African trypanosome antigens recognized during the course of infection in N''dama and Zebu cattle.

The humoral immune responses to Trypanosoma brucei infection were examined in N'dama and in Zebu, two breeds of cattle recognized for their differing susceptibility to trypanosomiasis. Regardless of the clinical course, animals of both breeds produced antibodies to nonsurface trypanosome antigen(s) detectable by both immunodiffusion and immune fluorescence. As a new approach to assessment of the humoral response to trypanosome infection, protein antigens responded to were isolated by immune precipitation, and their molecular weights were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This allowed the detection of differences in the immune response which correlated with the clinical course of the disease. All cattle of both breeds which exhibited a capacity to control the disease recognized at least one of three specific antigens: protein of 110,000, 150,000, and 300,000 daltons. The N'dama, which proved less susceptible to the disease, generally responded to more of the three identified trypanosome protein antigens than did the Zebu. Animals which died of trypanosomiasis failed to produce detectable antibodies to any of the three specific proteins, although they sometimes exhibited antibodies to another trypanosome antigen.     

Amplification of fluorescent in situ hybridisation signals in formalin fixed paraffin wax embedded sections of colon tumour using biotinylated tyramide.

Fluorescent in situ hybridisation (FISH) is a powerful tool for the evaluation of chromosomal alterations in formalin fixed paraffin wax embedded sections of colorectal cancer. However, initial experiments using a two-step detection system for digoxigenin labelled chromosome specific centromeric probes resulted in a complete lack of hybridisation signal from a number of colorectal tumour sections. This was due to high levels of background autofluorescence observed in this tissue, which masked any relatively weak hybridisations present. To overcome this problem, a biotinylated tyramide mediated amplification system was incorporated into the FISH detection protocol. This involves the use of horseradish peroxidase to activate the biotinylated tyramide, resulting in the deposition of a large number of biotin molecules at the site of bound peroxidase, which corresponds directly to the location of hybridised probe. Final detection was by means of a streptavidin-FITC conjugate. Using this technique, a panel of 11 colorectal tumour samples studied to date have shown strong, specific hybridisation signals to the nucleus of tumour cells. Amplification of FISH signals by biotinylated tyramide has the potential to improve weak hybridisation signals in cells from numerous sources, using a variety of probe types, including single copy gene probes as well as centromere specific probes.     

Topographical distribution and functional properties of cortically induced rhythmical jaw movements in the monkey (Macaca fascicularis)

1. The lateral part of the pericentral cortex of both hemispheres in three awake monkeys was explored with intracortical microstimulation (ICMS) using short trains (T/S; 200-microseconds pulses at 333 Hz for 35 ms, less than or equal to microA) and long trains (C/S; 200-microseconds pulses at 50 Hz for 3 s, less than or equal to 60 microA). In both hemispheres of one of these monkeys, the responsiveness of single cortical neurons to stimulation of the orofacial region was tested at the same intracortical sites where ICMS was applied. 2. Movements were evoked from four physiologically defined cortical regions: the primary face motor cortex (MI), the primary face somatosensory cortex (SI), the principal part of the cortical masticatory area (CMAp) which was located in the precentral gyrus lateral to MI, and a deep part of the cortical masticatory area (CMAd) which was located in the inferior face of the frontal operculum. 3. Two types of cortically induced movements were observed: a single twitch movement and EMG activity of the orofacial muscles that was evoked by T/S at a short latency (10-45 ms) and rhythmical jaw movements (RJMs) which were only evoked by C/S. 4. RJMs were evoked at C/S frequencies ranging from 20 to 300 Hz. At movement threshold, the frequency of the cortically induced RJMs varied from 0.7 to 1.5 Hz and usually increased with the increase of C/S intensity up to 2 times movement threshold. The vertical amplitude of RJMs was also stimulus dependent, and at movement threshold it ranged from 3 to 9 mm. 5. The movement patterns of the cortically induced RJMs remained constant during the course of C/S but could be differentiated in the frontal plane into ipsilateral- (RJMi), vertical-(RJMv), and contralateral- (RJMc) directed movements. These three different patterns of RJMs were associated with different patterns of masticatory muscle activity. 6. Each cortical region contained many sites from which RJMs could be induced (so-called RJM sites). The RJMi sites were more numerous than RJMc sites in all regions except SI and were located anterolateral or lateral to the RJMc sites in each region; the RJMv sites were scattered throughout each cortical region. 7. In MI, C/S elicited RJMs from 94 intracortical sites from which short-latency twitch movements could also be evoked by T/S.(ABSTRACT TRUNCATED AT 400 WORDS)     

Fornix transection impairs conditional visuomotor learning in tasks involving nonspatially differentiated responses.

