Natural or replacer sources of milk in lambs during feeding adaptation: influences on performance, metabolism of protein and lipid and oxidative/antioxidant status.

This study aimed to evaluate the effect of milk feeding using milk replacer (MR) or natural ewe's milk (NEM) for Lacaune lambs on performance, protein and lipid metabolism and oxidative/antioxidant status. These parameters were evaluated during the period of feeding adaptation, which corresponded to 12 days after ewe's separation. Fifteen lambs were selected and divided into two groups: Group A (n = 7) with lambs receiving natural milk; Group B (n = 8) with lambs receiving a milk replacer. Liquid food supply, for both groups, was set according to their body weight, given in two daily periods. Feed intake was also obtained by individual body weight and adjusted according leftovers. Blood samples were collected on three different days (0, 6 and 12) to perform the serum analysis of total protein, albumin, cholesterol, triglycerides, alanine aminotransferase (ALT) and urea. Additionally, the oxidative profile was analysed in sera samples through the assessment of TBARS (reactive substances thiobarbituric acid) and ROS (reactive oxygen species) which aims to identify lipid peroxidation and free radical levels, respectively, as well as the antioxidant profile evaluating glutathione S-transferase (GST). There was no statistical difference (p > 0.05) between groups on levels of total protein, albumin and globulin. However, cholesterol (day 6), triglycerides (days 6 and 12), urea (day 6) and ALT (day 12) levels differed (p < 0.05) between groups. Values of total protein, globulin, ALT and TBARS differed (p < 0.05) along the time. The average weight gain and body weight differed (p < 0.05) only on day 12, showing that weight gain was higher for lambs fed with NEM. The performance of lambs fed with MR was impaired. Biochemical parameters had a slight variation between groups and along the time, as well as oxidative/antioxidant status did not change in the different liquid diets.

Vertical distribution of methanogens in the anoxic sediment of Rotsee (Switzerland).

Anoxic sediments from Rotsee (Switzerland) were analyzed for the presence and diversity of methanogens by using molecular tools and for methanogenic activity by using radiotracer techniques, in addition to the measurement of chemical profiles. After PCR-assisted sequence retrieval of the 16S rRNA genes (16S rDNA) from the anoxic sediment of Rotsee, cloning, and sequencing, a phylogenetic analysis identified two clusters of sequences and four separated clones. The sequences in cluster 1 grouped with those of Methanosaeta spp., whereas the sequences in cluster 2 comprised the methanogenic endosymbiont of Plagiopyla nasuta. Discriminative oligonucleotide probes were constructed against both clusters and two of the separated clones. These probes were used subsequently for the analysis of indigenous methanogens in a core of the sediment, in addition to domain-specific probes against members of the domains Bacteria and Archaea and the fluorescent stain 4', 6-diamidino-2-phenylindole (DAPI), by fluorescent in situ hybridization. After DAPI staining, the highest microbial density was obtained in the upper sediment layer; this density decreased with depth from (1.01 +/- 0.25) x 10(10) to (2.62 +/- 0.58) x 10(10) cells per g of sediment (dry weight). This zone corresponded to that of highest metabolic activity, as indicated by the ammonia, alkalinity, and pH profiles, whereas the methane profile was constant. Probes Eub338 and Arch915 detected on average 16 and 6% of the DAPI-stained cells as members of the domains Bacteria and Archaea, respectively. Probe Rotcl1 identified on average 4% of the DAPI-stained cells as Methanosaeta spp., which were present throughout the whole core. In contrast, probe Rotcl2 identified only 0.7% of the DAPI-stained cells as relatives of the methanogenic endosymbiont of P. nasuta, which was present exclusively in the upper 2 cm of the sediment. Probes Rotp13 and Rotp17 did not detect any cells. The spatial distribution of the two methanogenic populations corresponded well to the methane production rates determined by incubation with either [14C]acetate or [14C]bicarbonate. Methanogenesis from acetate accounted for almost all of the total methane production, which concurs with the predominance of acetoclastic Methanosaeta spp. that represented on average 91% of the archaeal population. Significant hydrogenotrophic methanogenesis was found only in the organically enriched upper 2 cm of the sediment, where the probably hydrogenotrophic relatives of the methanogenic endosymbiont of P. nasuta, accounting on average for 7% of the archaeal population, were also detected.

