COVID-19 Induces Neuroinflammation and Suppresses Peroxisomes in the Brain.

OBJECTIVE: Peroxisome injury occurs in the central nervous system (CNS) during multiple virus infections that result in neurological disabilities. We investigated host neuroimmune responses and peroxisome biogenesis factors during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection using a multiplatform strategy. METHODS: Brain tissues from coronavirus disease 2019 (COVID-19) (n = 12) and other disease control (ODC) (n = 12) patients, as well as primary human neural cells and Syrian hamsters, infected with a clinical variant of SARS-CoV-2, were investigated by droplet digital polymerase chain reaction (ddPCR), quantitative reverse transcriptase PCR (RT-qPCR), and immunodetection methods. RESULTS: SARS-CoV-2 RNA was detected in the CNS of 4 patients with COVID-19 with viral protein (NSP3 and spike) immunodetection in the brainstem. Olfactory bulb, brainstem, and cerebrum from patients with COVID-19 showed induction of pro-inflammatory transcripts (IL8, IL18, CXCL10, NOD2) and cytokines (GM-CSF and IL-18) compared to CNS tissues from ODC patients (p < 0.05). Peroxisome biogenesis factor transcripts (PEX3, PEX5L, PEX11ß, and PEX14) and proteins (PEX3, PEX14, PMP70) were suppressed in the CNS of COVID-19 compared to ODC patients (p < 0.05). SARS-CoV-2 infection of hamsters revealed viral RNA detection in the olfactory bulb at days 4 and 7 post-infection while inflammatory gene expression was upregulated in the cerebrum of infected animals by day 14 post-infection (p < 0.05). Pex3 transcript levels together with catalase and PMP70 immunoreactivity were suppressed in the cerebrum of SARS-CoV-2 infected animals (p < 0.05). INTERPRETATION: COVID-19 induced sustained neuroinflammatory responses with peroxisome biogenesis factor suppression despite limited brainstem SARS-CoV-2 neurotropism in humans. These observations offer insights into developing biomarkers and therapies, while also implicating persistent peroxisome dysfunction as a contributor to the neurological post-acute sequelae of COVID-19. ANN NEUROL 2023;94:531-546.

Isotopic tracing of the impact of mobility on infectious disease: The origin of people with treponematosis buried in hull, England, in the late medieval period.

Treponematosis has been one of the most studied and debated infectious diseases in paleopathology, particularly from the standpoint of its origin, evolution, and transmission. This study links evidence for treponematosis in skeletons from the 14th-16th century AD cemetery of the Augustinian friary of Hull Magistrates Court, England, with data from stable isotope analysis to test the hypothesis that the people with treponemal disease buried at this site were not locally born and raised. The objective is to explore the potential of using stable isotope data to track the place of origin and extent of mobility of individuals with an infectious disease. Dental enamel samples of 12 skeletons were selected for strontium ((87) Sr/(86) Sr ratio) and oxygen (δ(18) O) stable isotope analysis based on the presence (six - diseased) or absence (six - controls) of bone changes associated with treponemal disease. The oxygen isotope ratios of all but three individuals (1047, 1121, 823) overlapped at two standard deviations with the inferred local precipitation range, and only one individual (1216) had a strontium isotope ratio outside the regional range. Two of the four had probable/possible treponemal bone changes. Those with treponemal bone changes were not demonstrably more likely to be migrants than those without such lesions. However, because of extensive documentary evidence for trade with the Baltic Sea area, and for merchants from towns such as Stralsund, Danzig and Elbing being in Hull, it is very plausible that the four migrants came from the Baltic area or even southern Sweden.

Viral cultures, cycle threshold values and viral load estimation for assessing SARS-CoV-2 infectiousness in haematopoietic stem cell and solid organ transplant patients: a systematic review.

