Ultrastructural studies on the early cuticular metamorphosis of adult female Lernaeocera branchialis (L.) (Copepoda,Pennellidae)

After its final moult and fertilization an adult female of the marine fish-parasitic copepod, Lernaeocera branchialis, begins an extensive metamorphosis. This commences while the parasite is still on the flounder intermediate host and is completed once the female has established itself on the whiting final host. One early component of the metamorphosis is a considerable elongation of the parasite's abdominal region. S.e.m. and t.e.m. studies have revealed that part of the mechanism of the elongation consists of a straightening out of a highly folded abdominal cuticle. Before fertilization, the epicuticle and outer procuticular layers of this integument are thrown into a series of transverse, 4–6 µm deep pleats or folds with a density of 1–1.2 folds/µm of abdominal length. Straightening these folds can generate an approximately 6-fold length increase. The folds are already present beneath the female chalimus IV cuticle when the epidermis of this development stage starts to secrete the adult cuticle. Immediately before the final moult, the adult cuticle is super-folded with the whole cuticle displaying second-order folds, 8–10 µm deep.The capacity of Lernaeocera to engage in extensive cuticular modifications without recourse to a moult is compared with similar abilities shown by some insect species.     

Fine structure of the spermatheca of the mealworm beetle (Tenebrio molitor L.)

The spermatheca of the female mealworm beetle is an inflorescence of branching cuticular ducts which is connected to the bursa copulatrix via a cuticular neck surrounded by a muscular coat. The infolded bursal cuticle consists of a distinct outer epicuticle, inner epicuticle, procuticle, and a subcuticular zone; the latter is rich in mucopolysaccharides. The cuticle of the neck lacks a distinct procuticle. The cuticle of the spermatheca itself is mostly inner epicuticle with two thin underlying lamellae of procuticle. The cells of the bursa are loosely coupled to the procuticle, whereas cuticular projections bind the epithelia of the "neck" and the spermatheca proper to the underlying epithelia. The apical plasma membranes of the spermathecal epithelium are sinuous and much infolded; we believe that this epithelium controls the micro-environment within the cuticular ducts.     

Ultrastructure of the nymphal integument in the camel tick Hyalomma (Hyalomma) dromedarii (Ixodoidea: Ixodidae)

The ultrastructure of the nymphal integument in the ixodid tick Hyalomma (Hyalomma) dromedarii is compared for stages of development during and after feeding, and up to the first step of molting, apolysis. The integument comprises a cuticular layer and underlying epidermal cells. The body cuticle, which consists of both sclerotized and non-sclerotized parts, is divided into an outer, thin epicuticle, and an inner, thick, fibrillar procuticle. Pore canals in the procuticle are continuous with wax canals which traverse the epicuticle. As feeding progresses, the parallel, extensible epicuticular folds disappear due to the gut filling with ingested blood. The procuticular zone, however, becomes subdivided into an exocuticle, similar to the previously seen procuticle, and a lamellate endocuticle. Pore canals lose their parallel pattern and appear to have become deformed by stretching of the cuticle. The flat epidermal cells grow asynchronously during feeding; their cytoplasm becomes packed with well-developed rough endoplasmic reticulum (RER), while the cell apices project long microvilli extending deep into the procuticle. The RER undergoes ultrastructural changes indicating synthetic activity. Dense material released through the microvilli may serve to lyse the endocuticle and thus cause separation of the cuticle from the epidermis during apolysis. The lysed area, the exuvial cavity, is filled with lysed components which are probably withdrawn by endocytosis into the apical coated vesicles which appear in epidermal cells. Two types of integumental glands, which may participate in wax production, are observed in this study. The ultrastructure of their previously undescribed cuticular ducts is described, in addition to other hypodermal structures including epidermis-muscle attachments and sensory receptors.     

