Local clinical quality monitoring for detection of excess operative deaths.

A monitoring system for cardiac surgery has been in use at Papworth Hospital for 10 years. We wished to determine whether this system would have detected an increase in deaths associated with a single practitioner, whether a poorly performing doctor or a serial killer such as Dr Harold Shipman, whose activities went undetected in the absence of a monitoring system for nearly a quarter of a century. Random extra deaths were artificially introduced into the practice of a surgeon and an anaesthetist in a way that broadly reproduced Shipman's pattern. The standard monitoring system was then used to analyse the hypothetical data thus generated. Using the current standard monitoring, the excess deaths would have been detected in less than 10 months. Suspicions would have been raised even earlier. Robust local quality monitoring of risk-adjusted outcomes is possible and, in our opinion, essential.

Low temperature acclimated populations of the grain aphid Sitobion avenae retain ability to rapidly cold harden with enhanced fitness.

In contrast to previous studies of rapid cold-hardening (RCH), which have investigated the responses of insects maintained under 'summer conditions' (20 degrees to 25 degrees C), this study focuses on the ability of low-temperature acclimated insects to undergo RCH. When the grain aphid Sitobion avenae Fabricus was low-temperature acclimated by rearing for three generations at 10 degrees C, the discriminating temperatures (temperature that results in approximately 20% survival after direct transfer from the rearing temperature to a sub-zero temperature for a period of 3 h), of first instar nymphs and adult aphids were -11.5 degrees and -12 degrees C, respectively. Maximum rapid cold-hardening was induced by cooling aphids at 0 degrees C for 2 h (nymphs) or 30 min (adults), resulting in survival at the respective discriminating temperatures increasing from 26% to 96% (nymphs) and 22% to 70% (adults). Cooling from 10 degrees to 0 degrees C at 1 degree, 0.1 degrees and 0.05 degrees C min-1 significantly increased survival of nymphs at the discriminating temperature, but not of adults. There were no ;ecological costs' associated with rapid cold-hardening at 0 degrees C, or with exposure of rapidly cold-hardened aphids to the discriminating temperatures; fecundity and longevity, in both nymphs and adults were either similar to control aphids or significantly increased. The study demonstrates that rapid cold-hardening ability is retained in aphids that have already undergone cold-acclimation, as would be the case in overwintering aphids. Both rapid cold-hardening and subsequent exposure at previously lethal temperatures can enhance fitness in surviving individuals.

Cold shock injury and ecological costs of rapid cold hardening in the grain aphid Sitobion avenae (Hemiptera: Aphididae).

The ability of first instar nymphs and newly moulted pre-reproductive adults of the grain aphid S. avenae to rapidly cold harden was investigated. When nymphs reared at 20 degrees C were transferred directly to -8 degrees C for 3 h, there was 18% survival. This exposure was selected as the discriminating temperature. Maximum increases in survival were achieved by acclimating nymphs for 2 h at 0 degrees C and adults for 3 h at 0 degrees C, resulting in survival of 83% and 68%, respectively. Cooling nymphs from 10 to 0 degrees C at different rates (1, 0.1 and 0.05 degrees C min(-1)) also increased cold hardiness, with the slowest rate of 0.05 degrees C min(-1) conferring the highest survival following exposure to the discriminating temperature. Adult aphids also expressed a rapid cold hardening response but to a lesser extent, with survival increasing from 16% to 68% following 3 h at 0 degrees C. There were no 'ecological costs' associated with rapid cold hardening in terms of development, longevity or fecundity. The data support the hypothesis that rapid cold hardening can be induced during the cooling phase of natural diurnal temperature cycles, allowing insects to track daily changes in environmental temperatures.

Resuscitation after cardiac surgery: are we ageist?

BACKGROUND AND OBJECTIVE: To study the effect of age and other risk factors on: (a) the incidence and outcome of cardiopulmonary resuscitation and (b) any decision to institute a 'Do Not Attempt Resuscitation' order following cardiac surgery. METHODS: Prospective audit of cardiac arrest calls following 6550 consecutive open-heart surgery cases and retrospective audit of all cardiac surgical deaths not preceded by cardiac arrest calls. RESULTS: One-hundred-and-seventy-four patients (2.7%) had audited cardiac arrests of whom 70 (40%) survived to discharge. Elderly patients (> or = 70 yr old) had higher incidence of cardiac arrest (3.8% vs. 2%, P < 0.001). Survival to discharge following cardiopulmonary resuscitation was lower in the elderly patients, 33% vs. 48%, the difference approaching statistical significance (P = 0.06). Cardiopulmonary resuscitation was withheld in 46% of elderly vs. 40% of younger deaths (P = 0.40) which represented 3.1% of elderly vs. 1.2% younger patients (P < 0.001). Similar proportions of elderly (62%) and younger (67%) patients had failure of > or = 3 organ systems on institution of the 'Do Not Resuscitate' order (P = 0.70). CONCLUSION: 'Do Not Resuscitate' orders appeared twice as frequently in elderly patients (> or = 70 yr). However, the proportions of deaths without cardiopulmonary resuscitation and the organ failure scores between age groups were similar suggesting that severity of illness was more important than age in determining resuscitation status.

