Evolution of asexual and sexual reproduction in the aspergilli.

Aspergillus nidulans has long-been used as a model organism to gain insights into the genetic basis of asexual and sexual developmental processes both in other members of the genus Aspergillus, and filamentous fungi in general. Paradigms have been established concerning the regulatory mechanisms of conidial development. However, recent studies have shown considerable genome divergence in the fungal kingdom, questioning the general applicability of findings from Aspergillus, and certain longstanding evolutionary theories have been questioned. The phylogenetic distribution of key regulatory elements of asexual reproduction in A. nidulans was investigated in a broad taxonomic range of fungi. This revealed that some proteins were well conserved in the Pezizomycotina (e.g. AbaA, FlbA, FluG, NsdD, MedA, and some velvet proteins), suggesting similar developmental roles. However, other elements (e.g. BrlA) had a more restricted distribution solely in the Eurotiomycetes, and it appears that the genetic control of sporulation seems to be more complex in the aspergilli than in some other taxonomic groups of the Pezizomycotina. The evolution of the velvet protein family is discussed based on the history of expansion and contraction events in the early divergent fungi. Heterologous expression of the A. nidulans abaA gene in Monascus ruber failed to induce development of complete conidiophores as seen in the aspergilli, but did result in increased conidial production. The absence of many components of the asexual developmental pathway from members of the Saccharomycotina supports the hypothesis that differences in the complexity of their spore formation is due in part to the increased diversity of the sporulation machinery evident in the Pezizomycotina. Investigations were also made into the evolution of sex and sexuality in the aspergilli. MAT loci were identified from the heterothallic Aspergillus (Emericella) heterothallicus and Aspergillus (Neosartorya) fennelliae and the homothallic Aspergillus pseudoglaucus (=Eurotium repens). A consistent architecture of the MAT locus was seen in these and other heterothallic aspergilli whereas much variation was seen in the arrangement of MAT loci in homothallic aspergilli. This suggested that it is most likely that the common ancestor of the aspergilli exhibited a heterothallic breeding system. Finally, the supposed prevalence of asexuality in the aspergilli was examined. Investigations were made using A. clavatus as a representative 'asexual' species. It was possible to induce a sexual cycle in A. clavatus given the correct MAT1-1 and MAT1-2 partners and environmental conditions, with recombination confirmed utilising molecular markers. This indicated that sexual reproduction might be possible in many supposedly asexual aspergilli and beyond, providing general insights into the nature of asexuality in fungi.

Speciation despite globally overlapping distributions in Penicillium chrysogenum: the population genetics of Alexander Fleming's lucky fungus.

Eighty years ago, Alexander Fleming described the antibiotic effects of a fungus that had contaminated his bacterial culture, kick starting the antimicrobial revolution. The fungus was later ascribed to a putatively globally distributed asexual species, Penicillium chrysogenum. Recently, the species has been shown to be genetically diverse, and possess mating-type genes. Here, phylogenetic and population genetic analyses show that this apparently ubiquitous fungus is actually composed of at least two genetically distinct species with only slight differences detected in physiology. We found each species in air and dust samples collected in and around St Mary's Hospital where Fleming worked. Genotyping of 30 markers across the genome showed that preserved fungal material from Fleming's laboratory was nearly identical to derived strains currently in culture collections and in the same distinct species as a wild progenitor strain of current penicillin producing industrial strains rather than the type species P. chrysogenum. Global samples of the two most common species were found to possess mating-type genes in a near 1:1 ratio, and show evidence of recombination with little geographic population subdivision evident. However, no hybridization was detected between the species despite an estimated time of divergence of less than 1MYA. Growth studies showed significant interspecific inhibition by P. chrysogenum of the other common species, suggesting that competition may facilitate species maintenance despite globally overlapping distributions. Results highlight under-recognized diversity even among the best-known fungal groups and the potential for speciation despite overlapping distribution.

Genotyping fetal DNA by non-invasive means: extraction from maternal plasma.

BACKGROUND AND OBJECTIVES: Identification of fetal DNA in maternal plasma may allow genetic analysis without the use of invasive techniques. The aim of this study was to extract DNA from maternal plasma, identify fetal material through the presence of SRY or RHD gene sequences and assess the reliability of these results. MATERIALS AND METHODS: A polymerase chain reaction (PCR) method of a commercial kit was used with primers for SRY or exon 10 of the RHD gene sequence. RESULTS: Multiple plasma samples were collected from 60 women who were evaluable for either SRY or RHD, or both, fetally derived DNA sequences. Fetal DNA was present in the plasma throughout the pregnancies and for some hours or days after delivery. CONCLUSION: Fetal DNA can be reliably detected in maternal plasma from early in pregnancy and normally is cleared within days of delivery.

