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Biomedical subjects

M J Hayden

Publications and source records attributed to M J Hayden.

At least 19 recordsLinked to original sources
PubMed2006

Development and genetic mapping of sequence-tagged microsatellites (STMs) in bread wheat (Triticum aestivum L.).

The density of SSRs on the published genetic map of bread wheat (Triticum aestivum L.) has steadily increased over the last few years. This has improved the efficiency of marker-assisted breeding and certain types of genetic research by providing more choice in the quality of SSRs and a greater chance of finding polymorphic markers in any cross for a chromosomal region of interest. Increased SSR density on the published wheat genetic map will further enhance breeding and research efforts. Here, sequence-tagged microsatellite profiling (STMP) is demonstrated as a rapid technique for the economical development of anonymous genomic SSRs to increase marker density on the wheat genetic map. A total of 684 polymorphic sequence-tagged microsatellites (STMs) were developed, and 380 were genetically mapped in three mapping populations, with 296 being mapped in the International Triticeae Mapping Initiative W7984 x Opata85 recombinant inbred cross. Across the three populations, a total of 479 STM loci were mapped. Several technological advantages of STMs over conventional SSRs were also observed. These include reduced marker deployment costs for fluorescent-based SSR analysis, and increased genotyping throughput by more efficient electrophoretic separation of STMs and a high amenability to multiplex PCR.

Chromosome Mapping↗
PubMed2004

Sequence tagged microsatellites for the Xgwm533 locus provide new diagnostic markers to select for the presence of stem rust resistance gene Sr2 in bread wheat ( Triticum aestivum L.).

The stem rust resistance gene Sr2 has provided durable broad-spectrum, adult-plant resistance to the fungal pathogen Puccinia graminis Pers. f. sp. tritici throughout wheat-growing regions of the world for more than 50 years. The ability to select for Sr2 in wheat breeding programs was recently improved by the identification of a tightly linked microsatellite marker gwm533. This marker typically amplifies a 120-bp polymerase chain reaction fragment from wheat lines carrying Sr2. In instances where the 120-bp fragment is not associated with the presence of Sr2, DNA sequence analysis has shown that a second allele was amplified, differing in the structure of the microsatellite repeat. To discriminate this allelic homoplasy (alleles identical in size, but not identical by descent), sequence-tagged microsatellites (STM) markers were developed for the Xgwm533 locus. These markers were shown to be diagnostic for the presence of Sr2 in a wide range of germplasm, representative of all major wheat varieties historically grown in Australia. The STMs will be particularly useful for marker-assisted selection in Southern Australian breeding programs, where the use of the marker gwm533 is often precluded by the presence of the non- Sr2-associated 120-bp allele in the pedigree of current breeding germplasm. The STMs also revealed a high incidence of previously undetected allelic homoplasy at the Xgwm533 locus and may have broader utility in genetic research and breeding, as this locus is also reported to be strongly associated with a major gene conferring resistance to Fusarium head blight.

Agriculture↗
PubMed2003

A new approach to extending the wheat marker pool by anchored PCR amplification of compound SSRs.

A study was undertaken to determine the utility in bread wheat of anchored PCR for the development of single locus SSR markers targeted at compound repeat motifs. In anchored PCR, microsatellite amplification is achieved using a single primer complementary to the flanking sequence, and one which anchors to the repeat junction of the compound SSR. The recovery rate of useable markers was found to be similar (43%) to that reported for conventionally generated SSRs. Thus, anchored PCR can be used to reduce the costs of marker development, since it requires that only half the number of primers be synthesised. Where fluorescence-based platforms are used, marker deployment costs are lower, since only the anchoring primers need to be labelled. In addition, anchored PCR improves the recovery of useful markers, as it allows assays to be generated from microsatellite clones with repeat sequences located close to their ends, a situation where conventional PCR amplification fails as two flanking primers cannot be designed. Strategies to permit the large-scale development of compound SSR markers amplified by anchored PCR are discussed.

