The hidden risks of hearing tests and programmable ventriculoperitoneal shunt valves.

Programmable variable pressure valves were introduced in the 1980s, providing a non-invasive solution to post-operative alterations of the valve opening pressure to address problems of under or overdrainage. Since their increased use in the treatment of hydrocephalus, there have been case reports of unintentional alterations of the valve opening pressure following exposure to magnetic fields in everyday environments, from televisions to rollercoasters. Here we describe two cases of patients' programmable valves being altered following audiology assessments. We subsequently discuss some of the available research on the safety of these valves in various magnetic field strengths, alongside interim and updated recommendations made by the British Society of Audiologists with regards to testing of children with programmable shunts in place.

Review of patient experience with bilateral sagittal split osteotomies as a day case procedure.

Historically, patients who received bilateral sagittal split osteotomies (BSSO) required an inpatient admission for at least one night. Since March 2015, the Oral and Maxillofacial Department at the Royal Gwent Hospital has performed bilateral sagittal split osteotomies (BSSO) as a day case procedure for their medically and socially fit patients. Our team's service evaluation by Davies et al (2018) for this procedure, demonstrated that this could be done both routinely and successfully, whilst conforming to national day case procedural standards. The aim of this satisfaction survey was to evaluate this procedure from a patient's perspective, to further consolidate our results from 2018. The forty-five patients who underwent day case BSSO (DCBSSO) between February 2015 and February 2020 were retrospectively identified and deemed eligible for inclusion. Participation involved completion of a 10-part questionnaire via telephone consultation. Patients were asked questions focussing on their experience of discharge timing, management of postoperative symptoms, and overall recovery at home. Twenty-four patients consented to partake in the survey (response rate of 73%). Twenty-three (96%) were extremely happy to be discharged the day of their surgery and felt that the timing of discharge was appropriate. Only 17% of patients experienced discomfort overnight and 96% of these stated they could manage their symptoms at home. From this survey, we can confirm that the majority of patients receiving DCBSSO at the Royal Gwent Hospital were happy to be discharged the day of their surgery and recover at home.

Day-case bilateral sagittal split osteotomy.

In the UK, patients who have bilateral sagittal split osteotomy (BSSO) have generally been thought to require inpatient admission and an overnight hospital stay. However, since the introduction of national standards on day case surgery in the UK in 2011, patients at the Royal Gwent Hospital, Newport, have been treated as day cases, and have been pleased with the results. The aim of this paper was to show that these procedures conform to current national standards, and can be done successfully and safely. We retrospectively reviewed all patients who had isolated BSSO planned as day cases between March 2015 and February 2017. Thirty-four were eligible. Of them, 32 were discharged on the day of operation and two were admitted postoperatively: one because of severe nausea and vomiting and the other because of bleeding. No patients were readmitted within 48hours of the procedure. BSSO can be done successfully and routinely as a day-case procedure. However, to reduce the rate of unplanned admissions, we recommended that operations start early in the morning.

Toxicology study assessing efficacy and safety of repeated administration of lipid/DNA complexes to mouse lung.

For gene therapy to improve lung function in cystic fibrosis (CF) subjects, repeated administration of the gene transfer agent over the lifetime of patients is likely to be necessary. This requirement limits the utility of adenoviral and adeno-associated viral vectors (both previously evaluated in CF gene therapy trials) because of induced adaptive immune responses that render repeated dosing ineffective. For CF gene therapy trials, non-viral vectors are currently the only viable option. We previously showed that the cationic lipid formulation GL67A is the most efficient of several non-viral vectors analysed for airway gene transfer. Here, we assessed the efficacy and safety of administering 12 inhaled doses of GL67A complexed with pGM169, a CpG-free plasmid encoding human CFTR complementary DNA, into mice. We show that repeated administration of pGM169/GL67A to murine lungs is feasible, safe and achieves reproducible, dose-related and persistent gene expression (>140 days after each dose) using an aerosol generated by a clinically relevant nebuliser. This study supports progression into the first non-viral multidose lung trial in CF patients.

Pre-clinical evaluation of three non-viral gene transfer agents for cystic fibrosis after aerosol delivery to the ovine lung.

