Comparative analysis of Tritrichomonas foetus (Riedmüller, 1928) cat genotype,T. foetus (Riedmüller, 1928) cattle genotype and Tritrichomonas suis (Davaine, 1875) at 10 DNA loci

The parasitic protists in the genus Tritrichomonas cause significant disease in domestic cattle and cats. To assess the genetic diversity of feline and bovine isolates of Tritrichomonas foetus (Riedmüller, 1928) Wenrich and Emmerson, 1933, we used 10 different genetic regions, namely the protein coding genes of cysteine proteases 1, 2 and 4–9 (CP1, 2, 4–9) involved in the pathogenesis of the disease caused by the parasite. The cytosolic malate dehydrogenase 1 (MDH1) and internal transcribed spacer region 2 of the rDNA unit (ITS2) were included as additional markers. The gene sequences were compared with those of Tritrichomonas suis (Davaine, 1875) Morgan and Hawkins, 1948 and Tritrichomonas mobilensis Culberson et al., 1986. The study revealed 100% identity for all 10 genes among all feline isolates (=T. foetus cat genotype), 100% identity among all bovine isolates (=T. foetus cattle genotype) and a genetic distinctness of 1% between the cat and cattle genotypes of T. foetus. The cattle genotype of T. foetus was 100% identical to T. suis at nine loci (CP1, 2, 4–8, ITS2, MDH1). At CP9, three out of four T. suis isolates were identical to the T. foetus cattle genotype, while the T. suis isolate SUI-H3B sequence contained a single unique nucleotide substitution. Tritrichomonas mobilensis was 0.4% and 0.7% distinct from the cat and cattle genotypes of T. foetus, respectively. The genetic differences resulted in amino acid changes in the CP genes, most pronouncedly in CP2, potentially providing a platform for elucidation of genotype-specific host-pathogen interactions of T. foetus. On the basis of this data we judge T. suis and T. foetus to be subjective synonyms. For the first time, on objective nomenclatural grounds, the authority of T. suis is given to Davaine, 1875, rather than the commonly cited Gruby and Delafond, 1843. To maintain prevailing usage of T. foetus, we are suppressing the senior synomym T. suis Davaine, 1875 according to Article 23.9, because it has never been used as a valid name after 1899 and T. foetus is widely discussed as the cause of bovine trichomonosis. Thus bovine, feline and porcine isolates should all be given the name T. foetus. This promotes the stability of T. foetus for the veterinary and economically significant venereal parasite causing bovine trichomonosis.     

Cattle pathogen tritrichomonas foetus (Riedmüller, 1928) and pig commensal Tritrichomonas suis (Gruby & Delafond, 1843) belong to the same species

A number of reports suggest that the sexually transmitted pathogen of cattle, Tritrichomonasfoetus, and a gastrointestinal commensal of pigs, Tritrichomonas suis, are very similar and may be co-specific. A conclusive review of the taxonomic and nomenclatural status of these species has not been presented so far. Toward this end, we reexamined and compared porcine and bovine trichomonads with regard to their morphology, pathogenic potential, and DNA polymorphism. Using light and electron microscopy, no distinguishing features between T. foetus and T. suis strains were found in size, general morphology, and karyomastigont structure. Both bovine and porcine trichomonads showed pathogenic potential in the subcutaneous mouse assays and did not separate into distinct groups according to strain virulence. Three DNA fingerprinting methods (i.e. RFLP, RAPD, and PCR-based analysis of variable-length DNA repeats) that produce species-specific DNA fragment patterns did not distinguish between the bovine and porcine strains. Sequencing of a variable 502-bp DNA fragment as well as comparison of 16S rRNA gene sequences did not reveal species-specific differences between the cattle and porcine strains. Therefore, we conclude that T. foetus and T. suis belong to the same species. To prevent confusion that may arise from T. foetus-T. suis synonymy, we propose to suppress the older name suis and maintain its accustomed junior synonym foetus as a nomen protectum for both cattle and porcine trichomonads. The case has been submitted to the International Commission on Zoological Nomenclature for ruling under its plenary power.     

Regional adaptation of Müller cells in the chick retina. A Golgi and electron microscopical study

We report the morphological differences of Müller cells in relation to their topography, using the Golgi method. Müller cells in the central retina are long and slender, with numerous inner prolongations. In the peripheral retina, the morphology of the Müller cells adapts to the reduced thickness of the retinal layers. In this zone, they are short and have thick inner prolongations which end in a large foot in the internal limiting membrane. In the optic disc margin, Müller cells have a particular morphology characterized by thick, arched prolongations that in general form a glial network between the retina and optic nerve. The ultrastructure of these cells is also described. The results are discussed with respect to the nature of Müller cells.     

