Ultrastructural changes during desiccation of the anhydrobiotic nematode Ditylenchus dipsaci

Ultrastructural changes during desiccation of the anhydrobiotic nematode Ditylenchus dipsaci were followed and quantified after preparation of material at different levels of hydration using freeze substitution techniques. Some shrinkage was caused by processing in the more hydrated specimens but the changes observed correspond to those observed in live nematodes by light microscopy, indicating that the technique is useful for following changes during desiccation. The overall pattern of changes was a rapid decrease in the magnitude of the measured parameter during the first 5 min of desiccation, followed by a slower rate of decrease upon further desiccation. This was observed in the cuticle, the lateral hypodermal cords and the muscle cells and is consistent with the pattern of water loss of the nematode. The contractile region of the muscle cells, however, proved an exception and the muscle fibres appear to resist shrinkage and packing until water loss becomes severe. The mitochondria swell and then shrink during desiccation, which may indicate disruption of the permeability of the mitochondrial membrane. A decrease in the thickness of the cortical zone was the most prominent change in the cuticle and this may be related to the permeability slump which occurs during the first 5 min of desiccation.     

Microtopography of Nippostrongylus brasiliensis (Nematoda: Heligmosomatidae): Free-living larval stages

Microtopographic features of the various growth stages of the three free-living larval stages of the rat hookworm Nippostrongylus brasiliensis (Nematoda) were surveyed by scanning electron microscopy. These worms have a rounded anterior end and an elongated tail. Cuticular annulations were observed along the body, which also bore two ribbon-like lateral alae. Two rings of six lip-like lappets were observed around the triradiate oral opening in all larval stages. The cephalic space contained two lateral amphidial pits. The excretory pore in the third anterior part was observed in a ventral view of the larvae. No deirids were observed. The anus with a crescent-shape opening was located posteriorly. Phasmidial apertures, only observed in the third-stage larvae, opened on the lateral alae in the tail region. © 1993 Wiley-Liss, Inc.     

Larval nematodes (Spiruroidea: Cystidicolidae) in Octopus vulgaris (Mollusca: Cephalopoda: Octopodidae) from the northeastern Atlantic Ocean.

Larval nematode parasites (Spiruroidea: Cystidicolidae) are recorded for the first time in Octopus vulgaris Cuvier, 1797 in the northeastern Atlantic Ocean. Prevalence was 16% and mean intensity was 1.46 worms/host. Body length of larval nematodes ranges from 8.3 to 9.3 mm, with a distance from the anterior end to nerve ring from 187.5 to 200 microm, and to excretory pore 194.6-350 microm. Anatomical characteristics, such as deirid, nerve ring, cephalic alae, excretory pore, pseudolabia amphids, sclerotized protuberance, and anus, examined using light microscopy (LM) or scanning electron microscopy (SEM), are illustrated. The nematode was designed as a cystidicolid "Type A" larva. The hemocytic infiltration present in the host tissue around the nematode capsule and the mechanical compression in the infected organs denote the pathogenicity of this nematode. In the study area, O. vulgaris may play the role of an intermediate or paratenic host in the nematode life cycle.     

Atomic Force Microscopy of Plant-Parasitic Nematodes

A simple method for atomic force microscopy (AFM) of nematode cuticle was developed to visualize the external topography of Helicotylenchus lobus, Meloidogyne javanica, M. incognita, and Xiphinema diversicaudatum. Endospores of two isolates of the nematode parasite, Pasteuria penetrans, adhering to M. incognita and X. diversicaudatum were also visualized and measured by this technique. Scanning procedures were applied to specimens killed and dehydrated in air or dehydrated and stored in glycerol. Atomic force microscopy scanning of nematodes in constant height mode yielded replicated high-resolution images of the cuticle showing anatomical details such as annulations and lateral fields. Submicrometer scale images allowed the identification of planar regions for further higher resolution scans.     