We use both large and small animal models in our pre-clinical evaluation of gene transfer agents (GTAs) for cystic fibrosis (CF) gene therapy. Here, we report the use of a large animal model to assess three non-viral GTAs: 25 kDa-branched polyethyleneimine (PEI), the cationic liposome (GL67A) and compacted DNA nanoparticle formulated with polyethylene glycol-substituted lysine 30-mer. GTAs complexed with plasmids expressing human cystic fibrosis transmembrane conductance regulator (CFTR) complementary DNA were administered to the sheep lung (n=8 per group) by aerosol. All GTAs gave evidence of gene transfer and expression 1 day after treatment. Vector-derived mRNA was expressed in lung tissues, including epithelial cell-enriched bronchial brushing samples, with median group values reaching 1-10% of endogenous CFTR mRNA levels. GL67A gave the highest levels of expression. Human CFTR protein was detected in small airway epithelial cells in some animals treated with GL67A (two out of eight) and PEI (one out of eight). Bronchoalveolar lavage neutrophilia, lung histology and elevated serum haptoglobin levels indicated that gene delivery was associated with mild local and systemic inflammation. Our conclusion was that GL67A was the best non-viral GTA currently available for aerosol delivery to the sheep lung, led to the selection of GL67A as our lead GTA for clinical trials in CF patients.

Long-term persistence of gene expression from adeno-associated virus serotype 5 in the mouse airways.

Recombinant adeno-associated virus vectors based on serotype 2 (rAAV2) have been used to deliver transgenes to the airways in a variety of pre-clinical and clinical studies. Gene transfer in these studies has been severely restricted by the basolateral localization of rAAV2 receptors. Here, we studied vectors constructed from the AAV5 genome and capsid, which utilize N-linked sialic acid-containing receptors found on the apical surface of airway epithelial cells. We investigated gene transfer efficacy and duration of transgene expression following delivery of rAAV5/5 vectors to the mouse respiratory tract. Robust, dose-dependent transgene expression was observed in the epithelium lining the nose for at least 32 weeks, and for at least 52 weeks in the lung. Importantly, in the lung, transgene expression mediated by rAAV5/5 was 40-fold greater than by rAAV2/2 vectors. A distinct cellular preference for rAAV5/5-mediated transduction was observed, with transgene expression being predominantly restricted to sustentacular cells of the olfactory epithelium in the nose and alveolar type II cells in the lung. Administration of rAAV5/5 vectors to both the nose and lungs led to the rapid development of rAAV5/5-neutralizing antibodies, suggesting that repeated administration may be severely hampered by host immune responses.

Optimising non-viral gene delivery in a tumour spheroid model.

BACKGROUND: Our current understanding of how the unique tumour microenvironment influences the efficacy of gene delivery is limited. The current investigation systematically examines the efficiency of several non-viral gene transfer agents to transfect multicellular tumour spheroids (MCTS), an in vitro model that displays a faithful three-dimensional (3D) representation of solid tumour tissue. METHODS: Using a luciferase reporter assay, gene transfer to MCTS was optimised for 22 kDa linear and 25 kDa branched polyethyleneimine (PEI), the cationic lipids Lipofectamine(trade mark) and DCChol : DOPE, and the physical approach of tissue electroporation. Confocal microscopy was used to take optical tissue slices to identify the tissue localisation of green fluorescent protein (GFP) reporter gene expression and the distribution of fluorescently labelled complexes. A MCTS model of quiescent tumour regions was used to establish the influence of cellular proliferation status on gene transfer efficiency. RESULTS: Of the polyplexes tested, 22 kDa linear PEI provided optimal gene delivery, with gene expression peaking at 46 h. Despite being the optimal vector tested, PEI-mediated transfection was limited to cells at the MCTS periphery. Using fluorescent PEI, it was found that complexes could only penetrate the outer 3-5 proliferating cell layers of the MCTS, sparing the deeper quiescent cells. Gene delivery in an MCTS model comprised entirely of quiescent cells demonstrated that in addition to being inaccessible to the vector, quiescent tumour regions are inherently less susceptible to PEI-mediated transfection than proliferating regions. This 'resistance' to transfection observed in quiescent cells was overcome through the use of electroporation. Despite the improved efficacy of electroporation in quiescent tissue, the gene expression was still confined to the outer regions of MCTS. The results suggest that limited access to central regions of an MCTS remain a significant barrier to gene delivery. CONCLUSIONS: This data provides new insights into tumour-specific factors affecting non-viral gene transfer and highlights the difficulties in delivering genes to avascular tumour regions. The MCTS model is a useful system for the initial screening of future gene therapy strategies for solid tumours.