The fine structure of the carapace integument of Daphnia magna straus (Crustacea branchiopoda)

Summary The structure of the two integumental layers comprising the carapace of female D. magna was examined at several points through the molt cycle. The epicuticle and procuticle are simple in organisation; pore canals are absent but intracuticular fibres are present, forming complexes with invaginations of the epidermal plasma membrane similar to such complexes described in the literature for other arthropods. The epidermis consists almost entirely of cuticle-secreting cells. Secretion of the new cuticle begins when 50–67% of the instar has elapsed by which time the epidermal cells have increased in height and their nuclei have become more rounded. However, other presumed secretory phenomena observed viz. the formation of dense core vesicles by Golgi bodies, and the occurrence of these and coated vesicles near the apical plasma membrane are not restricted to any particular period during the molt cycle. This suggests that the mechanisms of cuticle secretion do not undergo marked changes in activity as they do in decapods; presumably this relative continuity is related to the much shorter molt cycle of cladocerans.The technical assistance of G.A. Bance, and the financial support provided by the National Research Council of Canada are gratefully acknowledged     

An ultrastructural study of the larval integument of the midge,Chironomus riparius meigen (Diptera: Chironomidae)

Summary The larval integument of the midge, Chironomus riparius Mg., is unusually thin although it conforms with the normal insect pattern. The cuticle of the post-cephalic segments is about 3 m thick and overlies an epidermis which has an irregular basal plasma membrane resulting in spaces occurring between it and the basement membrane. The ventral tubuli have a similar epidermis but the cuticle is somewhat thinner. The anal papillae have the thinnest cuticular covering with a uniquely folded epicuticle of variable thickness, and their epidermis has the characteristics of a transporting epithelium. No evidence of pore canals could be found in the cuticle of any part except the head capsule which has a remarkably smooth epicuticle and a distinct layer which may represent the exocuticle. There are no spaces between the basement membrane and basal plasma membrane of the epidermis in the head. Ultrastructural evidence would suggest that gaseous exchange can occur across most of the post-cephalic integument.The author is indebted to Mrs. L. Rolph and Mr. R.L. Jones for their technical assistance     

Cuticular adaptations in two parasitic copepods in relation to their modes of life

The structure, histochemistry, and possible functional properties of the cuticle in two parasitic copepods Pennella elegans Gnanamuthu and Caligus savala Gnanamuthu have been studied: the former is partially embedded in the host while the latter is an ectoparasite capable of free swimming.In Pennella elegans the cuticle of the embedded anterior region of the body is soft, colourless, and lacks an outer epicuticle while that of the posterior exposed part is pigmented and hard. Conspicuous in the cuticle of the ventral region of the head are pore canals which, though not chitinized, are functional even in the intermoult stage: these canals may be involved in the transport of nutrient materials from the host. The horns, which serve to fix the parasite firmly in the host tissues, are covered by cuticle in which the epicuticle and outer layers of the procuticle are hardened by formation of disulphide linkages. The cuticle of the neck region is not hardened and the procuticle in this region shows transverse regions of dense and light zones probably related to the coiling of the neck during penetration. The epicuticle is two layered in the cuticle of the exposed posterior region, the inner epicuticle and outer region of the procuticle being partially hardened by phenolic tanning so confer rigidity and resistance. The cuticle of the plumes is soft and devoid of an outer lipid epicuticle and so possibly adapted for a respiratory function.In Caligus savala, the epicuticle is two layered, and the procuticle has pigmented, calcified, and uncalcified layers. The cuticle is hardened by phenolic tanning as well as by calcification thus recalling the cuticular organization of decapod crustaceans.     

