Effects of Extracellular Potassium on Acid Release and Motility Initiation in Toxoplasma gondii

The internal pH (pH_i) of Toxoplasma gondii was estimated by measuring the accumulation of the weak base 9-aminoacridine in buffers with various ionic compositions. The pH_i of the metabolizing parasite increased when the extracellular K⁺ was elevated in alkaline medium or when the external pH (pH_c) was substantially increased in medium employing high external K⁺ (90 mM). The parasite in mouse peritoneal fluid, or in potassium sulfate buffer (pH 8.2), where the pH_i was demonstrated to be increased to 7.9, became motile when acidic buffer was substituted for the original suspension medium. This acid-induced independent movement subsided within 5 min but was repeatedly induced if the pH_c was serially lowered to 6.0. Basic buffers, on the other hand, abolished motility when applied to the moving parasites. Nigericin, which is known to collapse pH gradients across the membrane, also abolished motility.     

Methane-flux-dependent lateral faunal changes in a Late Cretaceous chemosymbiotic assemblage from the Nakagawa area of Hokkaido, Japan

A Late Cretaceous carbonate body (2 m in maximum diameter) surrounded by clastic rocks, recently discovered in the Nakagawa area (Hokkaido, Japan), is interpreted as a methane‐seep deposit, on the basis of negative carbon isotopic composition (as low as ?43.5‰), variable sulphide sulphur isotopic composition, high carbonate content, and in situ fractures. It most likely formed owing to methane‐bearing pore‐water diffusion. We estimate that the concentration of methane decreased toward the margin of the carbonate body, and that only small carbonate concretions were precipitated at a certain distance from the methane‐seep centre. These spatial characteristics coincide well with the observed pattern of faunal distribution. The gastropod‐dominated association (indeterminate abyssochrysids and ataphrids and the acmaeid limpet Serradonta sp. are most common) co‐occurs with lucinid and thyasirid bivalves (Thyasira sp., Myrtea sp., and Miltha sp.), and was found within and just above the methane‐derived carbonate body. Acharax and Nucinella (solemyoid bivalves) are more typical of the peripheral part of the methane‐influenced sediments. We suggest that this pattern of faunal distribution reflects the decreasing concentration of methane and apparently also hydrogen sulphide when moving from the centre of discharge toward the periphery of the methane seep.     

Morphological difference and reproductive isolation: morphometries in the millipede Parafontaria tonominea and its allied forms

Thin-plate spline and other multivariate statistical methods were used to examine the variation in genital morphology and body size in 74 populations of the millipede Parafontaria tonominea and its allied forms in central Japan. Although the study populations included nine sympatric pairs of two reproductively isolated forms, the variations in genital morphology and body size observed were continuous across the study group. Analyses of the sympatric pairs using bootstrapping revealed that the degree of morphological difference between sympatric populations varied considerably from pair to pair. These results demonstrate that the degree of difference in genital morphology and body size may not correspond with the establishment of reproductive isolation. Significant correlation in the pattern of geographic variation was observed for many combinations of morphological components. However, it was not possible to detect a significant correlation between morphology and geographic distance in many of the morphological components studied. This study suggests that the incorporation of genital characters in classification should be done carefully, particularly for animal groups with low vagility, such as millipedes.     

Roles of Head-Activation and Head-Inhibition Potentials in Pattern Formation of Hydra: Analysis of a Multi-Headed Mutant Strain

SYNOPSIS. Lateral grafting of tissue was used to compare therelative head-activation and head-inhibition potentials of differentHydra strains. A small piece of tissue taken from one polyp,when grafted to another polyp, induces formation of a head structurewhen the relative head-activation potential of the donor tissueis sufficiently (i.e., more than some threshold value) higherthan the relative head-inhibition potential of the recipienttissue. It was found that a multi-headed mutant strain (mh-1),which produces many extra heads along its body column, has significantlyhigher head-activation and significantly lower head-inhibitionpotentials than the standard wild-type strain. This suggeststhat these potentials play important roles in hydra morphogenesis,and that an imbalance between the two potentials is responsiblefor the developmental abnormality of mh-1. The significanceof this finding is discussed in light of the "positional information"model proposed by Wolpert and his associates and the "lateralinhibition" model proposed by Gierer and Meinhardt.     

Biochemical assessment of evolution and taxonomy of the morphologically poorly diverged geckos, Gekko yakuensis and G. hokouensis (Reptilia: Squamata) in Japan, with special reference to their occasional hybridization

We assessed the validity of two gekkonid species, Gekko yakuensis and G. hokouensis, in southern Japan. We first assigned all 398 specimens into 18 samples merely on the basis of localities. By conducting significance test for deviations of genotype frequencies from Hardy‐Weinberg at 11 allozyme loci, we checked the reproductive unity of constituents in each of those local samples, and where necessary, rearranged them into subsamples on the basis of genetic markers so that we recognized minimum reproductively cohesive units. We then compared allele frequencies among all samples and subsamples examined. Results clearly indicated that all but two can be classified into two groups that can be discriminated from each other by remarkable allele frequency differences at four diagnostic loci, and by large genetic distances even between sympatric subsamples. Observations of morphological features of the samples and subsamples confirmed that the two groups correspond to G. yakuensis and G. hokouensis, supporting validities of these two species. Allele frequency comparisons, however, also revealed that the remaining two samples, both from southern Kyushu, possessed ‘marker alleles' of both G. yakuensis and G. hokouensis at all four diagnostic loci. These samples thus were considered to represent populations that have been derived through hybridization of the two species. Detailed analyses for genetic structures demonstrated that all hybrid genotypes in the two samples are post‐Fi generations with only one individual resulting from the back‐cross with a pure line population of G. yakuensis. This finding negates the possibility that the hybrid populations are maintained by a constant supply of newly produced Fj hybrids, but suggests that the hybrid genotypes constitute stable breeding populations. This implies that the genealogical independence of G. yakuensis and G. hokouensis in several other sympatric areas has been maintained by operations of some isolation mechanisms at a pre‐mating phase. Investigations of the morphological variation in each sample or subsample revealed that although the two species can be externally largely discriminated from each other by slight modifications of the currently used diagnoses, it is difficult to detect their hybrids based solely on the morphological features.     