A new genus of Cystidicolid nematode from the stomach of Galaxias maculatus (Osmeriformes: Galaxiidae) in Patagonia (Argentina)

During a parasitological survey of Galaxias maculatus (Jenyns) from Patagonian Andean Lakes, specimens of a new species of nematode were collected from the stomach of fish and studied by light and scanning electron microscopy. This species is described as the only member of a newly proposed genus of Cystidicolidae by having oral opening dorsoventrally elongated, 2 lateral pseudolabia, 4 cephalic papillae, stoma (vestibule) elongated, esophagus divided into anterior muscular and posterior glandular sections, and caudal alae in males. Placonema n. gen. (Habronematoidea, Cystidicolidae) is characterized by the combination of the following features: oral opening dorsoventrally elongated demarcated by 4 sclerotized plates and 2 well-developed pseudolabia projected to the buccal cavity, each pseudolabium with conspicuous, conical, anterior protuberances. Four cephalic papillae and deirids simple. Male with caudal alae, area rugosa absent, 4 pairs of preanal papillae, unpaired papilla present on anterior cloacal lip, and 6 pairs of postanal papillae. Larvigerous eggs without filaments. Placonema pataguense n. gen. n. sp. infects the stomach of G. maculatus from Lake Patagua (Patagonia, Argentina) and is the first species of Cystidicolidae described from G. maculatus.     

The tardigrade cuticle II. Evidence for a dehydration-dependent permeability barrier in the intracuticle

Tardigrades weighed during desiccation in high or low humidities show a short period of rapid transpiration followed by an abrupt decline in transpiration which virtually arrests water loss. The amount of water retained following this 'permeability slump' is greater at low rates of desiccation but the slump is not a metabolic phenomenon, being reproducible in dead or narcotised animals. Tardigrades rinsed in hot chloroform (62 degrees C) for 5 hr still show the characteristic permeability decline when desiccated in 80% RH. However, 25hr rinsing in hot chloroform apparently obliterates the slump. Estimates of the bound water content of tardigrades by DSC show that this can account for the dehydrated masses of these chloroform-rinsed animals and that all free water is probably transpired. Lipid analysis of the 25 hr chloroform extracts by GC-MS reveals several lipid classes, predominantly free fatty acids (C(12)-C(18)); these are not detectable in the 5 hr extracts. Control rinsing in hot water has no apparent effect on the permeability slump. TEM tracer studies with lanthanum show the lipid-rich intracuticle to serve as a transpiration barrier in dehydrated animals but not in fully hydrated specimens. There is thus strong support for the role of intracuticular lipids in effecting the permeability slump. A model to explain this phenomenon on the basis of lipid phase changes is postulated.     

Increased Water Activity Reduces the Thermal Resistance of Salmonella enterica in Peanut Butter

Increased water activity in peanut butter significantly (P < 0.05) reduced the heat resistance of desiccation-stressed Salmonella enterica serotypes treated at 90°C. The difference in thermal resistance was less notable when strains were treated at 126°C. Using scanning electron microscopy, we observed minor morphological changes of S. enterica cells resulting from desiccation and rehydration processes in peanut oil.     

Morphological response of a bdelloid rotifer to desiccation

We desiccated bdelloid rotifers (Macrotrachela quadricornifera), submitting the animals to four desiccation procedures (protocols A, B, C, D) that differed in the rate of water evaporation, in the time of desiccation, and in the substrates provided. We observed external morphological changes of the rotifer bodies during drying with scanning electron microscopy and, in parallel, assessed rates of recovery after a 7-day period of dormancy. Two protocols produced disorganized morphologies of the anhydrobiotic animals, with no (A) or very poor (B) recovery. Protocols C and D gave rather high rates of recovery and dry rotifers appeared unaltered and well organized. The different protocols affected rotifer morphology during the 7-day anhydrobiosis and rates of recovery after the 7-day anhydrobiosis; high recovery rates corresponded to well-organized morphologies of anhydrobiotic bdelloids, suggesting that a proper contraction of the body into a tun shape and probably a rigorous packing of internal structures are necessary for survival after anhydrobiosis. These features are affected by the time between water shortage and full desiccation, but also by the surrounding relative humidity and by the nature of the substrate. Possible adaptations of anhydrobiotic rotifers are discussed.     

A note on the development of a non-aldehyde fixation technique for examining spores of Bacillus subtilis under the electron microscope

Various techniques were studied for fixing spores of Bacillus subtilis prior to examining them by transmission or scanning electron microscopy. A non-aldehyde technique employing carbodiimide in cacodylate buffer produced excellent results and could be of value in studying the cytological changes produced in spores exposed to inimical treatments.     

Reliable diagnosis of Trichosomoides crassicauda in the urinary bladder of the rat

Two reliable methods are described for identifying infection of laboratory rats with the nematode Trichosomoides crassicauda. The first is a rapid method where cryostat sections of the rat urinary bladder are stained with acridine orange and viewed under a fluorescence microscope. The second involves the stabilization of the bladder surface prior to examination using scanning electron microscopy (SEM).     

