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M H Wilkinson

Publications and source records attributed to M H Wilkinson.

At least 19 recordsLinked to original sources
PubMed1995

DATAPLOT: a graphical display package for bacterial morphometry and fluorimetry data.

A microbiological image processing system has been developed, which is used to determine both morphometry and fluorimetry data on (mainly faecal) bacteria. A special program for display of these data has been developed for this system. Besides probability density estimates of any one or two parameters and scatter plots of any two or three parameters in the distribution, it can generate scatter plots using the actual bacterial shapes, rather than arbitrary symbols. A particularly powerful feature of the program is the possibility to overlay a contour plot of the probability density distribution of the bacteria in parameter space, with a scatter plot of the actual bacterial shapes involved. In this paper, the program's organization, options, and the algorithms used are discussed.

Algorithms↗
PubMed1995

Quantitative fluorescence in situ hybridization of Bifidobacterium spp. with genus-specific 16S rRNA-targeted probes and its application in fecal samples.

Three 16S rRNA hybridization probes were developed and tested for genus-specific detection of Bifidobacterium species in the human fecal flora. Variable regions V2, V4, and V8 of the 16S rRNA contained sequences unique to this genus and proved applicable as target sites for oligodeoxynucleotide probes. Determination of the genus specificity of the oligonucleotides was performed by whole-cell hybridization with fluorescein isothiocyanate-labelled probes. To this end, cells were fixed on glass slides, hybridized with the probes, and monitored by videomicroscopy. In combination with image analysis, this allowed quantification of the fluorescence per cell and objective evaluation of hybridization experiments. One of the probes developed was used to determine the population of Bifidobacterium spp. in human fecal samples. A comparison was made with results obtained by cultural methods for enumeration. Since both methods gave similar population estimates, it was concluded that all bifidobacteria in feces were culturable. However, since the total culturable counts were only a fraction of the total microscopic counts, the contribution of bifidobacteria to the total intestinal microflora was overestimated by almost 10-fold when cultural methods were used as the sole method for enumeration.

Base Sequence↗
PubMed1995

Fluorescence overlay antigen mapping of the epidermal basement membrane zone: III. Topographic staining and effective resolution.

In this third study on the fluorescence overlay antigen mapping (FOAM) technique, we have addressed the question of which differences of antigen distributions close to the resolving power of the light microscope can be distinguished. An answer to this question should provide clues to future applications of the technique aiming at the topographic differentiation of IgG deposits displayed at the epidermal basement membrane zone (EBMZ) in certain bullous skin disorders. For the present purpose we have developed a topographic staining model in human skin, using structural EBMZ antigens as topographic reference markers. The distribution of these markers relative to one another is visualized in FOAM images obtained by selective double immunofluorescence tracing and videomicroscopic overlay imaging. The theoretical resolution limit of the technique is discussed and suggests an effective lower limit of some 60-65 nm. Although this limit is not reached under present conditions, our results show that it is possible to distinguish topographic differences of antigen distributions with an upper resolution limit of 200 +/- 50 nm. Furthermore, our findings indicate that collagen Type VII and beta 4 integrin are the most suitable molecules to serve as topographic reference markers in future applications of the technique aiming at the differentiation of bullous pemphigoid (BP) and epidermolysis bullosa acquisita (EBA). Preliminary results on this topic are most promising indeed.

Antigens, Surface↗
PubMed1995

Fluorescence overlay antigen mapping of the epidermal basement membrane zone: II. Color fidelity.

In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight some of the errors that may influence faithful color rendition of slide preparations using triple antigen immunofluorescence staining. Reliable interpretation of multicolor fluorescence images requires that the observer can unambiguously assign each color in these images to the presence of a specific combination of the labeled antigens. This is possible only when the image fidelity meets certain standards. The present study concentrates on color fidelity which is easily undermined by spectral matching errors, image contrast errors, and exposure time errors. Evaluation of these errors, using the photomicrographic overlay variant of FOAM, showed the potential unreliability of the simultaneous use of multiple fluorophores for immunofluorescence microscopy. The procedures described here may serve as a solid starting point in formulating technical conditions that allow reliable color rendition in multicolor immunofluorescence microscopy. Furthermore, these procedures can be adapted to studies other than the analysis of basement membrane zone antigens, to which they have been first applied.

Basement Membrane↗
PubMed1994

Shading correction and calibration in bacterial fluorescence measurement by image processing system.