BACKGROUND: Solid organ and haematopoietic stem cell transplant recipients are more vulnerable to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) than non-transplant recipients due to immunosuppression, and may pose a continued transmission risk, especially within hospital settings. Detailed case reports including symptoms, viral load and infectiousness, defined by the presence of replication-competent viruses in culture, provide an opportunity to examine the relationship between clinical course, burden and contagiousness, and provide guidance on release from isolation. OBJECTIVES: To investigate the relationship between serial SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) cycle threshold (Ct) value or cycle of quantification value, or other measures of viral burden and the likelihood and duration of the presence of infectious virus based on viral culture, including the influence of age, sex, underlying pathologies, degree of immunosuppression, and/or vaccination on this relationship, in transplant recipients. METHODS: LitCovid, medRxiv, Google Scholar and the World Health Organization COVID-19 database were searched from 1st November 2019 to 26th October 2022. Studies reporting relevant data (results from serial RT-PCR testing and viral culture data from the same respiratory samples) for transplant recipients with SARS-CoV-2 infection were included in this systematic review: Methodological quality was assessed using five criteria, and the data were synthesized narratively and graphically. RESULTS: Thirteen case reports and case series reporting on 41 transplant recipients (22 renal, five cardiac, one bone marrow, two liver, one bilateral lung and 10 blood stem cell) were included in this review. A relationship was observed between proxies of viral burden and likelihood of shedding replication-competent SARS-CoV-2. Three individuals shed replication-competent viruses for >100 days after symptom onset. Lack of standardization of testing and reporting platforms precludes establishing a definitive viral burden cut-off. However, the majority of transplant recipients stopped shedding replication-competent viruses when the Ct value was >30 despite differences across platforms. CONCLUSIONS: Viral burden is a reasonable proxy for infectivity when considered within the context of the clinical status of each patient. Standardized study design and reporting are essential to standardize guidance based on an increasing evidence base.

Guideline for Technical Quality Assurance (TQA) of ultrasound devices (B-Mode)--version 1.0 (July 2012): EFSUMB Technical Quality Assurance Group--US-TQA/B.

The Technical Quality Assurance group was initiated by the EFSUMB Board in 2007 and met firstly in 2008 to discuss and evaluate methods and procedures published for performing technical quality assurance for diagnostic ultrasound devices. It is the aim of this group of experts to advise the EFSUMB Board of effective and efficacious methods for routine use and to make recommendations regarding the technical aspects of EFSUMB by-law 9, parts 11.6. & 11.7. The group's work focused on new developments and related European projects to establish a common guideline. There is a great need of a well established protocol and dedicated processing software for the performance testing of medical ultrasound equipment. The measurements should be user independent as much as physically possible. Only if these goals are achieved in an international (firstly European) context, the optimal quality of ultrasound imaging can be offered and maintained to the medical community. This guideline aims to offer and summarize suitable procedures and evaluation processes to lend support for an optimal Technical Quality Assurance (TQA) scheme. The content of this guideline was presented to the EFSUMB Board of Directors (delegates) and approved by the EFSUMB Executive Board (ExB) at the regular meeting during EUROSON 2012 in Madrid April 2012.

Viral cultures for assessing fomite transmission of SARS-CoV-2: a systematic review and meta-analysis.

BACKGROUND: The role of fomites in the transmission of SARS-CoV-2 is unclear. AIM: To assess whether SARS-CoV-2 can be transmitted through fomites, using evidence from viral culture studies. METHODS: Searches were conducted in the World Health Organization COVID-19 Database, PubMed, LitCovid, medRxiv, and Google Scholar to December 31st, 2021. Studies that investigated fomite transmission and performed viral culture to assess the cytopathic effect (CPE) of positive fomite samples and confirmation of SARS-CoV-2 as the cause of the CPE were included. The risk of bias using a checklist modified from the modified Quality Assessment of Diagnostic Accuracy Studies - 2 (QUADAS-2) criteria was assessed. FINDINGS: Twenty-three studies were included. The overall risk of bias was moderate. Five studies demonstrated replication-competent virus from fomite cultures and three used genome sequencing to match fomite samples with human clinical specimens. The mean cycle threshold (CT) of samples with positive viral culture was significantly lower compared with cultured samples that returned negative results (standardized mean difference: -1.45; 95% confidence interval (CI): -2.00 to -0.90; I2 = 0%; P < 0.00001). The likelihood of isolating replication-competent virus was significantly greater when CT was <30 (relative risk: 3.10; 95% CI: 1.32 to 7.31; I2 = 71%; P = 0.01). Infectious specimens were mostly detected within seven days of symptom onset. One study showed possible transmission of SARS-CoV-2 from fomites to humans. CONCLUSION: The evidence from published studies suggests that replication-competent SARS-CoV-2 is present on fomites. Replication-competent SARS-CoV-2 is significantly more likely when the PCR CT for clinical specimens and fomite samples is <30. Further studies should investigate the duration of infectiousness of SARS-CoV-2 and the frequency of transmission from fomites.

Vaccinia virus particles mix inefficiently, and in a way that would restrict viral recombination, in coinfected cells.