Volume and morphology changes of a bdelloid rotifer species (Macrotrachela quadricornifera) during anhydrobiosis

Following a study on the changes occurring in a bdelloid species (Macrotrachela quadricornifera, Rotifera, Bdelloidea) when entering anhydrobiosis, we investigated the changes in morphology, including weight and volume during the transition from the active hydrated to the dormant anhydrobiotic state by scanning electron microscopy, confocal microscopy and light microscopy. We compared sizes and morphologies of hydrated extended, hydrated contracted and anhydrobiotic specimens. Bdelloid musculature is defined: longitudinal muscles are contracted in the hydrated contracted animal (head and foot are retracted inside the trunk), but appear loose in the anhydrobiotic animal. When anhydrobiotic, M. quadricornifera appears much smaller in size, with a volume reduction of about 60% of the hydrated volume, and its internal organization undergoes remarkable modifications. Internal body cavities, clearly distinguishable in the hydrated extended and contracted specimens, are no longer visible in the anhydrobiotic specimen. Concomitantly, M. quadricornifera loses more than 95% of its weight when anhydrobiotic; this is more than expected from the volume reduction data and could indicate the presence of space-filling molecular species in the dehydrated animal. We estimate that the majority of body mass loss and volume reduction can be ascribed to the water loss from the body cavity during desiccation.     

Survival analyses of symbionts isolated from <Emphasis Type="Italic">Endocarpon pusillum</Emphasis> Hedwig to desiccation and starvation stress

This work deals with the survival analyses of the symbionts isolated from the lichen E. pusillum under desiccation and starvation stress. The mycobiont of the symbionts was under the desiccation in combination with starvation stress, and under starvation stress alone as well. The phycobiont of the symbionts was under desiccation stress alone. The experiments were detected by means of the biomass size, weight and cell density, deformity of the hyphae and cells, and metabolic activity through SEM (scanning electron microscopy), TEM (transmission electron microscopy), FM (fluorescence microscopy), spectrophotometry, and FCM (flow cytometry). The results show that the mycobiont can survive for seven months under desiccation stress in combination with starvation stress, and for eight months under starvation stress alone. The phycobiont can survive for two months under desiccation stress. It can provide a scientific basis for further research of the reproduction biology of lichens and arid desert biocarpet engineering to fix sand and carbon.     

Cemented annulation as an age criterion in the common marmoset (Callithrix jacchus)

Teeth from 18 marmosets, seven months to 5.8 years of age, were studied histologically. A minimum of one dark and one light band comprised one cemental annulation. Age was estimated by counting the number of annulations and adding four months to premolar and eight months to canine teeth for tooth development. Cemental annulations were most consistent near the gingival border on the labial aspect of the tooth. The method will be valuable in estimating the ages of marmosets and other nonhuman primates.     

Description of the glochidium of Margaritifera auricularia (Spengler 1793) (Bivalvia,Unionoidea)

The glochidium of Margaritifera auricularia is described for the first time by using light microscopy, scanning electron microscopy and histological techniques. The larval mantle is formed by only two layers of cells; the inner one being much thicker, with microvilli. All cell masses of the glochidium are temporary aggregations that are the rudiments of organs of the subsequent juveniles which will be released after metamorphosis in the host tissues. In the glochidium there are three main masses of cells: (i) the muscle, which is in an anterior position; (ii) the oral plate in the centre of the larva; and (iii) the more ventrally and posteriorly situated ventral plate, or foot rudiment, flanged with lateral pits all bearing dense cilia. No rudimentary organs such as the pericardium, the kidney, the heart or nerve ganglia have developed. There are no visible hooks in the valve margins, but by using light microscopy we observed minute teeth covered by a rim of the periostracum. Near the margin of the shell there are two pairs of sensory hair tufts only observable by scanning electron microscopy. The glochidium of M. auricularia is the largest of the family Margaritiferidae and intermediate between the glochidium of the known species of this family and those of Unionidae.     

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 μm long) of V. volvingentis is in sharp contrast to the minute vulva (± 6 μm 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.     