Measuring the immeasurable? Assessing the quality of anaesthetic care.

Quality can be viewed from three perspectives: the requirements of the healthcare system; issues related to delivery of care, such as access; and methods used to measure quality, such as outcome analysis. These viewpoints can be applied to the anaesthetic literature to analyse the assessment of quality of care.

Somatosensory evoked potential monitoring during cardiac surgery: an examination of brachial plexus dysfunction.

OBJECTIVE: To observe the effects of the Favoloro and sternal retractors on the ulnar and median nerve somatosensory evoked potentials (SSEPs) and to identify any relationship with postoperative brachial plexus injury. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Twenty cardiac patients. INTERVENTIONS: SSEPs were studied in patients undergoing cardiac surgery using normothermic cardiopulmonary bypass. Evoked potentials were obtained from bilateral median and ulnar nerves. MEASUREMENTS: The incidence of nerve-specific SSEP changes and their temporal relationship to retractor usage were determined. The overall incidence of SSEP changes was 75%. There were no differences (p > 0.05) between the group showing changes (n = 15) and the group with no changes (n = 5) with respect to age, body surface area, weight, cross-clamp or cardiopulmonary bypass times. There also were no differences (p > 0.05) between the frequencies of left- and right-sided changes, or in nerve-specific SSEP changes. Seventy-four percent of SSEP changes correlated with retractor usage. No SSEP changes were associated with the Favoloro retractor. Significant SSEP depression, assessed by either percentage reduction in amplitude or persistent amplitude reduction, occurred in the absence of postoperative neurological deficits. There were no detected postoperative brachial plexus injuries. CONCLUSIONS: SSEP changes correlate with the use of the sternal retractor but not the Favoloro retractor. It was not possible to replicate the results of previous investigators in predicting postoperative neurological deficits based on the SSEP changes, and therefore the routine application of SSEP as a monitor cannot be recommended on the basis on these data.

Report of the Anaesthetic Mortality Committee of Western Australia 1990-1995.

The structure and function of the Anaesthesia Mortality Committee of Western Australia are described. Reports of 500 deaths from 1990 to 1995 are analysed and discussed. The Committee determined that, in 21 cases, anaesthesia played a significant part in the death of the patient. These cases are further analysed. Deaths due mainly to anaesthetic factors are estimated to occur once in every 40,000 operations in Western Australia. One or two otherwise healthy people die each year in Western Australia from an anaesthetic mishap.

Anaesthetic management of a parturient with severe muscular dystrophy, lumbar lordosis and a difficult airway.

PURPOSE: This case describes the management of a 19-yr-old wheelchair bound primigravida with severe muscular dystrophy who presented for Caesarean section after spontaneous rupture of membranes. Anaesthesia was influenced by several features of her systemic disease which were impediments to both neuraxial and general anaesthesia. CLINICAL FEATURES: Other than for a prenatal record and the history obtainable from the patient, little additional medical information was available. Physical examination showed diffuse muscular weakness and an anatomically abnormal airway. Examination of the spine showed slight 10-15 degrees thoracolumbar scoliosis and > 45 degrees lumbar lordosis. Fetal assessment was normal. Echocardiography revealed mildly decreased left ventricular function and was consistent with pulmonary hypertension. After discussion with the patient and her obstetrician, elective Caesarean delivery was deemed the best management. Neuroaxial anaesthesia was at an increased risk of failure due to the profound lumbar lordosis. A plan for awake intubation and general anaesthesia was described to the patient in case regional anaesthesia could not be initiated. A fibreoptic bronchoscope and difficult intubation kit were made available. General anaesthesia was expected to have increased risk of postoperative pulmonary complications, hence epidural anaesthesia was attempted. After difficult catheter insertion, a sensory block was titrated to a T4 level. This was well tolerated by both mother and fetus. A healthy baby was delivered with Apgar scores of 9 and 9. Postoperatively the mother was transferred to the intensive care unit. After 72 hr, the patients respiratory status allowed transfer to the word. CONCLUSION: This case illustrates the use of epidural anaesthesia in the successful management of a severely compromised patient with limb-girdle muscular dystrophy undergoing elective Caesarean section.

Anaesthetic management of a patient with myasthenia gravis and tracheal stenosis.

