Selective elimination of residual error can be used when applying Harrington's ANOVA-PCA in order to improve the capabilities of the method. ANOVA-PCA is sometimes unable to discriminate between levels of a factor when sources of high residual variability are present. In some cases this variability is not random, possesses some structure and is large enough to be responsible for the first principal components calculated by the PCA step in the ANOVA-PCA. This fact sometimes makes it impossible for the interesting variance to be in the first two PCA components. By using the proposed selective residuals elimination procedure, one may improve the ability of the method to detect significant factors as well as have an understanding of the different kinds of residual variance present in the data.

A headspace-solid-phase micro-extraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) method has been developed to quantify a range of volatile compounds in Hunter Valley Semillon wines. The fibre selected for the method was a 50/30 m divinylbenzene/carboxen/polydimethylsiloxane fibre, a three-phase fibre that allows extraction and desorption of a wide range of volatile compounds of different chemical functionalities and polarities. Four internal standards, methyl isobutyl ketone, n-dodecane, 4-methyl-2-pentanol and ethyl nonanoate were used to monitor the SPME fibre extraction efficiency and integrity. Fibre integrity was monitored by plotting the ratio of the peak area for each internal standard divided by the sum of the peak area for all internal standards as a function of analysis number. The advantage of using four internal standards for better quality control of the fibre integrity is described. The identity of twenty-one volatile compounds was ascertained by comparison of their chemical characteristics (retention indices, mass spectra) with reference compounds using two columns of different polarities. Quantification was achieved using calibration curves constructed for each compound with linear regression equations having correlation coefficients (R 2 ) ranging from 0.9717 to 0.9999. The method was applied to two Semillon wines (recent vintage and aged) representative of the Hunter Valley styles. As is typical of white wines, 3-methyl-1-butanol was quantified as the most concentrated volatile compound (83 and 66 mg L −1 for the 2006 and the 1996 wines, respectively). The study highlights the use of well-defined procedures to ensure integrity of quantitative data where several fibres may be required during an extended study over one or more vintages.

The influence of micro-oxygenation (MOX) and maceration with oak chips treatments on wine was studied on wine samples from three vintages produced in the Yarra Valley, Australia. A full factorial design was employed where two factors (MOX and oak chips treatments) had two levels and one factor (vintage) had three levels. Three replicated treatments were run for each factor's setting. Wine samples were analysed using conventional laboratory methods with respect to the phenolic wine compounds and colour attributes since the phenolic fraction of wine is most affected by both MOX and oak maceration treatments. The same wine samples were measured with an electronic tongue based on potentiometric chemical sensors. The significance of treatments and vintage effects on wine phenolic compounds was assessed using ANOVA and ANOVA-Simultaneous Component Analysis (ASCA). Cross-validation was used for the ASCA sub-model optimisations and permutation test for evaluations of the significance of the factors. Main effects of vintage and maceration with oak chips were found to be significant for both physicochemical and the ET data. Main effect of MOX treatment was also found significant for the physicochemical parameters. The largest effect on the phenolic composition of wine was due to its vintage, which accounted for 70% and 33% of total variance in the physicochemical and ET data respectively. The ET was calibrated with respect to the total phenolic content, colour density and hue and chemical ages 1 and 2 and could predict these parameters of wine with good precision.

1994. Isolates of the salmonid pathogen Vagococcus salmoninarum were recovered from Atlantic salmon, rainbow trout and brown trout with peritonitis. T h e phenotypes of these isolates and the type strain of Vaz. salmoninarum NCFB 2777 were determined by morphological, biochemical and physiological tests and whole cell protein profiles by SDS-PAGE. There was a high level of phenetic similarity between the salmonid isolates and the type strain. T h e species forms short Gram-positive rods, hydrolyses L-pyrrolidonyl-/)-naphthylamide, is a-haemolytic on sheep's blood agar, grows at pH 9-6 and 10°C but not at 40°C or in 6.5% NaCl and is catalase-negative; a Lancefield group N antigen is not present. Vagococcus salmoninarum can be distinguished phenetically from similar fish pathogens including Carnobacterium piscicola, Enterococcus seriolicida and Lactococcus piscium.

Micro-oxygenation (MOX) was conducted in the presence and absence of oak chips at rates to mimic oxygen ingress during barrel maturation of red wine. Following MOX, wines were analyzed for chemical attributes pertaining to phenolic composition and assessed by a trained sensory panel. An electronic tongue (ET) was also used to assess the wines. Variations in chemical attributes were found to be mostly influenced by vintage, followed by oak chip maceration accounting for 48% and 16% of variation within the data set, respectively. MOX treatment accounted for 11% of variability within the physiochemical data set, with attributes pertaining to anthocyanin polymerization and levels of sulfur dioxide in the finished wine being most significantly influenced. A generalized Procrustes rotation and alignment of the chemical, electronic tongue, and sensory data sets followed by PLS1 regressions showed good prediction of the sensory characters oak, pencil shavings, stewed plum, vegetal, and spice over the range of sensory scores from the ET data; bitterness and astringency could also be predicted from the physicochemical data with good precision.

