The protelomerase of temperate Escherichia coli phage N15 has cleaving-joining activity.

Escherichia coli phage N15 encodes the slightly acidic, 630-residue protein of 72.2 kDa called protelomerase (TelN). TelN is a component of the N15 replication system proposed to be involved in the generation of the linear prophage DNA. This linear DNA molecule has covalently closed ends. The reaction converting circular plasmids into linear molecules was catalyzed in vitro. We demonstrate that the product of telN functions as the protelomerase in the absence of other N15-encoded factors. Purified TelN processes circular and linear plasmid DNA containing the proposed target site telRL to produce linear double-stranded DNA with covalently closed ends. The 56-bp telRL target site consists of a central telO palindrome of 22 bp and two 14-bp flanking sequences comprising inverted repeats. telO is separated from these repeats by 3 bp on each side. The telRL sequence is sufficient for TelN-mediated processing. The ends of the DNA molecules generated in vitro have the same configuration as do those observed in vivo. TelN exerts its activity as cleaving-joining enzyme in a concerted action.

Escherichia coli and Bacillus subtilis PriA proteins essential for recombination-dependent DNA replication: involvement of ATPase/helicase activity of PriA for inducible stable DNA replication.

The E. coli PriA protein, a DEXH-type DNA helicase with unique zinc finger-like motifs interrupting the helicase domains, is an essential component of the phiX174-type primosome and plays critical roles in RecA-dependent inducible and constitutive stable DNA replication (iSDR and cSDR, respectively) as well as in recombination-dependent repair of double-stranded DNA breaks. B. subtilis PriA (BsPriA) protein contains the conserved helicase domains as well as zinc finger-like motifs with 34% overall identity with the E. coli counterpart. We overexpressed and purified BsPriA and examined its biochemical properties. BsPriA binds specifically to both n'-pas (primosome assembly site) and D-loop and hydrolyzes ATP in the presence of n'-pas albeit with a specific activity about 30% of that of E. coli PriA. However, it is not capable of supporting n'-pas-dependent replication in vitro, nor is it able to support ColE1-type plasmid replication in vivo which requires the function of the phiX174-type primosome. We also show that a zinc finger mutant is not able to support recombination-dependent DNA replication, as measured by the level of iSDR after a period of thymine starvation, nor wild-type level of growth, cell morphology and UV resistance. Unexpectedly, we discovered that an ATPase-deficient mutant (K230D) is not able to support iSDR to a full extent, although it can restore normal growth rate and UV resistance as well as non-filamentous morphology in priA1::kan mutant. K230D was previously reported to be fully functional in assembly of the phiX174-type primosome at a single-stranded n'-pas. Our results indicate that ATP hydrolysis/ helicase activity of PriA may be specifically required for DNA replication from recombination intermediates in vivo.

Transsynovial kinetics of lonazolac and its hydroxy metabolite in synovitis patients.

OBJECTIVE: The study was designed to characterize the synovial distribution profiles and kinetics of the non-steroidal antiinflammatory agent, lonazolac, in patients with synovitis after multiple dosing with 300 mg tablets of lonazolac calcium salt. METHODS: Forty patients (36 male, 4 female) aged 21 to 50 years (mean: 38+/-9 years) undergoing arthroscopy of the knee joint for surgical reasons were given 7 total doses of drug administered as 300 mg oral tablets of lonazolac-calcium taken twice daily. Patients were assigned to one of 4 treatment groups (n = 10) in which arthroscopy was carried out 1, 2, 6, or 12 h after the seventh lonazolac dose. Samples of blood, synovial fluid, and synovial membrane were obtained during each operation and used to determine total concentrations of lonazolac and its main metabolite in plasma and synovial fluid by HPLC assay with UV detection. Free lonazolac concentrations in body fluids were determined after ultrafiltration by the same HPLC technique using a fluorescence detector. Tissue concentrations were assayed after additional steps using solvent and solid phase extractions. Total protein contents in plasma and synovial fluid were measured spectrophotometrically. RESULTS: Plasma drug levels were highest at 1 hour after dosing with mean peak concentrations of 1.8 mg/l total lonazolac, 1.2 mg/l total metabolite, and 9 microg/l free lonazolac. Profiles indicated a biphasic decline. Concentration vs. time profiles in synovial fluid were flattened compared to plasma profiles with mean peak values of 440 microg/l total lonazolac, 370 microg/l total metabolite, and 7 microg/l free lonazolac attained 2 hours after dosing. The mean unbound fraction of lonazolac was higher in synovial fluid (1.9%) compared to plasma (0.7%). Transsynovial partition coefficients increased continuously during a dosing interval from 0.16 to 3.15 for total lonazolac and from 0.56 to 5.05 for free lonazolac. Mean total protein contents for each group of patients ranged from 70 to 76 g/l for plasma and 32 to 42 g/l for synovial fluid. Total drug concentrations in synovial membrane were highest in tissues obtained 1 hour after dosing with mean values of approximately 1.0 microg/g dry weight. Tissue samples obtained at later times indicated that lonazolac profiles in tissue more closely resemble profiles obtained for plasma than for synovial fluid. Protein concentration ratios (synovial fluid : plasma) were between 0.45 and 0.58. Except for the absorption phase, transsynovial drug partition coefficients were always higher than the protein concentration ratios. CONCLUSIONS: Protein content is not an important factor for drug partition into inflamed joints after multiple dosing with lonazolac. Lonazolac distributes well into synovial fluid with therapeutically effective concentrations of unbound drug measured within 2 hours after dosing. Total lonazolac levels in synovial fluid exceed those measured in plasma at 6 to 12 hours after administration.