Reactivity of Monoclonal Antibodies to Species-Specific Antigens of Entamoeba histolytica

Twenty monoclonal antibodies were produced against trophozoites of Entamoeba histolytica strains HK-9 and HM-1: IMSS. When reactivity to various enteric protozoa was examined by an indirect fluorescence antibody test, 15 of the monoclonal antibodies were strongly reactive with E. histolytica trophozoites. Species-specific antigens recognized by these monoclonal antibodies were located on the plasma membrane, nucleus, cytoplasm, and cytoskeletal structures of the trophozoites. Two of the remaining five monoclonals reacted strongly with trophozoites of the E. histolytica -like Laredo strain. The determinant antigen was located in the cytoplasm. The three remaining monoclonal antibodies were found to recognize cross-reactive antigens between E. histolytica and E. histolytica -like Laredo, E. hartmanni, E. coli, Dientamoeba fragilis, Giardia lamblia , and Trichomonas hominis. These three antibodies were also reactive with T. vaginalis     

GENETIC ANALYSIS OF DEVELOPMENTAL MECHANISMS IN HYDRA I. SEXUAL REPRODUCTION OF HYDRA MAGNIPAPILLATA AND ISOLATION OF MUTANTS

Hydra magnipapillata strains collected from various localities in Japan were induced to reproduce sexually.
From the survival data of the progeny, it was calculated that H. magnipapillata contained an average of between 3.5 and 4.0 lethal equivalent units of recessive deleterious genes per gamate (between 7.0 and 8.0 per animal).
Various types of developmental mutants were found among the offspring of crosses made between strains isolated from the same ponds. The mutant types isolated included mini strains, maxi strains, multi-headed strains, nematocyst-deficient strains, regeneration-deficient strains and male sterile strains. The characters of these strains were stably transmissible to the successive progeny produced by budding. These strains therefore were propagated by budding and maintained as clonal lines to be used later for developmental studies.     

GENETIC ANALYSIS OF DEVELOPMENTAL MECHANISMS IN HYDRA VII. STATISTICAL ANALYSES OF DEVELOPMENTAL-MORPHOLOGICAL CHARACTERS AND CELLULAR COMPOSITIONS

Various strains belonging to Hydra magnipapillata are examined for their developmental-morphological characters and relative abundance of the six basic types of cells, and the results are statistically analysed.
Significant correlations are found between various (including seeming unrelated) characters. For example, budding rate, bud developmental rate and polyp size, which in theory can be all regulated by independent mechanisms, show strong correlations with each other. This suggests that the underlying mechanisms regulating these characters must be closely related to each other.
Similar significant coerelations are also found between the relative abundance of various cell types, but not between the developmental-morphological characters and the cellular composition. The significance of these findings are discussed.     

EFFECT OF ULTRAVIOLET LIGHT UPON VARIOUS PHYSIOLOGICAL ACTIVITIES OF CHLORELLA CELLS AT DIFFERENT STAGES IN THEIR LIFE CYCLE

1. Using Chlorella ellipsoidea as material, investigations weremade of the effects of ultraviolet irradiation upon variousactivities of cells at different developmental stages in theirlife cycle. Cell activities investigated were photosynthesis,respira tion, over-all growth, modes of synchronous growth andcell division as well as the formation of nucleic acids. Theu. v.- light applied was 30 µµW/cm²in intensityand 2537 Å in wavelength.
2. The most u. v.-sensitive wasthe over-all growth activity, andin this respect the irradiationapplied at the L₂-stage wasmore inhibitive than that givenat the D-stage. The next mostvulnerable was the photosyntheticactivity, the sensitivitybeing the same in the D- and L-cells.The most resistant towardu.v. was the endogenous respirationof D-cells followed by theirrespiration using exogenous glucoseas substrate. The L₂-cellsappeared to be unable to use exogenousglucose as substrateof respiration, but their endogenous respirationwas considerablystronger than that of D-cells, and its u. v.-sensitivitywasthe same as that of glucose respiration of D-cells.
3. WhenD-cells were u. v. irradiated immediately before the startofsynchronous culture, growth and cell division as well astheformation of DNA and RNA were retarded in proportion totheu. v.-dose applied. The division number (n) was normal (around4) at lower u.v.-doses (1-2 minute irradiation), but was reducedto a half (about 2) at a higher dose.
4. When, during the synchronousculture, 1-minute u.v.- irradiationwas applied at various stagesof the ripening phase, the divisionwas retarded, but the cells,after attaining an abnormally largesize, divided into about8. If the irradiation was given atthe L₄-stage, the divisionnumber was practically unmodified(n=4.5), although the divisionwas somewhat retarded comparedwith that of the control culture.When a 1-minute irradiationwas given at the L₂-stage, thereoccurred an apparent stimulationof DNA- and RNA-formation,a phenomenon which corresponds tothe production of a largernumber of daughter cells than itwas the case in control cultures.
5. Thus the cells which were moderately u.v.-irradiated at differentstages of synchronous culture were able to complete their lifecycle, but later a certain portion of irradiated cells becameunable to grow normally.

¹Present address: Department of Biochemistry, Dartmouth MedicalSchool, Hanover, New Hampshire, U.S.A. (Received March 6, 1961; )