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Two machine learning techniques were evaluated for automatic design of a rule-based control of functional electrical stimulation (FES) for locomotion of spinal cord injured humans. The task was to learn the invariant characteristics of the relationship between sensory information and the FES-control signal by using off-line supervised training. Sensory signals were recorded using pressure sensors installed in the insoles of a subject's shoes and goniometers attached across the joints of the affected leg. The FES-control consisted of pulses corresponding to time intervals when the subject pressed on the manual push-button to deliver the stimulation during FES-assisted ambulation. The machine learning techniques used were the adaptive logic network (ALN) [1] and the inductive learning algorithm (IL) [2]. Results to date suggest that, given the same training data, the IL learned faster than the ALN, while both performed the test rapidly. The generalization was estimated by measuring the test errors and it was better with an ALN, especially if past points were used to reflect the time dimension. Both techniques were able to predict future stimulation events. An advantage of the ALN over the IL was that ALN's can be retrained with new data without losing previously collected knowledge. The advantages of the IL over the ALN were that the IL produces small, explicit, comprehensible trees and that the relative importance of each sensory contribution can be quantified.

This study describes a prospective, randomized, clinical trial in patients infected with the protozoa Giardia lamblia. Patients received a 10-day treatment with twice a day doses of either 120,000 U (USP) of bacitracin zinc, 120,000 U (USP) of bacitracin, 120,000 U (USP) of neomycin, or 60,000 U (USP) of bacitracin zinc and 60,000 U (USP) of neomycin. At the first assessment (day 11), all 21 subjects (100%) treated with bacitracin zinc had ceased to show Giardia parasites in their stools compared with 19 (95%) of 20 receiving bacitracin, 20 (90.9%) of 22 subjects receiving neomycin, and 17 (89.5%) of 19 subjects receiving bacitracin zinc plus neomycin. During the two-week follow up period, one (5.3%) of the 19 subjects examined who received bacitracin zinc experienced a recurrence compared with one (6.7%) of 15 receiving bacitracin, one (5.0%) of 20 receiving neomycin, and 0 (0%) of 14 receiving the combination treatment. Final cure rates of 94.7% for bacitracin zinc, 87.5% for bacitracin, 86.4% for neomycin, and 87.5% for bacitracin zinc plus neomycin were obtained. No synergistic activity was noted between bacitracin zinc and neomycin. Side effects were generally limited to nausea, abdominal discomfort, and diarrhea in a small number of patients.

The activity of bacitracin in vitro against Giardia intestinalis, Trichomonas vaginalis and Tritrichomonas foetus was enhanced 5-10 times by equimolar concentrations of zinc. Bacitracin and bacitracin zinc activity was tolerant to either aerobic or anaerobic culture conditions and equally effective against T. vaginalis isolates sensitive or insensitive to metronidazole. This enhancement was not due to zinc toxicity and was zinc dose dependent.

This study extends the previous knowledge of the action of bacitracin zinc against xenic cultures of Entamoeba histolytica. Results presented here, using axenic E. histolytica, confirm the enhanced activity of bacitracin zinc as compared to native bacitracin and demonstrate that this enhancement is due to a direct effect upon the trophozoite. In the presence of zinc, amoebastatic concentrations of bacitracin became amoebacidal. The expression of the amoebacidal characteristics of 5 g/l bacitracin zinc following 24-35 h incubation coincided with the return of cell division by amoeba exposed to the same concentration of native bacitracin. The activity of both bacitracin and bacitracin zinc on trophozoites was unaffected by autoclaving, indicating a mode of action different from that exerted on Staphylococcus aureus. In contrast to previous clinical observations, no synergism could be detected between bacitracin and neomycin.

Twenty-one Entamoeba histolytica isolates from a range of geographical locations and representing 3 different zymodemes were tested under xenic anaerobic conditions for their sensitivities to bacitracin and bactracin zinc. Isolates were over 13 times more sensitive to bacitracin zinc than to bacitracin. Our results indicate that bacitracin zinc may form the basis of an alternative non-toxic treatment of luminal amoebiasis.

