Occurrence of Belonolaimus in Sinaloa, Northwestern Mexico: A New Report on Distribution and Host Range.

The present study reports the occurrence of the genus Belonolaimus in the state of Sinaloa, Mexico, associated with native plants (i.e., Ziziphus amole and Stenocereus alamosensis) in a natural coastal ecosystem. Both morphological and molecular approaches were employed to characterize the Sinaloa population. Notwithstanding of some morphological and morphometric variation between Belonolaimus from Sinaloa and other valid species, the characterization indicates that this population might belong to the Belonolaimus longicaudatus species complex. Molecular analyses based on the 28S gene and ITS1-5.8S-ITS2 regions of the ribosomal RNA (rRNA) identified four major clades within Belonolaimus; however, none of the species including B. longicaudatus, B. gracilis, and B. euthychilus were supported as monophyletic; yet monophyly is argued to be a basic requirement of species status. Sequence divergence among different Belonolaimus populations and species varied according to the rRNA dataset (i.e., ITS1-5.8S-ITS2 > 28S > 18S) used, thus showing the importance of using genes with different rates of evolution to estimate species relationships. The fact that Belonolaimus has not been found in other cultivated (including on suitable hosts) areas in Sinaloa and that this population is relatively distant from the common B. longicaudatus groups (i.e., clades A and B) suggests that its appearance was not due to a recent introduction associated with the local agriculture.

Periosteum tissue engineering in an orthotopic in vivo platform.

The periosteum plays a critical role in bone homeostasis and regeneration. It contains a vascular component that provides vital blood supply to the cortical bone and an osteogenic niche that acts as a source of bone-forming cells. Periosteal grafts have shown promise in the regeneration of critical size defects, however their limited availability restricts their widespread clinical application. Only a small number of tissue-engineered periosteum constructs (TEPCs) have been reported in the literature. A current challenge in the development of appropriate TEPCs is a lack of pre-clinical models in which they can reliably be evaluated. In this study, we present a novel periosteum tissue engineering concept utilizing a multiphasic scaffold design in combination with different human cell types for periosteal regeneration in an orthotopic in vivo platform. Human endothelial and bone marrow mesenchymal stem cells (BM-MSCs) were used to mirror both the vascular and osteogenic niche respectively. Immunohistochemistry showed that the BM-MSCs maintained their undifferentiated phenotype. The human endothelial cells developed into mature vessels and connected to host vasculature. The addition of an in vitro engineered endothelial network increased vascularization in comparison to cell-free constructs. Altogether, the results showed that the human TEPC (hTEPC) successfully recapitulated the osteogenic and vascular niche of native periosteum, and that the presented orthotopic xenograft model provides a suitable in vivo environment for evaluating scaffold-based tissue engineering concepts exploiting human cells.

Evolution of plant parasitism among nematodes.

Despite extraordinary diversity of free-living species, a comparatively small fraction of nematodes are parasites of plants. These parasites represent at least three disparate clades in the nematode tree of life, as inferred from rRNA sequences. Plant parasites share functional similarities regarding feeding, but many similarities in feeding structures result from convergent evolution and have fundamentally different developmental origins. Although Tylenchida rRNA phylogenies are not fully resolved, they strongly support convergent evolution of sedentary endoparasitism and plant nurse cells in cyst and root-knot nematodes. This result has critical implications for using model systems and genomics to identify and characterize parasitism genes for representatives of this clade. Phylogenetic studies reveal that plant parasites have rich and complex evolutionary histories that involve multiple transitions to plant parasitism and the possible use of genes obtained by horizontal transfer from prokaryotes. Developing a fuller understanding of plant parasitism will require integrating more comprehensive and resolved phylogenies with appropriate choices of model organisms and comparative evolutionary methods.

Hematopoietic function in the elderly.

Very little information is available on the effect of aging on human bone marrow function. In reviewing both murine and human studies on bone marrow function in the elderly, it may be concluded at present that there is an increasing incidence of anemia that develops with aging in humans. Extensive animal and limited human data support decreased marrow reserve capacity with decreased hormonal responsiveness to hematologic stress as the most likely cause. In normal human life spans, marrow stem cells do not appear to wear out, and, finally, clinicians are cautioned not to attribute anemia per se simply to aging. Any elderly individual with unexplained anemia must be fully evaluated for the possibility of occult gastrointestinal blood loss, marrow failure syndrome, or hemolytic state before his/her anemia is attributed to age alone. Much additional research remains to be done on hematopoietic function in aged individuals.