[Etiology of cerebral perfusion disorder in patients with severe craniocerebral trauma]. / Ursachen zerebraler Perfusionsstörungen bei Patienten mit schwerem Schädel-Hirn-Trauma.

In 53 patients with severe head injury, a decline of the cerebral perfusion pressure (CPP) was distinguished related to its cause, i.e., by a rise in intracranial pressure (ICP), a decline in the mean arterial pressure (MAP) or both. The mean Glasgow Outcome Scale (GOS) was 1.6 in patients with elevated ICP, 3.8 in those with decreased MAP and 2.7 when impaired CPP was due to both. It is concluded that in cases of decreased CPP, elevated ICP is the main cause of secondary brain injury and poor outcome, independent of the circulatory condition, whereas an isolated decline in the MAP does not show any influence on functional outcome.

[Prophylactic measures against infection in dairy cattle--necessity of hygiene management]. / Infektionsprophylaktische Massnahmen in Milchviehbeständen--Notwendigkeit eines Hygiene-managementes.

Prophylactic measures against infection are really essential to the preservation of the udder-health and the quality of the milk. When curing a dairy cattle population suffering from mastitis strict hygienic measures for the completion and maintenance of the therapy must be taken absolutely consequently. On the occasion of a study made in Bavaria for the period of one year 9948 dairy farmers were asked if hygienic measures against infection were taken. In detail it was evaluated if the secret was controlled during pre-milking, if the cleaning of the teats before milking and the post-milking teat end disinfection (teat dipping) was done, if dry cow therapy was practiced, if the milking machine was serviced and ist functioning checked regularly.

[Intracranial pressure monitoring in patients with severe craniocerebral injury]. / Hirndruck-Monitoring bei Patienten mit schwerem Schädel-Hirn-Trauma.

After severe head injury intracranial pressure (ICP) must be measured continuously for management to assess and maintain the cerebral perfusion. Therefore in our hospital epidural transducers are used. To prove the efficiency of this method in a 12-month period the clinical courses of 23 patients with intracranial pressure transducers were analysed retrospectively. Eighteen patients survived, 5 of them without residuals, 13 with residuals and 2 remained in coma. In 14 patients secondary rises of intracranial pressure were observed between days 3 and 6 post injury. The mean ICP value of the survivors revealed 25 mm Hg. whereas the expired showed 60 mm Hg. In 17 patients the measurements were considered as reliable, 6 measurements were not reliable, which included 1 of the 5 patients who died. One transduce was displaced, another one showed a hemorrhage at the drill hole. There was no infection.

[Measuring epidural intracranial pressure in patients with severe craniocerebral trauma]. / Epidurale Hirndruckmessung bei Patienten mit schwerem Schädel-Hirn-Trauma.

In 30 patients with severe head injury (SHI), intracranial pressure (ICP) was monitored using epidural transducers. In 22 patients, the measurements were reliable, with average values of 19.4 mmHg in the survivors and 64.6 mmHg in those who died. It is concluded that epidural measurement of ICP provides a helpful method for the management of SHI and to control the indication for CT scans.

Augmented repair and early mobilization of acute anterior cruciate ligament injuries.

This paper describes a technique for repair of the anterior cruciate ligament with protection of that repair by a synthetic augmentation device. Of 61 patients who underwent surgery 24 to 57 months (mean, 38.3) before data accumulation, 57 returned for followup. Subjectively, 53 (93%) patients reported a good or excellent functional result; however, only 29 (51%) of the patients returned to their preinjury sports level. Objectively, a radiographic Lachman test was performed on both the injured and the noninjured knee of all 57 patients. The preoperative mean difference between the knees was 8.6 mm (range, 4.6 to 17.2) and at followup it was 2.4 mm (range, -0.9 to 11.8). Radiographic abduction and adduction stress tests demonstrated stable healing of the (unrepaired) collateral ligament lesions (varus stress = 0.1 mm and valgus stress = 0.4 mm mean side-to-side difference), indicating that suturing of ruptured collateral ligaments is not necessary when the knee is centrally stabilized with the augmentation device. These results indicate that successful repair of the anterior cruciate ligament is frequently possible when enhanced with an augmentation device.

[The effect of environmental factors on the occurrence of udder diseases]. / Ein Beitrag zum Einfluss von Umweltfaktoren auf das Vorkommen von Eutererkrankungen.

The influence of housing factors on udder diseases was investigated in 2156 tiestall barns with 41893 cows and in 98 loose housed herds with 4126 cows. In loose housed herds less teat injuries were observed and also less mastitis than in tiestall barns. Within the different tiestall systems there was a trend to more udder diseases in short standings with a dung grid than in other systems. The ordinary neck chain and the "Grabner"-tethering system showed better results than the horizontal chain and the yoke tying system. Stall partitions between the cows caused a marked reduction of udder diseases. The installation of a cow trainer showed advantages regarding udder diseases. Inaccurate standing measurements (length and width) caused more udder diseases. When the metal grid was in poor condition, the incidence of teat injuries as well as udder diseases of heifers increased. Teat injuries and udder infections were more frequent when limb or digit injuries were found. Regular hoof care twice a year reduced the udder disease rate. Bedding material influenced the incidence of udder diseases positively. On farms where the cows were on pasture, the incidence of udder infections was higher than on farms where the cows were permanently housed. Heifers that were tied as calves or as juveniles showed significantly less udder diseases than those that were raised in groups. Faults in milking, defects in milking machines and deficits in milking hygiene showed a negative effect on udder diseases.