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R M Cooper

Publications and source records attributed to R M Cooper.

At least 37 records · Page 2Linked to original sources
PubMed1995

Cloning and characterisation of a gene encoding a cuticle-degrading protease from the insect pathogenic fungus Metarhizium anisopliae.

Metarhizium anisophilae (Ma) secretes a range of proteases when grown in vitro on insect cuticle. A trypsin-like serine protease, PR2, was purified from culture filtrates by anion exchange chromatography and the N-terminal sequence determined. Using oligodeoxyribonucleotide probes based on this sequence and that of the highly conserved trypsin active site, a gene was isolated from a lambda EMBL3 genomic library of Ma isolate ME1. Sequencing of the gene and RT-PCR revealed that the gene contains two introns which are 94 and 40 bp long. The deduced protein consists of 254 amino acids, has a putative signal sequence to allow transport into the endoplasmic reticulum and probably undergoes a second proteolytic processing step at its N terminus to yield the mature enzyme. The putative mature enzyme has extensive homology with other serine proteases of the trypsin subclass and, in particular, with the trypsin characterised from Fusarium oxysporum.

Amino Acid Sequence↗
PubMed1995

Superiority of conventional culture technique over rapid detection of group A Streptococcus by optical immunoassay.

An optical immunoassay (OIA) has been reported to be more sensitive than conventional culture for the detection of Group A Streptococcus, eliminating the need for culture. We attempted to confirm the sensitivity and specificity through a laboratory quantitation study and a clinical trial. OIA did not detect Group A Streptococcus below 10(5) colony forming units (CFU). Culture detected Streptococcus to 10(2) CFU from the inoculated swab. In the clinical study, throat swabs were obtained from 77 patients in an outpatient clinic. Compared with culture, the sensitivity of OIA was 78% and the specificity was 90%. These results demonstrate that OIA was less sensitive than culture in seeded experiments and missed 22% of positives in clinical practice. Our study, contrary to previous reports, suggests that OIA is not sensitive enough to be used as the sole assay for Group A Streptococcus pharyngitis.

Adult↗
PubMed1995

Metabolic activity in rat visual system during exposure to high and low intensities of patterned and diffuse light.

An examination of rat visual system activity, during exposure to either "pattern" (black and white stripes) or "diffuse" (eye covered by white mask) visual stimulation at high or low illumination intensities (1600 and 1 lux at cornea, respectively), was carried out using the 2-deoxyglucose (2-DG) autoradiographic technique. Pattern elevated 2-DG uptake in the dorsal and ventral lateral geniculate nuclei, in the lateral posterior nucleus, and in area 17, but was less effective at the high than at the low light intensity. Diffuse light also elevated 2-DC uptake in the thalamic nuclei but the increase was less impressive and the same at both intensities. Diffuse light at either high or low intensity had no effect on cortex. Like thalamus and cortex, pattern was a less effective stimulus for the colliculus at the high than at the low intensity, but, in contrast to thalamus and cortex, high intensity diffuse light suppressed 2-DG uptake in the colliculus to a level below that produced by darkness; low intensity diffuse light had no effect. These 2-DG findings are discussed in terms of how forebrain and midbrain divisions of the rat's rod-dominated visual system maintained their respective spatial processing and change-detecting functions over a considerable range of illumination intensity.

Animals↗
PubMed1995

Fifty years ago.

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History, 20th Century↗
PubMed1994

The use of an endotracheal ventilation catheter for jet ventilation during a difficult intubation.

