Mitochondrial DNA variation and population genetic structure of the northern redbelly dace (Phoxinus eos)

Two sections of the control region and the genes coding for NADH dehydrogenase sub-units 5 and 6 (ND-5/6) of mitochondrial DNA (mtDNA) were amplified from Phoxinus eos with the polymerase chain reaction. Both sections of the control region were sequenced directly while the ND-5/6 fragment was sequenced in from each end only. Additionally, the entire ND-5/6 fragment was examined for sequence variation using RFLP analysis. No sequence variation was detected in the control region among 70 individuals sampled from 18 populations across three Ontario regions (Spanish River, Madawaska R. and Cataraqui R.). To examine ND-5/6 variation, a total of 75 individuals were sampled from five populations representing two of the three regions (Madawaska River and Cataraqui R.). Six haplotypes were detected by direct sequencing and four by RFLP analysis. Estimates of population subdivision from RFLP data, sequence analysis, and the two data sets combined for the ND-5/6 fragment, suggest that gene flow is restricted within and between regions. However, estimates of sequence divergence for both sequence and RFLP analysis of this fragment suggested that populations were either founded by already differentiated populations or that populations were founded by a single stock and divergence between regions occurred prior to isolation of populations within regions. These estimates of population structure are much greater than those obtained from allozyme analysis. Additionally, high levels of heterozygosity in nuclear DNA, but low mtDNA diversity suggests that populations have experienced reductions in population size sufficient to reduce only mtDNA variation. Random lineage extinction and limited time for the accumulation of new mutations are likely responsible for low levels of mtDNA variation in ND-5/6 and the control region, while functional constraints may limit variation more than expected in the control region in dace and other fishes.     

Microhabitat selection by larval Chironomus tentans (Diptera: Chironomidae): effects of predators, food, cover and light

1. As part of a study designed to estimate the developmental costs of antipredator behaviour of larval chironomids, we used laboratory experiments to study effects of food and factors that could influence predation risk [presence of fish, cover from fish (simulated debris) and light level], on microhabitat selection by Chironomus tentans larvae in the third and fourth instar. 2. Larvae were more likely to build tubes where there was more food although their ability to move far to find food appeared limited. 3. Larvae did not avoid areas with fish and the presence of fish did not alter larval response to food. 4. Larvae avoided areas of cover (simulated debris) but cover did not alter larval response to food. 5. When provided with a choice between light and dark areas, larvae initially without tubes were found more often in the dark areas. Light level had no effect on location of larvae that had begun the experiment with tubes. 6. Results suggest the tubicolous life-style of larval Chironomus tentans limits their ability to select microhabitats that could alter their risk of predation.     

Characterization of Encephalitozoon (Septata) intestinalis Isolates Cultured from Nasal Mucosa and Bronchoalveolar Lavage Fluids of Two AIDS Patients

ABSTRACT. Microsporidia are obligate intracellular protozoan parasites that can cause opportunistic infections in AIDS patients. Species from five genera of microsporidia are presently known to infect man. One species, Septata intestinalis originally was detected in stool specimens of individuals with chronic diarrhea and subsequently was found to disseminate to the kidneys, lungs, and nasal sinuses. This organism has since been reclassified as Encephalitozoon and in this study, we report the culture of Encephalitozoon intestinalis from a bronchoalveolar lavage specimen and a nasal mucus aspirate of two AIDS patients living in the USA. The bronchoalveolar and nasal microsporidian isolates grew in several continuous cell lines including RK-13, MDCK, HT-29, Caco-2, Vero, and 1047. Transmission electron microscopy of the clinical and cell culture specimens revealed that the new isolates appeared to be E. intestinalis based on morphology and growth of organisms in septated membrane-bound parasitophorous vacuoles. The new E. intestinalis isolates were characterized and compared with the first isolated E. intestinalis that was cultured from stool to confirm their identity and to determine if there existed any minor differences, as seen in the closely related Encephalitozoon cuniculi strains. By the methods of sodium dodecyl sulfate-polyacrylamide gel electrophoresis staining for proteins and carbohydrates, Western blot immunodetection, and polymerase chain reaction-based methods with restriction endonuclease digestion, double-stranded DNA heteroduplex mobility shift analysis, and DNA sequencing of the ribosomal DNA intergenic spacer region, the new isolates were identical to each other and to the reference isolate of E. intestinalis. In addition, with any of these methods, the E. intestinalis organisms could be distinguished from the three E. cuniculi strains, Encephalitozoon hellem, and Vittaforma corneae, which is important for diagnostics, therapeutic strategies, and epidemiology.     