The development of gene therapy for diseases of the lung.

The development of a successful gene therapy has many stages, including preclinical testing in animal models and proof of principle clinical studies. A variety of diseases affect the lung, which are candidates for gene therapy; this review will mainly focus on the diseases that have attracted the most attention and have therefore yielded the most progress, namely lung cancer and the monogenic disorder cystic fibrosis. Knowledge gained from clinical studies could eventually be applied to more complex lung conditions such as acute respiratory distress syndrome and asthma. In addition, increased gene transfer efficiencies could be obtained by appropriate selection of the gene transfer vector and mode of delivery.

Effects of isoflurane, sevoflurane, and halothane on myofilament Ca2+ sensitivity and sarcoplasmic reticulum Ca2+ release in rat ventricular myocytes.

BACKGROUND: The aim of this study was to describe and compare the effects of isoflurane, sevoflurane, and halothane at selected concentrations (i.e., concentrations that led to equivalent depression of the electrically evoked Ca2+ transient) on myofilament Ca2+ sensitivity, sarcoplasmic reticulum (SR) Ca2+ content, and the fraction of SR Ca2+ released during electrical stimulation (fractional release) in rat ventricular myocytes. METHODS: Single rat ventricular myocytes loaded with fura-2 were electrically stimulated at 1 Hz, and the Ca2+ transients and contractions were recorded optically. Cells were exposed to each anesthetic for 1 min. Changes in myofilament Ca2+ sensitivity were assessed by comparing the changes in the Ca2+ transient and contraction during exposure to anesthetic and low Ca2+. SR Ca2+ content was assessed by exposure to 20 mm caffeine. RESULTS: Isoflurane and halothane caused a depression of myofilament Ca2+ sensitivity, unlike sevoflurane, which had no effect on myofilament Ca2+ sensitivity. All three anesthetics decreased the electrically stimulated Ca2+ transient. SR Ca2+ content was reduced by both isoflurane and halothane but was unchanged by sevoflurane. Fractional release was reduced by both isoflurane and sevoflurane, but was unchanged by halothane. CONCLUSIONS: Depressed myofilament Ca2+ sensitivity contributes to the negative inotropic effects of isoflurane and halothane but not sevoflurane. The decrease in the Ca2+ transient is either responsible for or contributory to the negative inotropic effects of all three anesthetics and is either primarily the result of a decrease in fractional release (isoflurane and sevoflurane) or primarily the result of a decrease in SR Ca2+ content (halothane).

Effects of halothane on the transient outward K(+) current in rat ventricular myocytes.

1. Halothane has been shown to affect several membrane currents in cardiac tissue including the L-type calcium current (I(Ca)), sodium current and a variety of potassium currents. However, little is known about the effects of halothane on the transient outward K(+) current (I(to)). 2. Single ventricular myocytes from rat hearts were voltage clamped using the whole cell patch configuration and an EGTA-containing pipette solution to record the Ca(2+)-independent, 4-aminopyridine sensitive component of I(to). 300 microM Cd(2+) or 10 microM nifedipine was used to block I(Ca). 3. At +80 mV, I(to) (peak current minus current at the end of the pulse) was 1.8+/-0.2 nA under control conditions which was reduced to 1.3+/-0.2 nA by 1 mM halothane (P:<0.001, mean+/-s.e.mean, n=9). The inhibition of I(to) by halothane was concentration-dependent (K(0.5), 1.1+/-0.2 mM). 4. One mM halothane led to a 16 mV shift in the steady-state inactivation curve towards negative membrane potentials (P:=0.005, n=8) but had no significant effect on the activation-voltage relationship (P:=0. 724). One mM halothane also increased the rate of inactivation of I(to); the dominant time constant of inactivation was reduced from 14+/-1 to 9+/-1 ms (P:=0.017, mean+/-s.e.mean, n=6). 5. These data show that halothane reduced I(to); 0.3 mM, close to the MAC(50) value for halothane, inhibited the current by 15% and as such, the inhibition of I(to) will be relevant to the clinical situation. Halothane induced a shift in the steady-state inactivation curve and accelerated the inactivation process of I(to) which could be responsible for its inhibitory effect. 6. Due to the differential transmural expression of I(to) in ventricular tissue, inhibition of I(to) would reduce the transmural dispersion of refractoriness which could contribute to the arrhythmogenic properties of halothane.