This case report describes the use of an endotracheal ventilation catheter (ETVC) to provide prolonged intraoperative jet ventilation, reintubation and the maintenance of tracheal access following extubation. It emphasizes that excellent oxygenation and ventilation can be achieved but such management can be complicated by a pneumothorax even when the risks are minimized. A 43-yr-old man presented for possible pulmonary sleeve resection. Placement of a double lumen endotracheal tube (DLT) by direct laryngoscopy was unsuccessful due to the inability to visualize the glottis. A 7.5 mm endotracheal tube (ETT) was successfully introduced over a fibreoptic bronchoscope (FOB). An ETVC was passed, permitting manually cycled jet ventilation while general intravenous anaesthesia and muscle relaxation were maintained. The ETT was then withdrawn over the ETVC and jet ventilation continued for approximately 90 min, while attempts at placing a DLT over a now malfunctioning FOB continued. These attempts were eventually abandoned and the patient was returned to the post-anaesthesia care unit (PACU) haemodynamically stable. The trachea was extubated over the ETVC, which remained in situ. A pneumothorax was noted on the postoperative chest x-ray. This case illustrates prolonged intraoperative jet injection via a "jet stylet" with satisfactory ventilation and oxygenation but complicated by a pneumothorax. Also it illustrates a strategy for the management of a "difficult extubation."

Adult↗
PubMed1994

Partial characterization of specific inducers of a cuticle-degrading protease from the insect pathogenic fungus Metarhizium anisopliae.

The insect pathogenic fungus Metarhizium anisopliae produces several extracellular cuticle-degrading proteases and evidence is consistent with one of these, PR1, which is a chymoelastase, being a determinant of pathogenicity. We have shown previously that PR1 production is regulated by both carbon catabolite and nitrogen metabolite repression and also by specific induction under derepressed conditions by insect cuticle. In the present work we have established that an enzymically released proteinaceous component(s) of insect cuticle is capable of inducing PR1 (based on appearance of extracellular activity). Cuticle of the desert locust Schistocerca gregaria treated with KOH to remove protein failed to induce PR1 production, whereas cuticle treated with either chloroform or ether to remove lipids still induced PR1. Cuticle digested with either PR1 or the trypsin-like PR2 of M. anisopliae released peptides mainly in the range 150-2000 Da; addition of these peptides generated by PR1 or PR2 at 3 micrograms alanine equivalents ml-1 induced PR1 production to a level similar (75%) to that obtained with untreated insect cuticle. Several amino acids and peptides which are abundant in insect cuticular protein (Ala, Gly, Ala-Ala, Ala-Ala-Ala, Ala-Pro and Pro-Ala) were tested at a range of concentrations and in restricted cultures for their ability to induce PR1. None induced the protease to the levels seen with cuticle or peptides enzymically released from cuticle, although some dimers and notably the monomers Ala and Gly gave 2-2.7-fold enhanced PR1 activity above depressed basal levels (up to 48-57% of that achieved with induced synthesis on cuticle).(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

Analysis of aminopeptidase and dipeptidylpeptidase IV from the entomopathogenic fungus Metarhizium anisopliae.

Analytical and preparative isoelectric focusing were used to separate extracellular isoenzymes of aminopeptidase (pI 4.51, M(r) 45,000, pH optimum 7.0) and prolyl-dipeptidylpeptidase (pI 4.01, M(r) 74,000, pH optimum 8.0) produced by the entomopathogenic fungus Metarhizium anisopliae during growth on locust cuticle. Production of both activities is repressed by readily utilized nitrogen sources, but unlike the aminopeptidase, the dipeptidylpeptidase was also excreted at high levels during growth on casein. Casein-grown cultures contained additional isoenzymes with activity against lysyl-alanyl-4-methoxy-2-naphthylamine indicating M. anisopliae possesses multiple peptidases as an adaptation to different nutrient conditions. The aminopeptidase hydrolysed alanyl-leucyl-alanine and showed a broad specificity versus monoaminoacyl beta-naphthylamine (beta NA) substrates with alanine beta NA being the most rapidly hydrolysed. Inhibition by both bestatin and amastatin indicated similarities to the class of alanyl aminopeptidases (aminopeptidase M). Metal complexing agents also inhibited the aminopeptidase indicating a metal ion requirement. A specific inhibitor for serine proteases [diisopropyl fluorophosphate (DFP)] was without effect. The dipeptidylpeptidase showed a strong preference for substrates having a penultimate proline residue including alanyl-prolyl-glycine and aa-prolyl-beta NA substrates. The enzyme showed a broad specificity at the N-terminal amino acid. Inhibition by diprotin A indicates similarities with mammalian prolyl-dipeptidylpeptidases. The enzyme was also inhibited by DFP, implying involvement of a serine residue in catalysis. The results are discussed in the context of cuticle degradation and the participation of exopeptidases as mediators in releasing amino acids necessary for pathogen growth.