A novel crutch, the harness crutch, is described which reduces loading on the arms during the swing phase of swing-through gait. The device was fabricated by attaching a modified mountaineering harness by two side straps, to modified axillary crutches. The harness crutch was compared with the saddle crutch, described by Taylor in 1883. The saddle crutch produced pressures in excess of 500 mmHg in the perineal area; no pressures were produced in this area with the harness crutch. Ischial pressures produced by both systems were similar. In six out of eight non-impaired subjects, significantly more force was transmitted to the harness crutch (an average 47% of body weight) than to the saddle crutch (an average 40% of body weight). A comparison of the oxygen cost of swing-through gait was made between the harness crutch and unmodified axillary crutches; with the harness crutch oxygen cost was significantly lower (p < 0.01) and there were no significant differences in speed and stride length.

The benefits of a functional electrical stimulation (FES) gait programme were assessed in a group of 6 incomplete spinal cord injured subjects. Measurements were made of quadriceps spasticity, lower limb muscle strength, postural stability in standing, spatial and temporal values of gait, physiological cost of gait and independence in activities of daily living. The subjects were assessed before commencement of the programme and after a period of gait training using FES. The benefits derived as a result of the FES gait programme included a reduction in quadriceps tone, an increase in voluntary muscle strength, a decrease in the physiological cost of gait and an increase in stride length.

One symbolic (rule-based inductive learning) and one connectionist (neural network) machine learning technique were used to reconstruct muscle activation patterns from kinematic data measured during normal human walking at several speeds. The activation patterns (or desired outputs) consisted of surface electromyographic (EMG) signals from the semitendinosus and vastus medialis muscles. The inputs consisted of flexion and extension angles measured at the hip and knee of the ipsilateral leg, their first and second derivatives, and bilateral foot contact information. The training set consisted of data from six trials, at two different speeds. The testing set consisted of data from two additional trials (one at each speed), which were not in the training set. It was possible to reconstruct the muscular activation at both speeds using both techniques. Timing of the reconstructed signals was accurate. The integrated value of the activation bursts was less accurate. The neural network gave a continuous output, whereas the rule-based inductive learning rule tree gave a quantised activation level. The advantage of rule-based inductive learning was that the rules used were both explicit and comprehensible, whilst the rules used by the neural network were implicit within its structure and not easily comprehended. The neural network was able to reconstruct the activation patterns of both muscles from one network, whereas two separate rule sets were needed for the rule-based technique. It is concluded that machine learning techniques, in comparison to explicit inverse muscular skeletal models, show good promise in modelling nearly cyclic movements such as locomotion at varying walking speeds.(ABSTRACT TRUNCATED AT 250 WORDS)

In the restoration of gait for paraplegics using functional electrical stimulation, the method most commonly used to produce hip flexion for the swing phase of gait has been the elicitation of the flexion withdrawal response. Several problems have been noted with the response: there is a decrease in the magnitude of the hip flexion to repeated stimuli (habituation); long latency; and inhibition of the response when stimulated bilaterally. These have been characterized and methods for overcoming the problems tested. Results show that increasing stimulation frequency reduces latency. Habituation can be reduced in some subjects by multiplexing two sites of stimulation. Habituation can further be reduced by applying single high-intensity pulses and this has been used in a one-step-ahead controller for regulating hip flexion angle. Inhibition due to bilateral stimulation had been significantly reduced by altering the timing of the stimulation to the two legs.

In budding yeast, two transcription factors, Swi4 and Swi6, control the expression of important cell cycle regulatory proteins (the G1 cyclins, Cln1 and Cln2, and the cyclin-like Hcs26) as well as the HO gene, whose product initiates mating-type switching. Both Swi4 and Swi6 are components of a protein complex that forms at a repeated sequence element, SCB (SWI4, -6-dependent cell cycle box), found in the upstream regulatory sequences of target genes. We show, by using proteins synthesized in vitro, a direct association between Swi4 and Swi6. The cdc10-Swi6 or ankyrin motifs present in both Swi4 and Swi6 are dispensable for their association, which is mediated instead by a region near the C terminus of each protein. Furthermore, we show that interaction with Swi6 is not necessary for specific recognition of the SCB sequence by the Swi4 protein; we propose that Swi4 is responsible for binding to the SCB sequence while Swi6, through its association with Swi4, regulates activity of the complex.

