Stage-specific expression and genomic organization of the actin genes of the malaria parasite Plasmodium falciparum.

Two different actin transcripts are found in the human malaria parasite Plasmodium falciparum. One of these is a 2.5-kb-long RNA found both in asexual blood stages and in the sexual stages (i.e., gametes/zygotes) of the parasite. This transcript is encoded by the P. falciparum (pf)-actin I gene. The second malarial actin gene, the pf-actin II gene, yields a 1.9-kb-long transcript which is formed solely in the sexual stages. Elucidation of the genomic organisation of these two Plasmodium actin genes showed that the pf-actin I gene does not possess any introns whereas the coding region of the pf-actin II gene is interrupted by a 368-bp intron. This intron has consensus splice junction sequences. Nucleotide sequence analysis of the 3' non-coding regions of the pf-actin genes revealed that these regions are quite long (pf-actin I, 250 bp; pf-actin II, 331 bp) and that these trailers do not share sequence similarity. Furthermore, the poly(A)+ addition sites of both actin mRNAs have now been identified. The 5' untranslated regions are also rather long; the sequenced areas lack sequence similarity and have, as do the 3' untranslated regions, a very high A + T content.

Endogenous gene and amplifiable cDNA construct both produce unstable t-PA mRNA in Bowes melanoma cells.

Bowes melanoma cells, which naturally produce tissue-type plasminogen activator (t-PA), were transfected with a plasmid containing a human t-PA cDNA under transcriptional control of the promoter/enhancer of the major immediate early gene of human cytomegalovirus (CMV) plus genes expressing geneticin (G418) resistance and dihydrofolate reductase (DHFR). In one of the initial geneticin-resistant transformants, t-PA mRNA transcribed from the chromosomally integrated plasmid had the same short half-life, 20-30 min, as did mRNA transcribed from the endogenous t-PA gene compared to 7-8 h for total poly(A)+ mRNA. After subsequent selection of such cells with methotrexate, a cell line was obtained in which the t-PA cDNA construct was co-amplified with the DHFR gene and which produced 10 times more t-PA protein than the original Bowes melanoma cells.

The applicability of ambulatory electroencephalography (AEEG) in healthy horses and horses with abnormal behaviour or clinical signs of epilepsy.

BACKGROUND: Short-duration electroencephalography (EEG) recordings in horses are helpful in diagnosing intracranial disorders. Potentially, long-duration ambulatory EEG (AEEG) recordings in horses will enhance the chance of detecting abnormal brain activity independent of the presence of an insult. OBJECTIVE: The objective of this study was to test if AEEG recordings in unsedated horses can be acquired and benefit diagnosing abnormal brain activity. ANIMALS AND METHODS: Recordings were taken from 8 adult control horses and 10 patients suspected of intracranial abnormalities. Self-adhesive electrodes and the 'Porti-5' recording system were used. Filter settings were 0.5 Hz high pass and 35 Hz low pass. The records were analysed offline at a 50-200 µV/division and 10 seconds/division scale. Abnormal activity was defined as a spike or sharp wave, a period of generalised slow wave rhythmical activity or a generalised fast rhythmical discharge. The recording time ranged from 5 to 49 hours. RESULTS: In the control group, one horse showed pathological activity. In the patient group, six out of nine horses showed abnormal activity during the recordings. Magnetic resonance imaging confirmed the presence of an intracranial mass in one patient. Long-term recordings of high quality can be obtained in unsedated horses by allowing daily activity using AEEG, resulting in a reasonable chance of recording (inter)ictal abnormal brain activity indicating epileptic or seizure-like activity in the absence of clinical signs or seizures. CONCLUSIONS: It is concluded that abnormal behaviour can be expressed intermittently, and with the availability of AEEG a useful tool is added to the diagnostic scenario for horses.

Increased susceptibility of transfected prokaryotic and eukaryotic cells to antibiotic selection.

At 39 to 40 degrees C, selection of antibiotic-resistant transfected mammalian cell lines or Escherichia coli required lower aminoglycoside antibiotic concentrations than at 37 degrees C. The thermosensitivity of antibiotic susceptibility was much more manifest during genetic selection experiments than in conventional growth inhibition assays.

Influence of recording site on CMAP amplitude on its variation over a length of nerve.

The distinctions between blocking, abnormal temporal dispersion, and normal conduction require delineation of the normal change in amplitude of the compound muscle action potential (CMAP) over a length of nerve. Effects of the recording site on CMAP amplitude and on its variation were studied in median and ulnar nerves of 13 healthy subjects. CMAPs were recorded from three sites: halfway along the muscles and 1 cm distal and proximal. Elbow-wrist amplitude percentages (CMAP%) were calculated. CMAP amplitudes varied considerably between sites and subjects. Amplitudes were maximal at the middle site in only 16 of 26 nerves. The site of maximal amplitude could not be identified on the basis of thumb anatomy. CMAP% was not related to CMAP amplitude, and differed by up to 32% between adjacent sites. CMAP formation involves spatial factors (electrode site, limb position, and limb anatomy), temporal factors (dispersion), and their interaction, explaining why CMAP% can exceed 100%. The site of the recording electrode affects CMAP amplitude and CMAP% to clinically relevant degrees. Standardization of the recording site may improve reliability of CMAP% studies.

A dynamic quaternary structure of bovine alpha-crystallin as indicated from intermolecular exchange of subunits.

The structural bovine eye lens protein alpha-crystallin was dissociated in 7 M urea and its four subunits, A1, A2, B1, and B2, were separated by means of ion-exchange chromatography. Homopolymeric reaggregates of these subunits were prepared by removal of the denaturant via dialysis. It was found that subunits were exchanged upon incubation of mixtures of two homopolymers under native conditions. New hybrid species were formed within 24 h as demonstrated by isoelectric focusing. Moreover, native alpha-crystallin molecules also exchanged subunits when incubated with homopolymeric aggregates of B2 subunits. Subunit exchange between native alpha-crystallin molecules is postulated, and a "dynamic quaternary structure" is presented that allows the polydisperse protein to adapt to changes in cytoplasmic conditions upon aging of the lens tissue.

Molecular fingerprinting defines a strain of Salmonella enterica serotype Anatum responsible for an international outbreak associated with formula-dried milk.

Molecular analyses based on plasmid profile typing and pulsed-field gel electrophoresis have defined a strain of Salmonella enterica serotype Anatum associated with the consumption of a particular brand of formula-dried milk responsible for an outbreak in late 1996/early 1997 involving 15 infants and 2 relatives in the UK, and 2 infants in France. The study has demonstrated the value of laboratory-based surveillance involving identification of the outbreak strain at the molecular level coupled with food microbiology and targeted epidemiological investigations, and has highlighted the importance of rapid communication and subsequent international collaboration through the European Union-funded Salm-Net salmonella surveillance network.