Methanogenic archaea and CO2-dependent methanogenesis on washed rice roots.

Washed excised roots of rice (Oryza sativa) immediately started to produce CH4 when they were incubated in phosphate buffer under anoxic conditions (N2 atmosphere), with initial rates varying between 2 and 70nmolh(-1)g(-1) dry weight of root material (mean +/- SE: 20.3 +/- 5.9 nmol h(-1) g(-1) dry weight; n = 18). Production of CH4 continued for at least 500 h, with rates usually decreasing slowly. CH4 production was not significantly affected by methyl fluoride, an inhibitor of acetoclastic methanogenesis. Less than 0.5% of added [2-14C]-acetate was converted to 14CH4, and conversion of 14CO2 to 14CH4 indicated that CH4 was almost exclusively produced from CO2. Occasionally, however, especially when the roots were incubated without additional buffer, CH4 production started to accelerate after about 200h reaching rates of > 100 nmol h(-1) g(-1) dry weight. Methyl fluoride inhibited methanogenesis by more than 20% only in these cases, and the conversion of 14CO2 to 14CH4 decreased. These results indicate that CO2-dependent rather than acetoclastic methanogenesis was primarily responsible for CH4 production in anoxically incubated rice roots. Determination of most probable numbers of methanogens on washed roots showed highest numbers (10(6)g(-1) dry roots) on H2 and ethanol, i.e. substrates that support CH4 production from CO2. Numbers on acetate (10(5) g(-1) dry roots) and methanol (10(4)g(-1) dry roots) were lower. Methanogenic consortia enriched on H2 and ethanol were characterized phylogenetically by comparative sequence analysis of archaeal small-subunit (SSU) ribosomal RNA-encoding genes (rDNA). These sequences showed a high similarity to SSU rDNA clones that had been obtained previously by direct extraction of total DNA from washed rice roots. The SSU rDNA sequences recovered from the H2/CO2-using consortium either belonged to a novel lineage of methanogens that grouped within the phylogenetic radiation of the Methanosarcinales and Methanomicrobiales or were affiliated with Methanobacterium bryantii. SSU rDNA sequences retrieved from the ethanol-using consortium either grouped within the genus Methanosarcina or belonged to another novel lineage within the phylogenetic radiation of the Methanosarcinales and Methanomicrobiales. Cultured organisms belonging to either of the two novel lineages have not been reported yet.

Isolation of acidophilic methane-oxidizing bacteria from northern peat wetlands.

Acidic northern wetlands are an important source of methane, one of the gases that contributes to global warming. Methane oxidation in the surface of these acidic wetlands can reduce the methane flux to the atmosphere up to 90 percent. Here the isolation of three methanotrophic microorganisms from three boreal forest sites is reported. They are moderately acidophilic organisms and have a soluble methane monooxygenase. In contrast to the known groups of methanotrophs, 16S ribosomal DNA sequence analysis shows that they are affiliated with the acidophilic heterotrophic bacterium Beijerinckia indica subsp. indica.

Diversity and structure of the methanogenic community in anoxic rice paddy soil microcosms as examined by cultivation and direct 16S rRNA gene sequence retrieval.

A dual approach consisting of cultivation and molecular retrieval of partial archaeal 16S rRNA genes was carried out to characterize the diversity and structure of the methanogenic community inhabiting the anoxic bulk soil of flooded rice microcosms. The molecular approach identified four groups of known methanogens. Three environmental sequences clustered with Methanobacterium bryantii and Methanobacterium formicicum, six were closely related but not identical to those of strains of Methanosaeta concilii, two grouped with members of the genus Methanosarcina, and two were related to the methanogenic endosymbiont of Plagiopyla nasuta. The cultivation approach via most-probable-number counts with a subsample of the same soil as an inoculum yielded cell numbers of up to 10(7) per g of dry soil for the H2-CO2-utilizing methanogens and of up to 10(6) for the acetate-utilizing methanogens. Strain VeH52, isolated from the terminal positive dilution on H2-CO2, grouped within the phylogenetic radiation characterized by M. bryantii and M. formicicum and the environmental sequences of the Methanobacterium-like group. A consortium of two distinct methanogens grew in the terminal positive culture on acetate. These two organisms showed absolute 16S rRNA gene identities with environmental sequences of the novel Methanosaeta-like group and the Methanobacterium-like group. Methanosarcina spp. were identified only in the less-dilute levels of the same dilution series on acetate. These data correlate well with acetate concentrations of about 11 microM in the pore water of this rice paddy soil. These concentrations are too low for the growth of known Methanosarcina spp. but are at the acetate utilization threshold of Methanosaeta spp. Thus, our data indicated Methanosaeta spp. and Methanobacterium spp. to be the dominant methanogenic groups in the anoxic rice soil, whereas Methanosarcina spp. appeared to be less abundant.

