Host-pathogen dynamics of squirrelpox virus infection in red squirrels (Sciurus vulgaris).

To improve our understanding of squirrelpox virus (SQPV) infection in the susceptible host, three red squirrels were challenged with wild-type SQPV via scarification of the hind-limb skin. All squirrels seroconverted to the infection by the end of the experiment (17 days post-challenge). Challenged animals suffered disease characterised by the development of multiple skin and oral lesions with rapid progression of skin lesions at the infection site by day 10 post-challenge. No internal pathological changes were found at post-mortem examination. A novel SQPV Taqman(®) Real-time PCR detected viral DNA from multiple organs, with the largest amounts consistently associated with the primary and secondary skin and oral lesions where viral replication was most likely occurring. Immunohistochemistry clearly detected viral antigen in the stratified squamous epithelium of the epidermis, tongue and the oropharyngeal mucosa-associated lymphoid tissue and was consistently associated with histological changes resulting from viral replication. The lack of internal pathological changes and the detection of relatively low levels of viral DNA when compared with primary and secondary skin lesions argue against systemic disease, although systemic spread of the virus cannot be ruled out. This study allowed a comprehensive investigation of the clinical manifestation and progression of SQPV infection with a quantitative and qualitative analysis of virus dissemination and shedding. These findings suggest two separate routes of SQPV transmission under natural conditions, with both skin and saliva playing key roles in infected red squirrels.

First cases of squirrelpox in red squirrels (Sciurus vulgaris) in Scotland.

Squirrelpox, caused by a poxvirus, is a major threat to the remaining UK red squirrel population. The spread of antibody-positive grey squirrels has been monitored in the UK for the past decade. In 2005 grey squirrels that had been exposed to the virus appeared in the south of Scotland for the first time, followed approximately two years later by the appearance of squirrelpox disease in the local red squirrels. Four squirrels were examined. They all had gross external lesions and histological lesions typical of squirrelpox disease, but no significant internal lesions. The diagnosis was confirmed by PCR, electron microscopy and serology.

Piloting new ways of working: evaluation of the WILSTAAR Programme.

The current study aims to investigate the reliability and effectiveness of the WILSTAAR screening assessment and intervention programme. It also explores parental perceptions and evaluations of their involvement in the intervention programme. At this stage data collection is incomplete however interim results are presented. False positive rates are slightly higher than expected raising questions regarding the screen's reliability. Numbers are too small at present to report with confidence on the effectiveness of the intervention programme. Results indicate that parents rate their involvement in the WILSTAAR programme in a highly positive manner. The need for further research in this area is highlighted to ensure that early identification and intervention programmes meet the needs of parents and their children.

Evaluation of the Hanen Early Childhood Educators Programme.

The current study is an evaluation of the Hanen Early Childhood Educators Programme (Weitzman 1992). It explores outcomes for both staff and children following staff participation in the training programme. Using a comparative study design and pre- and post-training measures the changes in staff and children were investigated. Results revealed that the training programme did not bring about change in children's receptive and expressive language skills. Changes were observed in children's social interaction skills. This was reinforced by staff reports of changes in children's interaction. Staff changes included increased skills and confidence in identifying and supporting children with speech and language difficulties alongside positive changes in their interaction styles. The methodological difficulties inherent in evaluating training effects and the need for further research are highlighted.

Submandibular gland transfer: a new method of preventing radiation-induced xerostomia.

OBJECTIVE: Radiation-induced xerostomia is a significant morbidity of radiation therapy in the management of patients with head and neck cancers. We have recently reported a method of transfer of one submandibular gland to the submental space in a small pilot series of eligible surgical patients. The submental space was shielded during postoperative radiation therapy. The transferred gland continued to function after the completion of radiation therapy and none of the patients developed xerostomia. The purpose of this article is to present the technique of submandibular gland transfer in detail and to evaluate the postoperative survival and function of the transferred submandibular glands. DESIGN: Prospective clinical trial. METHODS: The submandibular gland was transferred on eligible patients as part of their surgical intervention. The patients were followed clinically, with salivary flow and radioisotope studies. RESULTS: We performed the surgical transfer of the submandibular salivary gland in 24 of 25 patients placed on the protocol. All the glands survived transfer and functioned well postoperatively as demonstrated on the salivary flow and the radioisotope studies. The surgical transfer was relatively simple and added 45 minutes to the surgical procedure. There were no complications attributed to the submandibular gland transfer. CONCLUSIONS: We have successfully demonstrated that the submandibular gland can be surgically transferred to the submental space with its function preserved. The gland seems to continue functioning even after radiation therapy with the appropriate shielding. This surgical transfer procedure has the potential to change the way we currently manage patients with head and neck cancer.

