic part of thehaustorium and the cut face of the host root develop balloon-likeoutgrowths which intrude into the lumina of severed xylem vesselsof the host. Experiments feeding 0.05% (w/v) basic fuchsin tofreshly cut ends of host root segments distal to terminally-attachedmature haustoria demonstrate an apoplastic pathway from hostxylem elements fractured at the interface into haustorial parenchyma,and thence through vascular tissue to the haustorium into thetranspiring plant of Nuytsia. Application of labelled water(D₂O) to uncut basal roots of potted plants ofAcacia acuminataparasitized by Nuytsia results in labelling of leafy shootsof parasite and host, indicative of haustorial uptake of waterby Nuytsia from host root xylem in the intact association. Measurementsof xylem water potentials of pot-cultured seedling Nuytsia associatedwith a range of hosts, or of mature trees of Nuytsia and partnerwoody hosts in the native habitat, demonstrate consistentlymore negative potentials in the parasite than host, suggestingthat the parasite may regularly obtain xylem water through itshaustorial apparatus. Copyright 2000 Annals of Botany Company Root hemiparasite, Nuytsia floribunda, Loranthaceae, haustorial structure, host–parasite water relations     

Estimation of the sensitivity to photoinhibition in Striga hermonthica-infected sorghum

Sensitivity to photoinhibition was assessed in sorghum infectedwith the angiosperm root parasite Striga her-monthica and inuninfected sorghum plants, at four times during the developmentof the host-parasite association. Photoinhibition was inducedby exposing either leaf discs or intact leaves to a photosyntheticphoton flux density of 2000 µmol m^–2 s^–1 for4 h. The inhibition of apparent quantum yield (_a) and photosynthesisin high light (A₁₅₀₀) were assessed in leaf discs using an oxygenelectrode and the recovery of these from photoinhibition wasfollowed in intact leaves using an infra-red gas analyser. Fromsoon after attachment of the parasite, infected sorghum plantshad a lower A₁₅₀₀. During the period when Striga induced a loweringof A₁₅₀₀, _a was more sensitive to photoinhibition in Striga-infectedplants. However, at the same time, the high-light-induced inhibitionof A₁₅₀₀ was similar in Striga-infected and uninfected plants.Recovery of both _a and A₁₅₀₀ was incomplete after 6 h and thetime-course of recovery was similar in Striga-infected and uninfectedplants. The results indicate that Striga-infected plants weremore sensitive to photoinhibition and that photoinhibition wasprimarily due to damage to electron transport/photo-phosphorylationand not disablement of the recovery processes. Key words: Photoinhibition, quantum yield, recovery from photoinhibition, parasitic plants     

Chemical Induction and Repression of Haustoria in Orthocarpus Purpurascens (Scrophulariaceae)

A preliminary survey of compounds that induce haustoria in thehemiparasite Orthocarpus purpurascens suggests that cellulosefibre is one source of the active, water soluble factor(s).Haustoria are induced by extracts of soya bean seeds, flax seeds,plant roots, cotton string, cotton, paper towel, tissue paperand yeast extract. Sugars, hormones, nitrogenous compounds,microbial exudates, and citric and ascorbic acid do not inducehaustoria. Citric acid has a positive synergistic affect inthe presence of an inducing factor. Terramycin applied to seedlings 3 days of age had a detrimentaleffect on root development, while penicillin did not. Penicillinreduced the numbers of haustoria formed between 19 and 25 daysof age. The combined use of penicillin and inducing factorsmay provide an excellent experimental system for determiningthe temporal and developmental pattern of haustoria formation. haustoria, chemical induction and repression, Orthocarpus purpurascens, hemiparasite     

Molecular Specificity of Haustorial Induction in Agalinis purpurea (L.) Raf. (Scrophulariaceae)