It is well established that poxviruses are subjected to genetic recombination, but attempts to map vaccinia virus genes using classical genetic crosses were historically confounded by high levels of experimental noise and a poor correlation between physical and genetic map distances. These virus-by-virus crosses also never produced the 50% recombinant progeny that should be seen in experiments involving distant markers. Poxviruses replicate in membrane-wrapped cytoplasmic structures called virosomes (or factories) and we have developed a method for tracking the development of these structures using live cell imaging and cells expressing phage lambda Cro protein fused to enhanced green fluorescent protein (EGFP). The EGFP-cro protein binds nonspecifically to DNA and permits live cell imaging of developing vaccinia virus factories. Using this method, we see virosomes first appearing about 4 to 5 h postinfection. The early virosomes exhibit a compact appearance and then, after a period of exponential growth lasting several hours, blur and start to dissipate in a process presumably linked to viral packaging. During the growth period, the virosomes migrate toward the nuclear periphery while colliding and fusing at a rate dependent upon the numbers of infecting particles. However, even at high multiplicities of infection (10 PFU/cell), we estimate approximately 20% of the virosomes never fuse. We have also used fluorescence in situ hybridization (FISH) methods to study virosomes formed by the fusion of viruses carrying different gene markers. FISH showed that DNA mixes rather poorly within fused virosomes and the amount of mixing is inversely dependent on the time between virosome appearance and fusion. Our studies suggest that the intracellular movement and mixing of virosomes create constraints that reduce opportunities for forming recombinants and that these phenomena create outcomes reflected in classical poxvirus genetics.

Colour flow and motion imaging.

Colour flow imaging (CFI) is an ultrasound imaging technique whereby colour-coded maps of tissue velocity are superimposed on grey-scale pulse-echo images of tissue anatomy. The most widespread use of the method is to image the movement of blood through arteries and veins, but it may also be used to image the motion of solid tissue. The production of velocity information is technically more demanding than the production of the anatomical information, partly because the target of interest is often blood, which backscatters significantly less power than solid tissues, and partly because several transmit-receive cycles are necessary for each velocity estimate. This review first describes the various components of basic CFI systems necessary to generate the velocity information and to combine it with anatomical information. It then describes a number of variations on the basic autocorrelation technique, including cross-correlation-based techniques, power Doppler, Doppler tissue imaging, and three-dimensional (3D) Doppler imaging. Finally, a number of limitations of current techniques and some potential solutions are reviewed.

Identifying the high-risk patient with clinically relevant embolisation after carotid endarterectomy.

OBJECTIVES: Sustained embolisation after carotid endarterectomy (CEA) predicts an increased risk of stroke due to post-operative carotid thrombosis (POCT). Progression towards stroke can be prevented by transcranial Doppler (TCD) directed intravenous Dextran therapy. However, TCD monitoring is extremely labour intensive. The aim of this study was to see whether a small cohort of high-risk patients could be identified following a 30-min period of monitoring in the Recovery Area of the operating theatre so as to reduce the overall burden of monitoring for the majority of patients. METHODS: Retrospective audit of prospectively acquired data in 821 patients with an accessible temporal window who had undergone 3h of TCD monitoring after CEA. Patients with >25 emboli in any 10 min period or large emboli distorting the waveform received Dextran. RESULTS: Group 1: 694 patients (85%) with

Immunolocalization of Na+/K+-ATPase and Na+/K+/2Cl- cotransporter in the tubular epithelia of sea snake salt glands.

The sublingual salt gland is the primary site of salt excretion in sea snakes; however, little is known about the mechanisms mediating ion excretion. Na(+)/K(+)-ATPase (NKA) and Na(+)/K(+)/2Cl(-) cotransporter (NKCC) are two proteins known to regulate membrane potential and drive salt secretion in most vertebrate secretory cells. We hypothesized that NKA and NKCC would localize to the basolateral membranes of the principal cells comprising the tubular epithelia of sea snake salt glands. Although there is evidence of NKA activity in salt glands from several species of sea snake, the localization of NKA and NKCC and other potential ion transporters remains unstudied. Using histology and immunohistochemistry, we localized NKA and NKCC in salt glands from three species of laticaudine sea snake: Laticauda semifasciata, L. laticaudata, and L. colubrina. Antibody specificity was confirmed using Western blots. The compound tubular glands of all three species were found to be composed of serous secretory epithelia, and NKA and NKCC were abundant in the basolateral membranes. These results are consistent with the morphology of secretory epithelia found in the rectal salt glands of marine elasmobranchs, the nasal glands of marine birds and the gills of teleost fishes, suggesting a similar function in regulating ion secretion.