Ultrastructure of the post-corpus of Zeldia punctata (Cephalobina) for analysis of the evolutionary framework of nematodes related to Caenorhabditis elegans (Rhabditina).

The ultrastructure of the post-corpus of Zeldia punctata (Cephalobina) was compared with previous observations of Caenorhabditis elegans (Rhabditina) and Diplenteron sp. (Diplogastrina) with the goal of interpreting the morphological evolution of the feeding structures in the Secernentea. The post-corpus of Z. punctata consists of six marginal, 13 muscle, five gland and seven nerve cells. The most anterior of four layers of muscle cells consists of six mononucleate cells in Z. punctata. The homologous layer in C. elegans and Diplenteron consists of three binucleate cells, suggesting a unique derived character (synapomorphy) shared between the Rhabditina and Diplogastrina. Contrary to Diplenteron sp. where we observed three oesophageal glands, Z. punctata and C. elegans have five oesophageal glands. We question this shared character as reflecting a common evolution between the Cephalobina and Rhabditina, because there are strong arguments for functional (adaptive) convergence of the five glands in these bacterial feeders. Convergence is further suggested by the mosaic distribution of three versus five glands throughout the Nemata; this distribution creates difficulties in establishing character polarity. Although morphological data are often laborious to recover and interpret, we nevertheless view 'reciprocal illumination' between molecular and morphological characters as the most promising and robust process for reconstructing the evolution of the Secernentea and its feeding structures.

True anemia: incidence and significance in the elderly.

The incidence of anemia increases as humans age, but in healthy aging individuals followed longitudinally, significant anemia does not develop in the absence of disease. Mild lymphopenia, as well as abnormal platelet function tests and increased coagulability when measured in vitro, also tend to develop in aging individuals. Hematologic neoplasia in the elderly behaves very similarly to the way it does in young individuals, but elderly individuals with hematologic neoplasms have a poor response to therapy.

Fine structure of body wall cuticle of females of Meloidodera charis, Atalodera lonicerae, and Sarisodera hydrophila (Heteroderidae).

The body wall cuticle of adult females of Meloidodera charis, Atolodera lonicerae, and Sarisodera hydrophila is examined by transmission electron and light microscopy for comparison with Heterodera schachtii and previous observations of additional species of Heterodera, Globodera, and Punctodera. The cuticle of M. charis is least complex, consisting of layers A, B, C (with A outermost), and varies in overall thickness from 3 to 8 mum. As in other species, the cuticle is thickest in mature specimens. The cuticle of A. lonicerae is 6-9 mum thick; unlike M. charis it has an innermost layer, D, in addition to A, B, and C. The cuticle of S. hydrophila varies from 14 to 30 mum thick and includes a D layer similar to A. lonicerae; layer C is subdivided into additional zones relative to other heteroderids, and the external portion of the cuticle is infused with an electron-dense material. The presence of a D layer in A. lonicerae and S. hydrophila is a character state which is shared with Globodera spp. and Punctodera sp. The electron-dense material in the outer layers of S. hydrophila also occurs in Globodera spp. and Punctodera sp. On the other hand, H. schachtii resembles other Heterodera spp. as well as M. charis by the absence of a D layer and lack of electron-dense material in the outer layers. The pattern of occurrence of shared character states, including those of the cuticle, may be useful for phylogenetic analysis of Heteroderidae.

Fine Structure of the Esophagus of Males of Sarisodera hydrophila (Heteroderoidea).