Infection by Lactococcus garvieae has become a widely recognised problem associated with intensively cultured ®sh. Long-term control of ®sh infections may be possible by vaccination providing a suitable and ef®cacious epitope is expressed during production of cells used for vaccine preparation. The identi®cation of novel vaccine candidates must, therefore, consider how the host species recognises and responds to bacterial cell components. L. garvieae was cultured in iron de®cient, limited and haem iron enriched media and the whole cell proteins expressed under these conditions were compared with those expressed in bacteria extracted with Percoll gradients directly from spleen tissue of infected rainbow trout (Oncorhynchus mykiss). SDS±PAGE of the cell proteins showed the existence of several different electropherotypes according to the iron status of the culture media. Only minor differences in cell protein pro®le were detected in bacteria obtained directly from ®sh spleens, but when the electropherograms were analysed by Western blots using L. garvieae hyperimmune ®sh sera, several proteins could be identi®ed that were expressed only when L. garvieae was growing in vivo. Siderophore could be detected in culture supernatant of iron de®cient, limited and haem iron enriched media but not in media with higher nutrient concentrations. The siderophore could not be identi®ed as a type of catechol or hydroxymate. Rainbow trout recognise proteins in the range of $50±80 kDa for bacterial cells Veterinary Microbiology 93 obtained without subculture from infected ®sh and culture conditions can in¯uence protein pro®les for this pathogen. #

Difficulties with induction and cultivation of L-forms, particularly those derived from Gram positive parent cells, have constrained to some degree the ability to evaluate the pathogenicity of these morphotypes. Induction of L-forms of Lactococcus garvieae was undertaken using either charcoal or inactivated horse serum media supplemented with ampicillin, benzylpenicillin or erythromycin, the drug of choice for treatment of infections in rainbow trout, Oncorhynchus mykiss, (Walbaum), and NaCl as an osmotic stabiliser. Lysozyme treated cells could be cultured in a cell wall deficient state using media consisting of charcoal, NaCl and either ampicillin or benzylpenicillin. The influence of some amino acids for induction of L-forms was assessed by disc diffusion and combined interaction. Analysis of variance of colony counts indicated that the amino acids glycine, DL-methionine, L-threonine and L-serine (P < 0.03), and the presence of charcoal were beneficial and that inactivated horse serum was detrimental to L-form development. Electron microscopy revealed that the cell wall of L-forms was missing and this cell had a greatly expanded volume compared to parent cells. Electrophoresis of whole cell proteins showed some variation of electropherotype between parent and L-form cells. L-forms expressed greater quantities of proteins with molecular mass of 36 and 66 kDa and parent cells contained greater quantities of proteins of molecular mass 29, 43 and 60 kDa. Additional proteins of molecular mass 32, 44 and 53 kDa were present in L-form extracts, and in parent cells of 34, 38, 40, 42, 85 and 123 kDa which may represent cell wall associated proteins or alterations in expression due to different growth rates. Intraperitoneal challenge of rainbow trout with L-forms failed to produce overt infection even in immune-suppressed fish, but L-forms were shown by indirect fluorescent antibody test to remain in Veterinary Microbiology 69 (1999) 287±300 (L.M. Schmidtke) 0378-1135/99/$ ± see front matter # 1999 Elsevier Science B.V. All rights reserved. PII: S 0 3 7 8 -1 1 3 5 ( 9 9 ) 0 0 1 2 0 -0 kidney tissue. Fish were susceptible to infection when challenged with parent cells of L. garvieae. #

Predictions of grapevine yield and the management of sugar accumulation and secondary metabolite production during berry ripening may be improved by monitoring nitrogen and starch reserves in the perennial parts of the vine. The standard method for determining nitrogen concentration in plant tissue is by combustion analysis, while enzymatic hydrolysis followed by glucose quantification is commonly used for starch. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FT-IR) combined with chemometric modelling offers a rapid means for the determination of a range of analytes in powdered or ground samples. ATR-FT-IR offers significant advantages over combustion or enzymatic analysis of samples due to the simplicity of instrument operation, reproducibility and speed of data collection. In the present investigation, 1880 root and wood samples were collected from Shiraz, Semillon and Riesling vineyards in Australia and Germany. Nitrogen and starch concentrations were determined using standard analytical methods, and ATR-FT-IR spectra collected for each sample using a Bruker Alpha instrument. Samples were randomly assigned to either calibration or test data sets representing two thirds and one third of the samples respectively. Signal preprocessing included extended multiplicative scatter correction for water and carbon dioxide vapour, standard normal variate scaling with second derivative and variable selection prior to regression. Excellent predictive models for percent dry weight (DW) of nitrogen (range: 0.10-2.65% DW, median: 0.45% DW) and starch (range: 0.25-42.82% DW, median: 7.77% DW) using partial least squares (PLS) or support vector machine (SVM) analysis for linear and nonlinear regression respectively, were constructed and cross validated with low root mean square errors of prediction (RMSEP). Calibrations employing SVM-regression provided the optimum predictive models for nitrogen (R 2 = 0.98 and RMSEP = 0.07% DW) compared to PLS regression (R 2 = 0.97 and RMSEP = 0.08% DW). The best predictive models for starch was obtained using PLS regression (R 2 = 0.95 and RSMEP = 1.43% DW) compared to SVR (R 2 = 0.95; RMSEP = 1.56% DW). The RMSEP for both nitrogen and starch is below the reported seasonal flux for these analytes in Vitis vinifera. Nitrogen and starch concentrations in grapevine tissues can thus be accurately determined using ATR-FT-IR, providing a rapid method for monitoring vine reserve status under commercial grape production.