Alleles↗
PubMed2002

Sequence tagged microsatellite profiling (STMP): improved isolation of DNA sequence flanking target SSRs.

Sequence tagged microsatellite profiling (STMP) enables the rapid development of large numbers of co-dominant DNA markers, known as sequence tagged microsatellites (STMs). Each STM is amplified by PCR using a single primer specific to the conserved DNA sequence flanking the microsatellite repeat in combination with a universal primer that anchors to the 5'-ends of the microsatellites. It is also possible to convert STMs into conventional microsatellite, or simple sequence repeat (SSR), markers that are amplified using a pair of primers flanking the repeat sequence. Here, we describe a modification of the STMP procedure to significantly improve the capacity to convert STMs into conventional SSRs and, therefore, facilitate the development of highly specific DNA markers for purposes such as marker-assisted breeding. The usefulness of this technique was demonstrated in bread wheat.

Conserved Sequence↗
PubMed2001

Sequence-tagged microsatellite profiling (STMP): a rapid technique for developing SSR markers.

We describe a technique, sequence-tagged microsatellite profiling (STMP), to rapidly generate large numbers of simple sequence repeat (SSR) markers from genomic or cDNA. This technique eliminates the need for library screening to identify SSR-containing clones and provides an approximately 25-fold increase in sequencing throughput compared to traditional methods. STMP generates short but characteristic nucleotide sequence tags for fragments that are present within a pool of SSR amplicons. These tags are then ligated together to form concatemers for cloning and sequencing. The analysis of thousands of tags gives rise to a representational profile of the abundance and frequency of SSRs within the DNA pool, from which low copy sequences can be identified. As each tag contains sufficient nucleotide sequence for primer design, their conversion into PCR primers allows the amplification of corresponding full-length fragments from the pool of SSR amplicons. These fragments permit the full characterisation of a SSR locus and provide flanking sequence for the development of a microsatellite marker. Alternatively, sequence tag primers can be used to directly amplify corresponding SSR loci from genomic DNA, thereby reducing the cost of developing a microsatellite marker to the synthesis of just one sequence-specific primer. We demonstrate the utility of STMP by the development of SSR markers in bread wheat.

Australia↗
PubMed2001

Targeted development of informative microsatellite (SSR) markers.

We describe a novel approach, selectively amplified microsatellite (SAM) analysis, for the targeted development of informative simple sequence repeat (SSR) markers. A modified selectively amplified microsatellite polymorphic loci assay is used to generate multi-locus SSR fingerprints that provide a source of polymorphic DNA markers (SAMs) for use in genetic studies. These polymorphisms capture the repeat length variation associated with SSRs and allow their chromosomal location to be determined prior to the expense of isolating and characterising individual loci. SAMs can then be converted to locus-specific SSR markers with the design and synthesis of a single primer specific to the conserved region flanking the repeat. This approach offers a cost-efficient and rapid method for developing SSR markers for predetermined chromosomal locations and of potential informativeness. The high recovery rate of useful SSR markers makes this strategy a valuable tool for population and genetic mapping studies. The utility of SAM analysis was demonstrated by the development of SSR markers in bread wheat.

Alleles↗
PubMed1998

Parental attitudes toward infant pulmonary function testing.

Infant pulmonary function tests (PFTs) have proven increasingly popular and useful for clinical and research purposes. Informed consent requires accurate information on side effects. Our aim was to quantify minor side effects from a parental point of view by means of a questionnaire. The parents of 97 infants attending for PFTs were asked to complete a simple questionnaire. Eighty-one parents (84%) returned the questionnaire. Forty-one percent felt that their infants were not troubled by the process of administering the sedative chloral hydrate, whereas 55% suffered mild to moderate distress. In contrast, 94% of infants were not distressed by the actual PFTs. Similarly, 46% of parents were not distressed by the administration of sedative to their infant, with 49% expressing distress to a mild or moderate degree. Although 73% of parents were not distressed by watching their infants undergo the PFTs, 27% were to a mild to moderate degree. Seventy-three percent of infants were untroubled on waking. Seventy percent of infants had a good nights sleep after the PFTs. The vast majority of parents (94%) were happy to recommend that others allow their infants to undergo similar testing. We noted that most problems caused by infant PFTs relate to the administration of the sedative. Most infants awake from the tests not distressed and sleep normally the following night.