The fine structure of the esophagus, including procorpus, metacorpus, isthmus, gland lobe, and esophago-intestinal junction, is examined in males of Sarisodera hydrophila. A cuticle-lined lumen extends most of the length of the esophagus, broadens to form a pump chamber in the metacorpus, and posteriorly is continuous with junctional complexes among four esophago-intestinal cells. These four cells are partially enveloped by the gland lobe which basically consists of three gland cells, one dorsal and two subventral. Each gland cell has an anterior process which opens into the lumen of the esophagus through a cuticle-lined duct. The dorsal gland joins the lumen in the anterior portion of the procorpus, whereas ducts of the subventral glands terminate at the base of the metacorpus pump chamber. The subventral glands are predominant in the posterior portion of the gland lobe and are partially ensheathed by a narrow portion of the dorsal gland which extends to within 5 mum of the posterior terminus of the gland lobe. Contents of the dorsal gland include primarily electron dense granules, although rough endoplasmic reticulum (RER) is predominant posteriorly. Secretory granules within the subventral glands vary in morphology and are evenly distributed throughout the two ceils among other organelles, including RER and a large Golgi apparatus. Innervation of the esophagus includes nerve processes which originate from several perikaryons (cell bodies) located in the anterior portion of the gland lobe. The esophagus of males of S. hydrophila is compared with that of other Heteroderoidea, Heterodera glycines and Meloidogyne incognita.

Host-Parasite Relationships of Atalodera spp. (Heteroderidae).

Atalodera ucri, Wouts and Sher, 1971, and A. lonicerae, (Wouts, 1973) Luc et al., 1978, induce similar multinucleate syncytia in roots of golden bush and honeysuckle, respectively. The syncytium is initiated in the cortex; as it expands, it includes several partially delimited syncytial units and distorts vascular tissue. Outer walls of the syncytium are relatively smooth and thickest near the feeding site of the nematode; inner walls are interrupted by perforations which enlarge as syncytial units increase in size. The cytoplasm of the syncytium is granular and includes numerous plastids, mitochondria, vacuoles, Golgi, and a complex network of membranes. Nuclei are greatly enlarged and amoeboid in shape. Although more than one nucleus sometimes occur in a given syncytial unit, no mitotic activity was observed. Syncytia induced by species of Atalodera chiefly differ from those of Heterodera sensu lato by the absence of cell wall ingrowths; wall ingrowths increase solute transport and characterize transfer cells. In syncytia of Atalodera spp., a high incidence of pits and pit fields in walls adjacent to vasctdar elements suggests that in this case plasmodesmata provide the pathway for increased entry of sohttes. The formation of a syncytium by species of Atalodera and Heterodera sensu lato, but a single uninucleate giant cell by Sarisodera and Hylonema, indicates a pattern of host responses that may be useful, with other characters, for phylogenetic inference for Heteroderidae.

Host Response to Sarisodera hydrophila Wouts and Sher, 1971.

The histopathology of two populations of Sarisodera hydrophila Wouts and Sher, 1971 was examined on Salix lasiolepis Benth. (willow), Populus fremontii Wats. (cottonwood), and Lyonothamnus floribundus Gray (ironwood) using light microscopy as well as scanning and transmission electron microscopy. Sarisodera hydrophila induces formation of a single uninucleate hypertrophied cell (giant cell) which varies only slightly among the three hosts. The giant cell is enclosed by the root stele and contacts phloem, vascular cambium, and xylem. The single hypertrophied nucleus of the giant cell is ameboid or lobulate in shape, generally with a single nucleolus. The cell is characterized by a wall which is separated into two distinct regions about 2 mum and 13 mum thick; the thicker region occurs adjacent to the nematode head. Cell wall ingrowths, such as those associated with host responses to certain other plant-parasitic nematodes, were not observed in giant cells induced by S. hydrophila. However, a high frequency of pit fields with plasmodesmata occurred in the thinner portion of the cell wall which is adjacent to vascular elements. Roots of the three hosts simultaneously infected with S. hydrophila and Meloidogyne sp. resulted in adjacent responses characteristic of each nematode, supporting the view that the specific type of host response is a function of the nematode rather than the host. The varying expressions of host responses among Heteroderoidea may be useful in testing congruency with existing interpretations of phylogeny.

Host Response to Meloidodera spp. (Heteroderidae).