Scanning electron microscopy of viable and calcified Onchocerca gibsoni

Holdsworth P. A. 1987. Scanning electron microscopy of viable and calcified Onchocerca gibsoni. International Journal for Parasitology17: 957–964. Scanning electron microscopy (SEM) of adult Onchocerca gibsoni revealed that the cephalic ends of male and female specimens possessed features of little value for differentiation from other species. The marked cuticular ridges of the mid-body region of female worms were highlighted using SEM, as were the cuticular annulations and the body constriction at the anal region. Male specimens revealed the uniform annulations of the worm body along with the caudal papillae and the spicules. Microfilariae (mff) possessed cephalic papillae, lateral lines and cuticular annulations similar to adult worms. The cephalic hook was the most prominent feature at the anterior end of the mff. Calcification of adult worms was found to be mainly internal, however, one specimen was recorded with external calcification. Energy dispersive X-ray analysis revealed the composition of the calcification to be of calcium phosphate.     

Geissleria gereckei sp. nov. (Bacillariophyta) from leaf-litter covered stones of very shaded carbonate mountain springs with extremely low discharge

A new benthic freshwater diatom, Geissleria gereckei sp. nov., was identified from light and scanning electron micrographs. The most characteristic morphological features are the valve outline (elliptical, but with lateral margins that can range from slightly convex to slightly concave and also straight, and protracted, almost capitate ends), the size and length/width ratio, and the pattern of the striae around the central area. The new species was found to be epilithic, and characteristic of leaf-litter covered stones of very shaded carbonate mountain springs of the Italian Alps with extremely-low discharge (likely to be affected by seasonal desiccation). Very-low-discharge springs host a highly specialized diatom microflora, and are likely to increase in number in the near future due to the reduction of water, which is predicted to be caused by climate change in many regions.     

First Record of Cosmocephalus obvelatus (Acuariidae) in Common Gulls (Larus canus) from Gangneung,Korea

A nematode species belonging to the genus Cosmocephalus was collected from the stomach of 2 common gulls, Larus canus. The common gulls were found dead on the seaside of Gangneung City, the Republic of Korea. The worms were identified and classified by light (LM) and scanning electron microscopy (SEM) on the basis of important taxonomic characters. The nematodes were characterized by a body length 9.1-9.3 mm (males) and 15.5-15.9 mm (females) and cordons recurrent in anterior direction and anastomosing laterally at about the level of anterior quarter of the buccal cavity. The salient bicuspid deirids were located on the posterior to the cordons. Lateral alae were well-developed, extending from the level just posterior of deirids to the level about middle of the body. LM and SEM observations identified the worms as C. obvelatus. This is the first reported case of C. obvelatus infection in common gulls in Korea.     

Terminal electron acceptors influence the quantity and chemical composition of capsular exopolymers produced by anaerobically growing Shewanella spp

Bacterial exopolymers perform various roles, including acting as a carbon sink, a protective layer against desiccation or antimicrobial agents, or a structural matrix in biofilms. Despite such varied roles, little is known about the heterogeneity of bacterial exopolymer production under varying growth conditions. Here we describe experiments designed to characterize the quantity and quality of exopolymers produced by two commonly studied members of the widely distributed genus Shewanella. Electrokinetic, spectroscopic, and electron microscopic techniques were employed to demonstrate that cell surfaces of Shewanella oneidensis MR-1 (electrophoretic softness, lambda(-1), range from 0.4 to 2.6 nm) are associated with less extracellular polymeric material than surfaces of Shewanella putrefaciens 200R (lambda(-1) range from 1.6 to 3.0 nm). Both species exhibit similar responses to changes in electron acceptor with nitrate- and fumarate-grown cells producing relatively little exopolymer compared to trimethylamine N-oxide (TMAO)-grown cells. In S. oneidensis, the increase in exopolymers has no apparent effect upon cell-surface fixed charge density (-7.7 to -8.7 mM), but for S. putrefaciens a significant drop in fixed charge density is observed between fumarate/nitrate-grown cells (-43 mM) and TMAO-grown cells (-20.8 mM). For both species, exopolymers produced during growth on TMAO have significant amide functionality, increasing from approximately 20-25% of C-containing moieties in nitrate-grown cells to over 30% for TMAO-grown cells (determined from X-ray photoelectron spectroscopy). The increased exopolymer layer associated with TMAO-grown cells appears as a continuous, convoluted layer covering the entire cell surface when viewed by low-temperature, high-resolution scanning electron microscopy. Such significant changes in cell-surface architecture, dependent upon the electron acceptor used for growth, are likely to influence a variety of cell interactions, including aggregation and attachment to surfaces, and the binding of aqueous metal species.