Zinc, Lead and Cadmium Tolerance, Uptake and Accumulation by the Common Reed,Phragmites australis(Cav.) Trin. ex Steudel

Zinc (Zn), lead (Pb) and cadmium (Cd) tolerance in populationsof seedlings ofPhragmites australisraised from seeds collectedfrom a mine site (Plombières, Belgium) contaminated withZn, Pb and Cd and three ‘clean’ sites (Felixstowe,UK; Wisbech, UK; and Mai Po, Hong Kong) were studied under glasshouseconditions. Small differences were found between the metal-contaminatedpopulation and the three ‘clean’ populations whenseedlings were grown in 1.0 µg  ml^-1Zn and 10.0 µgml^-1Pb treatment solutions. In general, however, different populationsof seedlings showed similar growth responses, metal uptake andindices of Zn, Pb and Cd tolerance when cultured in the samemetal-contaminated media for 89 d or in the same metal treatmentsolutions (ZnSO₄:1.0 and 4.0 µg ml^-1Zn; Pb(NO₃)₂: 10.0and 25.0 µg ml^-1Pb; CdSO₄: 0.5 and 1.0 µg ml^-1Cd)for 3 weeks. There was insufficient evidence to support thehypothesis that the metal-contaminated population has evolvedto a Zn-, Pb- or Cd-tolerant ecotype but the results indicatedsome differentiation between the populations with that fromHong Kong being the least productive under the experimentalconditions used. The implications of the findings on selectionof provenances for use in constructed wetlands for wastewatertreatment are discussed. Metal accumulation; heavy metal tolerance; Phragmites australis; population differentiation     

Comparative floral structure and systematics of Pelagodoxa and Sommieria (Arecaceae)

Floral structure is compared in Pelagodoxa and Sommieria (Arecaceae, Arecoideae). Male flowers have three free, imbricate sepals, three basally congenitally united and apically valvate petals, and six stamens. Anthers are dorsifixed and dehiscence introrse. The sterile gynoecium is tricarpellate. Female flowers have three free, imbricate sepals and three free, imbricate petals, which are slightly fused with the sepals at the base. Four to six staminodes are congenitally united at the base and fused with the ovary for a short distance. The gynoecium is syncarpous. Carpels are almost equal in early development; later the gynoecium becomes pseudomonomerous. The three stigmatic branches are equally developed, apical and sessile. The carpels are (syn-)ascidiate up to the level of the placenta and (sym-)plicate above. Each carpel has one ovule, in the sterile carpels it is aborted at anthesis. The fertile ovule is erect up to anthesis and pendant afterwards because of the bulging out of the ovary. Pollen tube transmitting tracts (PTTT) encompass the secretory epidermis of the ventral slits of each carpel. Floral structure in Pelagodoxa and Sommieria supports the sister group relationship between the two genera suggested in recent molecular phylogenies and reflects their close relationships to a major clade of pseudomonomerous arecoid palms from the Indo-Pacific region.  © 2004 The Linnean Society of London, Botanical Journal of the Linnean Society , 2004, 146 , 27–39.     

A comparative study on efficiency of adult fibroblasts and amniotic fluid-derived stem cells as donor cells for production of hand-made cloned buffalo (Bubalus bubalis) embryos

