Dense Crithidia Growth and Heme Sparing: Relation to Fe,Cu, Mo Chelation*

SYNOPSIS. Heme, intrinsically required by Trypanosomatidae, is unstable, especially in conventional alkaline (pH 7.2–8.0) media. Low solubility of heme in a pH 6.5 basal medium (developed to assay biopterin with Crithidia fasciculata) posed a problem: in media acidified during growth because of glycolysis, heme precipitated, perhaps contributed to acid-limited growth and interfered with densitometric estimation of growth. The remedy was to: replace glucose with less rapidly metabolized mannitol; distribute media in thin layers to promote oxidation of acetate, fumarate, and malate (presumably leaving an alkaline residue); and buffer heavily with histidine + Good zwitterionic buffers, and superimpcse physiological buffering by arginine + asparagine whose catabolism appeared to yield an excess of NH⁺₄ over acid. Thereupon, Fe and Cu deficiencies sharply limited growth in the medium whose main chelators were: (a) 2,3–dihydroxybenzoic + 5-sulfosalicylic acids (which preferentially bind transitional elements at their higher valences; (b) malic and gluconic acids; and (c) histidine. With unconventionally heightened concentrations of Fe, Cu, and Mo (the latter serving as Cu buffer as well as nutrient per se), the hemin concentration could be lowered, widening the margin of safety for heme solubility. Growth then reached 1.4 × 10⁸ cell/ml. This medium may serve to screen for ligands promoting uptake or release of Fe and Cu. The increased growth is a step towards improving the assay medium for biopterin and practical use of Crithidia to assay several B vitamins and essential amino acids for metazoa.     

Electron Microscopic Studies of Skeletal and Cardiac Muscle of a Benthic Fish. I. Myofibrillar Structure in Resting and Contracted Muscle

SYNOPSIS. Electron microscopic studies are reported on glycerinatedskeletal and cardiac muscle of a benthic fish, Coryphaenoidesspecies. In white skeletal muscle, the sarcomeres have a restinglength of approximately 1.8 µ, with thick filaments 1.4µ and thin filaments 0.75 µ in length. These dimensionsare somewhat shorter than filament lengths of oilier vertebratemuscles, possibly due to the elfect of volume increase duringassembly of thick and thin filaments at high hydrostatic pressure.During ATP-induced contraction of Coryphaenoides muscle fromsarcomere lengths of 1.8 µ to 1.6 µ, there is acharacteristic interdigitation of thick and thin filaments,with decrease in I band length and no change in length of thickor thin filaments. However, in sarcomeres contracted to lengthsof 1.5 µ. to 1.2 µ, there is a slight shorteningof the A band, apparently due to shortening of thick filaments,that occurs despite the presence of residual I band in the samesarcomeres. There is no obvious crumpling or distortion of thickfilaments during contraction to sarcomere lengths as low as1.0 µ, but filament organization undergoes extensive disarrayat sarcomere lengths approaching 0.7 µ. Although effectsfrom heterogeneity of filament length cannot be excluded withcertainty, the present evidence does suggest that contractionot Coryphaenoides muscle from 1.6 µ to 1.0 µ sarcomerelengih is accompanied by shortening of thick filaments consequentto a structural change within the thick filament core.     

Biogeochemistry of microbial mats under Precambrian environmental conditions: a modelling study