Molecular dynamics simulation of four-alpha-helix bundles that bind the anesthetic halothane.

The mutation of a single leucine residue (L38) to methionine (M) is known experimentally to significantly increase the affinity of the synthetic four-alpha-helix bundle (Aalpha(2))(2) for the anesthetic halothane. We present a molecular dynamics study of the mutant (Aalpha(2)-L38M)(2) peptide, which consists of a dimer of 62-residue U-shaped di-alpha-helical monomers assembled in an anti topology. A comparison between the simulation results and those obtained for the native (Aalpha(2))(2) peptide indicates that the overall secondary structure of the bundle is not affected by the mutation, but that the side chains within the monomers are better packed in the mutant structure. Unlike the native peptide, binding of a single halothane molecule to the hydrophobic core of (Aalpha(2)-L38M)(2) deforms the helical nature of one monomer in a region close to the mutation site. Increased exposure of the cysteine side chain to the hydrophobic core in the mutant structure leads to the enhancement of the attractive interaction between halothane and this specific residue. Since the mutated residues are located outside the hydrophobic core the observed increased affinity for halothane appears to be an indirect effect of the mutation.

The concentration-dependent effects of propofol on rat ventricular myocytes.

UNLABELLED: Whether propofol contributes a direct negative inotropic effect is controversial. Our principal aim in this study was to determine whether negative inotropic effects of propofol occur at clinically relevant concentrations. We constructed the concentration-response relationship for the negative inotropic effects on intact, isolated, stimulated rat ventricular myocytes. Contraction was measured as cell shortening by using an optical system. Propofol was applied as dilutions of the commercial preparation in physiological saline solution. The drug vehicle had a minimal effect on myocyte contractility. Propofol produced a concentration-dependent reduction in evoked contraction at concentrations greater than 5 microM. The maximum effect was observed at >100 microM, with the K(0.5) calculated to be 34.5 microM (95% CI, 21.8-54.7 microM). In further experiments, we investigated the relationship between changes in contractility and changes in Ca(2+) transient (measured by using fura-2 fluorescence) after the application of propofol. By using the shift in the relationship of the cell length to fura-2 fluorescence ratio in the relaxation phase of a contraction as an index of Ca(2+) response of the myofilaments, we demonstrated that some of the negative inotropic effect of propofol may be caused by a reduction in myofilament Ca(2+) sensitivity. We confirmed this by comparing the reduction in contractility in the presence of propofol with that caused by reducing the extracellular Ca(2+) concentration. We observed that, for a decrease in the fura-2 fluorescence ratio of 21%, propofol caused a 12% (95% CI, 2% to 22%) greater reduction in contractility than predicted from reducing the extracellular Ca(2+) concentration. However, the K(0.5) for the negative inotropic effect of propofol we observed is more than 80 times the 50% effective concentration value for anesthesia. The potential relevance of these findings for clinical use of propofol in humans is discussed. IMPLICATIONS: By using intact, isolated rat heart ventricle cells, we investigated the mechanisms and concentration dependence of the depressant effect of propofol on contractility of the heart. We conclude that direct effects of propofol on the heart are unlikely to be of significance at the clinical dosage usually given.

A designed four-alpha-helix bundle that binds the volatile general anesthetic halothane with high affinity.

The structural features of volatile anesthetic binding sites on proteins are being examined with the use of a defined model system consisting of a four-alpha-helix bundle scaffold with a hydrophobic core. Previous work has suggested that introducing a cavity into the hydrophobic core improves anesthetic binding affinity. The more polarizable methionine side chain was substituted for a leucine, in an attempt to enhance the dispersion forces between the ligand and the protein. The resulting bundle variant has an improved affinity (K(d) = 0.20 +/- 0.01 mM) for halothane binding, compared with the leucine-containing bundle (K(d) = 0.69 +/- 0.06 mM). Photoaffinity labeling with (14)C-halothane reveals preferential labeling of the W15 residue in both peptides, supporting the view that fluorescence quenching by bound anesthetic reports both the binding energetics and the location of the ligand in the hydrophobic core. The rates of amide hydrogen exchange were similar for the two bundles, suggesting that differences in binding affinity were not due to changes in protein stability. Binding of halothane to both four-alpha-helix bundle proteins stabilized the native folded conformations. Molecular dynamics simulations of the bundles illustrate the existence of the hydrophobic core, containing both W15 residues. These results suggest that in addition to packing defects, enhanced dispersion forces may be important in providing higher affinity anesthetic binding sites. Alternatively, the effect of the methionine substitution on halothane binding energetics may reflect either improved access to the binding site or allosteric optimization of the dimensions of the binding pocket. Finally, preferential stabilization of folded protein conformations may represent a fundamental mechanism of inhaled anesthetic action.