An exogenous signal normally contained in host root exudateis required for initiation of the haustorium by the root parasiteAgalinis purpurea (L.) Raf. (Scrophulariaceae). Two flavonoidsthat induce haustoria have been isolated from gum tragacanthand a number of structural analogues have been synthesized.The results show that a high degree of molecular specificityis required for haustorial induction. Both isolated flavonoidscontain substituted 3-methoxyphenol functionality, and syntheticanalogues have shown that 4-substituted 3-methoxyphenol functionalityis critical for high levels of haustorial induction. These dataprovide a model for understanding host recognition at the levelof haustorial induction in parasitic angiosperms. Agalinis purpurea (L.) Raf. Scrophulariaceae, haustorial induction, flavonoids, molecular specificity, parasitic angiosperms, xenognosin     

Structure and Development of the Mature Secondary Haustorium in Alectra vogelii Benth

Anatomical investigations were carried out on the structureand development of the mature secondary haustorium in Alectravogelii growing on Arachis hypogaea or Vigna unguiculata. Followingthe formation of the young secondary haustorium, both the cambiumand pericycle of the host root directly opposite the young secondaryhaustorium are stimulated to divide and form new tissues andorgans including haustorial roots. Further proliferations ofthe host root pericycle and the haustorial cortex give riseto a large, tuberous and complex mature secondary haustoriumwithin which the tissues of the host and parasite remain inintimate contact forming a perfect graft union with a wide zoneof contact. Apart from the haustorial axial xylcm strand whichnormally connects the xylem of the parasite secondary root withthat of the host, direct xylary connections are also establishedbetween the axial xylem of the haustorium and the xylem of thehaustorial roots. The entire surface of the mature secondaryhaustorium of Alectrais covered with these haustorial rootsas was previously observed in its mature primary haustorium. Alectra vogelii Benth, secondary haustorium, haustorium, haustorial roots, root parasite, hemiparasitism, Arachis hypogaea, Vigna unguiculata     

Signaling Organogenesis in Parasitic Angiosperms: Xenognosin Generation, Perception, and Response

Abstract Parasitic strategies within the angiosperms generally succeed by tightly coupling developmental transitions with host recognition signals in a process referred to as xenognosis. Within the Scrophulariaceae, Striga asiatica is among the most studied and best understood parasitic member with respect to the processes of host recognition. Specific xenognosins regulate seed germination, the development of the host attachment organ, the haustorium, and several later stages of host-parasite integration. Here we discuss the signals regulating the development of the haustorium, the critical vegetative/parasitic transition in the life cycle of this obligate parasite. We provide evidence for the localized production of H₂O₂ at the Striga root tip and suggest how this oxidant is used to exploit host peroxidases and cell wall pectins to generate a simple benzoquinone signal. This benzoquinone xenognosin proves to be both necessary and sufficient for haustorial induction in cultured seedlings. Furthermore, evidence is provided that benzoquinone binding to a redox active site completes a ``redox circuit' to mediate signal perception. This redox reaction regulates the time-dependent expression of specific marker genes critical for the development of the mature host attachment organ. These studies extend the emerging series of events necessary for the molecular regulation of organogenesis within the parasitic plants and suggest novel signaling features and molecular mechanisms that may be common across higher plants.     

Control of Germination in Striga asiatica: Chemistry of Spatial Definition

Striga asiatica (Scrophulariaceae), a member of a heterogeneous group known as the parasitic plants, is totally dependent on host root attachment for survival. In agar, Striga seeds germinated in high percentages within 5 millimeters of a sorghum (Sorghum bicolor (L.) Moench) host root surface, and no germination was observed at distances greater than 1 centimeter. This spatially restricted germination may be explained by the chemistry of a single compound, 2-hydroxy-5-methoxy-3-[8′Z, 11′Z)-8′, 11′, 14′ -pentadecatriene]-p-hydroquinone, structure 1, which is exuded by sorghum roots. The presence of the compound was chemically imaged with pigments such as methylene blue. The use of methylene blue suggested that structure 1 was exuded along the entire surface of the root for long periods. This exudation and the inherent instability of structure 1 together establish an apparent steady state concentration gradient of the germination stimulant around the sorghum root. The Striga seed must be exposed to micromolar concentrations of 1 for ≥5 hours before high germination percentages were observed. Such a requirement for a long term exposure to a steady state concentration of an inherently labile, exuded compound would provide an extra degree of resolution to signal detection and host commitment in Striga parasitism.