Infraciliature and morphogenesis in three rumen Diplodinium ciliates, Diplodinium polygonale, Diplodinium leche, and Diplodinium nanum, observed by light microscopy

Infraciliature and morphogenesis of three rumen ophryoscolecid ciliates, Diplodinium polygonale Dogiel, 1925, Diplodinium leche Imai et al., 1992, and Diplodinium nanum Imai, 1988, are described from pyridinated silver carbonate-impregnated specimens. These three species have two polybrachykineties in the buccal area and a polybrachykinety in the dorsal ciliary zone. The vestibular polybrachykinety (VP) of D. polygonale and D. leche arises from the dorsal extremity of the adoral polybrachykinety (AP) as in Entodinium species, extending toward the left in D. polygonale and toward the left posterior in D. leche. The VP of D. nanum arises from the inner side of the AP, separate from its dorsal extremity, as in other Diplodinium species and extends toward the left posterior. These series of the polybrachykinety arrangements in D. polygonale, D. leche, and D. nanum can be regarded as transitional forms in the evolution of an Entodinium-like ancestor to Diplodinium. Morphogenesis of these three Diplodinium species is not different from that of other Diplodinium species.     

The new higher level classification of eukaryotes with emphasis on the taxonomy of protists

This revision of the classification of unicellular eukaryotes updates that of Levine et al. (1980) for the protozoa and expands it to include other protists. Whereas the previous revision was primarily to incorporate the results of ultrastructural studies, this revision incorporates results from both ultrastructural research since 1980 and molecular phylogenetic studies. We propose a scheme that is based on nameless ranked systematics. The vocabulary of the taxonomy is updated, particularly to clarify the naming of groups that have been repositioned. We recognize six clusters of eukaryotes that may represent the basic groupings similar to traditional "kingdoms." The multicellular lineages emerged from within monophyletic protist lineages: animals and fungi from Opisthokonta, plants from Archaeplastida, and brown algae from Stramenopiles.     

A huge,undescribed soil ciliate (Protozoa: Ciliophora) diversity in natural forest stands of Central Europe

We investigated 12 natural forest stands in eastern Austria for soil ciliate diversity, viz., eight beech forests and two lowland and Pinus nigra forests each. The stands span a wide range of climatic (e.g., 543–1759 mm precipitation, 160–1035 m above sea-level) and abiotic (e.g., pH 4–7.4) factors. Samples were taken twice in autumn and late spring and analysed with the non-flooded Petri dish method. Species were identified in vivo, in silver preparations, and in the scanning electron microscope. A total of 233 species were found, of which 30 were undescribed, a surprising number showing our ignorance of soil ciliate diversity, even in Central Europe. Species number varied highly from 45 (acidic beech on silicate) to 120 (floodplain forest) and was strongly correlated with pH and overall habitat quality, as measured by climate, the C/P quotient (ratio of r-selected colpodean and k-selected polyhymenophorean ciliates), and the proportion of mycophagous ciliate species; multivariate analysis showed further important variables, viz., the general nutrient status (glucose, nitrogen, C/ N ratio) and microbial (urease) activity. The highest species number occurred in one of the two floodplain soils, supporting the intermediate disturbance hypothesis. The three main forest types could be clearly distinguished by their ciliate communities, using similarity indices and multidimensional scaling. Individual numbers varied highly from 135^–1 (lowland forest) to 10,925 ml^–1 (beech on silicate) soil percolate and showed, interestingly, a weak correlation with soil protozoan phospholipid fatty acids. Eight of the 30 new species found and a forgotten species, Arcuospathidium coemeterii (Kahl 1943) nov. comb., are described in detail, as examples of how species were recognized and soil protozoan diversity should be analyzed: Latispathidium truncatum bimicronucleatum, Protospathidium fusioplites, Erimophrya sylvatica, E. quadrinucleata, Paragonostomum simplex, Periholosticha paucicirrata, P. sylvatica, and Australocirrus zechmeisterae.     