Entry of budding yeast cells into the mitotic cell cycle requires the activity of a conserved regulatory kinase encoded by the CDC28 gene. The kinase is thought to trigger entry into the cell cycle or START, through association with a number of regulatory subunits known as G1 cyclins. A number of genes whose transcription is dependent on CDC28 and thus linked to START are controlled by two transcription factors, SWI4 and SWI6. The genes controlled by SWI4 and SWI6 include two known G1 cyclins (CLN1 and CLN2), a putative new G1 cyclin (HCS26), and the HO gene whose product initiates cell type switching. SWI4 and SWI6 act through a repeated sequence element, SCB (SWI4,6-dependent cell cycle box), found 2-10 times in the upstream regulatory sequences of target genes. We have constructed a library of mutants in the SCB using doped oligonucleotide mutagenesis. All single base pair changes examined compromised the ability of the SCB to activate transcription in vivo. Analysis of the behaviour of the mutant SCBs in an in vitro DNA binding assay shows that the inability to activate transcription can be explained by reduced binding of SWI4 and SWI6 to the mutant SCBs. This analysis, together with a consideration of the SCBs found upstream of known SWI4,6-dependent genes, leads to the proposal of a revised consensus sequence for this important regulatory element.

The aim of this study was to investigate the use of functional electrical stimulation (FES) as a means of pressure sore prevention in seated spinal cord injured (SCI) subjects. Nine SCI subjects took part in tests in which electrical stimulation was applied to the quadriceps with the lower legs restrained. Ischial pressures were measured during periods of quiet sitting and FES application. A strain gauged lever arm was used to measure the knee moment during quadriceps stimulation. The average pressure drop at the right and left buttocks was 44 mmHg and 27 mmHg respectively. In general the greatest reductions occurred in subjects with larger knee moments; however, there was no direct relationship between the pressure reduction obtained and the quadriceps strength. This form of FES may be useful as a prophylactic aid in the management of pressure sores in SCI subjects.

The flexion withdrawal reflex, evoked by surface electrical stimulation, has been used to provide hip flexion for the restoration of gait in paraplegics. A major limitation to its use has been the decrease in the magnitude of the response to repeated stimulation (habituation). In this study it was found that by using high intensity stimulation the response could be dishabituated. It was demonstrated that sufficient hip flexion for functional electrical stimulation-assisted gait was maintained using high intensity pulses in a one-step-ahead controller.

SWI4 of budding yeast codes for a component of a transcription factor (cell cycle box factor, or CCBF) necessary for G1-specific expression of HO. We show that SWI4 is essential for haploid cell viability at high temperature and in a/alpha cells at all temperatures: SWI4-deficient cells arrest as large unbudded cells. Eight high copy number plasmids were identified that allow swi4- strains to grow under nonpermissive conditions. Two carry G1 cyclin genes, CLN1 and CLN2; another carries HCS26, coding for a putative cyclin, a/alpha swi4- mutants exhibit 3- to 20-fold reductions in the levels of CLN1, CLN2, and HCS26 transcripts. The requirement of SWI4 for transcription appears to be direct: each gene contains sites similar to the CCBF-binding site; CCBF binds to the upstream region of HCS26. We propose that SWI4 participates in a positive feedback loop by which CLN1, CLN2, and possibly HCS26 promote their own transcription in G1.

The biotinylation of protein antigens for use in immunoprecipitation studies with parasite antigens is described. This technique was standardized to produce maximal labelling without compromising either the viability of the parasite or the antigenicity of labelled proteins. Live axenic Entamoeba histolytica trophozoites were optimally labelled by incubation of 10(7) parasites in 1 ml phosphate-buffered saline for 20 min at 37 degrees C with a final concentration of 10 mM N-hydroxy succinimido-biotin (NHS-B). Examination of antigens recognized by immunoglobulin from convalescent amoebic liver abscess patients and precipitated by protein A-Sepharose showed a general increase in immune response to amoebic antigens with time following treatment. Mass ratio mol. wts of immunoprecipitated antigens concur with those presented by other authors using alternate technology, in addition to recognizing antigens previously not identified by human sera. We therefore recommend biotinylation as a viable alternative to radiolabelling techniques for the study of parasite antigens and the humoral immune responses raised against them.