Ultrastructure of the integument during moulting of the quiescent tritonymphal instar of trombiculid mite Hirsutiella zachvatkini (Acariformes: Trombiculidae)

The ultrastructure of the integument of the quiescent reduced tritonymph of the trombiculid mite Hirsutiella zachvatkini (Schluger) was investigated by means of transmission electron microscopy. Mites were investigated daily during the 14–16 day tritonymphal period (imagochrysalis). This period includes the deutonymphal moult (1–3 days), the quiescent tritonymph period (2–4 days), and the tritonymphal moult into the adult mite (6–10 days). A distinct recognizable feature of the tritonymphal moulting cycle is a sequence of events independent of precise time intervals. This process involves partial destruction and reorganization of the hypodermis of the previous instar, and formation of a new hypodermis of the subsequent instar from islands of rudimentary hypodermal cells. The integument of the reduced tritonymph differs greatly from that of both larva and active deutonymph and adult. It consists of a simply organized hypodermal layer of varying thickness and a thick clear poorly lamellate cuticle with curved pore canals, and lacking setae. The epicuticle is very thin and without a clear protein layer. The tritonymphal instar as such with its own cuticle situated near the hypodermis is encased within the detached covering of the previous active deutonymph, and may be considered a calyptostasic and entirely pharate instar. There is a tendency for reduced tritonymphal stage to be eliminated from ontogenesis and this stage is not homologous to the pupa of insects.     

Etudes sur l'épicuticule des insectes

Résumé L'épicuticule de l'adulte de Tenebrio molitor est composée de deux couches distinctes dénommées épicuticule externe et épicuticule interne. L'épicuticule externe est la première couche cuticulaire sécrétée sous forme de petites plaques s'agrandissant par leurs bords pour recouvrir toute la surface cellulaire. Au moment de sa sécrétion, cette couche est formée de quatre lames denses A, B, C1 et C2. La lame B, très fine, disparaît par la suite et les lames C1 et C2 deviennent très nettes. L'épicuticule externe de l'adulte est donc formée de trois lames denses séparées par deux lames claires.L'épicuticule interne est formée de lames superposées denses et claires de structure complexe, qui sont masquées pendant la sécrétion des premières couches de cuticule lamellée (procuticule). Cette structure correspond à un arrangement moléculaire hautement organisé.La forme de la surface cuticulaire des sternites est déterminée par la forme de la surface cellulaire avant le dépôt de l'épicuticule.

---

The development of the epicuticle in the adult Tenebrio molitor L.

Summary The epicuticle of adult Tenebrio consists of two distinct layers named outer and inner epicuticle.The outer epicuticle is the first cuticular layer to be deposited in form of small patches on top of the microvilli. These initial patches are composed of four dense laminae (A, B, C1 and C2) separated by three light spaces. The outer epicuticle grows by densification of diffuse material at the edges of the patches until the entire area is covered.The thickness of outer epicuticle remains constant (175 Å) during the development of the pharate adult, lamina B however rapidly disappears. Thus, the adult outer epicuticle is fivelayered (three dense laminae: A, C1 and C2).After being deposited, the inner epicuticle shows a complex laminar structure interpreted to represent a highly organized molecular system. The laminae are masked during the formation of the first procuticle lamellae.During the deposition of the epicuticle, lamina A is covered by a component of the moulting fluid, forming an irregular dense layer which disappears after the resorption of this fluid. Perhaps this layer protects the new epicuticle from lytic enzymes of the moulting fluid.In adult animals, there is an additional superficial layer, the signification of which is not clear. The possibility of remains of cement or wax is discussed.The development of the surface patterns of the sternal and pleural cuticle is determined before the epicuticle formation by the shape of the epidermal surface. The rate of outer epicuticle deposition appears to depend on the size of the microvilli: epicuticle deposition seems to proceed faster over high microvilli.

Nous tenons à remercier notre directeur de recherche, le Professeur Noirot pour ses encouragements et ses conseils et Madame Curie pour son aide technique efficace.     