Regeneration of cryoresistance of in vitro rumen ciliate cultures

The purpose of this study was to investigate factors affecting mechanical- and cryo-resistance of the rumen ciliates Entodinium caudatum (E.c.), Entodinium furca monolobum (E.f.m.), Entodinium simplex (E.s.), Diplodinium denticulatum (two clones, D.d.01 and D.d.02), Diploplastron affine (D.a.) and Epidinium ecaudatum forma caudatum (E.e.c.) after long-term in vitro cultivation. Following prolonged in vitro cultivation (more than six months), the ciliates were very sensitive to both centrifugation and 5% (v/v) dimethylsulphoxide, with motility decreased to: 39 and 23% for E.c., 66 and 32% for E.f.m., 46 and 27% for D.d. 01, 64 and 41% for D.a., and 44 and 28% for E.e.c., respectively. Thus, cryopreservation was unsuccessful. The effect of supplementing the ciliate growth medium with rumen fluid, glycine-betaine, proline, myo-inositol, linoleic acid, Sel-Plex or insulin, together with the effect of the source of rumen fluid on ciliate resistance to centrifugation, dimethylsulphoxide and freezing was also tested. The omission of rumen fluid from the growth medium resulted in the loss of cryoresistance after one-month cultivation. Supplementing the growth environment with a combination of glycine-betaine, proline, linoleic acid, Sel-Plex, insulin plus improved quality rumen fluid significantly enhanced survival of the ciliates after the freezing-thawing procedure (from 1 to 33% survival in un-supplemented vs. supplemented for E.c., P < 0.01; 4-40% E.f.m., P < 0.01; 0-17% D.d., P < 0.05; 5-7% D.a. and 4-36% E.e.c., P < 0.01).     

Detritus as a potential food source for protozoans: utilization of fine particulate plant detritus by a heterotrophic flagellate, <Emphasis Type="Italic">Chilomonas paramecium</Emphasis>, and a ciliate, <Emphasis Type="Italic">Tetrahymena pyriformis</Emphasis>

We investigated the direct utilization of fine particulate detritus (dried and homogenized plant material in the size range of bacteria) as a food source for protozoans using axenic cultures of the cryptomonad, heterotrophic flagellate, Chilomonas paramecium, and the hymenostome ciliate, Tetrahymena pyriformis. When fed media containing only particulate detritus, these species revealed growth rates similar to those reported for field populations. The growth rates of Chilomonas fed exclusively particulate detritus were similar to those obtained on a bacterial diet. Considering the high percentage of detritus particles in the size range of bacteria in lakes, our results imply that direct utilization of detritus by protozoans may form an additional pathway of carbon in aquatic food webs that has generally been overlooked.     

The role of prey nutritional status in governing protozoan nitrogen regeneration efficiency

Laboratory experiments were conducted to study nitrogen (N) regeneration by the heterotrophic marine dinoflagellate Oxyrrhis marina when ingesting phytoplankton prey of two different species and of two alternative carbon:nitrogen (C:N) ratios. Experiments were conducted in the presence of L-methionine sulfoximine (MSX) which acts as a glutamine synthetase inhibitor. Utilisation by phytoplankton of N regenerated by protozoans and other organisms drives secondary production in marine food webs. However, the rapid utilisation of this N by phytoplankton has previously hampered accurate assessment of the efficiency of protozoan N regeneration. This phenomenon is particularly problematic when the phytoplankton are nutrient stressed and most likely to rapidly utilise N. The use of MSX prevented significant utilisation by phytoplankton of protozoan regenerated N. Hence, by removing the normal pathway of N cycling, we were able to determine the N regeneration efficiency (NRE) of the protozoan. The results suggested that predator NRE could be explained in terms of the relative CN stoichiometry of prey and predator. Using a mathematical model we demonstrated that changing the method used to simulate the NRE of the protozoan trophic level has the potential to markedly modify the predicted dynamics of the simulated microbial food web.     