Six-year prospective audit of chest reopening after cardiac arrest.

OBJECTIVE: To identify which patients benefit from chest reopening after cardiac arrest. SETTING: Cardio-thoracic hospital undertaking full range of adult cardio-thoracic surgery. METHODS: In-hospital arrests were prospectively audited over a 6-year period. Information was collected for every patient whose chest was reopened following cardiac arrest: location of arrest, type of arrest, specialty, time since surgery, time to chest reopening, location of chest opening, surgical findings on reopening, time to cardiopulmonary bypass (if used) and patient outcomes. EXCLUSIONS: Arrests in theatre and chest openings for reasons other than cardiac arrest. RESULTS: There were 818 confirmed in-hospital arrests following 'cardiac arrest calls'. Chest reopening was undertaken in 79 surgical patients. Overall survival to discharge was 20/79 (25%). Favourable determinants of outcome were: arrest on intensive care unit (ICU), arrest within 24 h of surgery and reopening within 10 min of arrest. Nineteen of 58 (33%) chest openings following arrests on the ICU survived to discharge compared to one of 21 (5%) patients whose initial arrest was outside the ICU (P=0.017). One of nine ward arrests scooped to ICU for chest reopening survived whereas all 12 patients reopened on the ward died. Fifteen of 40 patients (38%) reopened within 24 h surgery survived compared to five of 39 patients where reopening was undertaken more than 24 h after surgery (P=0.02). Fourteen of 29 (48%) patients opened within 10 min of arrest survived to discharge compared to six of 50 (12%) patients where time to reopening was more than 10 min (P=<0.001). Seven of 22 patients (32%) patients where emergency bypass was utilised survived to discharge. CONCLUSION: This study strongly confirms the benefit of chest reopening after cardiac arrest in the cardiac surgical ICU. Patients who arrest within 24 h of surgery and in whom reopening is instituted within 10 min are particularly likely to benefit. The value of chest reopening in arrests outside the ICU remains unresolved. All patients reopened on the ward died, suggesting that this practice should be discontinued. Early 'scoop and run' resulted in one solitary survivor though it should probably be restricted to patients who arrest within 72 h of surgery as surgically remediable problems are unlikely after this time.

POSSUM and Portsmouth POSSUM for predicting mortality. Physiological and Operative Severity Score for the enUmeration of Mortality and morbidity.

BACKGROUND: There is a need for an accurate measure of surgical outcomes so that hospitals and surgeons can be compared properly regardless of case mix. POSSUM (Physiological and Operative Severity Score for the enUmeration of Mortality and morbidity) uses a physiological score and an operative severity score to calculate risks of mortality and morbidity. In a previous small study it was found that Portsmouth POSSUM (P-POSSUM; a modification of the POSSUM system) provided a more accurate prediction of mortality. METHODS: Some 10000 general surgical interventions (excluding paediatric and day cases) were studied prospectively between August 1993 and November 1995. The POSSUM mortality equation was applied to the full 10000 surgical episodes. The 10000 patients were arranged in chronological order and the first 2500 were used as a training set to produce the modified P-POSSUM predictor equation. This was then applied prospectively to the remaining 7500 patients arranged chronologically in five groups of 1500. RESULTS: The original POSSUM logistic regression equation for mortality overpredicts the overall risk of death by more than twofold and the risk of death for patients at lowest risk (5 per cent or less) by more than sevenfold. The P-POSSUM equation produced a very close fit with the observed in-hospital mortality. CONCLUSION: P-POSSUM provides an accurate method for comparative surgical audit.

Cloning, sequencing and functional expression of a guinea pig lung bradykinin B2 receptor.

Kinin receptors are classified as B1 and B2 based upon agonist and antagonist potencies and cloning and expression studies. Using sequences from human and rat bradykinin B2 receptors, polymerase chain reaction (PCR) was utilized to isolate cDNA from guinea pig lung. The receptor obtained is predicted to have 372 amino acids and shares > 80% sequence homology with human, rat, rabbit and mouse B2 receptors. In competition binding experiments in Chinese hamster ovary (CHO-K1) cells in which the guinea pig cDNA was expressed, [3H]bradykinin was displaced by kinin receptor ligands with an order of potency consistent with a B2 subtype. In CHO cells expressing the guinea pig receptor, bradykinin caused a concentration 45Ca2+ efflux. A B1 receptor agonist, desArg9-bradykinin, also caused 45Ca2+ efflux but with a potency several orders of magnitude lower than bradykinin. Curiously, several B1 and B2 receptor antagonists induced 45Ca2+ efflux, indicating that this receptor may be coupled differently in CHO cells than in native tissues.