The efficiency of two cell types, namely adult fibroblasts, and amniotic fluid stem (AFS) cells as nuclear donor cells for somatic cell nuclear transfer by hand-made cloning in buffalo (Bubalus bubalis) was compared. The in vitro expanded buffalo adult fibroblast cells showed a typical “S” shape growth curve with a doubling time of 40.8 h and stained positive for vimentin. The in vitro cultured undifferentiated AFS cells showed a doubling time of 33.2 h and stained positive for alkaline phosphatase, these cells were also found positive for undifferentiated embryonic stem cell markers like OCT-4, NANOG and SOX-2, which accentuate their pluripotent property. Further, when AFS cells were exposed to corresponding induction conditions, these cells differentiated into osteogenic, adipogenic and chondrogenic lineages which was confirmed through alizaran, oil red O and alcian blue staining, respectively. Cultured adult fibroblasts and AFS cells of passages 10–15 and 8–12, respectively, were used as nuclear donors. A total of 94 embryos were reconstructed using adult fibroblast as donor cells with cleavage and blastocyst production rate of 62.8 ± 1.8 and 19.1 ± 1.5, respectively. An overall cleavage and blastocyst formation rate of 71.1 ± 1.2 and 29.9 ± 2.2 was obtained when 97 embryos were reconstructed using AFS cells as donor cells. There were no significant differences (P > 0.05) in reconstructed efficiency between the cloned embryos derived from two donor cells, whereas the results showed that there were significant differences (P < 0.05) in cleavage and blastocyst rates between the cloned embryos derived from two donor cell groups. Average total cell numbers for blastocyst generated using AFS cells (172.4 ± 5.8) was significantly (P < 0.05) higher than from adult fibroblasts (148.2 ± 6.1). This study suggests that the in vitro developmental potential of the cloned embryos derived from AFS cells were higher than that of the cloned embryos derived from adult fibroblasts in buffalo hand-made cloning.     

A histological and immunohistochemical study of the effects of N-acetyl cysteine on retinopathy of prematurity by modifying insulin-like growth factor-1

Retinopathy of prematurity (ROP) is a vasoproliferative disorder that occurs in premature infants and may lead to permanent visual impairment. We investigated both the possible protective role of N-acetyl cysteine (NAC) for preventing ROP and the role of IGF-1 in the disorder. Forty-five newborn rats were divided into three groups. Group 1 was raised in room air as controls. Group 2 was exposed to 60% oxygen for 14 days after birth, then transferred to room air. Group 3 was exposed to the same conditions as group 2, but received intraperitoneal injections of NAC on postnatal days 7–17. After 35 days, both eyes of all rats were processed for histology. Some sections were stained with hematoxylin and eosin to assess structural changes and other sections were immunostained to determine the location of IGF-1. Frozen sections also were prepared and stained for adenosine triphosphatase to detect retinal blood vessels. Compared to the controls, more blood vessels, many of which were abnormal, and increased IGF-1 expression were observed in group 2. In group 3, abnormal blood vessels and IGF-1 expression were less evident. NAC appeared to be an effective vascular-protective agent for ROP by decreasing IGF-1 expression.     

Hydrolytic Enzymes of Euglena gracilis: Characterization and Activity as a Function of Culture Age and Carbon Deprivation*†

SYNOPSIS. Optimal assay conditions are described for 8 hydrolases of Euglena gracilis var. bacillaris, SM-L1 (streptomycinbleached) strain, 7 of which have an acid pH-optimum. Acid phosphatase, β-galactosidase, β-glucosidase, β-fucosidase, cathepsin D, RNase, DNase, and an esterase are active in cell homogenates. Amylase has very low activity, and β-glucuronidase, arylsulfatase, β, N-acetyl-glucosaminidase, α-fucosidase, and α- and β-mannosidase are inactive. Hydrolase activity increases as a culture proceeds from the midexponential to the late stationary-phase of growth, being most pronounced in the case of β-glucosidase. In cultures deprived of a utilizable carbon source, the specific activities of the hydrolases (per mg total protein or dry weight) increase. When expressed on a per cell basis, however, the activities of DNase decrease while those of β-galactosidase, cathepsin D, and RNase increase. The hydrolases appear to be involved in the adaptation of Euglena to the metabolic demands imposed by different conditions of growth.     

Sedimentation of Bovine Rhodopsin—Digitonin Micelles

RHODOPSIN, the photo-sensitive pigment of vertebrate vision receptors, consists of the lipoprotein opsin bound to the 11-cis isomer of retinal. Light isomerizes the 11-cis configuration to the all-trans, which makes the pigment unstable, leading eventually to the dissociation of the retinal from the lipoprotein. The belief that these dark steps involve conformational changes in the lipoprotein moiety stems from spectroscopic measurements which show the disappearance of lipoprotein-chromophore interactions and from kinetic and thermodynamic considerations^1–5, but more direct evidence has come from the changes in circular dichroism and optical rotatory dispersion which occur with bleaching^6–9. There is also an increase of Stokes radius on bleaching¹¹.