Concentration-dependent inotropic effects of halothane, isoflurane and sevoflurane on rat ventricular myocytes.

We have described the concentration-dependent inotropic effects of halothane, isoflurane and sevoflurane on rat ventricular cells and investigated the role of the sarcoplasmic reticulum (SR) in these inotropic actions. Single ventricular myocytes, isolated from rat hearts, were stimulated electrically at 1 Hz and contractions recorded optically. Cells were exposed to a range of concentrations of halothane, isoflurane or sevoflurane for a period of 1 min to determine the concentration-dependency of their inotropic actions. For each anaesthetic, the peak negative inotropic action was determined early during an exposure, and sustained negative inotropic action was measured at steady-state just before wash-off. In some experiments, cells were equilibrated with ryanodine 1 mumol litre-1 to investigate the role of the SR in these intropic effects. Halothane caused a concentration-dependent initial increase in contractions (to mean 130 (SEM 28)% at 10 mmol litre-1) followed by rapid onset of a negative inotropic effect (K0.5 0.34 mmol litre-1 for peak effect; K0.5 0.46 mmol litre-1 for sustained effect). Exposure to isoflurane induced a small potentiation of contractions in some cells, followed by a concentration-dependent decrease in contraction in all cells (K0.5 0.85 mmol litre-1 for peak effect; K0.5 1.92 mmol litre-1 for sustained effect); contractions recovered partially during a 1-min exposure. On wash-off, contractions were increased transiently above control. Sevoflurane caused a large initial decrease in contraction which then returned rapidly towards control (K0.5 0.2 mmol litre-1 for peak effect; K0.5 2.57 mmol litre-1 for sustained effect). In common with isoflurane, removal of sevoflurane caused a transient increase in contractions above control. After exposure to ryanodine, the positive inotropic effects of halothane and isoflurane did not occur, and recovery of contractions during exposure to isoflurane and sevoflurane was abolished as was the transient increase in contractions seen on wash-off, indicating that these effects were mediated via the SR. Halothane had the most potent sustained negative inotropic effect but there was little difference between the negative inotropic effects of isoflurane and sevoflurane at clinically relevant concentrations. At higher concentrations, sevoflurane caused a less potent negative inotropic effect than isoflurane. The SR plays a major role in the effects of all three anaesthetics. One possible mechanism underlying the initial potentiation of contraction by halothane (and isoflurane) may be sensitization of the Ca(2+)-induced Ca(2+)-release process of the SR.

Mechanisms underlying the inotropic action of halothane on intact rat ventricular myocytes.

The mechanisms contributing to the negative inotropic effect of halothane were studied in isolated rate ventricular myocytes. Contraction and intracellular Ca2+ transients were measured optically in these cells. The initial application of halothane (2% or 0.5 mmol litre-1) led to short-lived increases in the Ca2+ transient and contraction, which were abolished by ryanodine. Continued application of halothane led to a sustained decrease in contraction: this resulted from: (i) a decrease in myofilament Ca2+ sensitivity; (ii) a decrease in the Ca2+ transient; and (iii) a decrease in the Ca2+ content of the sarcoplasmic reticulum. Although halothane reduced action potential duration, the sustained negative inotropic effect was similar when action potentials or voltage clamp pulses of constant duration were used to trigger contractions. In cells exposed to nifedipine 0.5 mumol litre-1 (which decreases the L-type Ca2+ current, ICa), Ca2+ transients, sarcoplasmic reticulum Ca2+ content and fractional release (the fraction of sarcoplasmic reticulum Ca2+ content released during each stimulus) were reduced. Halothane 0.5 mmol litre-1 (which also decreases ICa) decreased Ca2+ transients to a lesser extent and reduced sarcoplasmic reticulum Ca2+ content to a greater extent than nifedipine, whereas fractional release was unchanged compared with control. These data suggest that halothane sensitizes Ca(2+)-induced Ca2+ release from the sarcoplasmic reticulum in addition to reducing ICa.