PURPOSE: The combination of myasthenia gravis and tracheal obstruction presents a number of difficulties for anaesthetic management. This case illustrates the advantages of careful planning. CLINICAL FEATURES: A 66-yr-old man with myasthenia gravis required resection of a stenosis at the site of an old tracheostomy. The primary goal was to accomplish safe management of the airway, a task made more difficult because the airway was shared with the surgeon. Awake fibreoptic examination of the tracheal stenosis performed in the operating room provided useful information in planning the subsequent anaesthetic. From this examination, it was found that the trachea could be intubated by a normal endotracheal tube passed through the stenosis over the fibreoptic bronchoscope. Intraoperatively, the orotracheal tube was withdrawn temporarily and replaced with an endotracheal tube placed by the surgeon into the distal trachea. Extubation was carried out judiciously and a plan for reintubation prepared in advance. The anaesthetic plan was modified because of the myasthenia gravis. Following careful investigation of the extent of the patient's disease and its treatment, an assessment was made of the patient's need for postoperative ventilation. The anaesthetic plan included maintenance of anticholinergic medications until the time of surgery and their early resumption postoperatively, avoidance of neuromuscular blocking agents, and careful monitoring of neuromuscular function during the anaesthetic. CONCLUSION: Careful examination of the area of tracheal stenosis and a carefully considered plan for reintubation are prerequisites for this type of surgery. Clinically well controlled myasthenia gravis was managed successfully using familiar principles.

Developing a "clinical presentation" curriculum at the University of Calgary.

Currently, medical curricula are structured according to disciplines, body systems, or clinical problems. Beginning in 1988, the faculty of the University of Calgary Faculty of Medicine (U of C) carefully evaluated the advantages and disadvantages of each of these models in seeking to revise their school's curriculum. However, all three models fell short of a curricular structure based on current knowledge and principles of adult learning, clinical problem solving, community demands, and curriculum management. By 1991, the U of C had formulated a strategic plan for a revised curriculum structure based on the way patients present to physicians, and implementation of this plan has begun. In creating the new curriculum, 120 clinical presentations (e.g., "loss of consciousness/syncope") were defined and each was assigned to an individual or small group of faculty for development based on faculty expertise and interest. Terminal objectives (i.e., "what to do") were defined for each presentation to describe the appropriate clinical behaviors of a graduating physician. Experts developed schemes that outlined how they differentiated one cause (i.e., disease category) from another. The underlying enabling objectives (i.e., knowledge, skills, and attitudes) for reaching the terminal objectives for each clinical presentation were assigned as departmental responsibilities. A new administrative structure evolved in which there is a partnership between a centralized multidisciplinary curriculum committee and the departments. This new competency-based, clinical presentation curriculum is expected to significantly enhance students' development of clinical problem-solving skills and affirms the premise that prudent, continuous updating is essential for improving the quality of medical education.

Anaesthetic management of a patient with a descending thoracic aortic aneurysm and severe bilateral bullous pulmonary parenchymal disease.

The anaesthetic management of the surgical repair of a descending aortic aneurysm in a patient with large, bilateral, pulmonary bullae is described. Anaesthesia for descending aortic surgery normally involves unilateral, positive-pressure ventilation, an option which poses some risk of barotrauma in the presence of bilateral bullae. Patients with bullous disease commonly have severe lung disease and thorough preoperative assessment and preparation are necessary. Intraoperatively, bilateral rupture of the bullae could be catastrophic and preparations should be made for this possibility. In order to diminish this risk, a surgical technique including preemptive collapse of the bulla by minithoracotomy and tube drainage, with use of a bronchial blocker to the affected part of the lung may be used. If rupture occurs, then high frequency jet ventilation may be effective. Use of a double lumen endobronchial tube may be advantageous for patients with either unilateral and bilateral bullae. Anaesthesia for patients with bullae should avoid positive-pressure ventilation and nitrous oxide in order to limit the risk of barotrauma from a ball valve mechanism. In this case, the risk of barotrauma was reduced by performing an inhalational induction of anaesthesia and limiting peak inflation pressures during thoracotomy. It was elected to use positive-pressure ventilation through a double lumen endobronchial tube following chest incision. A high frequency jet ventilator was available but not employed. Anaesthetic management was complicated by the presence of pleural adhesions, surgical approach directly through a bulla, and the requirement for one lung ventilation.

Anaesthesia for coronary artery bypass surgery supplemented with subarachnoid bupivacaine and morphine: a report of 18 cases.