Microbial mats have arguably been the most important ecosystem on Earth over its 3.5 Gyr inhabitation. Mats have persisted as consortia for billions of years and occupy some of Earth's most hostile environments. With rare exceptions (e.g. microbial mats developed on geothermal springs at Yellowstone National Park, USA), today's mats do not exist under conditions analogous to Precambrian habitats with substantially lower oxygen and sulphate concentrations. This study uses a numerical model of a microbial mat to investigate how mat composition in the past might have differed from modern mats. We present a numerical model of mat biogeochemistry that simulates the growth of cyanobacteria (CYA), colourless sulphur bacteria (CSB), and purple sulphur bacteria (PSB), with sulphate‐reducing bacteria (SRB) and heterotrophic bacteria represented by parameterized sulphate reduction rates and heterotrophic consumption rates, respectively. Variations in the availability of light, oxygen, sulphide, and sulphate at the upper boundary of the mat are the driving forces in the model. Mats with remarkably similar biomass and chemical profiles develop in models under oxygen boundary conditions ranging from 2.5 × 10^?13 to 0.25 mm and sulphate boundary concentrations ranging from 0.29 to 29 mm , designed to simulate various environments from Archean to modern. The modelled mats show little sensitivity to oxygen boundary conditions because, independent of the overlying oxygen concentrations, cyanobacterial photosynthesis creates similar O₂ concentrations of 0.45–0.65 mm in the upper reaches of the mat during the photoperiod. Varying sulphate boundary conditions have more effect on the biological composition of the mat. Sulphide generated from sulphate reduction controls the magnitude and distribution of the PSB population, and plays a part in the distribution of CSB. CSB are the most sensitive species to environmental change, varying with oxygen and sulphide.     

How cold‐adapted flightless flies dispersed over the northern hemisphere: phylogeny and biogeography of the snow fly genus Chionea Dalman (Diptera: Limoniidae)

A phylogeny of the 37 known species and subspecies of the micropterous snow fly genus Chionea Dalman is presented using adult morphological characters. The genus contains two major clades: a strictly Palaearctic clade, and a combined Nearctic‐Palaearctic clade with representatives in the Nearctic and Western Palaearctic regions. As there is little congruence between the recovered phylogeny of Chionea and the currently used subgeneric division in Chionea s.s. and Sphaeconophilus Becker, we propose to abandon the use of subgeneric taxa in Chionea. A strictly morphological analysis appears to be insufficient to fully resolve the phylogeny of the genus at the species level, and future molecular work should provide additional evidence for the establishment of relationships among the members of Chionea. The large‐scale historical biogeography of Chionea was analysed using dispersal‐vicariance analysis. The initial distribution area of the genus probably extended in the Eastern Palaearctic, and the Nearctic and the origin of Chionea could be dated in the Late Cretaceous. The various dispersal and vicariance events that led to the major speciation events in the genus are set against major paleogeographic developments. The ancestor of the Western Palaearctic group in the second major clade originated from the Nearctic. The presence of the cold‐adapted Chionea in currently temperate to warm climatic zones in the southern parts of its distribution was analysed using ecological niche modelling. It appears that prolonged periods of climate cooling, as occurred during the Last Glacial Maximum, enabled Chionea to cover large parts of central and southern Europe and reach the southern distribution areas where the genus is present today. A similar biogeographic pattern was less evident in the Nearctic region.     

Effects of Cyclophosphamide on Visceral Leishmaniasis in the Mouse*

SYNOPSIS Cyclophosphamide (Cy), 125 or 200 mg/kg body weight, injected intraperitoneally (i.p.) into BALB/c mice one day before infection with amastigotes of Leishmania donovani, by the 8th day postinfection caused a significant decrease in the mean numbers of parasites in spleens and livers when compared to mice injected with phosphate buffered saline (PBS). When 125 mg/kg was injected into chronically infected mice (on day 34 of infection), however, within 2 days (day 36) mean parasite levels in both the spleens and livers were statistically greater than in PBS-treated controls. Similarly, when a series of 6 Cy injections, 50 mg/kg each, was injected over a period of 8 days during the chronic stage of infection, the mean parasite levels in both spleens and livers were significantly increased over those of PBS-treated controls. Druing the chronic stage of infection, Cy injections suppressed the immunity to superinfection. Neither plasma nor parasitized peritoneal macrophages obtained from Cy-treated mice, when compared to PBS-treated mice, differed in their respective capacities to influence the growth of intracellular amastigote of L. donovani in vitro. Passive transfer of hyperimmune mouse serum could not reverse the immunosuppressive effects of Cy upon chronic leishmaniasis in the mouse. It is suggested that neither readily demonstrable anti-leishmanial humoral factors nor “immune” macrophages per se, plays a major role in acquired immunity to leishmaniasis in the mouse.