Molecular dynamics simulation of a synthetic four-alpha-helix bundle that binds the anesthetic halothane.

The structural features of binding sites for volatile anesthetics are examined by performing a molecular dynamics simulation study of the synthetic four-alpha-helix bundles (Aalpha2)2, which are formed by association of two 62-residue di-alpha-helical peptides. The peptide bundle (Aalpha2)2 was designed by Johansson et al. [Biochemistry 37 (1998) 1421-1429] and was shown experimentally to have a high affinity for the binding of the anesthetic halothane (CF3CBrCIH) in a hydrophobic cavity. Since (Aalpha2)2 can exhibit either the anti or syn topologies, the two distinct bundles are simulated both in the presence and in the absence of halothane. Nanosecond length molecular dynamics trajectories were generated for each system at room temperature (T = 298 K). The structural and dynamic effects of the inclusion of halothane are compared, illustrating that the structures are stable over the course of the simulation; that the (Aalpha2)2 bundles have suitable pockets that can accommodate halothane; that the halothane remains in the designed hydrophobic cavity in close proximity to the Trp residues with a preferred orientation; and that the dimensions of the peptide are perturbed by the inclusion of an anesthetic molecule.

Analysis of five tylosin biosynthetic genes from the tyllBA region of the Streptomyces fradiae genome.

The tyllBA region of the tylosin biosynthetic gene cluster of Streptomyces fradiae contains at least five open reading frames (ORFs). ORF1 (tylI) encodes a cytochrome P450 and mutations in this gene affect macrolide ring hydroxylation. The product of ORF2 (tylB) belongs to a widespread family of proteins whose functions are speculative, although tylB mutants are defective in the biosynthesis or addition of mycaminose during tylosin production. ORFs 3 and 4 (tylA1 and tylA2) encode delta TDP-glucose synthase and delta TDP-glucose dehydratase, respectively, enzymes responsible for the first two steps common to the biosynthesis of all three deoxyhexose sugars of tylosin via the common intermediate, delta TDP-4-keto, 6-deoxyglucose. ORF5 encodes a thioesterase similar to one encoded in the erythromycin gene cluster of Saccharopolyspora erythraea.

Expansion of the Candida albicans cell envelope in different morphological forms of the fungus.

Modes of cell envelope expansion were monitored in developing cells of Candida albicans 73/055 to which polystyrene beads were attached. Eight different conditions of culture medium, pH and temperature were used to promote growth in a variety of morphological forms. The cells were observed microscopically during growth in Sykes-Moore perfusion chambers, and sequential measurements of distances between the bead and the parent cell, and the bead and the apical tip were used to distinguish apical envelope expansion from general envelope expansion. Morphology index (Mi) was determined at each time point as an estimate of each cell's morphology. Calculations based on the measurements showed that general envelope expansion was inversely proportional to Mi, but that general expansion greater than 20% occurred only in cells with a final Mi less than 2.0, indicating that regulation of apical and general envelope expansion alone may be insufficient to determine the different morphologies seen in cells with higher Mi. The rate of expansion of the perimeter of cells was linearly proportional to the final Mi. This observation suggests that commitment to morphological development in C. albicans may in part involve commitment to a rate of envelope expansion, which itself helps determine the final morphology of a cell.

Reaction of Candida albicans cells of different morphology index with monoclonal antibodies specific for the hyphal form.

Two monoclonal antibodies (MAbs), 3D9 with reported specificity for Candida albicans hyphae, and 3B7 with reported specificity for morphological forms of C. albicans found in vivo, were tested by indirect immunofluorescence with C. albicans cells that were grown in 12 different environments (four different culture media incubated at various temperatures) and whose cellular morphology was estimated in terms of morphology index (Mi). Both MAbs reacted strongly with cells with Mi greater than 3.0, i.e., with pseudohyphal and hyphal forms, but in Eagle's medium at 26 degrees C and in a modified Sabouraud's broth medium at 30 degrees C, some reactivity was also found with cells of lower Mi (i.e., yeast forms). Therefore, it was concluded that the hyphal phenotype and the epitopes reactive with the MAbs were co-expressed but that the epitopes could also be expressed independently of the hyphal phenotype. The results confirm the propensity of C. albicans for variation of its surface antigenic composition.