An electron microscope study of spermiogenesis in Spirorbis (Laeospira) mörchi Levinsen (Polychaeta)

Summary 1. In the hermaphroditic polychaete Spirorbis (Laeospira) mörchi, early spermatids develop in clusters within the coelom of the male segments. The cells within a single cluster are all in the same stage of development and are connected by an extensive cytoplasmic bridge system. 2. The acrosome forms in a single lamella of the Golgi apparatus which bears a close association to the plasma membrane. The final position of the acrosome is at a point considerably removed from the site of formation. 3. The nuclear changes culminating in condensation and elongation of the head are described. A rearrangement of cytoplasmic microtubules occurs simultaneously with nuclear elongation. 4. Redundant nuclear envelope, resulting from nuclear volume reduction, is pinched off in the form of four vesicles. The latter structures are lost with the residual cytoplasm. 5. Pour spherical mitochondria elongate to become incorporated into the middle-piece. A rearrangement of microtubules also occurs simultaneously with mitochondrial elongation. Cytoplasmic microtubules are absent from the fully formed sperm.This investigation was made possible through a Postdoctoral Fellowship from the National Science Foundation, 44150. — I wish to express my gratitude to Dr. John Luft for the training I received in the techniques of electron microscopy. Dr. James Koehler and Dr. Daniel Szollosi are thanked for their many helpful suggestions.     

Morphological differentiation of the Müller cell: Golgi and electron microscopy study in the chick retina

The sequence of morphological differentiation of Müller cells in the chick retina was investigated in relation to the differentiation of the retinal neurons using the Golgi method. From the beginning of differentiation, the Müller cell develops spurs and lateral processes. Some of these glial processes become transformed into accessory prolongations of the Müller cell. From the 17th or 18th day of incubation, the morphology of the Müller cells is similar to that of the adult retina. On the basis of their inner prolongation, two types of Müller cells were identified. The first type, with diffuse and abundant descending processes, is identical to that described classically. The second type is a cell characterized by sparse and scanty inner ramifications. This report also describes electron microscopic observations of Müller cells and their enwrapping relationship with the axons of the optic nerve fiber layer.     

A cytogenetics study of Hydrodroma despiciens (Müller, 1776) (Acari: Hydrachnellae: Hydrodromidae)

The karyotypes of water mites (Acari: Hydrachnellae: Hydrodromidae) are largely unknown. The present investigation is the first report of a study designed to characterize the chromosomes of water mites. The study was carried out with specimens of Hydrodroma despiciens collected from Eber Lake in Afyon, Turkey. Several different methods were tried to obtain chromosomes of this species. However, somatic cell culture proved to be the most effective for the preparation of chromosomes. In the present study, we determined the diploid chromosome number of Hydrodroma despiciens to be 2n = 16. However, a large metacentric chromosome was found in each metaphase, which we believed to be the X chromosome. We could not determine the sex chromosomes of this species. This study is the first approach to the cytogenetic characterization of this water mite group. Furthermore, these cytogenetic data will contribute to the understanding of the phylogenetic relationship among water mites. To our knowledge, this is the first report on the cytogenetics of water mites.     

A stereologic electron microscope study of “tubular myelin figures” in alveolar fluids of rat lungs

Summary The three-dimensional structure of a composite material found in alveolar exudate of oxygen poisoned lungs but also present in normal lungs is stereologically analysed. It is composed of tubules of 450 Å diameter which are tightly packed in a quadratic lattice. The wall of the tub vile is formed by four-winged osmiophilic filaments which are located in the corners of the quadratic lattice; their interior is made up of a hydrophilic substance which contains either a tubule or a filament of moderate electron density. The osmiophilic substance of the walls is continuous with associated myelin figures which can be resolved into lamellae with a periodicity of 42 Å and can thus be considered to be water crystals of phospholipids. The nature of the content of the tubules, which presumably exerts the formative force on the phospholipid lamellae to form tubules, remains undetermined.Dedicated to Prof. W. Bargmann in honor of his 60th birthday.The research reported here has been sponsored by the Schweizerischer Nationalfonds zur Förderung der wissenschaftlichen Forschung (Nr. 2569); by the Stiftung für wissenschaftliche Forschung an der Universität Zürich; by the National Institutes of Health, USPHS, through grant RF-57; and the 6570th Aerospace Medical Research Laboratories under contract AF 61(052)-784 through the European Office of Aerospace Research (OAR), United States Air Force.     