Ultrastructure and cytochemistry of the integument of Modiolicola gracilis,parasitic copepod in mussel gills (Mytilus galloprovincialis and Mytilus edulis)

Of mussels taken from the Ebro Delta River (E. Spain), 3% have a nonmodified copepod, Modiolicola gracilis, in the gill tissues. The cuticle of different segments of the body has an epicuticle with two layers, which show external microvilli-like projections. Weakly positive reactivity to the PTA technique has been detected in the external region. The procuticle has the helicoidal architecture of the chitinous tegument in arthropods, whereas the cuticle shows discontinuities in the regions of ducts in tegumental glands. The integument is comprised of three types of cells. Epidermal cells are flat with numerous mitochondria. Muscle cells show well-developed mitochondria with several longitudinally distributed cristae. A third and secretory cell shows a well-developed rough endoplasmic reticulum and Golgi complex in the basal zone. Its apical portion is full of secretory granules. Through the cuticle, these integumental glands open directly to the cuticular surface via a short duct coated by epicuticle. The composition and specializations of this complex cuticular architecture differ markedly from those shown by an endoparasitic copepod detected in the digestive gland of the mussel. It does not appear that the specializations detected in the cuticle of M. gracilis lead to any histopathological alteration in host tissues. © 1994 Wiley-Liss, Inc.     

The formation of the epicuticle and associated structures inOniscus asellus (Crustacea,Isopoda)

Summary The integument of the woodlouse,Oniscus asellus, consists of a two-layered epicuticle, a largely lamellate procuticle — itself divided into two regions (pre-and postecdysial cuticles), and the epidermis. At the initiation of new cuticle production the epidermal cells become vacuolated and retract away from the cuticle. Apolysis occurs immediately after the cessation of postecdysial cuticle production. The formation of the epicuticle is unique among the arthropods since material aggregates along the distal epidermal membrane. By indenting, doubling back on itself, and incorporating septa, the epicuticle forms surface structures such as plaques and tricorns.The innervation, and so the receptive function of the tricorns is confirmed, but since there is no connection between the old and new receptors during premoult, sensory information from these exoreceptors must be severely curtailed. This may explain the biphasic moult in all isopods since it ensures that only half the body experiences this sensory deprivation at any one time. In terrestrial species there is the additional advantage of restricting the area of permeable new cuticle. The frequency of moulting may be due to the need to renew disrupted receptor surfaces.Tricorns do not appear to be the mechanoreceptors involved in the marked thigmotactic response of woodlice since they do not have the typical internal structure of such receptors; rather, the dendrite —which extends into the lumen of the tricorn —is protected from deformation by the previously unreported combination of a dendritic sheath and a cuticular tube. The modality of tricorns is possibly one of hygro-perception. One of the behavioural responses of woodlice to desiccation is aggregation. The numerical distribution of tricorns over the body surface is admirably suited to assist in the formation and maintenance of such aggregates during desiccation and to their observed dispersal when the relative humidity rises.     

Micromechanics of smooth adhesive organs in stick insects: pads are mechanically anisotropic and softer towards the adhesive surface

Animals have evolved adhesive structures on their legs to cling to the substrate during locomotion. Here we characterise the ultrastructure and mechanical properties of adhesive pads in Carausius morosus (Phasmatodea) using atomic force microscopy (AFM) as well as transmission and scanning electron microscopy (TEM, SEM). The smooth adhesive arolium has a soft cuticle consisting of principal rods, which branch into finer fibres near the surface. Indentation experiments showed that the pad material consists of distinct layers with different mechanical properties. The 100–300 nm thick outermost layer consisting of the cuticulin envelope and the epicuticle is extremely soft and resilient (mean effective Young’s modulus 12 kPa), while the subjacent procuticle is a much stiffer material (mean effective Young’s modulus 625 kPa). AFM contact mode imaging revealed that the cuticle is mechanically anisotropic, which can be explained by its fibrillar inner structure. We propose that the described layered structure of smooth adhesive pads, consisting of materials decreasing in stiffness towards the outer surface, represents a superior design to conform and adhere to substrates with roughnesses at different length scales. This design principle could be easily implemented in technical adhesives, and thus has a potential to inspire biomimetic applications.     