Amino Acid Sequence↗
PubMed1993

Penicillin-binding proteins from Erwinia amylovora: mutants lacking PBP2 are avirulent.

Radiolabelled penicillin G was used to examine penicillin-binding proteins (PBPs) from Erwinia amylovora (OT1). This procedure identified seven PBPs with molecular masses ranging from 22 to 83 kDa. E. amylovora PBPs were compared with those from Escherichia coli (JM101) and from two spherical, avirulent TnphoA mutants derived from OT1. Radiolabelled penicillin G bound to only six proteins from the spherical mutants which lacked a 69-kDa PBP. The spherical mutants could be complemented by the cloned E. coli pbpA-rodA operon, which restored both cell shape and virulence to apple seedlings. This suggested that the E. amylovora 69-kDa PBP is probably the functional equivalent of the E. coli PBP2 protein. Southern blot analysis using the E. coli rodA and pbpA genes as radiolabelled probes showed that TnphoA had inserted into the E. amylovora equivalent of the E. coli rodA-pbpA operon. Southern blots to chromosomal DNAs of the two spherical mutants, using the cloned hrp and dsp genes from E. amylovora as radiolabelled probes, confirmed that the TnphoA insertions were not located in the region of the E. amylovora chromosome postulated to encode known virulence factors. Both of the spherical TnphoA mutants synthesized amounts of extracellular polysaccharide equivalent to those synthesized by the wild-type strain (OT1), were resistant to lysis in distilled water and to lysozyme, and elicited the hypersensitive response on nonhost plants. These results indicate a possible role for cell shape in the virulence of this plant pathogen.

Bacterial Proteins↗
PubMed1993

An intergroup phase III randomized study of doxorubicin and dacarbazine with or without ifosfamide and mesna in advanced soft tissue and bone sarcomas.

PURPOSE AND METHODS: Doxorubicin alone or with dacarbazine (DTIC; AD) is considered the best available therapy for metastatic adult sarcomas. Ifosfamide is active in sarcomas that have failed to respond to a doxorubicin-based regimen. This study was designed to determine if ifosfamide added to doxorubicin and DTIC (ADI) significantly effects toxicity, response rate, and survival. Patients with measurable metastatic or unresectable sarcoma were randomized to receive AD or ADI. Patients with chondrosarcomas, fibrosarcomas, and other sarcomas of bone were eligible, although those with osteosarcoma, rhabdomyosarcoma, Ewing's sarcoma, Kaposi's sarcoma, and mesothelioma were excluded, as were patients with prior chemotherapy for sarcoma or prior doxorubicin. RESULTS: Between 1987 and 1989, 340 eligible patients were randomized. Significantly more myelosuppression, a higher response rate (17% v 32%; P < .002) and longer time to progression (4 v 6 months; P < .02) were observed for patients who received ifosfamide. An overall survival advantage for the two-drug regimen (12 v 13 months; P = .04) was not significant by multivariate analysis. CONCLUSION: In all three randomized trials of doxorubicin with and without ifosfamide (Eastern Cooperative Oncology Group [ECOG], European Organization for Research and Treatment of Cancer [EORTC], and this study), the response rate was higher for the ifosfamide-containing arm, significantly so in this and the ECOG studies. An improved response rate may be particularly important for the preoperative management of high-grade, borderline resectable lesions or pulmonary metastases, particularly in younger patients. In older patients, or for low-to intermediate-grade lesions, doxorubicin and DTIC followed by ifosfamide on progression is preferred.

Adolescent↗
PubMed1991

Effects of flashing-diffuse light on [2-14C]deoxyglucose uptake in the visual system of the black-hooded rat.