Submandibular salivary gland transfer prevents radiation-induced xerostomia.

BACKGROUND: Xerostomia is a significant morbidity of radiation therapy in the management of head and neck cancers. We hypothesized that the surgical transfer of one submandibular salivary gland to submental space, outside the proposed radiation field, prior to starting radiation treatment, would prevent xerostomia. METHODS: We are conducting a prospective clinical trial where the submandibular gland is transferred as part of the surgical intervention. The patients are followed clinically, with salivary flow studies and University of Washington quality of life questionnaire. RESULTS: We report early results of 16 patients who have undergone this procedure. Seven patients have finished and 2 patients are currently undergoing radiation treatment. In 2 patients, no postoperative radiation treatment was indicated. Two patients are waiting to start radiation treatment and 2 patients refused treatment after surgery. The surgical transfer was abandoned in 1 patient. All of the transferred salivary glands were positioned outside the proposed radiation fields and were functional. The patients did not complain of any xerostomia and developed only minimal oral mucositis. There were no surgical complications. CONCLUSIONS: Surgical transfer of a submandibular salivary gland to the submental space (outside the radiation field) preserves its function and prevents the development of radiation-induced xerostomia.

A novel gene encoding a ras-like GTP-binding protein from Trypanosoma brucei: an evolutionary ancestor of the ras and rap genes of higher eukaryotes?

The ras superfamily of GTP binding proteins encompasses a wide range of family members, related by conserved amino-acid motifs, and act as molecular binary switches that play key roles in cellular processes. Gene duplication and divergence has been postulated as the mechanism by which such family members have evolved their specific functions. We have cloned and sequenced a ras-like gene, tbrlp, from the primitive eukaryote Trypanosoma brucei. The gene encodes a protein of 227 amino acids and contains the six conserved subdomains that designate it as a ras/rap subfamily member. However, the presence of key diagnostic residues characteristic of both the ras and rap families of GTP confuse the familial classification of this gene. Phylogenetic analysis of the GTP binding domain places its origins at the divergence point of the ras/rap families and suggests that tbrlp is an ancestral gene to the ras/rap genes of higher eukaryotes.

Semantic pragmatic disorder with application of selected pragmatic concepts.

Children with semantic pragmatic disorder have been described as having difficulties in conversational interaction. The content of their conversations has been described as 'odd', 'loose', tangential', 'irrelevant' and 'inappropriate' (Stubbs 1986). A diagnosis of semantic pragmatic disorder is currently made based on whether or not a child displays certain surface characteristics. The present study attempts to apply precision to the intuitive terms which have been used to describe these children's conversations. To this end conversation samples from three children identified by speech and language therapists as fitting the clinical diagnosis of semantic pragmatic disorder were analysed. Aspects of pragmatic theory which best explained the strengths and weaknesses which emerged in the conversations are discussed. Therefore this represents an essentially data driven study.

Encouraging speech and language development in exceptional circumstances.

An evaluation of the accessibility of the speech and language therapy service within Down Lisburn Trust and a needs assessment of the levels of speech and language difficulties in preschool children was conducted between October 1996 and March 1997. Results highlighted that 42% of preschool children had speech and language difficulties, however only 17.8% were able to access the speech and language therapy service for remediation of their difficulties. A further 2-3 year project is now planned. This aims to tackle the high levels of speech and language difficulties in children through prevention, early identification and remediation programmes. It also aims to ensure equitable access to the speech and language therapy service for all children.

Identification of a new class of exonic splicing enhancers by in vivo selection.