Cuticular morphogenesis during continuous growth of the final instar larva of a moth

The larvae of the tobacco hornworm, Manduca sexta, grow continuously. During the feeding period of the fifth larval instar their weight increases ten-fold (ca. 1·2–12 g) accompanied by a four-fold expansion of the surface area of the abdominal cuticle. We have found that this cuticle contains structures which facilitate its expansion. Folds in the epicuticle (papillae) flatten as the cuticle expands. The endocuticle, in contrast, does not unfold but rather is plastically deformed. This plastic deformation is assisted by vertical structures in the cuticle (cuticular columns) which are more easily deformed than the surrounding lamellate cuticle. The head capsule cuticle, which does not expand as the larva grows, lacks papillae and cuticular columns. Thus, these are specialized structures that are reserved for cuticle that must expand as the larva grows.     

Hydrocarbon distribution and colony odour homogenisation in <Emphasis Type="Italic">Pachycondyla apicalis</Emphasis>

Summary Within and between individuals hydrocarbon (HC)-circulation was studied in Pachycondyla apicalis workers, using radioactive labeling. Newly synthesized HCs occurred both in the PPG and on the epicuticle in appreciable amounts, lesser quantities were found in the crop. The front basitarsal brush contained a greater amount of radiolabeled HCs than could be predicted from its surface area, suggesting preferential secretion to these organs. We propose that the newly synthesized HCs are secreted primarily to the front basitarsal brushes and are thereafter either distributed throughout the body surface, or cleared via the PPG and the alimentary canal.Using labeled HCs as a model, we tracked the time-dependent dispersion of cuticular lipids among 11 workers, one of which was prelabeled for 24 hours. Distribution among the recipients became progressively uniform, reaching near homogenization between 5–10 days. The mean HCs transfer of P. apicalis to the PPG was substantially lower compared to that of Camponotus fellah or Aphaenogaster senilis. In contrast, transfer to the cuticle in this species was superior. We attribute the low transfer to the PPG to the inefficacy of passive body contact characteristic of P. apicalis, as opposed to trophallaxis and/or allogrooming that typify the other two species. The higher occurrence of radiolabeled HCs in P. apicalis cuticle can be attributed to their accumulation in the basitarsal brushes. The impact of cuticular lipid transfer and formation of uniform colony odour, as opposed to the maintenance of an idiosyncratic caste-specific composition, are discussed.Received 5 September 2002; revised 17 January 2003; accepted 10 February 2003.     

Cuticle differentiation in the embryo of the amphipod crustacean Parhyale hawaiensis

The arthropod cuticle is a multilayered extracellular matrix produced by the epidermis during embryogenesis and moulting. Molecularly and histologically, cuticle differentiation has been extensively investigated in the embryo of the insect Drosophila melanogaster. To learn about the evolution of cuticle differentiation, we have studied the histology of cuticle differentiation during embryogenesis of the amphipod crustacean Parhyale hawaiensis, which had a common ancestor with Drosophila about 510 million years ago. The establishment of the layers of the Parhyale juvenile cuticle is largely governed by mechanisms observed in Drosophila, e.g. as in Drosophila, the synthesis and arrangement of chitin in the inner procuticle are separate processes. A major difference between the cuticle of Parhyale and Drosophila concerns the restructuring of the Parhyale dorsal epicuticle after deposition. In contrast to the uniform cuticle of the Drosophila larva, the Parhyale cuticle is subdivided into two regions, the ventral and the dorsal cuticles. Remarkably, the boundary between the ventral and dorsal cuticles is sharp suggesting active extracellular regionalisation. The present analysis of Parhyale cuticle differentiation should allow the characterisation of the cuticle-producing and -organising factors of Parhyale (by comparison with the branchiopod crustacean Daphnia pulex) in order to contribute to the elucidation of fundamental questions relevant to extracellular matrix organisation and differentiation. This work was supported by the German Research Foundation (DFG, grant number MO 1714/1-1).     