Human bradykinin B2 receptor: nucleotide sequence analysis and assignment to chromosome 14.

Functional cDNA clones for human bradykinin B2 receptor were isolated from uterus RNA by a polymerase chain reaction (PCR)-based method and by screening a human cosmid library with rat bradykinin B2 receptor probe. We isolated several overlapping clones from the cosmid library, each of which encodes the entire protein coding sequence. The human bradykinin B2 receptor gene codes for a 364-amino-acid protein with a molecular mass of 41,442 Da that is highly homologous to rat bradykinin B2 receptor cDNA (81%). The entire human cDNA sequence was cloned into an expression vector and mRNA was synthesised by in vitro transcription. Applications of bradykinin caused membrane current responses in Xenopus oocytes injected with the in vitro-synthesized mRNA. Preincubation with the potent B2 antagonist, HOE140, prevented this response. The genomic clone is intronless, and we have identified an upstream promoter region and a downstream polyadenylation signal. The human bradykinin B2 receptor gene has been mapped to chromosome 14 using PCR to specifically amplify DNA from somatic cell hybrids.

Inhibition of biofilm populations of Nitrosomonas europaea.

The influence of surface attachment and growth on inhibition of the ammonia oxidizing bacterium, Nitrosomonas europaea, by nitrapyrin was investigated in liquid culture in the presence and absence of glass slides. Significant attachment to glass slides occurred in the absence of ammonia, but the extent of attachment was not affected by nitrapyrin, nor by previous culture of cells in medium containing nitrapyrin. The presence of glass slides affected neither the specific growth rate of N. europaea, measured by changes in nitrite concentration, nor inhibition by nitrapyrin. Inhibitory effects of nitrapyrin on increases in nitrite concentration and in free cell concentration were similar, but greater effects were observed on changes in attached cell concentration. Established biofilms on glass slides grew at a lower specific growth rate than freely suspended cells. Both biofilm cells, and those detached from the biofilm, were protected from inhibition. A mechanism for protection of biofilm populations is proposed involving reduced sensitivity of slowly growing cells producing extracellular polymeric material.

Human NK-2 receptor gene maps to chromosome region 10q11-21.

The human NK-2 receptor gene has been mapped to chromosome 10 using the polymerase chain reaction to amplify specifically the human NK-2 receptor sequence in hamster/human hybrid DNA and also in mouse/human monochromosome hybrids. The assignment to chromosome 10 was confirmed by in situ hybridisation to human metaphase chromosomes, giving a regional localisation of 10q11-21.

Isolation and characterisation of the human lung NK-1 receptor cDNA.

Functional cDNA clones for human NK-1 receptor were isolated from human lung RNA using the polymerase chain reaction (PCR). We have screened a human cosmid library and isolated a clone which appeared to contain the entire NK-1 receptor gene. From the published rat NK-1 receptor cDNA sequence we designed primers within the protein coding sequence, but outwards towards both the 5' and 3' ends of the putative human protein sequence. By this method we derived DNA sequence from the 3' end of the human gene. In order to determine the 5' end of the gene we used a PCR based method called Rapid Amplification of cDNA Ends (RACE). From the derived human sequences amplimers were designed upstream of the ATG initiation codon and downstream of the stop codon. The entire cDNA was obtained by RNA-PCR from human lung RNA. The sequence obtained was 407 amino acids in length, encoding an open-reading frame that was highly homologous to the rat NK-1 receptor cDNA (89%). The entire human cDNA was then cloned into a mammalian expression vector and mRNA was synthesized by in vitro transcription. Applications of tachykinins caused membrane current responses in Xenopus oocytes injected with the in vitro synthesized mRNA. The most potent of the three tachykinin peptides tested was Substance P. The human NK-1 receptor gene has been mapped to chromosome 2 using the polymerase chain reaction to specifically amplify the human sequence in hamster/human hybrid DNA and also in mouse/human monochromosome hybrids.

Protection of Nitrosomonas europaea colonizing clay minerals from inhibition by nitrapyrin.