In vitro selection strategies have typically been used to identify a preferred ligand, usually an RNA, for an identified protein. Ideally, one would like to know RNA consensus sequences preferred in vivo for as-yet-unidentified factors. The ability to select RNA-processing signals would be particularly beneficial in the analysis of exon enhancer sequences that function in exon recognition during pre-mRNA splicing. Exon enhancers represent a class of potentially ubiquitous RNA-processing signals whose actual prevalence is unknown. To establish an approach for in vivo selection, we developed an iterative scheme to select for exon sequences that enhance exon inclusion. This approach is modeled on the in vitro SELEX procedure and uses transient transfection in an iterative procedure to enrich RNA-processing signals in cultured vertebrate cells. Two predominant sequence motifs were enriched after three rounds of selection: a purine-rich motif that resembles previously identified splicing enhancers and a class of A/C-rich splicing enhancers (ACEs). Individual selected ACEs enhanced splicing in vivo and in vitro. ACE splicing activity was competed by RNAs containing the purine-rich splicing enhancer from cardiac troponin T exon 5. Thus, ACE activity is likely to require a subset of the SR splicing factors previously shown to mediate activity of this purine-rich enhancer. ACE motifs are found in two vertebrate exons previously demonstrated to contain splicing enhancer activity as well as in the well-characterized Drosophila doublesex (dsx) splicing enhancer. We demonstrate that one copy of the dsx repeat enhances splicing of a vertebrate exon in vertebrate cells and that this enhancer activity requires the ACE motif. We suggest the possibility that the dsx enhancer is a member of a previously unrecognized family of ACEs.

The cloning and sequencing of a ribosomal L18 protein from an evolutionary divergent eukaryote, Trypanosoma brucei.

Eukaryotic ribosomal proteins are highly conserved across widely divergent species, suggesting that strong functional constraints prevent divergence of important amino acid motifs. Using this as a basis, an evolutionary approach could be used to identify putative functional motifs. We obtained the DNA sequence of the ribosomal protein L18 from the evolutionary divergent protozoan parasite, Trypanosoma brucei. Analysis of this sequence showed that it had 46% and 43% identity with the human and yeast sequences, respectively, and 30% of amino acid residues were identical across all the species analysed. Using these data, amino acids essential to the structure and function of ribosomal protein L18 can be inferred and could provide valuable information for molecular modelling and mutational studies.

Systemic absorption and toxicity from topically administered lidocaine during transesophageal echocardiography.

This report describes the cases of three patients who had central nervous system toxicity from locally administered lidocaine during transesophageal echocardiography. Parenteral sedation was not employed. Serum levels of lidocaine were obtained after the procedure in all three patients, who recovered spontaneously and suffered no permanent ill effects. Medical conditions such as congestive heart failure and diminished hepatic function or concomitant use of lidocaine analogs may predispose patients to the toxic side effects of locally administered lidocaine. Extreme care and close monitoring of patients are warranted when topical lidocaine anesthetic is employed in the setting of delayed lidocaine clearance.

Site-specific comparison of p53 immunostaining in squamous cell carcinomas.

The p53 tumor suppressor gene is believed to be the most commonly mutated gene in human cancer. p53 is thought to function as a negative regulator of the cell cycle, arresting cells in the G1 phase. This study examined the effects of different mutagenic environments on the incidence of p53 overexpression in squamous cell carcinomas (SCCs) from sun exposed and non-sun exposed squamous epithelium. An immunohistochemical analysis was undertaken in an attempt to assay SSC for p53 overexpression, an indirect measure of missense mutant p53. Positive nuclear staining for p53 was observed in 14 of 21 sun exposed SCCs, two of 19 vulvar/perianal SCCs, and 15 of 20 oral cavity SCCs. The number of positive anogenital tumors was low compared with that of both sun exposed (chi-squared, 1 df, P = .0004) and oral (chi-squared, 1 df, P < .0001) sites. It was concluded that p53 protein accumulation is common in sun-exposed cutaneous SCC and oral SCC compared with anogenital SCC, and thus it is hypothesized that the nature of the mutagenic environment in which SCC develops directly affects the incidence of immunohistochemically detectable p53-positive cells.

Trypanosoma brucei: characterisation of a life cycle stage-specific G-protein.