Attitude↗
PubMed1998

Bronchodilator responsiveness testing using raised volume forced expiration in recurrently wheezing infants.

We hypothesized that a new test of infant lung function, less affected by shifts in lung volume, might better detect bronchodilator effects. Using the raised volume forced expiration technique (RVFET), the effect of a bronchodilator on lung function was studied in 22 infants with a history of recurrent wheeze and five healthy infants. Forced expiratory volume in 0.75 s (FEV0.75), forced expiratory vital capacity (FVC), and forced expiratory flow at 75% of FVC (FEF75%) were measured by forcing expiration, using an inflatable jacket from a lung volume set by an inspiratory pressure of 20 cm H2O. A minimum of five measurements were made at baseline and following the administration of 500 microg of salbutamol from a metered dose inhaler via a small volume metal spacer. Changes in lung function in the group of 25 infants who received salbutamol were compared to seven infants who received placebo aerosol. No significant changes occurred in measures of lung function following salbutamol administration when compared to baseline or placebo despite a significant increase in heart rate. A shift in lung volume is unlikely the reason why infants do not demonstrate a change in forced expiration following bronchodilator administration.

Albuterol↗
PubMed1998

Using low-frequency oscillation to detect bronchodilator responsiveness in infants.

The potential of the low-frequency forced oscillation technique (FOT) to measure the response to inhaled salbutamol was studied in 13 infants with a history of recurrent wheeze and nine healthy infants. The input impedance of the respiratory system (Zrs) between 0.5 and 20 Hz was measured at a transrespiratory pressure of 20 cm H2O during a brief Hering-Breuer reflex-induced pause in breathing. Parameters representing the airway resistance (Raw) and inertance (law), and a constant-phase tissue damping (G) and elastance (H) were estimated from the Zrs spectra. Lung function was measured before and after the administration of 500 microg of salbutamol via a small-volume metal spacer. Six of these infants also received a placebo aerosol. A fall in Raw (13% for the entire group) occurred following treatment with salbutamol (p < 0.008) but not placebo. There was no significant difference in the response to salbutamol between the normal infants (7.65% +/- 5.49%) and those with recurrent wheeze (17.58% +/- 8.67%). On grouped data, the fall in G just failed to reach statistical significance (p = 0.05) after correcting the significance level for multiple tests. No significant change occurred in law or H. We conclude that the low-frequency FOT is a suitable methodology for studying bronchodilator responsiveness in infants.

Administration, Inhalation↗
PubMed1998

Salbutamol delivery from a hydrofluoroalkane pressurized metered-dose inhaler in pediatric ventilator circuits: an in vitro study.

STUDY OBJECTIVES: The aim of our study was to determine the in vitro delivery of salbutamol from a pressurized metered-dose inhaler (pMDI) containing hydrofluoroalkane (HFA) propellant through various delivery devices to four models of a pediatric lung. DESIGN: To determine the effect of electrostatic charge, delivery of salbutamol was initially assessed with a multistage liquid impinger (MSLI) through an inline nonchamber device (Baxter MDI Adapter) and a small (Aerochamber MV) and a large (Nebuhaler) inline chamber device. Following this, the delivery was assessed to four lung models appropriate for a child of 70 kg, 50 kg, 15 kg, and 4 kg, with the same three reduced static devices inserted directly into a pediatric ventilator circuit. MEASUREMENTS AND RESULTS: Reduction of electrostatic charge improved small particle delivery through holding chambers to the MSLI by 12 to 14%. In the ventilator model, the mean delivery was between 1.9% and 5.4% for the nonchamber device, between 14.3% and 27.2% for the small holding chamber, and between 7.2% and 25.7% for the large holding chamber. Delivery was the least efficient in the 4-kg model compared to the 70-kg, 50-kg, and 15-kg models. CONCLUSIONS: Salbutamol from an HFA pMDI is delivered efficiently through inline holding chambers with reduced static in pediatric ventilator settings. A large holding chamber has no advantage over a small holding chamber. In addition, salbutamol delivery is more efficient through a holding chamber than through a nonchamber device.