Hindgut microbes, fermentation and their seasonal variations in Hokkaido native horses compared to light horses

Fecal bacteria and protozoa of Hokkaido native horses and light horses were enumerated to compare seasonal variation in hindgut microbes and fermentation between the two breeds. Fecal samples were collected in winter and summer from eight horses (four for each breed) that had been reared together under the same conditions after birth (on woodland pasture in winter and on grassland pasture for the rest of the year). Total fecal bacteria counts for both breeds showed temporal variation, with the highest levels occurring in summer (P<0.05). For both breeds, Gram-negative rods were the major constituents (58–69%) and showed higher counts in winter (P<0.05) than in summer. Total protozoa counts in both breeds were lower in winter than in summer (P<0.05). The proportion of large cellulolytic protozoa such as Cochliatoxum periachtum was increased (P<0.05) in winter, and this tended to be more pronounced in native horses. Although total volatile fatty acids (VFA) in feces were lower in winter (P<0.05), the reduction was smaller in native horses (P<0.05). Fecal VFA pattern showed a shift toward more acetate and less propionate production in winter regardless of the horse breed. Evaluation of digestive tract organs in 12 animals showed that the relative weight of the colon in body weight or total digestive tract weight is larger in native horses than in light horses (P<0.05). The present results suggest that hindgut microbial adaptation to winter diets occurs to a greater extent in native horses, as partly characterized by advantages in anatomy.     

Mycobacterium avium ssp. paratuberculosis and its potential survival tactics

Mycobacterium avium ssp. paratuberculosis (Map) is an important animal pathogen with a potential, but as yet unproven, role in human disease. This review briefly describes the characteristics of Map that distinguish it from other Mycobacterium spp., presenting new information arising from completion of the sequencing of the Map genome. It then focuses on the potential mechanisms Map might employ to survive and disseminate in the environment, including interaction with protozoa and insects, dormancy, biofilm formation and aerosolization.     

Toll-like receptor recognition of Toxoplasma gondii

Toxoplasma gondii potently stimulates IFN-gamma production by both the innate and adaptive immune system as part of its host adaptation. This response is known to be dependent on an Myeloid Differentiation factor 88 signaling pathway used by Toll-like receptors (TLRs), a family of proteins involved in the recognition of microbial molecular patterns. In the following review, we summarise the evidence for specific TLR function in host resistance to T. gondii focusing on the recent discovery in the parasite of a profilin-like ligand that potently stimulates TLR11 and regulates the production of IL-12, a cytokine necessary for the protective IFN-gamma response. In addition, we discuss the hypothesis that TLR11 may have evolved as a general pattern recognition receptor for apicomplexan protozoa and that as highly conserved proteins associated with actin-based motility, profilins are logical ligand targets for this form of pathogen detection. Finally, we review the evidence for involvement of other TLR and TLR ligands in host resistance to T. gondii and discuss how such receptors might synergise with TLR11 in the innate response to the parasite.     

Global change belowground: impacts of elevated CO2, nitrogen,and summer drought on soil food webs and biodiversity

The world's ecosystems are subjected to various anthropogenic global change agents, such as enrichment of atmospheric CO₂ concentrations, nitrogen (N) deposition, and changes in precipitation regimes. Despite the increasing appreciation that the consequences of impending global change can be better understood if varying agents are studied in concert, there is a paucity of multi‐factor long‐term studies, particularly on belowground processes. Herein, we address this gap by examining the responses of soil food webs and biodiversity to enrichment of CO₂, elevated N, and summer drought in a long‐term grassland study at Cedar Creek, Minnesota, USA (BioCON experiment). We use structural equation modeling (SEM), various abiotic and biotic explanatory variables, and data on soil microorganisms, protozoa, nematodes, and soil microarthropods to identify the impacts of multiple global change effects on drivers belowground. We found that long‐term (13‐year) changes in CO₂ and N availability resulted in modest alterations of soil biotic food webs and biodiversity via several mechanisms, encompassing soil water availability, plant productivity, and – most importantly – changes in rhizodeposition. Four years of manipulation of summer drought exerted surprisingly minor effects, only detrimentally affecting belowground herbivores and ciliate protists at elevated N. Elevated CO₂ increased microbial biomass and the density of ciliates, microarthropod detritivores, and gamasid mites, most likely by fueling soil food webs with labile C. Moreover, beneficial bottom‐up effects of elevated CO₂ compensated for detrimental elevated N effects on soil microarthropod taxa richness. In contrast, nematode taxa richness was lowest at elevated CO₂ and elevated N. Thus, enrichment of atmospheric CO₂ concentrations and N deposition may result in taxonomically and functionally altered, potentially simplified, soil communities. Detrimental effects of N deposition on soil biodiversity underscore recent reports on plant community simplification. This is of particular concern, as soils house a considerable fraction of global biodiversity and ecosystem functions.