G-proteins have important and diverse roles in cellular signaling in higher eukaryotes but as yet little is known about the involvement of these GTP-binding proteins in protozoan parasites. Bacterial toxin-catalysed ADP-ribosylation, routinely used in the characterisation of G-proteins in higher eukaryotes, was used to determine whether functional G-protein homologues could be detected in Trypanosoma brucei. A 43-kDa polypeptide that was ADP-ribosylated in the presence of pertussis toxin was identified in membrane fractions. This polypeptide was also recognised by an antibody generated against a peptide sequence representing a mammalian GO alpha G-protein subunit. GTP and GTP gamma S (a nonhydrolysable analogue of GTP) abolished the ADP-ribosylation reaction, indicating that the 43-kDa polypeptide also binds GTP. Furthermore, this T. brucei G-protein is present in bloodstream but not procyclic life cycle stages. Using an in situ GTP binding assay, a further group of small-molecular-size GTP-binding proteins which are present in both bloodstream and procyclic stages have been identified. This demonstration of G-proteins in T. brucei suggests possible intracellular mechanisms that may be involved in the transduction of host or other parasite signals.

A randomized comparison of intravenous amrinone versus dobutamine in older patients with decompensated congestive heart failure.

OBJECTIVE: To compare the hemodynamic effects of amrinone and dobutamine in patients 75 years of age or older who have severe congestive heart failure requiring invasive hemodynamic monitoring and inotropic support. DESIGN: Prospective, randomized, double-blind clinical trial. SETTING: Coronary care unit of a university teaching hospital. PARTICIPANTS: Fourteen patients > or = 75 years of age (mean 80.3 +/- 5.7 years) with refractory New York Heart Association class IV congestive heart failure. All patients had a cardiac index < 2.5 L/min/M2 (mean 1.8 +/- 0.3 L/min/M2), pulmonary capillary wedge pressure > or = 18 mm Hg (mean 26 +/- 10 mm Hg), and left ventricular ejection fraction < 40% (mean 26 +/- 10%). INTERVENTION: Patients were randomly assigned to treatment with 2-hour infusions of amrinone (n = 7) or dobutamine (n = 7) at fixed dosages of 5 and 10 micrograms/kg/min. MEASUREMENTS: Complete hemodynamic data were obtained at baseline and after each 2-hour medication infusion. Transthoracic two-dimensional echocardiography was performed at baseline and after the 10 micrograms/kg/min medication dose. The primary analysis compared the effects of the two drugs on cardiac index and stroke volume index at each of the two dosages. RESULTS: Both amrinone and dobutamine had salutary hemodynamic effects, as indicated by improvements in cardiac index, stroke volume index, pulmonary capillary wedge pressure, and systemic vascular resistance (all P < .05 except effect of amrinone on stroke index and wedge pressure). Although the overall hemodynamic effects of amrinone and dobutamine were similar, stroke volume index was higher with dobutamine at the 10 micrograms/kg/min dose (35 +/- 7 ml/M2 vs 26 +/- 6 mL/M2; P = .045). Two dobutamine patients were withdrawn from the study after the 5 micrograms/kg/min dose due to adverse effects (tachycardia, increased ventricular ectopy). One additional patient in each group was noted to have ventricular arrhythmias not requiring termination of the protocol. CONCLUSIONS: Both amrinone and dobutamine are efficacious in improving hemodynamics in older patients with severe congestive heart failure caused by left ventricular contractile dysfunction. Despite the effect of aging on beta-adrenergic responsiveness, dobutamine is at least as effective as amrinone but may be associated with a higher incidence of arrhythmic side effects.

Anosognosia in Alzheimer's disease: relationships to depression, cognitive function, and cerebral perfusion.

Awareness of memory loss was rated in 57 patients with clinically diagnosed Alzheimer's disease, and analyzed in relation to neuropsychological tests and presence of depression. Single photon emission computed tomography measures of regional cerebral blood flow were obtained on an unselected subsample of 20. Anosognosia was associated with diminished relative right dorsolateral frontal lobe perfusion and with high rates of false positive errors on recognition memory testing. Less aware patients did not differ from others on learning or recall scores, or on dementia severity as measured by mental status scores. Neither anosognosia nor depression was associated with disease duration or dementia severity and patients who were aware of their memory loss were no more likely than others to be depressed. This is further evidence that dementia severity alone does not account for anosognosia in Alzheimer's disease; frontal lobe involvement and the presence of specific memory impairments may be important determining factors.