A chromosome study ofSchistocephalus solidus (Müller, 1776) (Cestoda: Pseudophyllidea)

Analysis of Giemsa-stained metaphase plates of plerocercoids ofSchistocephalus solidus revealed the mode diploid number of chromosomes for this species as 2n=18. Chromosome pairs 1, 5, 6, 8 and 9 are metacentric, pairs 2, 4 and 7 are subtelocentric, and pair 3 is subtelocentric to acrocentric. The karyotype is characterised by mean absolute chromosome length between 1.9 and 5.9 m and a total chromosome length of the haploid complement of 32 m. A comparison with other known karyotypes of related species of pseudophyllidean cestodes was made.     

Morphological study of muscular tissue collagen of wild and reared Cornu aspersum (Müller, 1774)

Muscular tissue collagen fibrils’ diameter and period of reared and wild snails (Cornu aspersum) were measured in order to investigate the role of nutrition on the collagen. Wild adult individuals were collected from Crete (Greece) and compared with adult individual snails reared under laboratory conditions fed a formulated diet. Transmission electron microscope and image analysis algorithms were used in the study. Statistically significant differences between the diameters of the collagen fibrils in the reared and the wild snails were found, and the D-period of the fibrils differed.     

A protein phosphatase 1 gamma (PP1γ) of the human protozoan parasite Trichomonas vaginalis is involved in proliferation and cell attachment to the host cell

In this work, evidence for a critical role of Trichomonas vaginalis protein phosphatase 1 gamma (TvPP1γ) in proliferation and attachment of the parasite to the mammalian cell is provided. Firstly, proliferation and attachment of T. vaginalis parasites to HeLa cells was blocked by calyculin A (CA), a potent PP1 inhibitor. Secondly, it was demonstrated that the enzyme activity of native and recombinant TvPP1γ proteins was inhibited by CA. Thirdly, reverse genetic studies confirmed that antisense oligonucleotides targeted to PP1γ but not PP1α or β inhibited proliferation and attachment of trichomonads CA-treated parasites underwent cytoskeletal modifications, including a lack of axostyle typical labelling, suggesting that cytoskeletal phosphorylation could be regulated by a CA-sensitive phosphatase where the role of PP1γ could not be ruled out. Analysis of subcellular distribution of TvPP1γ by cell fractionation and electron microscopy demonstrated the association between TvPP1γ and the cytoskeleton. The expression of adhesins, AP120 and AP65, at the cell surface was also inhibited by CA. The concomitant inhibition of expression of adhesins and changes in the cytoskeleton in CA-treated parasites suggest a specific role for PP1γ -dependent dephosphorylation in the early stages of the host-parasite interaction. Molecular modelling of TvPP1γ showed the conservation of residues critical for maintaining proper folding into the gross structure common to PP1 proteins. Taken together, these results suggest that TvPP1γ could be considered a potential novel drug target for treatment of trichomoniasis.     

A review of the reproductive behavior of Polyphemus pediculus (L.) Müller (Crustacea: Branchiopoda)

The reproductive behavior of Polyphemus pediculus (L.) includes mating, laying of resting eggs and giving birth to juveniles. All these forms of behavior are performed in a population daily and simultaneously by means of individual and social actions. Individual behavior is manifested by certain sets of stereotyped movements which can be observed at any time of the day and at any location in a water body. Individual behavior has an episodic, stable character and low intensity. Social behavior is accomplished by specimens of similar physiological state who form transitory swarms. They form within the main continually existing main swarms of the population. Social behavior is rhythmic and of high intensity. Reproductive parthenogenetic swarms first form around dawn and later during the day: about noon in May–July and before sunset in Autumn. As swarms of parthenogenetic females disintegrate, swarms of old gamogenetic females form and mass laying of resting eggs begins. Young gamogenetic females and males swarm only when resting-egg-laying swarms disintegrate. Parthenogenetic reproduction peaks in northern Russia in May and August and persists until late September. Swarms of resting-egg-laying females form once a day around dawn, from May to September. Mating swarms form in summer in midmorning, at twilight and on moonlit nights. In Autumn, there is only one peak of mating, in midmorning. Intensity, duration and time of manifestation of different forms of social behavior change from spring to autumn. Location and character of their manifestation are predictable.     