The Leicester Cough Monitor: preliminary validation of an automated cough detection system in chronic cough.

Chronic cough is a common condition that presents to both primary and secondary care. Assessment and management are hampered by the absence of well-validated outcome measures. The present study comprises the validation of the Leicester Cough Monitor (LCM), an automated sound-based ambulatory cough monitor. Cough frequency was measured with the LCM and compared with coughs and other sounds counted manually over 2 h of a 6-h recording by two observers in nine patients with chronic cough in order to determine the sensitivity and specificity of the LCM. Automated cough frequency was also compared with manual counts from one observer in 15 patients with chronic cough and eight healthy subjects. All subjects underwent 6-h recordings. A subgroup consisting of six control and five patients with stable chronic cough underwent repeat automated measurements > or = 3 months apart. A further 50 patients with chronic cough underwent 24-h automated cough monitoring. The LCM had a sensitivity and specificity of 91 and 99%, respectively, for detecting cough and a false-positive rate of 2.5 events x h(-1). Mean+/-SEM automated cough counts x patient x h(-1) was 48+/-9 in patients with chronic cough and 2+/-1 in the control group (mean difference 46 counts x patient x h(-1); 95% confidence interval (CI) 20-71). The automated cough counts were repeatable (intra-subject SD 11.4 coughs x patient x h(-1); intra-class correlation coefficient 0.9). The cough frequency in patients undergoing 24-h automated monitoring was 19 coughs x patient x h(-1); daytime (08:00-22:00 h) cough frequency was significantly greater than overnight cough frequency (25 versus 10 coughs x patient x h(-1); mean difference 15 coughs x patient x h(-1), 95% CI 8-22). The Leicester Cough Monitor is a valid and reliable tool that can be used to assess 24-h cough frequency in patients with cough. It should be a useful tool to assess patients with cough in clinical trials and longitudinal studies.

Objective selection of signals for assessment of cerebral blood flow autoregulation in neonates.

A number of different system identification techniques have been proposed to assess dynamic cerebral autoregulation in critically ill patients. From these methods, the response to a standard stepwise change in blood pressure can be estimated. Responses lacking physiological consistency are a common occurrence and could be the consequence of particular system identification procedures or, alternatively, caused by measurements with a poor signal-to-noise ratio. A multi-observer approach was adopted in this paper to classify cerebral blood flow velocity (CBFV) step responses to spontaneous changes in arterial blood pressure in a group of 43 neonates with a mean gestational age of 33.7 weeks (range 24-42 weeks) and a mean birthweight of 1,980 g (range 570-3,910 g). Three experienced observers independently analysed the estimated step responses in 191 recordings each lasting 100 s; for an autoregressive (ARX) model, 124 (65%) of the step responses were accepted by at least two of the three observers. Two other system identification methods, transfer function analysis and the moving average Wiener-Laguerre model, gave 90 (45%) and 98 (51%) acceptable responses, respectively. Only 54 epochs (28%) were accepted with all three methods. With 88 (46%) responses rejected by at least two methods, it can be concluded that signal quality was the main reason for nonphysiological step responses. To avoid the need for subjective visual selection, an automatic procedure for classifying step responses was implemented leading to sensitivities and specificities in the range 85-90%, with respect to the agreement with subjective evaluations. Objective selection of CBFV step responses is thus feasible and could also be adapted for other physiological measurement techniques relying on system identification methods.

Cerebral critical closing pressure estimation from Finapres and arterial blood pressure measurements in the aorta.

Estimates of cerebral critical closing pressure (CrCP) and resistance-area product (RAP) are often derived using noninvasive measurements of arterial blood pressure (ABP) in the finger, but the errors introduced by this approach, in relation to intra-vascular measurements of ABP, are not known. Continuous recordings of ABP (Finapres and solid-state catheter-tip transducer in the ascending aorta), cerebral blood flow velocity (CBFV, bilateral Doppler), ECG and transcutaneous CO(2) were performed following coronary catheterization. CrCP and RAP were calculated for each of 12,784 cardiac cycles from 27 subjects using the classical linear regression (LR) of the instantaneous CBFV-ABP relationship and also the first harmonic (H(1)) of the Fourier transform. There was a better agreement between LR and H(1) for the aortic measurements than for the Finapres (p < 0.000,01). For LR there were no significant differences for either CrCP or RAP due to the source of ABP measurement, but for H(1) the differences were highly significant (p < 0.000,03). The coherence functions between either CrCP or RAP values calculated with aortic pressure (input) or the Finapres (output) were significantly higher for H(1) than for LR for most harmonics below 0.2 Hz. When using the Finapres to estimate CrCP and RAP values, the LR method produces similar results to intra-arterial measurements of ABP for time-averaged values, but H(1) should be preferred in applications analysing beat-to-beat changes in these parameters.