Nitrate production by Nitrosomonas europaea in inorganic liquid medium containing ammonium was limited by reduction in pH. In the presence of montmorillonite and vermiculite, expanding clays with high cation-exchange-capacity (CEC), nitrite yield was increased, ammonia oxidation continued at pH values below those which inhibited growth in the absence of clays and growth was biphasic. The first phase was similar to that in the absence of clays, while the second was characterized by a lower rate of nitrite production. Illite, a non-expanding clay with low CEC, had no significant effect on ammonia oxidation, while oxidation of ammonia-treated vermiculite (ATV) occurred with no significant change in the pH of the medium. ATV, montmorillonite and vermiculite, but not illite, protected cells from inhibition by nitrapyrin at concentrations inhibitory to cells growing in suspended culture. This protection was maintained in ATV homo-ionic to Al3+, but montmorillonite made homo-ionic to Al3+ did not provide protection from inhibition. Attachment of cells to clays with high CEC is therefore advantageous in providing exchange at the clay surface of NH+4 and H+ produced by ammonia oxidation, in reducing pH toxicity, and in protecting cells from inhibition.

Isolation and characterisation of the human lung NK-2 receptor gene using rapid amplification of cDNA ends.

Functional cDNA clones for human NK-2 receptor were isolated from human lung RNA using a polymerase chain reaction (PCR) based method (RACE-PCR). In this method the cDNA was isolated as 5' end and 3'-end fragments; the entire cDNA was obtained by RNA-PCR. The sequence derived was 398 amino acids in length encoding an open-reading frame that was highly homologous to both the bovine and rat NK-2 receptor. The entire human cDNA sequence was cloned into a mammalian expression vector and mRNA was synthesised by in vitro transcription. Applications of tachykinins caused membrane current responses in Xenopus oocytes injected with the in vitro synthesised mRNA. The most potent of the three tachykinin peptides tested was neurokinin A. We have screened a human cosmid library and isolated a clone which contains the entire NK-2 receptor gene. The gene contains five exons and we have determined the complete sequence of the exons and the intron-exon junctions.

A yeast artificial chromosome contig encompassing the cystic fibrosis locus.

The gene responsible for cystic fibrosis (CF) has recently been identified. Coding sequence for the cystic fibrosis transmembrane conductance regulator (CFTR) spans at least 230 kb of the human genome. Although all 27 exons of the gene are represented in cosmid or bacteriophage clones, there are still several gaps in the physical map of this region. It should be possible to complete the map and to clone the entire CFTR gene in a single fragment of DNA using a yeast artificial chromosome (YAC) vector. Herein we describe the construction and physical mapping of a 1.5-Mb YAC contig which encompasses D7S8 (J3.11) and D7S23 (KM19), two genetic loci flanking the CF locus. One of the clones in the contig, 37AB12, contains a 310-kb YAC which includes the entire CFTR gene and flanking sequence in both the 5' and 3' directions.

The epsilon-subunit of ATP synthase from bovine heart mitochondria. Complementary DNA sequence, expression in bovine tissues and evidence of homologous sequences in man and rat.

The epsilon-subunit of ATP synthase from bovine heart mitochondria is assembled into the extrinsic membrane sector, F1-ATPase. The mature protein is 50 amino acid residues in length and its function is unknown. It is a nuclear gene product that is imported into the organelle. A mixture of 64 oligonucleotides 17 bases long, designed on the basis of the known protein sequence, was synthesized and used as a hybridization probe to isolate a cognate cDNA clone from a bovine library. The DNA sequence of this clone was determined, and the protein sequence of the epsilon-subunit deduced from it agrees exactly with that determined by direct sequence analysis of the protein isolated from bovine hearts. The bovine cDNA was used as a hybridization probe to examine the expression of the epsilon-subunit in various bovine tissues. mRNAs related to the cDNA are found in all of these tissues, and no evidence was obtained of the presence of mRNAs for the epsilon-subunit with similar coding sequences and dissimilar 3' non-coding regions. By hybridization experiments with digests of DNA from cow, man and rat it has been shown that sequences related to the bovine cDNA are present in the genomes of all three species. More than one related sequence was detected in all cases, indicating the presence in all three genomes of more than one gene and/or pseudogenes.

Molecular characterisation of three alpha-1-antitrypsin deficiency variants: proteinase inhibitor (Pi) nullcardiff (Asp256----Val); PiMmalton (Phe51----deletion) and PiI (Arg39----Cys).

Three mutations causing alpha-1-antitrypsin deficiency have been identified by gene amplification and direct DNA sequencing. In the Pi (proteinase-inhibitor) nullcardiff gene, the codon for aspartate at position 256 has mutated to encode valine. In PiMmalton and Pi I, the respective mutations are the deletion of the codon for a phenylalanine residue at position 51 or 52, and a single base substitution resulting in arginine being replaced by cysteine at position 39. Examination of the protein tertiary structure suggests that all of these mutations are likely to result in folding abnormalities that may explain the deficiency states.