The size,density, and relative area of chromatic granules (“hydrogenosomes”) in Trichomonas vaginalis donne from cultures in logarithmic and stationary growth

Summary Electron micrographs of Trichomonas vaginalis cells were examined morphometrically by means of an electronic image analyser. Cells from cultures in logarithmic growth contained smaller chromatic granules (hydrogenosomes ) and a higher proportion of sausage- or dumb-bell-shaped granules than did cells in stationary growth.These findings support the view that chromatic granules replicate by division of pre-existing granules and that this involves elongation and attenuation of their mid-region.The total area of cross sectioned granules occupied the same percentage of the cytoplasm of cells from cultures in logarithmic and stationary growth. The density of chromatic granules in the cytoplasm was identical for the two groups of cells.     

A general-purpose system for the rapid acquisition and computer processing of optical and scanning electron microscope images—II. Software

The design and implementation of software for the operation of a general-purpose optical and electron microscope image processing system is described. The software is a group of programs, controlled by a command-line interpreter called IPR (Image PRocessing). The interpreter may be used interactively, or groups of commands may be issued indirectly after placing them in files. Programs for specialized image processing applications may obtain the services of the system's memory-resident programs, through the same interprogram communication methods that are used by the command-line interpreter.     

Purification and characterisation of glutathione S-transferases from the freshwater clam Corbicula fluminea (Müller)

Glutathione S-transferases (GSTs) are involved in the phase II detoxification metabolism. To provide a molecular basis for their use as biomarkers of pollution, cytosolic GSTs from the freshwater clam Corbicula fluminea have been purified by glutathione-Sepharose affinity chromatography, anion-exchange chromatography (AEC) and reversed-phase (RP) HPLC. SDS-PAGE of visceral mass (VM) affinity-purified extracts revealed four subunits with apparent molecular masses (MW) of 30.2, 29.2, 28.5 and 27.2 kDa. Analysis by non-denaturing PAGE revealed three acidic dimeric proteins with apparent MW of 64, 55 and 45 kDa, named GSTc1, GSTc2 and GSTc3, respectively, based on their elution order by AEC. Only GSTc2 and GSTc3 exhibited GST activity towards 1-chloro-2,4-dinitrobenzene. A tissue-specific subunit pattern was obtained by RP-HPLC of affinity-purified extracts from VM and gills (GI): three major peaks were resolved, one of which was common to both tissues. MW of each VM subunit was determined by electrospray ionisation-mass spectrometry: 23602+/-1 Da for the major subunit and 23289+/-1 Da for the minor ones. Immunoblot analysis revealed all subunits from both tissues were related to the Pi-class GSTs. In addition, minor VM subunits were slightly related to the Mu-class ones. The interest of such molecular studies in biomonitoring programs is discussed.     

Biological studies and phenology of the slug Deroceras reticulatum (Müller, 1774) (Pulmonata: Stylommatophora)

Summary

In the southeast of Buenos Aires Province (Argentina), slugs cause reductions in crop yield by killing seeds and seedlings, by destroying stems and growing points, and by reducing leaf area. Deroceras reticulatum (Müller, 1774) was introduced into Argentina a long time ago, but the biology and ecology of this species in this country are still unknown. The aim of this study was to determine how temperature affects growth rate, reproduction and survival of D. reticulatum and also to assess the seasonal phenology of this species. The life cycle of D. reticulatum was investigated at 12°C, 20°C and at a temperature which alternated between 12°C for 16 h and 20°C for 8 h. The light/dark regime during the experiment was LD: 8/16 h. Slug populations were sampled from grassland located at the Experimental Station of INTA Balcarce. Eggs hatched after 16.3 to 39.8 days and fertility varied between 83.9% and 91.1%. The mean time from hatching to oviposition was longer and fecundity lower at 20°C than 12°C and 12/20°C (p <0.05). The net reproductive rate was 1.49 to 70.53 and the mean generation time varied between the 27 and 30 weeks. In natural conditions, eggs of D. reticulatum were found from early winter until the end of the spring, when soil temperature was 8–15°C and humidity was above 15%. These results indicate that D. reticulatum has one generation per year, univoltine phenology, with peak densities of slugs from the beginning of winter to the end of spring, which coincides with the sowing and emergence of winter and summer crops.