Administration, Inhalation↗
PubMed1997

Aerosol delivery to wheezy infants: a comparison between a nebulizer and two small volume spacers.

Inhalation therapy for wheezy infants with either a nebulizer or a pressurized metered-dose inhaler (pMDI) through a spacer is common practice. The aim of our study was to compare aerosol delivery to wheezy infants from a nebulizer and from a pMDI via two small volume spacers. Twenty wheezy infants (aged 4-12 months) were recruited. They inhaled salbutamol from a Pari-Baby nebulizer, from a detergent-coated Babyhaler, and from a Nebuchamber in random order. A filter was placed between the inhalation systems and the patients. The amount of salbutamol deposited on the filter was measured using an ultraviolet spectrophotometer and was expressed as a percentage of the total nebulized or actuated doses. The mean total nebulized dose for the Pari-Baby (1030 micrograms) was higher (P < 0.001) than the mean actuated dose from a pMDI for the Babyhaler (374 micrograms) and for the Nebuchamber (378 micrograms). Mean drug deposition on the filter was 40.2% (150 micrograms) of the total actuated dose for the detergent-coated Babyhaler and 40.7% (154 micrograms) of the total actuated dose for the Nebuchamber. There was no significant difference in drug deposition on the filter between the two spacers. Mean drug deposition on the filter was 25.3% (260 micrograms) of the total nebulized dose for the Pari-Baby nebulizer. There was no weight dependence in drug deposition on the filter for the two spacers, but, drug deposition increased with the subject's weight for the nebulizer. We have shown that aerosol delivery to wheezy infants from a pMDI through small volume spacers is effective and that a higher percentage of the total amount of salbutamol is delivered than from a nebulizer. The weight dependence in drug deposition for the nebulizer can be of clinical relevance.

Administration, Inhalation↗
PubMed1997

Methacholine responsiveness using the raised volume forced expiration technique in infants.

Infant lung function can be assessed with the tidal volume "squeeze" technique or, over an extended volume range, with the newer raised volume forced expiration technique (RVFET). We assessed methacholine responsiveness in 11 infants, measuring both maximal expiratory flow at functional residual capacity (V.max,FRC)with the tidal volume technique, and forced expiratory volume/time (FEV(t)) with RVFET. We used a standard methodology for the former. FEV(t) was measured by inflating the infant's lungs to 20 cm H2O and forcing expiration using a jacket setup to transmit a pressure of 20 cm H2O to the airway. Lung function was measured at baseline and after methacholine inhalations, increasing from 0.1 g/L to 10 g/L in half log dosage increments (DI). The provocative concentrations (PC) of methacholine leading to a 40% fall in V.max,FRC and a 15 or 20% fall in FEV(t) were calculated. The mean provocative concentration of methacholine required to produce a 40% fall in V.max,FRC was less than that required to produce a 20% fall in FEV0.5 by 0.39 DI (95% CI, -0.60 to 1.38) and less than that required to produce a 20% fall in FEV0.75 by 0.42 DI (95%, CI, -0.54 to 1.39). Similarly, the provocative concentration of methacholine required to produce a 40% fall in V.max,FRC was less than that required to produce a 15% fall in FEV0.5 by 0.14 DI (95% CI, -0.99 to 1.28) or a 15% fall in FEV0.75 by 0.13 DI (95% CI, -0.80 to 1.08), but the differences were small and not significant. Despite these differences the agreement between the two methods was good, and bronchoconstriction was not attenuated by the forced inspiration delivered by the raised volume maneuver. We conclude that the raised volume forced expiration technique is able to detect methacholine-induced bronchoconstriction.