We report our experience with general anaesthesia (GA) supplemented with subarachnoid bupivacaine and morphine for coronary artery bypass surgery (CABG) in 18 patients. Fifteen patients were male, and mean age was 62 yr. Anaesthesia (GA) was induced with alfentanil 97 +/- 22 micrograms.kg-1 and midazolam 0.04 +/- 0.02 mg.kg-1 supplemented with a muscle relaxant, and maintained with isoflurane (0.25-0.5%) in oxygen throughout surgery. Spinal anaesthesia (SA) was then performed at a lumber level using hyperbaric bupivacaine (23-30 mg) and/or lidocaine (150 mg) with morphine (0.5-1 mg). Pooled data showed the following haemodynamic results (P < 0.05). Induction of GA produced a decrease in mean arterial pressure (MAP). Addition of SA produced a decrease in heart rate. Heart rate and MAP did not change with sternotomy. Phenylephrine support of arterial blood pressure was used at some time during operation in 17 patients. Supplementation of GA was minimal. Patients received 2.7 +/- 0.7 coronary grafts. Operating room time was 3.9 +/- 0.6 hr. Postoperative analgesic requirements were minimal, and in half of the patients tracheal extubation occurred on the day of surgery. Complications included one myocardial infarction, one resternotomy, a metabolic encephalopathy in a dialysis-dependent patient, and one case of herpes labialis. No patient recalled intraoperative events. Combined GA with SA may be an effective technique for CABG surgery. Further study of the cardiovascular, neurological and metabolic effects of the technique is required.

The cost of an established quality assurance programme: is it worth it?

Although the literature concerning quality assurance (QA) is voluminous, little information exists about the costs or benefits of departmentally based QA programmes. We measured the direct costs and then investigated the financial and nonfinancial benefits derived from a well-funded QA programme over a period of five years. Data were obtained from departmental budgets, annual reports of the QA programme, and several databases used by the programme. The average annual cost was $79,900, with salaries being the largest component, while $14,300 each year were recovered through the activities of the programme. True costs were higher than those calculated since time volunteered by medical staff and resources shared with other programmes could not be determined. Some of the costs encountered at the outset of this programme were later offset by the use of commercial software and employment of volunteers and casual staff. Fifty-three projects were identified over the five-year period. Most lacked directly measurable financial outcomes (because they were based on education, research, patient or practitioner satisfaction). The benefit of the programme has been greater to the department than suggested from cost analysis alone. Although this programme could not be justified on a simple cost recovery basis, the authors felt it to be worthy of continued support because of the nonfinancial benefits. However, modification is required to minimize costs.

A patient presenting for pharyngeal biopsy.

A 68-yr-old man presented for pharyngeal biopsy under general anaesthesia. Coincidentally he was found to have a large mediastinal mass. The evaluation of this patient is described. The exact risk of catastrophic airway collapse on induction of anaesthesia in patients with mediastinal masses is controversial but probably small. As there is no test to prevent airway collapse, it is suggested that attempts at biopsy be performed with regional anaesthesia after radiotherapy.

Simple narcotic kits for controlled-substance dispensing and accountability.

Operating rooms require a storage, dispensing and accounting system for restricted drugs which satisfies narcotics control authorities and is compatible with efficient care of patients. We describe narcotic kits containing fentanyl-morphine-midazolam, alfentanil-midazolam and sufentanil-midazolam, for general operating rooms, and two kits with larger quantities of fentanyl and sufentanil for cardiac operating rooms. The container for each kit is a video cassette holder which has a foam-rubber liner with sculpted depressions for each ampoule. Sealed kits are delivered each morning from pharmacy to the locked narcotics cupboard in the recovery room. On request, the recovery room nurse unlocks the cupboard and the anaesthetist signs out the required kit(s) for the day. A drug utilization form is enclosed with each kit, on which the anaesthetist records the amount of drug administered to each patient, and before returning the kit to the locked narcotics cupboard, the total amount of each drug used, discarded, and returned. Used kits are collected the following morning by a pharmacy technician who reconciles the contents and drug form of each kit. More than 40 staff anaesthetists and a similar number of residents have used the system for seven years, during which time 130,000 patients have passed through the operating rooms. Detection of one case of drug diversion by a staff anaesthetist was made partly by the control system, but mainly by behavioural changes. The system is simple, inexpensive, and effective and has been well received by the departments of pharmacy, anaesthesia, and nursing.

Use of refractometry to identify opioid-containing solutions.

The purpose of this laboratory study was to assess the value of refractometry in identifying the contents of a variety of opioid-containing solutions. A hand-held refractometer was used to document the refraction produced by the undiluted contents of alfentanil, fentanyl, morphine, sufentanil ampoules and by solutions of Ringer's lactate, 0.9% saline, 3.3% dextrose in 0.3% saline, and distilled water. Each opioid was then serially diluted in serial 1:2, 1:4, and 1:8 dilutions in each of these solutions and the refractions of each determined. Based on this information, blinded identification of various diluted opioid solutions was attempted. Refractometer values for undiluted fentanyl and sufentanil were identical with those for distilled water. Those for undiluted alfentanil and morphine were almost identical with each other and with 1:2 and 1:4 dilutions of either drug in Ringer's lactate or 0.9% saline. We conclude that refractometry is an unreliable screening method to detect tampering with opioid solutions.