Bioactivation of aromatic amines by recombinant human cytochrome P4501A2 expressed in Ames tester strain bacteria: a substitute for activation by mammalian tissue preparations.

The most widely used bioassay in genetic toxicology is the Ames test, which combines a bacterial mutagenicity assay (reversion of Salmonella typhimurium histidine-auxotrophic tester strains) with an exogenous bioactivation system (hepatic postmitochondrial supernatant or "S9"). The enzymatic activities of S9 prepared from the tissues of experimental animals are difficult to control. We show that the requirement for S9 can be obviated by the engineered expression of enzymes of bioactivation within the bacterial cell. With this strategy, reactive metabolites are produced inside the bacterial cell, proximate to the genetic target. Species boundaries can be crossed, and chimeric or mutant enzymes can be studied. We have constructed an Ames tester strain, expressing both aromatic amine N-acetyltransferase and human cytochrome P4501A2, which detects aromatic amine mutagenicity in the absence of S9.

Effect of DNA structure and nucleotide sequence on Holliday junction resolution by a Saccharomyces cerevisiae endonuclease.

Previous studies have demonstrated that mitotic Saccharomyces cerevisiae cells contain an endonuclease that cleaves Holliday junctions. In this paper, the cleavage of a number of model branched substrates has been characterized in detail. Three-armed Y-branched molecules were not substrates for the enzyme. Holliday junction substrates constructed from wild-type lambda att sites were resolved in a concerted reaction by paired single-strand breaks that contained 5'-phosphate and 3'-hydroxyl groups and were often symmetrically related. Holliday junctions were also constructed using DNAs derived from lambda safG and safT mutants to alter the nucleotide sequence immediately flanking the cross-strand exchange. These one to six base-pair changes in nucleotide sequence were observed to have dramatic effects on both the directionality and rate of resolution. More than 90% of wild-type junctions were cleaved in only one direction, while Holliday junctions composed of safT DNA were cleaved equally in both possible directions. Hybrid junctions composed of half wild-type DNA and half safG DNA were cleaved in the same orientation as the wild-type junction but at one-seventh of the rate, while junctions constructed completely from safG DNA were not cleaved at all. The cleavage sites were mapped at the nucleotide level and the locations of the paired nicks made by the endonuclease were also found to be affected by the sequence of the substrates and in such a way as to account for the directionality of cleavage. These results have important consequences for the interpretation of genetic experiments, since they provide biochemical evidence that some of the non-random nature of genetic recombination might be due to non-randomly distributed resolution processes.

Shope fibroma virus DNA topoisomerase catalyses holliday junction resolution and hairpin formation in vitro.

The telomeres of poxviral chromosomes comprise covalently closed hairpin structures bearing mismatched bases. These hairpins are formed as concatemeric replication intermediates and are processed into mature, unit-length genomes. The structural transitions and enzymes involved in telomere resolution are poorly understood. Here we show that the type I topoisomerase of Shope fibroma virus (SFV) can promote a recombination reaction which converts cloned SFV replication intermediates into hairpin-ended molecules resembling mature poxviral telomeres. Recombinant SFV topoisomerase linearised a palindromic plasmid bearing 1.5 kb of DNA encoding the SFV concatemer junction, at a site near the centre of inverted-repeat symmetry. Most of these linear reaction products bore hairpin tips as judged by denaturing gel electrophoresis. The resolution reaction required palindromic SFV DNA sequences and was inhibited by compounds which block branch migration (MgCl2) or poxviral topoisomerases. The resolution reaction was also slow, needed substantial quantities of topoisomerase, and required that the palindrome be extruded in a cruciform configuration. DNA cleavage experiments identified a pair of suitably oriented topoisomerase recognition sites, 90 bases from the centre of the cloned SFV terminal inverted repeat, which may mark the resolution site. These data suggest a resolution scheme in which branch migration of a Holliday junction through a site occupied by covalently bound topoisomerase molecules, could lead to telomere resolution.

Heteroduplex DNA formation is associated with replication and recombination in poxvirus-infected cells.