The use of functional electrical stimulation (FES) of muscle for paraplegic locomotion, or grasp augmentation in tetraplegia, is limited by the variability in muscle response to stimulation as a result of several external and internal factors. Previous approaches to this problem have used position-servo controllers, which have been shown to function satisfactorily in the laboratory. However, such systems will fail should obstacles be encountered or should the stimulation hardware develop a fault. To prevent such potentially dangerous failures some form of sensory feedback is required. This paper describes the first application of a technique known as extended physiological proprioception (EPP) to the control of FES to compensate for muscle response variability and provide proprioceptive feedback via the appropriate sensory pathways. In the experimental system described, a paraplegic subject controlled the extension of his paralysed knee by shoulder protraction. A Bowden cable linked the two joints, and a dynamometer in this cable was used to derive the control signal for a computer-controlled stimulator which delivered surface stimulation to the quadriceps muscle group. Modelling and parameter identification were performed by analysis of the step response, and the controller was designed from consideration of the root locus. The advantages of the system, in terms of improved proprioceptive feedback and reduced limb-positioning error were assessed in a test of joint positioning accuracy with vision occluded. The EPP system showed improvements over both open and closed-loop position-servo controllers.

A non-cloned and a cloned isolate of non-pathogenic (zymodeme I) Entamoeba histolytica derived from homosexual males were incubated in Diamonds' TP-S-1 medium in the presence of Crithidia for 52 d. A single addition of Escherichia coli irradiated at 1000 Gy rendered the culture axenic. Analysis of zymodeme patterns established that both isolates had transformed to zymodeme II. The re-establishment of cultures in the presence of microflora from the original cultures resulted in a return to zymodeme I, whereas incubation with microflora from an amoebic isolate exhibiting a zymodeme II pattern maintained the pathogenic pattern.

We have shown previously that baboons (Papio anubis) develop high levels (greater than 80%) of protection against challenge infection following immunization with Schistosoma haematobium cercariae irradiated with 20 krad. In the present study baboons were immunized with schistosomula irradiated with either 20 krad or 60 krad, with variations in the timing and number of larvae comprising each vaccination. Baboons immunized 2 or 3 times with schistosomula irradiated with 20 krad were significantly more protected (85-90%) against challenge infection than baboons similarly immunized with larvae receiving 60 krad (56-50% protection). Baboons immunized with schistosomula irradiated with 20 krad were better protected against challenge infection at 8 weeks after immunization than at 28 weeks after immunization. Protection was manifest by a reduction in worm numbers, tissue and excreta egg counts, gross pathology and, to a lesser extent, by stability of body weight and haematological indices following challenge. Enzyme-linked immunosorbent assay (ELISA) results of selected baboon sera showed few differences related to irradiation dose alone, but titres were higher in baboons receiving booster immunizations, and there was a significant correlation between titres immediately preceding challenge and the degree of resistance. Examination of responses to individual schistosomular surface antigens by immunoprecipitation and sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed no correlation between the pattern of antigens recognized and resistance status. As with the ELISA assay, an anamnestic response was detected after vaccination, while the amount of antibody present declined markedly with increasing time after individual immunizations.

Most laboratory strains of the yeast Saccharomyces cerevisiae contain many copies of an autonomously replicating plasmid called 2-micron circle DNA. This plasmid codes for a site-specific recombinase, the FLP protein which promotes recombination across two 599-base pair inverted repeats of the plasmid DNA. We have cloned the FLP gene under the control of a strong Escherichia coli promoter and have hyperproduced the protein in that organism. Cell-free extracts from this source promote highly efficient site-specific recombination in vitro and we have used this activity to purify the FLP protein substantially. The enzyme acts efficiently on circular and linear substrates and requires only monovalent or divalent cations for activity.