Host responses to Meloidodera floridensis Chitwood et al., 1956, M. charis Hooper, 1960, and M. belli Wouts, 1973 were examined on loblolly pine, peony, and sage, respectively, with light, scanning, and transmission electron microscopy. In each case the nematodes induce a single uninucleate giant cell. The giant cell is initiated in the pericycle and expands as it matures. The mature giant cell induced by M. floridensis is surrounded by vascular parenchyma, whereas that caused by M. charts and M. belli coutacts xylem and phloem. The cell wall of giant cells induced by all three Meloidodera spp. is generally thicker than that of surrounding cells, with the thickest part adjacent to the lip region of the nematode. The thinner portion of the wall includes numerous pit fields with plasmodesmata, but wall ingrowths were not detected in a thorough examination of the entire wall. The nucleus of a giant cell induced by M. goridensis is highly irregular in shape with deep invaginations, whereas those caused by M. charis and M. belli include a cluster of apparently interconnected nuclear units. Organelles, including mitochondria, endoplasmic reticulum, and plastids of giant cells caused by Meloidodera, are typical of those reported in host responses of other Heteroderidae. The formation of a single uninucleate giant cell by Meloidodera, Cryphodera, Hylonerna, and Sarisodera, but a syncytium by Atalodera and Heterodera sensu lato, might be considered in conjunction with additional characters to determine the most parsimonious pattern of phylogeny of Heteroderidae.

Cactodera eremica n. sp., Afenestrata africana (Luc et al., 1973) n. gen., n. comb., and an Emended Diagnosis of Sarisodera Wouts and Sher, 1971 (Heteroderidae).

Systematic contributions to Heteroderidae include description of Cactodera eremica n. sp., an emended diagnosis of Sarisodera Wouts and Sher, 1971, and proposal of a new genus and new combination, Afenestrata africana (synonym Sarisodera africana Luc et al., 1973). Cactodera eremica, from the roots of shadscale in Utah, most closely resembles Cactodera thornei (Golden and Raski, 1977) but differs by the presence of a finely striated cuticle, a fine surface pattern on eggs, a shorter female stylet, distance of the DGO from the stylet, vulval slit, and smaller diameter of circumfenestra, as well as a shorter tail in second-stage juveniles. The response of the host to C. eremica is similar to other Heterodera sensu lato including a large syncytium with wall ingrowths. The diagnosis of Sarisodera is emended to exclude cysts, which do not form in the type species, S. hydrophila. Afenestrata africana differs from S. hydrophila by the formation of cysts, the dorsal position of the anus in females, the shorter stylet, and a pore-like phasmid opening in second-stage juveniles. In addition, the lip pattern of males and juveniles is characterized by a greater degree of fusion of lip parts, the host response is a syncytium (versus a single uninucleate giant cell in S. hydrophila), and the cuticle is thinner and lacks a D layer. Unlike Heterodera, the cyst of Afenestrata lacks fenestrae.

Fine structure of body wall cuticle of females of eight genera of heteroderidae.

Body wall cuticle of adult females of eight genera within the Heteroderidae was examined by transmission electron microscopy for comparison with previously studied species within the family. Cuticle structure was used to test some current hypotheses of phylogeny of Heteroderidae and to evaluate intrageneric variability in cuticle layering. Verutus, Rhizonema, and Meloidodera possess striated cuticle surfaces and have the simplest layering, suggesting that striations have not necessarily arisen repeatedly in Heteroderidae through convergent or parallel evolution. Atalodera and Thecavermiculatus possess similar cuticles with derived characteristics, strengthening the hypothesis that the two genera are sister groups. Similarly, the cuticle of Cactodera resembles the specialized cuticle of Globodera and Punctodera in having a basal layer (D) and a surface layer infused with electron-dense substance. Heterodera betulae has a unique cuticle in which the thickest layer (C) is infiltrated with an electron-dense matrix. Little intrageneric difference was found between cuticles of two species of Meloidodera or between two species of Atalodera. However, Atalodera ucri has a basal layer (E) not found in other Heteroderidae. The most striking intrageneric variation in cuticle structure was observed between the thin three-layered cuticle of Sarisodera africana and the much thicker four-layered cuticle of Sarisodera hydrophila; results do not support monophyly of Sarisodera.

Comparative Morphology of Meloidodera spp. and Verutus sp. (Heteroderidae) with Scanning Electron Microscopy.