Poxviruses are large DNA viruses that replicate in the cytoplasm of infected cells and recombine at high frequencies. Calcium phosphate precipitates were used to cotransfect Shope fibroma virus-infected cells with different DNA substrates and the recombinant products assayed by genetic and biochemical methods. We have shown previously that bacteriophage lambda DNAs can be used as substrates in these experiments and recombinants assayed on Escherichia coli following DNA recovery and in vitro packaging. Using this assay it was observed that 2-3% of the phage recovered from crosses between point mutants retained heteroduplex at at least one of the mutant sites. The reliability of this genetic analysis was confirmed using DNA substrates that permitted the direct detection of heteroduplex molecules by denaturant gel electrophoresis and Southern blotting. It was further noted that heteroduplex formation coincided with the onset of both replication and recombination suggesting that poxviruses, like certain bacteriophage, make no clear biochemical distinction between these three processes. The fraction of heteroduplex molecules peaked about 12-hr postinfection then declined later in the infection. This decline was probably due to DNA replication rather than mismatch repair because, while high levels of induced DNA polymerase persisted beyond the time of maximal heteroduplex recovery, we were unable to detect any type of mismatch repair activity in cytoplasmic extracts. These results suggest that, although heteroduplex molecules are formed during the progress of poxviral infection, gene conversion through mismatch repair probably does not produce most of the recombinants. The significance of these observations are discussed considering some of the unique properties of poxviral biology.

A WD repeat protein, Rec14, essential for meiotic recombination in Schizosaccharomyces pombe.

Mutations in the Schizosaccharomyces pombe rec14 gene reduce meiotic recombination by as much as a factor of 1000 in the three intervals tested on chromosomes I and III. A DNA clone complementing the rec14 mutation was shown by genetic and physical analysis to contain the rec14 gene, which was functional in plasmid-borne inserts as small as 1.4 kb. The rec14 gene contains two exons separated by a 53-bp intron, which was confirmed by analysis of rec14 transcripts. The spliced transcript encodes a protein product of 302 amino acids, which contains six WD repeat motifs found in the G-beta transducin family of proteins and other proteins, including the Saccharomyces cerevisiae Ski8 (Rec103) protein. Although the rec14 transcripts were present in mitotically dividing cells, rec14 mutations had no detectable effect on mitotic recombination. The pattern of expression of rec14 differes from that of previously analyzed S. pombe rec genes. Based upon mutant phenotypes and amino acid sequence similarities, we propose that S. pombe Rec14 is a functional homologue of S. cerevisiae Rec103.

Ultrasonic measurement of prosthetic heart valve action.

An in-vitro study has been made of the echo patterns obtained during ultrasonic Time-Motion (T-M) scanning of commonly used prosthetic heart valves. Studies have been undertaken of Starr-Edwards, Beal, and Björk-Shilley valves. From the recorded traces information has been obtained regarding the orientations of the ultrasound beam to the central axes of the valves for which accurate measurements can be made. The orientations have been determined for which the opening and closing times of the valves can be measured to within 20 ms. Characteristics of the T-M trances have been noted which allow the angle between the direction of an ultrasound beam and a central valve axis to be reduced to less than 20 degrees. This makes the error due to angular misalignment of the beam less than --6% when the range of movement of a ball, disc, or flap is being measured. Finally a number of artefacts are considered which can arise when prosthetic valves are examined with ultrasound.

The accuracy of caridac function indices derived from ultrasonic time-position scans.

A study has been made of Time-Position ultrasonic scanning as a means of obtaining measurements related to cardiac performance. The errors involved in determining these measurements have been estimated. It has been shown that the greatest accuracy can be achieved for quantities which depend on action rather than shape, such as ejection fractions and rates of myocardial fibre shortening. Attention is drawn to the fact that the ultrasonic method, unlike angiography, in no way alters the quantities being measured.

Changes in Doppler ultrasound sonagrams at varying distances from stenoses.

The effect of different degrees of arterial stenosis on Doppler ultrasound sonagrams recorded from various distances downstream has been investigated in a canine model. It has been found that even mild stenoses give rise to considerable disturbances close to the stenosis, and that more severe stenoses give rise to greater disturbances which propagate further downstream. The distance the disturbances travelled was not related to stroke volume, and it was only for stenoses of 88% area reduction and greater that such disturbances ever propagated beyond one stroke-length. Quantitative assessment of moderate degrees of proximal stenosis using continuous wave Doppler ultrasound will not be reliable unless the distance between the major stenosis and the recording site is known.