Rhesus monkeys learned a series of conditional visuomotor associations involving two-dimensional "objects" that instructed one of three responses: tapping a touch screen, steady contact with the screen for a brief period, or steady contact for a longer period. Relative to controls, fornix-transected monkeys were impaired in the acquisition of new associations and in the retention of preoperatively learned ones. These findings challenge the view that the hippocampal system participates in associative learning only when spatial information is relevant to either the stimulus or the response.     

Comparison of the lysis-centrifugation and agitated biphasic blood culture systems for detection of fungemia.

Although the detection of fungemia has been improved by the use of vented or biphasic blood culture bottles, the best recovery and earliest detection have been reported in the Isolator lysis-centrifugation system. It was recently demonstrated that improved detection of both bacteria and fungi was accomplished by mechanically agitating blood culture bottles for the first 24 h of incubation. In this study the detection of fungemia by use of the Isolator system was compared with that of an agitated biphasic system. A total of 182 fungi were isolated from blood specimens inoculated into both culture systems. No difference in the overall recovery of fungi or individual species of yeasts was observed between the two systems. However, all seven isolates of Histoplasma capsulatum were recovered in the Isolator system only. The time required to detect fungemia with each of the two systems was also compared. No statistically significant difference was observed. From the data collected during this 18-month study, it can be concluded that the overall recovery and time of detection of yeasts are equivalent in the lysis-centrifugation system and the agitated biphasic blood culture system. The lysis-centrifugation system is still superior for the detection of filamentous fungi such as H. capsulatum.     

Identification of a galactose-binding lectin on Fusobacterium nucleatum FN-2.

A previous study has suggested that Fusobacterium nucleatum FN-2 contains a galactose-binding protein (lectin) on the cell surface (P. A. Murray, V. Matarese, C. I. Hoover, and J. R. Winkler, FEMS Microbiol. Lett. 40:123-127, 1987). In the present study, the molecular specificity and size of this lectin were investigated by several techniques. Whole-cell affinity chromatography with asialofetuin covalently coupled to Sepharose 6MB demonstrated that 81% of 3H-labeled F. nucleatum were specifically eluted by 0.5 M galactose. Specific binding was calcium dependent and did not occur in the presence of calcium chelators. Binding was inhibited by preincubation with galactose. Agglutination of human parotid saliva by F. nucleatum was also inhibited by galactose and its structural analogs. Inhibition by lactose was 2 times that of galactose, inhibition by p-aminophenyl galactosides was 4 times that of galactose, and inhibition by asialoglycopeptides was 100 times that of galactose. Similar inhibition results were obtained for hemagglutination of neuraminidase-treated erythrocytes. These findings suggest that the binding specificity of F. nucleatum FN-2 is more complex than simply the recognition of the monosaccharide galactose. This is consistent with the concept that lectins considered identical in terms of monosaccharide specificity can recognize fine differences in more complex structures. To identify the specific bacterial component(s) involved in galactose recognition, proteins of F. nucleatum FN-2 were separated on a 4 to 11% gradient sodium dodecyl sulfate slab gel, transferred to nitrocellulose paper to renature bacterial binding sites, and then incubated with 125I-labeled asialofetuin. Autoradiographs of the nitrocellulose revealed a band at a range of Mr 300,000 to 330,000 which was not present when the blots were preincubated with galactose. These data support the concept that F. nucleatum FN-2 possesses a lectin that recognizes galactose and galactose-containing substrates.     

Translocation of Enterococcus faecalis strains across a monolayer of polarized human enterocyte-like T84 cells

We used a two-chamber system to study transcytosis of Enterococcus faecalis across monolayers of human colon carcinoma-derived T84 cells, which show structural resemblance to the native intestine. Among 16 E. faecalis isolates from different sources, the well-characterized strain OG1RF and 8 other isolates (2 endocarditis isolates, 1 urine isolate, and all 5 fecal isolates) showed translocation in this assay, while 6 clinical isolates (3 endocarditis and 3 urine isolates), the recipient strain JH2-2, and the control, Escherichia coli DH5alpha, had no detectable translocation. Of two OG1RF mutants involving the previously studied epa (enterococcal polysaccharide antigen) gene cluster, known to be needed for virulence and resistance to killing by polymorphonuclear leukocytes, one epa mutant (TX5179) was unable to translocate, while TX5180, with an epa disruption farther downstream, showed a moderate decrease in translocation relative to that of the wild-type strain OG1RF (P < 0.01), indicating that the epa gene cluster is important for translocation across a T84 monolayer. This observation was confirmed by complementation of the epa mutant (TX5179) with epa genes and restoration of its translocation ability. In conclusion, we have demonstrated translocation of at least some strains of E. faecalis across T84 monolayers, although strains differ considerably in this ability, and we have demonstrated that epa mutations can cause marked changes in successful translocation. These results suggest that this model may be a useful in vitro system for studying the process of translocation from the intestinal tract.     