Bronchial Provocation Tests↗
PubMed1997

Volume dependence of respiratory impedance in infants.

We previously studied low-frequency respiratory impedance (Zrs) data at an elevated lung volume to separate airway and tissue mechanical properties in normal infants (Am. I. Respir. Crit. Care Med. 1996; 154:161-166). The aim of the present study was to determine the volume dependence of the airway and tissue mechanics by extending Zrs measurements to lower lung volumes. Zrs spectra between 0.5 and 21 Hz were measured in supine sleeping infants (n = 8; 7 to 26 mo of age) at mean transrespiratory pressures (Ptr[mean]) of 20, 10, and 0 cm H2O, during periods of apnea induced by inflating the infants' lungs to a pressure of 20 cm H2O through a face mask. At each inflation pressure, a model containing airway resistance (Raw) and inertance (law) and tissue damping (G) and elastance (H) was fitted to Zrs data. At FRC, the values of Raw, law, G, and H were 20.6+/-4.9 (SD) cm H2O x s/L, 0.037+/-0.014 cm H2O x s2/L, 39.6+/-10.3 cm H2O/L, and 147+/-35 cm H2O/L, respectively. Increase of Ptr(mean) caused a monotonous decrease in Raw (42+/-7% of the value at FRC), while law remained constant. The tissue parameters were minimal at a Ptr(mean) of 10 cm H2O (68+/-10% and 78+/-6% in G and H, respectively) and significantly higher at both 0 and 20 cm H2O. Although Zrs measurements can be made in most infants at lung volumes as low as FRC, an inflation pressure of 20 cm H2O provides a higher success rate and is therefore a more suitable condition for general use.

Airway Resistance↗
PubMed1997

Influence of driving pressure on raised-volume forced expiration in infants.

The raised-volume forced-expiration technique measures infant lung function over an extended volume range. To improve comparisons between individuals and populations, we investigated the influence of jacket pressure on outcome variables in 21 infants. To quantify pressure transmitted from the jacket to the pleural space at a given lung volume, the jacket was inflated against an occluded airway, and the increase in pressure at the mouth was measured. Flow-volume curves were recorded at transmitted pressure (Ptrans) values ranging from 0 to 41.9 cm H20. The effect of Ptrans on the FEV measures of FEV0.5, FEV0.75, and FVC, and on the forced expiratory flow measures of FEF25%, FEF50% and FEF75% was assessed. At Ptrans values between 0 to 20 cm H20, a significant positive relationship existed between transmitted pressure (Ptrans) and all outcome variables except FVC. At higher Ptrans values, all outcome variables demonstrated pressure independence, with the exception of FEF25% (which remained positive) and FVC (which was negative in a subgroup of wheezy infants). FEF75% values tended to decrease at Ptrans values > 25 cm H20. At Ptrans values between 20 and 25 cm H20, most outcome variables are pressure independent. This range is therefore the most suitable for use with the raised-volume forced expiration technique.

Female↗
PubMed1996

Effect of electrostatic charge, flow, delay and multiple actuations on the in vitro delivery of salbutamol from different small volume spacers for infants.