Two new restriction endonucleases DraII and DraIII from Deinococcus radiophilus.

In addition to recently characterized DraI (1), two new Type II restriction endonucleases, DraII and DraIII, with novel site-specificities were isolated and purified from Deinococcus radiophilus ATCC 27603. DraII and DraIII recognize the hepta- and nonanucleotide sequences (sequence in text) The cleavage sites within both strands are indicated by arrows. The recognition sequences were established by mapping of the cleavage sites on pBR322 (DraII) and fd109 RF DNA (DraIII). The sequence specifities were confirmed by computer-assisted restriction analyses of the generated fragment patterns of the sequenced DNA's of the bacteriophages lambda, phi X174 RF, M13mp8 RF and fd109 RF, the viruses Adeno2 and SV40, and the plasmids pBR322 and pBR328. The cleavage positions within the recognition sequences were determined by sequencing experiments.

Analysis of a DNA segment from rat liver mitochondria containing the genes for the cytochrome oxidase subunits I, II and III, ATPase subunit 6, and several tRNA genes.

The sequence of a 6.24 kb DNA segment of the mitochondrial genome from rat liver has been determined. It comprises several genes coding for mitochondrial protein subunits and five tRNA genes in the following order: cytochrome oxidase subunit I - tRNA (UCN) (Ser) -tRNA(Asp) - cytochrome oxidase subunit II - tRNALys -ATPase subunit - cytochrome oxidase subunit III -tRNA(Gly) - potential open reading frame - tRNA(Arg) -two potential open reading frames. The tRNA genes were detected by a computer search programme. The assignments for the protein coding sequences were made through comparison with known sequences, mainly from the yeast mitochondrial proteins (e.g. Bonitz et al. 1980). Our data are discussed with regard to the features of gene arrangement, codon usage, and tRNA structure in mammalian mitochondria (Anderson et al. 1981).

tRNA genes in rat liver mitochondrial DNA.

Continuing the analysis of rat liver mitochondria) DNA, we have determined the sequences of two segments containing several tRNA genes. One of these segments also comprises the L-strand origin of replication. The characteristics of the 15 rat liver mitochondrial tRNA structures known at present are compared with those from other mammalian mitochondria.

A detailed restriction endonuclease cleavage map of rat liver mitochondrial DNA.

A restriction endonuclease cleavage map of rat liver mitochondrial DNA is presented for the following enzymes: Xba I, Bgl II, Hae II, Bam HI, Hpa I, Hha I, Bcl I, Hind II, Hind III, Eco RI, Hpa II, Hae III, and Sau 3A. It was derived from complete and partial digestions with these enzymes, double digestions, and redigestions of defined fragments obtained with one enzyme with other restriction enzymes. By the use of these and further enzymes (Taq YI, Alu I) the mitochondrial DNA (ca. 15.6 Kb) can be dissected into a large number of defined fragments.

[A method of microphotometry with TV-image analysing systems (author's transl)]. / Verfahren für photometrische Messungen mit Fernseh-Bildanalysegeräten

In scanning microphotometry only a small spot in the specimen is illuminated at the same time. However, lamp and optical system are designed to transfer 10(5) to 10(6) times more information at once. It is shown, that correct and reproducible results can be attained with stimultaneous illumination of the field received by a TV-camera. It is necessary to calibrate for stray light. Thus the high speed of measurement of TV-Systems can be used for microphotometry.