Who should give lifestyle advice in general practice and what factors influence attendance at health promotion clinics? Survey of patients' views.

BACKGROUND: Health promotion activity in general practice has increased greatly since 1990. A large proportion of this work is undertaken by practice nurses. Little is known about patients' views about the providers of health promotion or their views about general practice health promotion clinics. AIM: A study was carried out in 1992 to determine patients' views about the provision of health promotion advice by general practitioners and practice nurses and their views about attending health promotion clinics. METHOD: A postal questionnaire was sent to a random sample of 1750 patients aged 16 years and over from five general practices in south Tyneside. The questionnaire explored patients' preferences regarding health promotion advice from the general practitioner or practice nurse in relation to four areas of lifestyle advice and factors that might encourage patients to attend a health promotion clinic. RESULTS: A response rate of 75% was obtained from 1639 eligible patients. Receiving health promotion advice from either the general practitioner or the practice nurse was the most commonly preferred option expressed by patients overall. The ability of health promotion clinic staff to deal with patients' concerns about their illness and short waiting times were more likely to influence patients' attendance at health promotion clinics than the presence of a general practitioner or practice nurse. CONCLUSION: In the present study, many patients found health advice received from practice nurses and general practitioners equally acceptable. However, it was the ability of health professionals to respond to patients' health concerns in the health promotion clinic rather than the type of health professional running the clinic that was important for patients.     

Clinical evaluation of three urine screening tests

Evaluations of screening tests for bacteriuria have traditionally compared the test results with those of quantitative urine cultures. However, many patients with symptomatic urinary tract infections can have less than 10(5) CFU/ml in their urine. Therefore, the results of urine culture and three screening tests (Bac-T-Screen, Chemstrip LN [which tests for leukocyte esterase and nitrate reductase], and Gram stain) were correlated with the clinical classification of urinary tract infection. The Bac-T-Screen test detected 98, 93, and 100% of the infections classified as probable, possible, and asymptomatic, respectively. In contrast, the Gram stain, leukocyte esterase, and nitrate reductase tests were all insensitive screening tests for infection. Additionally, only 45% of the patients with probable infections had greater than or equal to 10(5) CFU/ml. Thus, the majority of infected patients would not have been detected if quantitative urine cultures were used alone.     

Adrenocortical adenoma and carcinoma: histopathological and molecular comparative analysis.

We compared histomorphological features and molecular expression profiles of adrenocortical adenomas (ACAd) and carcinomas (ACCa). A critical histopathological review (mean, 11 slides per patient) was conducted of 37 ACAd and 67 ACCa. Paraffin-embedded tissue cores of ACAd (n = 33) and ACCa (n = 38) were arrayed in triplicate on tissue microarrays. Expression profiles of p53, mdm-2, p21, Bcl-2, cyclin D1, p27, and Ki-67 were investigated by immunohistochemistry and correlated with histopathology and patient outcome using standard statistical methodology. Median follow-up period was 5 years. Tumor necrosis, atypical mitoses, and >1 mitosis per 50 high-power fields were factors that were highly specific for ACCa (P <.001). Number (0 to 4) of unfavorable markers [Ki-67 (+), p21 (+), p27 (+), mdm-2(-)] expressed was significantly associated with mitotic activity and morphologic index (i.e., number of adverse morphologic features) and highly predictive of malignancy (P <.001). Ki-67 overexpression occurred in 0 ACAd and 36% ACCa (P <.001) and was significantly associated with mitotic rate and unfavorable morphologic index (P <.001). Tumor necrosis, atypical mitoses, >5 mitoses per 50 high-power fields, sinusoidal invasion, histologic index of >5, and presence of more than two unfavorable molecular markers were associated significantly with metastasis in ACCa. Well-established histopathologic criteria and Ki-67 can specifically distinguish ACCAd from ACCa. Tumor cell proliferation (Ki-67) correlates with mitotic activity and morphologic index. Tumor morphology is a better predictor of metastatic risk in ACCa than current immunohistochemistry-detected cell cycle regulatory and proliferation-associated proteins.