An image processing system with applications in bacterial (immuno-)fluorescence measurement has been developed. To reach quantitative results, correction for non-uniformities in system sensitivity, both as a function of time (calibration for drifts) and as a function of image coordinates (shading correction), is essential. Both problems can be handled simultaneously by acquiring images of a uniformly fluorescent, solid standard as a reference image. To measure bacterial fluorescence, the average fluorescence intensity of isolated areas of interest (the bacteria) is computed, and corrected using the reference images. Two shading correction methods are theoretically and experimentally compared: direct averaging in the corrected image, and (weighted and unweighted) averaging using the raw image and a separate shading image to determine the weights and correct for shading during the averaging. The latter method proved computationally 3.5-6.5 times faster on average and reduced propagation of truncation errors during computation, resulting in 40% less noise, for 8-bits/pixel images.

Bacteria↗
PubMed1994

A simple data compression scheme for binary images of bacteria compared with commonly used image data compression schemes.

A run length code compression scheme of extreme simplicity, used for image storage in an automated bacterial morphometry system, is compared with more common compression schemes, such as are used in the tag image file format. These schemes are Lempel-Ziv and Welch (LZW), Macintosh Packbits, and CCITT Group 3 Facsimile 1-dimensional modified Huffman run length code. In a set of 25 images consisting of full microscopic fields of view of bacterial slides, the method gave a 10.3-fold compression: 1.074 times better than LZW. In a second set of images of single areas of interest within each field of view, compression ratios of over 600 were obtained, 12.8 times that of LZW. The drawback of the system is its bad worst case performance. The method could be used in any application requiring storage of binary images of relatively small objects with fairly large spaces in between.

Bacteria↗
PubMed1994

Computer processing of microscopic images of bacteria: morphometry and fluorimetry.

Several techniques that use computer analysis of microscopic images have been developed to study the complicated microbial flora in the human intestine, including measuring the shape and fluorescence intensity of bacteria. These techniques allow rapid assessment of changes in the intestinal flora and could apply equally to other complex microbial ecosystems.

Bacteria↗
PubMed1994

Fluorescence overlay antigen mapping of the epidermal basement membrane zone: I. Geometric errors.

To identify in tissue sections the relative positions of antigen distributions close to the resolving power of the microscope, we have developed the fluorescence overlay antigen mapping (FOAM) procedure. As this technique makes high demands on the geometric fidelity of the overlay image, it is essential to recognize geometric errors resulting from optical imperfections. This applies in particular to the image shift difference (ISD) that may routinely occur during fluorescence overlay. We describe here procedures for assessment and mechanical correction of the ISD in tissue sections. Furthermore, we describe an alignment verification test to assess the accuracy of the ISD correction procedure, using collagen Type VII as the geometric verification marker. These procedures should enable reliable evaluation of relative antigen distributions in tissue sections using photomicrographic multicolor fluorescence overlay. Further details of the FOAM technique, such as color fidelity and its utility for diagnostic and research purposes, will be published separately.

Basement Membrane↗
PubMed1992

Circulating antibodies against faecal bacteria assessed by immunomorphometry: combining quantitative immunofluorescence and image analysis.

A new technique to study the prevalence of circulating antibodies directed against different morphological groups ('morphotypes') of bacteria in fresh faeces is presented. The technique combines quantitative indirect immunofluorescence with digital image analysis. Plasma antibody titres and patterns of IgA, IgG and IgM isotype against morphotypes of faecal bacteria were determined in ten healthy individuals.

Adult↗
PubMed1991

Objective quantitation of serum antibody titres against Enterobacteriaceae using indirect immunofluorescence, read by videocamera and image processing system.

A new way of measuring indirect immunofluorescence (IIF) of microscopic bacterial slide preparations by videocamera and an image processing system is presented. This method is compared with the conventional method of reading the slides by eye. The advantages of this new approach are objective reading, greater accuracy and easier performance. We have applied the method to measure serum antibody titres against endogenous Enterobacteriaceae. The method offers the opportunity to combine IIF with automatic morphological analysis, thereby maximally exploiting the possibilities of the immunofluorescence technique.

Antibodies, Bacterial↗
PubMed1991

Electromyographic changes in the isolated rat diaphragm during the development of fatigue.