Levels of antibody a binding to the schistosomulum surface and b mediating in vitro eosinophil-dependent cytotoxicity of schistosomula were studied and compared to the in vivo levels of resistance to cercarial challenge in mice infected with irradiated cercariae, unirradiated cercariae of single or mixed sex, or injected with eggs. Antibody-binding was assessed by counting the number of IgG-Fc-receptor bearing cells (P388D1 cells) adhering to mechanically-transformed schistosomula. Significant levels of adherence occurred with sera taken from 1--2 weeks following exposure to irradiated cercariae, the level increasing gradually thereafter and being enhanced by repeated exposure. Sera from the bisexual infection showed a dramatic increase in binding activity between weeks 5--8, and with the single sex infections there was a steady rise up to week 10 followed by a sharp rise between weeks 10--12. Weekly injections of eggs produced a steady rise in serum binding activity. Sera taken just before challenge were also tested for their ability to mediate killing of schistosomula by eosinophil-enriched preparations of heterologous rat peritoneal exudate cells in vitro. Significant levels of killing occurred with all sera, but the greatest lethal activity was found in sera from the egg-injected, chronically-infected and 200 male cercariae-infected groups. This ranking did not correlate with in vivo resistance against challenge as assessed by worm recovery, the egg-injected and single sex-infected groups failing to manifest significant resistance.

We have characterized various anti-schistosomular surface MoAbs previously shown to partially block in-vitro killing of schistosomula by human sera and eosinophils (Dunne et al. 1987). Immunodiffusion analysis showed that four IgM and one IgG3 MoAbs recognized periodate sensitive epitopes on the same molecular species present in schistosomular antigen but their patterns of reactivity with soluble egg antigen demonstrated that at least three distinct epitopes were involved. SDS-PAGE analysis showed the IgMs to react with 125I-labelled surface antigens of Mr 35,000-38,000 and Mr 20,000, and the IgG3 to react with an Mr 38,000 antigen. In spite of their effect in vitro, transfer of the IgM MoAbs at the time of challenge of mice vaccinated with irradiated cercariae or of mice injected with an unrelated (anti-Mr 16,000) protective MoAb failed to produce in-vivo blocking. Similarly, injection of Schistosoma mansoni eggs, prior infection with worms of one sex, or passive transfer of serum from single-sex infected mice failed to influence the resistance conferred by vaccination with irradiated cercariae.

We have studied the interaction of purified FLP protein with restriction fragments from the substrate 2mu circle DNA of yeast. We find that FLP protects about 50 bp of DNA from nonspecific nuclease digestion. The protected site consists of two 13 bp inverted repeat sequences separated by an 8 bp spacer region. A third 13 bp element is also protected by binding of the FLP protein. We demonstrate that FLP introduces single- and double-strand breaks into the substrate DNA. This site-specific cleavage occurs at the margins of the spacer region, generating 8 bp 5' protruding ends with 5'-OH and 3'-protein-bound termini. Binding to mutant sites and half-sites demonstrates that the third symmetry element is not important for binding and cleavage by the FLP protein. The integrity of the core region is important for the cleavage activity of FLP.

A hybrid functional electrical stimulation (FES) orthosis is described, comprising a rigid ankle-foot brace, a multi-channel FES stimulator with surface electrodes, body mounted sensors, a 'rule-based' controller and an electro-cutaneous display for supplementary sensory feedback. The mechanical brace provides stability, without FES activation of muscles, for standing postures normally adopted by patients. This avoids inducing muscle fatigue during prolonged upright activity. However, stability is conditional upon the position of the ground reaction vector (GRV) relative to the knee joint. The finite state FES controller reacts automatically to destabilizing shifts of the GRV by stimulating appropriate anti-gravity musculature to brace the leg. The FES system also features a control mode to initiate and terminate flexion of the leg during forward progression. A simple mode of supplementary sensory feedback was used during the laboratory standing tests to assist the patient in maintaining a set posture. Preliminary results of laboratory tests for two spinal cord injured subjects are presented.

Patients with hemiplegia frequently have difficulty in walking due to lack of eversion and dorsiflexion capability of the foot. One method of treating these patients utilizes functional electrical stimulation (FES). The effect of FES on locomotion, co-ordination, proprioception and balance sense was assessed using instrumented gait analysis and a postural sway test. In general patients treated with FES showed either a marked improvement or very little change. Any improvement was reflected in postural sway and ankle control during locomotion. Changes in hip and knee control were insignificant.