Electron microscopy of infection of nematodes byDactylaria haptotyla

Infection of nematodes byDactylaria haptotyla, a nematode-trapping hyphomycete, was studied by electron microscopy. The cytoplasm of the adhesive knob in the fungus contained a number of electron-dense, membrane-bound vesicles, 0.2–0.5 µm in diam. The vesicles were rarely seen in the stalk cell or vegetative cell cytoplasm. When the adhesive knob came into contact with the nematode's cuticle, it secreted an adhesive which was seen in ultrathin sections between the knob and the cuticle as an amorphous mass. At the same time, electron-dense vesicles in the cytoplasm were reduced in number and many small vacuoles developed. Soon after capture of a nematode, the cell wall of the adhesive knob became obscure at the prospective site of penetration, where a vesicle, 0.7 µm in diam, was found in serial thin sections of the knob's cytoplasm. At the site facing the vesicle, the peripheral part of the nematode's cell exhibited a high electron density. The vesicle, which appeared to be derived from smaller electron-dense vesicles coalesced with each other, released its enzymic contents toward the captured nematodes before penetration by the fungus.     

Rotifers as predators on small ciliates

Clearance rates of Synchaeta pectinata, Brachionus calyciflorus and Asplanchna girodi on Tetrahymena pyriformis (46 µm in length) at a density of 10 cells ml^–1, in the presence of algal food, were 2.5 to 6.1 ml rot.^–1 day^–1. Clearance rates of these rotifers were, respectively, about 2, 3, and 13 times lower on Strobilidium gyrans (58 µm in length) than on T. pyriformis, indicating that the saltations of S. gyrans are an effective escape response. Clearance rates of S. pectinata were considerably lower on Colpidium striatum (81 µm) than on S. gyrans, suggesting that S. pectinata may not be able to ingest ciliates of this size. S. pectinata had a clearance rate of 19 ml rot.^–1 day^–1 on S. gyrans at a density of 1.2 cells ml^–1, in the absence of edible algal food. Rotifers may prey extensively on ciliates in natural plankton communities, ingesting 25 to 50 individuals in the 45–60 µm size range day^–1.     

Development of plant cuticles: fine structure and cutin composition of Clivia miniata Reg. leaves

The fine structure and monomeric composition of the ester-cutin fraction (susceptible to BF₃/CH₃OH transesterification) of the adaxial leaf cuticle of Clivia miniata Reg. were studied in relation to leaf and cuticle development. Clivia leaves grow at their base such that cuticle and tissues increase in age from the base to the tip. The zone of maximum growth (cell expansion) was located between 1 and 4 cm from the base. During cell expansion, the projected surface area of the upper epidermal cells increased by a factor of nine. In the growth region the cuticle consists mainly of a polylamellate cuticle proper of 100–250 nm thickness. After cell expansion has ceased both the outer epidermal wall and the cuticle increase in thickness. Thickening of the cuticle is accomplished by interposition of a cuticular layer between the cuticle proper and the cell wall. The cuticular layer exhibits a reticulate fine structure and contributes most of the total mass of the cuticle at positions above 6 cm from the leaf base. The composition of ester cutin changed with the age of cuticles. In depolymerisates from young cuticles, 26 different monomers could be detected whereas in older ones their number decreased to 13. At all developmental stages, 9,16-/10,16-dihydroxyhexadecanoic acid (positional isomers not separated), 18-hydroxy-9-octadecenoic acid, 9,10,18-trihydroxyoctadecanoic acid and 9,10-epoxy-18-hydroxyoctadecanoic acid were most frequent with the epoxy alkanoic acid clearly predominating (47% at 16 cm). The results are discussed as to (i) the age dependence of cutin composition, (ii) the relationship between fine structure and composition, (iii) the composition of the cuticle proper, the cuticular layer and the non-depolymerizable cutin fraction, and (iv) the polymeric structure of cutin.Abbreviations CL cuticular layer - CP cuticle proper - MX cutin polymer matrix     