To investigate whether the power spectrum of the electromyogram of a fatiguing muscle can be used to infer the degree to which the muscle is fatigued, we recorded isometric tension and two monopolar electromyograms from eight isolated rat diaphragm preparations suspended in an organ bath containing a balanced salt solution. Each preparation was excited with a fixed phrenic nerve impulse pattern made up of a 70-Hz train of impulses of supramaximal voltage delivered for 170 ms with a 500-ms recovery period. Tension fell rapidly over the first 60 s of the fatigue run and more slowly for the remaining 60 s analysed. The duration of extracellular action potentials increased and their amplitude decreased as the tension developed by the diaphragm decreased; conduction velocity along muscle fibres also decreased. The centroid frequency (fcen) of the power spectrum of the first action potential elicited by each train of stimuli decreased rapidly until tension fell to approximately 70% of the initial value; thereafter little change in fcen occurred, although tension continued to fall to 33% of its initial value. Our results demonstrated that under controlled conditions, fcen provided a sensitive index of fatigue in its early stages, but provided no information once fatigue was pronounced.

Animals↗
PubMed1991

Morphometrical parameters of gut microflora in human volunteers.

The morphology of faecal microflora of nine healthy human volunteers was studied by digital image analysis of microscopic slides. Weekly specimens were collected during an 8-week period. Seven morphometrical parameters were derived: the means and medians of components 1, 2 and 3, and morphometrical entropy. Statistically significant differences among subjects were found for means of components 1 and 2, medians of components 1, 2 and 3, and entropy. The stability in normal circumstances provides an excellent basis for the detection of pathological change in gut flora balance.

Analysis of Variance↗
PubMed1991

Effects of ceftriaxone on faecal flora: analysis by micromorphometry.

In order to elucidate the effect of ceftriaxone therapy on the morphology of gut microflora, 11 human volunteers were treated with ceftriaxone, 1 g daily, given intramuscularly in one dose. Treatment continued for 5 days. Faecal microflora was analysed by digital image processing before, during and after the treatment period. We derived simple numerical parameters which describe the morphologic composition of the flora. They were significantly influenced by the antibiotic, and returned to their baseline values more than 7 days after treatment was stopped. The procedure holds promise for clinical application.

Adult↗
PubMed1987

Progressive shortening of the periodic breathing cycle duration in normal infants.

Ten normal full-term neonates were monitored by oxycardiorespirography in hospital during the 1st wk of life, and subsequently at home at 4, 8, and 12 wk of age in a longitudinal study of the duration of the periodic breathing cycle during spontaneous sleep. Periodic breathing was observed in 25 of the 40 studies, with a total of 168 epochs of periodicity noted. Eighty-four percent of the periodic epochs occurring during nonrapid eye movements were preceded by a sigh, body movements, or sleep state transition, compared with 41% during rapid eye movements (p less than 0.005). There was a progressive reduction in the periodic breathing cycle duration, which fell significantly between the 1st and 4th wk, in both sleep states, (p less than 0.001). There was a further significant fall between 4 and 12 wk in nonrapid eye movement sleep (p less than 0.05), and there was no significant difference between sleep states at any postnatal age. The overall change in the duration of the periodic cycle, for both sleep states combined, was from 15.0 +/- 3.6 s at less than 1 wk to 12.4 +/- 1.8 s at 12 wk of age. The periodic breathing cycle duration progressively shortens over the first 3 months of life. We postulate that this shortening may be useful indicator of peripheral chemoreceptor maturation over this time period.

Apnea↗
PubMed1986

Phasic respiratory activity in the fetal lamb during late gestation and labour.

We quantified the respiratory activity of 9 fetal lambs using computer-analysis of the diaphragmatic electromyogram (EMG) obtained during 2 h recording sessions interspersed over the last 13 days of gestation. The fetuses delivered unassisted at an average gestational age of 145 days (term = 147 days). During the last 2 h of labour the number of phasic EMG bursts (breaths) averaged 3% of the peak recorded earlier in the study. This decline in breathing began at least 2 days before labour and resulted predominantly from the fetus spending an increasing proportion of time in apnoea. Respiratory rate within epochs of breathing also fell significantly 1 day before labour, and the proportion of time spent in the low voltage electrocortical state declined once labour commenced. No significant change occurred in arterial PO2, PCO2 or pH over the study period. We conclude that fetal respiratory activity falls well before the onset of labour, largely as a result of increased apnoea, and that the decline does not result from the development of a progressive hypoxaemia associated with labour.

Animals↗
PubMed1983

Computerised nutritional data management in neonatal intensive care.

We report a minicomputer based recording, reporting, and research system for patient data management in the neonatal intensive care unit (NICU). It is moderately priced, transportable, and independent of a central computer. It requires only a desk top and standard power supply, and successfully meets the medical documentation, communication, and research needs of the NICU. The first phase of programme development has enabled us to process growth and nutritional data from infants on complex parenteral and enteral nutrition. The system is well suited to medical users and provides a practical interactive data management system with many potential applications in the NICU.

Australia↗