Scanning electron microscopy (SEM) of second-stage juveniles (J2), males, and females of Meloidodera floridensis, M. charis, M. belli, and Verutus volvingentis reveals detailed characteristics of the head region, lateral field, phasmid, body striae, vulva, and perineal region. In M. charis and M. belli the en face pattern conforms to a basic pattern in which the labial disc is surrounded by six lips (sectors) of the first head annulation. In J2 the head has additional annulations, whereas in males annulation is replaced by longitudinal blocks. Conversely, J2 and males of M. floridensis and V. volvingentis each have a unique derived face pattern with fusion of various lip components and with head annulation. All six lips of females of M. charis and M. belli are fused, whereas females of M. floridensis and V. volvingentis have distinct lateral lips. Lateral fields vary among species, with only slight differences at the anterior and posterior ends of the lateral lines and in the spatial relation of the lines to phasmid openings. Phasmid openings are present in adults of Meloidodera spp., but were not observed in adults of V. volvingentis; in this respect, the female perineal pattern of Verutus is different from Meloidodera spp, The very large vulva (+/- 48 mum long) of V. volvingentis is in sharp contrast to the minute vulva (+/- 6 mum long) in a population of M. charis from San Bernardino. Morphological characters revealed by SEM will be most informative when investigated throughout Heteroderidae and incorporated with additional characters for a phylogenetic analysis of the family.

Comparative fine structure of the stomatal region of males of Meloidogyne incognita and Heterodera glycines.

The fine structure of the stylet, "guiding apparatus," anti protractor muscles of males of Meloidogyne incognita and Heterodera glycines is ehtcittated and compared. In both nematodes, the stylet cone is of greater electron density than the shaft. The cone is heterogeneous; shaft material extends into it for most of its length, whereas the shaft proper and knob regions are relatively homogeneous. The stylet lumnen is round throughout its length in H. glycines, but in M. incognita changes from round, posteriorly, to irregular, triradiate, and oblong near the styler tip. The various layers of the lining of the stylet shaft are continuous with those of the cuticular lining of the esophageal lumen. Similarly, cuticular layers of the stomatal lining and vestibule extension are continuous with the body wall cuticle. The stylet "guiding apparatus" is formed by linings of the stomatal opening, vestibule, and vestibule extension, together with transversely folded membranes, which extend further posteriorly and attach near the junction of shaft and cone. Noncontractile regions of the three stylet protractor muscles originate in the esophagus. Contractile portions extend anteriorly from the stylet knobs and branch into a total of 10 elements that attach near the basal ring of the cephalic framework and the vestibule extension. A second, noncontractile region is present at the anteriormost part of the protractor muscles in H. glycines.

Fine Structure of Cephalic Sense Organs in Heterodera glycines Males.

Cephalic sense organs of Heterodera glycines males were examined in detail by electron microscopy. Each amphid basically consists of an amphidial gland, a nerve bundle, and an amphidial duct. The amphidial gland consists of a microvillous region, and laterally is closely associated with a large secretory cell. The nerve bundle penetrates the microvillous region, and further anteriorly individual nerve processes (dendrites) separate from one another, thus forming a sensilla pouch which is enveloped by the microvillous region of the gland. Anterior to the pouch, the cilia-like dendrites converge as they enter and eventually terminate in the amphidial duct. Heterodera glvcines males have the innervation basis for a full complement of sixteen papillae, although surface manifestations are present for only six minute inner labial papillae. In addition, four outer labial and four cephalic receptors terminate beneath the surface, and another two dendrite pairs end further posteriorly beneath the basal plate of the cephalic framework. Papillary receptors which terminate beneath the surface are probably mechanoreceptive, whereas inner labial papillae have pore-like openings to the exterior and may be chemoreceptive. Amphids and papillae of H. glycines are fundamentally similar to those of Meloidogyne incognita, although certain striking differences exist.

Body Wall Fine Structure of the Anterior Region of Meloidogyne incognita and Heterodera glycines Males.