Two monoclonal antibodies against the surface of S. mansoni schistosomula were found to confer significant passive protection to mice (M7B3A, range 28-70%; M22H12C, range 14-58%). No additive effect was observed when both were transferred together. Neither McAb bound to the cercarial surface but both bound to the surface of in vitro derived schistosomula and schistosomula recovered from mouse skin up to 3 days after infection. The McAbs were species specific, but not S. mansoni strain specific. M22H12C immunoprecipitated an 125I-labelled surface antigen of relative molecular weight (mol. wt) 32 000. In Western blotting of an NP40 schistosomular extract, M7B3A recognized an antigen smear of 13 000-18 000 with a dominant band at 16 000. This 16 000 antigen was recognized by serum from demonstrably immune mice and rats vaccinated with highly irradiated carcariae but not by sera from mice with chronic single sex or bisexual infections.

A single dose of Ro 11-3128 was found to be 98-100% effective against Schistosoma mansoni infections at intervals of 3 h to 15 days following infection, and apparently killed the schistosomula stages soon after administration, thus allowing an assessment of the immunizing potential of progressive larval stages. Following infection with 500 unirradiated cercariae, optimum resistance was manifest by groups drug-treated at 48-96 h (60-75%). Significantly lower levels of resistance occurred with early (3 h) or later (6-15 day) treatments. Superimposition of an infection treated at 15 days on a prior infection which had been treated at 2 days did not reduce the level of resistance caused by the latter, indicating that the infection plus delayed treatment had not induced suppression. Thus the peak resistance manifest during the 48-96 h period may reflect optimum expression of protective antigens. Comparison of irradiated (20 krad.) with unirradiated infections showed that, when drug-terminated 24, 48 or 96 h post-infection, irradiated cercariae induced significantly less resistance than unirradiated cercariae, perhaps indicating a delayed appearance of protective antigens following radiation treatment.

Transcription of the HO gene of Saccharomyces cerevisiae, which encodes a site-specific endonuclease that initiates cell-type switching (reviewed in refs. 1,2), is restricted to a short window of the cell cycle in late G1 (refs 3,4). A repeated element in the upstream region of HO (the cell-cycle box, CCB) and two regulatory proteins, SWI4 and SWI6, are required for cell-cycle-dependent expression of HO. Biochemical experiments have identified a factor, CCBF (cell-cycle box factor), that binds to the CCB elements and that presumably plays a key part in cell-cycle regulation of HO. The SWI4 and SWI6 genes are required for formation of the CCBF-DNA complex. Here we report the nucleotide sequence of the SWI4 gene and show that it contains two copies of the conserved SWI6-cdc10 motif observed in SWI6 of budding yeast, the Schizosaccharomyces pombe cdc10 gene required for progression through G19, the Drosophila Notch gene, and in the Caenorhabditis elegans lin-12 and glp-1 genes. We demonstrate by using antibodies to the SWI4 protein in gel-shift assays that the protein is present in the CCBF-DNA complex.

One of the problems which occurs in the development of a control system for functional electrical stimulation of the lower limbs is to detect accurately specific events within the gait cycle. We present a method for the classification of phases of the gait cycle using the artificial intelligence technique of inductive learning. Both the terminology of inductive learning and the algorithm used for the analyses are fully explained. Given a set of examples of sensor data from the gait events that are to be detected, the inductive learning algorithm is able to produce a decision tree (or set of rules) which classify the data using a minimum number of sensors. The nature of the redundancy of the sensor set is examined by progressively removing combinations of sensors and noting the effect on both the size of the decision trees produced and their classification accuracy on 'unseen' testing data. Since the algorithm is able to calculate which sensors are more important (informative), comparisons with the intuitive appreciation of sensor importance of five researchers in the fields were made, revealing that those sensors which appear intuitively most informative may, in fact, provide the least information. Comparison results with the standard statistical classification technique of linear discriminant analysis are also presented, showing the relative simplicity of the inductively derived rules together with their good classification accuracy. In addition to the control of FES, such techniques are also applicable to automatic gait analysis and the construction of expert systems for diagnosis of gait pathologies.