What components of the visual system process diffuse light information? A [2-14C]deoxyglucose (2-DG) autoradiographic analysis revealed that exposure of freely moving rats (wearing light-diffusing masks) to flashing-diffuse light consistently elevated 2-DG uptake in the lateral geniculate nucleus and superior colliculus to levels rivalling those occurring in rats exposed to flashing-gratings. Uptake in visual cortex (area 17) in response to flashing-diffuse light, however, varied as a function of early contour experience, i.e. lower than that produced by darkness in rats reared with high contrast patterns, higher than darkness in rats which had been lid-sutured from the time of eye opening, and falling between these two extremes in 'ordinary' cage-reared rats. The findings point to subcortical mediation of discriminations based on diffuse light information. Cortex might participate in the processing of diffuse light information in the special case of animals lacking contour experience during development.

Animals↗
PubMed1991

Metabolic activation of the rat visual system by patterned light and footshock.

Autoradiography with [14C]2-deoxy-D-glucose was used to examine metabolic changes in the visual system of hooded rats exposed to patterned light or to darkness following footshock. Primary retinorecipient structures (superficial layer of the superior colliculus and the dorsal lateral geniculate nucleus) showed a response to light but not to shock. Higher visual sites showed two different shock effects. First, in darkness the intermediate grey layer of the superior colliculus was suppressed by the shock. Second, in the lateral posterior nucleus and primary visual cortex, the footshock led to significant enhancement of the metabolic responses to the patterned light. The findings suggest that footshock-induced arousal has significant modulatory effects on the operations of higher visual centers of behaving rats.

Analysis of Variance↗
PubMed1991

Activity-dependent changes in eye influence during monocular blockade: increases in the effects of visual stimulation on 2-DG uptake in the adult rat geniculostriate system.

We examined the effects of loss of monocular retinal activity on 2-deoxyglucose (2-DG) uptake in the adult rat geniculostriate system. Of particular interest was whether the influence of the normally functioning eye changed during long-term contralateral retinal silence. Group 1 rats were subjected to short-term (24 hours) and group 2 rats to long-term (21-90 days) monocular tetrodotoxin (TTX) blockade, and metabolic activity was assessed during exposure to square-wave gratings. Group 1 rats exhibited patterns of cortical glucose utilization commensurate with complete monocular loss of retinal activity: minimal 2-DG uptake in contralateral monocular area 17 and dorsal lateral geniculate nucleus (LGN), and a bilateral depression in the binocular regions; 2-DG uptake was highest in the monocular regions fed by the stimulated normal eye (in both area 17 and the LGN) and these regions appeared unaffected by the monocular blockade. After repeated injections of TTX (group 2), metabolic activity in binocular area 17 and binocular LGN increased bilaterally relative to the metabolically active monocular regions contralateral to the normal eye. Group 3 rats were monocularly TTX-injected for 30 or 60 days, and, 24 hours before 2-DG, all retinal activity was eliminated by means of binocular TTX injections or binocular enucleation. Glucose utilization in the binocular regions of both area 17 and the LGN in these rats was depressed to levels seen in monocular area 17 after complete and recent loss of activity from the contralateral eye, indicating that the metabolic increase which occurred in the binocular regions during long-term monocular retinal blockade was dependent upon the neuronal processing of retinal information from the non-TTX eye. We conclude that, in the adult rat, an activity-dependent, physiologically based shift in ocular influence occurred in the binocular geniculostriate system during long-term monocular retinal inactivation.

Animals↗
PubMed1991

[2- 14C]deoxyglucose uptake in rat visual system during flashing-diffuse and flashing-pattern stimulation over a 6 log range of luminance.

The 2-deoxyglucose autoradiographic technique was used to assess the metabolic activity of cortical area 17, the dorsal and ventral lateral geniculate nuclei, the lateral posterior nucleus, and the superior colliculus, during 5-Hz flashing-pattern (montage of black and white square-wave gratings) and flashing-diffuse (eye covered with white mask) stimulation at three intensities over a 6 log range. In area 17 flashing-pattern was found to be equally effective at elevating uptake of the functional label over the photopic-scotopic range of luminance levels tested, whereas flashing-diffuse was ineffective. In subcortical nuclei, however, flashing-diffuse was no less effective than flashing-pattern and uptake of the label correlated positively with intensity level. The results suggest that the subcortical components of the visual system do play an important role in the processing of intensity information and that primary visual cortex does not.

Animals↗