The body wall fine structure including the cuticle, hypodermis, and somatic muscles is similar in males of Meloidogyne incognita and Heterodera glycines. The cuticle can be regarded as basically three-layered in both species, but is much thicker in M. incognita than in H. glycines, and differences occur in surface markings. The chordal and interchordal hypodermis is syncytial. Hypodermal tissue pervades the lip region, and lines the stomatal cavity and stylet shaft. Various organelles and structures, some previously undescribed, are concentrated in the chords. Their possible role in lipid metabolism is considered, as well as the probable function of the hypodermis in fornlation of the cephalic framework and stylet. The interchordal hypodermis which encloses peripheral nerves, is periodically transversed by bundles of fibrils which are homologous with the subcuticular striation previously observed in the light microscope. The somatic musculature is meromyarian, and the muscle cells are of the platymyarian type with I, A, and H bands, but without Z bands or T tubules. Thin dense bands are present in the H bands, and appear to be associated with sarcoplasmic reticulum.

Ultrastructure of Phasmid Development in Meloidodera floridensis and M. charis (Heteroderinae).

Phylogenetic characters for Heteroderinae Luc. et al., 1988 are evaluated in Meloidodera which is believed to have primarily ancestral characters. Phasmid ultrastructure is observed in second-stage juveniles (J2), third-stage juvenile males, fourth-stage juvenile males, and fifth-stage males of Meloidodera floridensis and M. charis. Phasmid secretion occurs inside the egg before the J1-J2 molt. Before J2 hatch, concentric lamellar membranes occur within the sheath and socket cells. Some membranes become lamellae of the sheath cell plasma membrane; others become multilamellar bodies. During early molting, plasma membrane lamellae disappear and a distal dendrite segment appears in a rudimentary canal. After the molt, the distal dendrite is not present within the canal. The phylogenetic utility of phasmid features is discussed. In both species the ampulla shape and size between molts are stable features in juveniles and males. The posthatch J2 sheath cell receptor cavity may vary in a species specific manner, but comparative morphology requires precise timing after hatch.

Phylogenetic implications of phasmid absence in males of three genera in heteroderinae.

Absence of the phasmid was demonstrated with the transmission electron microscope in immature third-stage (M3) and fourth-stage (M4) males and mature fifth-stage males (M5) of Heterodera schachtii, M3 and M4 of Verutus volvingentis, and M5 of Cactodera eremica. This absence was supported by the lack of phasmid staining with Coomassie blue and cobalt sulfide. All phasmid structures, except the canal and ampulla, were absent in the postpenetration second-stage juvenile (J2) of H. schachtii. The prepenetration V. volvingentis J2 differs from H. schachtii by having only a canal remnant and no ampulla. This and parsimonious evidence suggest that these two types of phasmids probably evolved in parallel, although ampulla and receptor cavity shape are similar. Absence of the male phasmid throughout development might be associated with an amphimictic mode of reproduction. Phasmid function is discussed, and female pheromone reception ruled out. Variations in ampulla shape are evaluated as phylogenetic character states within the Heteroderinae and putative phylogenetic outgroup Hoplolaimidae.

Fine Structure of Sperm of Ekphymatodera thomasoni (Heteroderinae, Nemata).

Fine structure of developing sperm of the monospecific genus, Ekphymatodera, was compared with other Heteroderinae as part of a study to recognize diversity and phylogenetically informative characters within the subfamily. Sperm of Ekphymatodera originate from germ cells connected to a central rachis, a character which is shared with Globodera, but not with other Heteoderinae. In Ekphymatodera, and cyst-forming genera, a layer of cortical microtubules lies just beneath the surface of the plasma membrane. Sperm of Ekphymatodera are unique among Heteroderinae examined by the presence of spiral surface elevations on the filopodia, a character that may prove to be a synapomorphy for Sarisoderini. Fibrous bodies are abundant in spermatids; however, they do not persist in sperm of Ekphymatodera as they do in Meloidodera and Verutus. The male gonad of Ekphymatodera is lined by epithelial cells, which are greatly enlarged near the ejaculatory canal. These enlarged cells contain vesicles with concentric lamellar inclusions, not observed in other genera of the subfamily. Sperm of Heteroderinae are rich in diversity, and examination of additional representative species may indicate new phylogenetically informative characters.