BACKGROUND: A study was undertaken to determine the influences of electrostatic charge, flow, delay, and multiple actuations on the in vitro delivery of salbutamol generated by a pressurised metered dose inhaler (pMDI) from small volume spacers used in infants. METHODS: Ten actuations from a salbutamol pMDI were drawn at different flow rates after either single or multiple actuations, with or without delay, through either static or reduced static spacers. An ionic detergent was used to reduce the charge of plastic spacers (Babyhaler, Babyspacer, Aerochamber, Nebuhaler). Electrostatic charge was measured using an electrometer. A multistage liquid impinger was used to determine the particle size distribution of the output of the pMDI through the spacers. RESULTS: Electrostatic charge on the surface of plastic spacers had the greatest influence on delivery, causing a decrease in drug delivery. Reducing charge by coating the surface with ionic detergent resulted in an increase of 46.5-71.1% (p < 0.001) in small (< 6.8 microns) particle delivery from small volume plastic spacers. Lower flow, delay, and multiple actuations resulted in decreased delivery from static spacers. Lower flow resulted in a decrease of 15% in small (< 9.6 microns) particle delivery. Delay and multiple actuations resulted in a decrease of 40.7% and 76.0%, respectively, in small (< 6.8 microns) particle delivery. The influences of lower flow, delay, and multiple actuations were greatly reduced or even eliminated by reducing charge. However, multiple actuations still resulted in a significant decreased delivery (p < 0.05). The reduced static Nebuhaler had a higher delivery than all small volume spacers. CONCLUSIONS: Electrostatic charge has a major influence on the delivery of salbutamol from small volume spacers. Using a metal spacer or ionic detergent coating of plastic spacers resulted in no or reduced charge and hence in improved delivery. Lower flow, delay, and multiple actuations played a major part only in static spacers.

Albuterol↗
PubMed1996

Measurement of low-frequency respiratory impedance in infants.

Low-frequency respiratory impedance (Zrs) data permit the separate estimation of the mechanical properties of the airways and the tissues, but they are difficult to collect in humans because of the need for apneic conditions. We exploited the apneic phase produced by invoking the Hering-Breuer reflex with end-inspiratory airway occlusion in five sedated infants aged 9 to 16 mo. A computer-controlled pump and solenoid valves were used to inflate the supine infants through a face mask to a transrespiratory pressure of 20 cm H2O and to affect the airway occlusion. A loudspeaker-in-box system was connected to the mask through a side-arm, and small-amplitude pseudorandom oscillations containing 23 frequency components between 0.5 and 20.75 Hz were applied for 6 s. Four consecutive measurements were made in each infant, and the averaged Zrs spectra were evaluated on the basis of a model containing the frequency-independent resistance (Raw) and inertance (law) of the airways, and the viscous damping (G) and elastance (H) parameters of the constant-phase compartment of the chest wall and parenchymal tissues. The measured Zrs values were consistent with the model up to 15 Hz, and the average fitting error was 0.89 +/- 0.11 (SD) cm H2O.s/L. The following parameter values were obtained: Raw = 10.0 +/- 2.1 cm H2O.s/L, law = 0.061 +/- 0.014 cm H2O.s2/L, G = 28.6 +/- 4.9 cm H2O/L, H = 141 +/- 55 cm H2O/L. The tissue hysteresivity (G/H) values were 0.218 +/- 0.061. Our results indicate that, in short apneic periods evoked by the Hering-Breuer reflex, reliable low-frequency Zrs data can be collected to partition the tissue and airway impedances in sedated infants.

Airway Resistance↗
PubMed1996

Electrostatic charge on a plastic spacer device influences the delivery of salbutamol.

The aim of this study was to determine whether electrostatic charge on a plastic spacer decreases the delivery of salbutamol from a pressurized metered-dose inhaler (pMDI) and, if so, to find an optimal and practical treatment to remove the charge. Ten single actuations from a salbutamol pMDI were drawn through different Volumatic spacers at a constant flow of 60 L.min-1. The efficacies of different methods of removing charge were tested, including detergent coating of the spacers. A multistage liquid impinger was used to determine the particle size distribution of the output of the pMDI through the Volumatic spacers. The electrostatic charge on the inner surface of the spacers was measured both quantitatively with an electrometer, and qualitatively by the attraction of a thin strip of cellulose membrane to the wall of the spacer. Each experiment was repeated four times. Ionic detergent coating of the spacers removed the charge for at least 24 h. This resulted in an increase of 55-70% in small particle (< 6.8 microns) delivery compared to delivery from new spacers with high charge. We have demonstrated that electrostatic charge plays a major role in the delivery of salbutamol through plastic spacers. Adequate treatment with ionic detergent removes the charge and improves drug delivery.

Albuterol↗