This paper describes further characteristics of the immunization of sheep against schistosomes using live, irradiation schistosomula. Sheep immunized with a non-virulent strain of Schistosoma mattheei were protected against a more virulent strain of the same species for over a year. As there was no evidence that the irradiated parasites were able to persist this long, it was concluded that the vaccine had induced a sterile resistance. Heterologous vaccination, using irradiated S. mattheei schistosomula to immunize against S. bovis or irradiated S. mansoni schistosomula to immunize against S. mattheei, failed to induce any protection.

Most strains of the yeast Saccharomyces cerevisiae harbor several copies of a 2-micron plasmid circle DNA termed "2 micron." This circular plasmid contains two 599-base-pair precise inverted repeats across which a site-specific inversion event occurs in vivo. This inversion is promoted by a plasmid-encoded function called "FLP." We have cloned the FLP gene of 2-micron DNA under control of a strong yeast promoter and transformed yeast cells with a plasmid containing the cloned FLP gene. Cell-free extracts from such a transformant promote highly efficient inversion of 2-micron DNA in vitro. The reaction requires a cation and works efficiently on supercoiled, relaxed circular, or linear DNA. The FLP activity bears certain similarities to the cre protein, a site-specific recombinase encoded by bacteriophage P1.

A microcomputer was used to analyse uterine activity during spontaneous labour in 30 nulliparous and 30 parous patients. The intrauterine mean active pressure (MAP) in the nulliparous group was 1.51 kPa (SD 0.45) in the first stage and 2.71 kPa (SD 0.77) in the second stage. For the parous group, the MAP in the first stage was 1.22 kPa (SD 0.37) and in the second stage was 2.92 kPa (SD 0.98). A significant parity difference in MAP and contraction frequency was observed in the first stage but not in the second stage. Epidural analgesia did not appear to influence uterine activity in the first stage but was associated with a lower MAP, contraction frequency and intensity in the second stage. During the first stage, MAP increased by 21% in nulliparas and by 39% in multiparas. The rise in MAP was mainly due to an increase in contraction frequency.

The 2 micron plasmid of Saccharomyces cerevisiae codes for a site-specific recombinase, the FLP protein, that catalyzes efficient recombination across two 599-base-pair (bp) inverted repeats of the plasmid DNA both in vivo and in vitro. We analyzed the interaction of the purified FLP protein with the target sequences of two point mutants that exhibit impaired FLP-mediated recombination in vivo. One mutation lies in one of the 13-bp repeat elements that had been previously shown to be protected from DNase digestion by the FLP protein. This mutation dramatically reduces FLP-mediated recombination in vitro and appears to act by reducing the binding of FLP protein to its target sequence. The second mutation lies within the 8-bp core region of the FLP target sequence. The FLP protein introduces staggered nicks surrounding this 8-bp region, and these nicks are thought to define the sites of strand exchange. The mutation in the core region abolishes recombination with a wild-type site. However, recombination between two mutated sites is very efficient. This result suggests that proper base pairing between the two recombining sites is an important feature of FLP-mediated recombination.

The Oswestry 'Parawalker' orthosis has been supplied to 15 adult thoracic level complete paraplegic patients enabling them to achieve a reciprocal gait with an inherent low energy demand. In order to further off-load the work demands on the upper limb girdle musculature during ambulation, three of these patients have undergone electrical stimulation of the gluteal muscles in stance phase. This was shown to increase the stability of the adduction and also provided forward propulsion by driving the stance-leg into extension. Bilateral stimulation of the quadriceps muscles has been shown to facilitate standing and sitting in the orthosis. Using a Kistler force platform, the crutch impulse can be seen to be reduced by as much as 36%. We speculate that this leads to a similar order of reduction in the work done by the upper limbs in these patients.