Cuticular and non-cuticular substrate influence on expression of cuticle-degrading enzymes from conidia of an entomopathogenic fungus,Nomuraea rileyi

Larval cuticle fromTrichoplusia ni, Helicoverpa (=Heliothis)zea, andHeliothis virescens and a cellulose substrate were used to quantify release of proteolytic, chitinolytic, and lipolytic enzymes by germinating conidia of the entomopathogenic fungus,Nomuraea rileyi. There was no significant difference in conidial viability incubated withT. ni, H. zea or cellulose substrates. Conidial viability onH. virescens cuticle, however, was significantly lower (ca. 19–25%) than the other three substrates. The presence of cuticle substrates, especially cuticle ofT. ni, stimulated germination. The nature of the substrate influenced both the time and quantity of the enzymes expressed. Specific proteases (aminopeptidase, chymoelastase, trypsin) generally were expressed earlier and/or in greater quantities on cuticular than on the cellulose substrate. Although both chitinolytic enzymes (endochitinase, N-acetylglucosaminidase) were detected on all three cuticular substrates, their activity was substantially lower than that of the proteolytic enzymes. Lipase activity was only minimally present. Early concurrent release of both proteases and chitinases suggested that both may be important in the penetration of the larval integument by germinating conidia ofN. rileyi. Expression of proteases and chitinases, especially aminopeptidase and endochitinase was probably a specific response to cuticle, because little or no activity was expressed on the non-host, cellulose substrate.This article reports the results of research only. Mention of a proprietary product in this paper does not constitute a recommendation for use by the US Department of Agriculture.     

The improvement and utilization in forestry of nitrogen fixation by actinorhizal plants with special reference toAlnus in Scotland

Summary Alnus species are used widely in Britain for land reclamation, forestry and other purposes. Rapid juvenile growth of the AmericanAlnus rubra makes it an attractive species for planting on N-deficient soils, particularly those of low organic content. In small plot trials, this species is nodulated by indigenous soil frankiae as effectively asAlnus glutinosa. Over a three year period both species return similar amounts of N to the ecosystem, estimated at up to 10–12 kg N ha^–1. Several strains ofFrankia have been isolated from local (Lennox Forest)A. rubra nodules. These differ morphologically and in their growth on different culture media, both from each other and fromA. glutinosa nodule isolates. AllAlnus isolates, however, have a total cellular fatty acid composition qualitatively similar to some other Group B frankiae. Glasshouse tests in N free culture suggest thatA. rubra nodules formed after inoculation of seedlings with American spore (–) isolates are three times more effective in N fixation than those inoculated with LennoxA. rubra spore (+) nodule homogenates. By contrast, the early growth of seedlings inoculated with spore (–)Frankia strains suggests at best a 35% improvement in N fixing activity over seedlings inoculated with LennoxA. rubra nodule isolates. Nevertheless, this improvement in activity, together with the better performance of seedlings inoculated with isolates compared with those treated with crushed nodule preparations, suggest that it would be worthwhile commercially to inoculate nursery stock with a spore (–)Frankia strain.     

Pollen morphology ofSonchus and related genera,and a general discussion

The pollen morphology of the taxa belonging to the generaAetheorhiza Cass.,Launaea Cass.,Reichardia Roth andSonchus L. in the Iberian Peninsula has been studied with light and electron microscopy. The pollen is 3(-4)-zonocolporate and echinolophate (without polar lacunae, but in general with prelacunae), with equatorial ridges and 15–20 lacunae: 3–4 poral, 6–8 abporal and 6–8 paraporal. Small to medium size, P × E = 19–36 × 23–42 µm; sometimes two different sizes have been found. Exine 3–9 µm thick and ornamentation microreticulate and echinate. The results clearly show the relationships between genera. Moreno-Socías, E., Mejías, J. A., Díez, M. J., 1994: Morfología polínica deLactuceae (Asteraceae) en la Península Ibérica, I.Lactuca y géneros relacionados. — Acta Bot. Malacitana.19: 103–113.