Three sheep were vaccinated with two doses of 3 krad-irradiated cryopreserved Schistosoma bovis schistosomula containing 20,000 and 17,000 organisms respectively, injected intramuscularly 23 days apart after storage in liquid nitrogen for between 9 and 46 days. A challenge of 5360 S. bovis cercariae was administered percutaneously approximately four weeks after the last vaccine dose to these animals and to three controls. Post-challenge the vaccinated animals gained significantly more weight (27% v. 9%), produced fewer eggs in their faeces, showed a smaller reduction in PCV values (-18% v. -27%) and were over-all in better condition than control animals. At perfusion 49.1% fewer adult worms were found in the vaccinated sheep than in controls. The tissue egg burdens were similar in both groups. Histopathologically both groups were similar except that fewer and smaller egg lesions were observed in the livers of vaccinated animals.

Transcription of the HO gene is controlled by at least eleven trans-acting regulators, which are responsible for limiting expression to mother cells and to one period in the cell cycle. A subset of these regulators and a repeated element (the cell-cycle box, CCB) are required for expression of HO during late G1. We describe gel retardation and footprinting experiments that identify a factor, CCBF (cell-cycle box factor), that binds to multiple cell-cycle box elements. We show that the SWI4 and SWI6 genes are required for formation of the CCBF-promoter complex in vitro, either as components of CCBF or as modulators of CCBF activity. The other SWI genes (SWI1, 2, 3, and 5) are not required. We also show that SWI4 and SWI6 are the only SWI genes required for expression in vivo from the CCB sequences. These observations indicate that the CCBF is responsible for cell-cycle regulation of HO and lead to the view that two of the SWI genes, SWI4 and SWI6, are specifically involved in this regulatory event.

The biosynthesis and efflux of sterols from cells into the medium were investigated in skin fibroblasts from a control, a patient with obligate heterozygous familial hypercholesterolaemia and a patient with the homozygous condition. The behaviour of the cells was studied in two lipid free media (lipoprotein deficient and delipidated serum), with and without the addition of low density lipoproteins (LDL) in order to find experimental conditions which showed maximum differences between the three cell lines. Incorporation of [14C]acetate into sterols in the presence (repression) and absence (induction) of LDL was similar in the normal and heterozygous cells, whereas the homozygous cells showed reduced repression and increased induction. In all three cell lines induction of sterol synthesis was greater with delipidated than lipoprotein deficient serum. The efflux of sterols in both the presence and absence of LDL did not differ between the three cell lines, but it was greater when LDL was added to the medium and when delipidated serum was used. Sterol biosynthesis and efflux from the cells of the heterozygote did not differ significantly from those of the control.

In order to develop a control system for electrical stimulation of paralysed muscle and improve muscle resistance to fatigue, it is useful to investigate the possibilities of simulating the control systems of the normal body. One way is the periodic shifting of stimulation from one muscle to another. This technique is called sequential stimulation and allows sufficient rest time for each muscle to reduce fatigue and consequently prolong muscle strength. It can also be seen to improve the muscle recovery time. In the following study, the muscles rectus femoris, vastus lateralis and vastus medialis were used to keep the knee locked and extended during stimulation. Several experiments were carried out using a three-channel computer controlled stimulator. The results for three-phase sequential stimulation (33% duty cycle per muscle) were most effective and significantly improved the muscle fatigue characteristics.

The immune-dependent elimination of a challenge infection in rats vaccinated with highly-irradiated cercariae of Schistosoma mansoni was analysed by passive transfer of serum, recovery of the challenge from the lungs and liver and by transferring lung-stage schistosomula. Recipients of serum from rats immunized with either unirradiated, 20 or 40 krad.-irradiated cercariae, most of which die in the liver, lungs and skin, respectively, were equally resistant if the serum was injected on the day of infection or 5-7 days after infection. In addition, vaccinated rat serum transferred to mice and vaccinated rabbit serum transferred to rats conferred comparable protection when injected on day 0 or 5 days after infection of the recipients. This apparent susceptibility of the lung schistosomula to immune attack was confirmed by challenging 20 or 40 krad.-irradiated cercariae vaccinated rats with lung-stage schistosomula derived either from mice or rats. All the detectable attrition of a cercarial challenge in vaccinated rats occurred between 7 and 10 days post-challenge, before the parasites reached the liver. Since there was no evidence of damage or attrition in the skin or lungs before day 7 it was concluded that immune-dependent elimination occurred rapidly following a 'window of sensitivity' coinciding with the migration of the parasites from the lungs to the liver.