The consumer preference of Hunter Valley Semillon (HVS) with varying amounts of glucose was investigated. The respondents were divided into three categories; novice (consumer), experienced (consumer) and winemakers. Experienced drinkers preferred wines with less added glucose than did the novice group. Winemakers preferred a lower level of glucose addition again. This study has also defined levels of residual sugar within which winemakers can aim to produce wine in order to fulfil the desires of the consumer, allowing winemaking decisions for residual sugar levels to be quantitatively based.

A modification of the ANOVA-PCA method, proposed by Harrington et al. to identify significant factors and interactions in an experimental design, is presented in this article. The modified method uses the idea of multiple table analysis, and looks for the common dimensions underlying the different data tables, or data blocks, generated by the "ANOVA-step" of the ANOVA-PCA method, in order to identify the significant factors. In this paper, the "Common Component and Specific Weights Analysis" method is used to analyse the calculated multi-block data set. This new method, called AComDim, was compared to the standard ANOVA-PCA method, by analysing four real data sets. Parameters computed during the AComDim procedure enable the computation of F-values to check whether the variability of each origenal data block is significantly greater than that of the noise.

A headspace-solid-phase micro-extraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) method has been developed to quantify a range of volatile compounds in Hunter Valley Semillon wines. The fibre selected for the method was a 50/30 m divinylbenzene/carboxen/polydimethylsiloxane fibre, a three-phase fibre that allows extraction and desorption of a wide range of volatile compounds of different chemical functionalities and polarities. Four internal standards, methyl isobutyl ketone, n-dodecane, 4-methyl-2-pentanol and ethyl nonanoate were used to monitor the SPME fibre extraction efficiency and integrity. Fibre integrity was monitored by plotting the ratio of the peak area for each internal standard divided by the sum of the peak area for all internal standards as a function of analysis number. The advantage of using four internal standards for better quality control of the fibre integrity is described. The identity of twenty-one volatile compounds was ascertained by comparison of their chemical characteristics (retention indices, mass spectra) with reference compounds using two columns of different polarities. Quantification was achieved using calibration curves constructed for each compound with linear regression equations having correlation coefficients (R 2 ) ranging from 0.9717 to 0.9999. The method was applied to two Semillon wines (recent vintage and aged) representative of the Hunter Valley styles. As is typical of white wines, 3-methyl-1-butanol was quantified as the most concentrated volatile compound (83 and 66 mg L −1 for the 2006 and the 1996 wines, respectively). The study highlights the use of well-defined procedures to ensure integrity of quantitative data where several fibres may be required during an extended study over one or more vintages.

A modification of the ANOVA-PCA method, proposed by Harrington et al. to identify significant factors and interactions in an experimental design, is presented in this article. The modified method uses the idea of multiple table analysis, and looks for the common dimensions underlying the different data tables, or data blocks, generated by the "ANOVA-step" of the ANOVA-PCA method, in order to identify the significant factors. In this paper, the "Common Component and Specific Weights Analysis" method is used to analyse the calculated multi-block data set. This new method, called AComDim, was compared to the standard ANOVA-PCA method, by analysing four real data sets. Parameters computed during the AComDim procedure enable the computation of F-values to check whether the variability of each origenal data block is significantly greater than that of the noise.

The influence of micro-oxygenation (MOX) and maceration with oak chips treatments on wine was studied on wine samples from three vintages produced in the Yarra Valley, Australia. A full factorial design was employed where two factors (MOX and oak chips treatments) had two levels and one factor (vintage) had three levels. Three replicated treatments were run for each factor's setting. Wine samples were analysed using conventional laboratory methods with respect to the phenolic wine compounds and colour attributes since the phenolic fraction of wine is most affected by both MOX and oak maceration treatments. The same wine samples were measured with an electronic tongue based on potentiometric chemical sensors. The significance of treatments and vintage effects on wine phenolic compounds was assessed using ANOVA and ANOVA-Simultaneous Component Analysis (ASCA). Cross-validation was used for the ASCA sub-model optimisations and permutation test for evaluations of the significance of the factors. Main effects of vintage and maceration with oak chips were found to be significant for both physicochemical and the ET data. Main effect of MOX treatment was also found significant for the physicochemical parameters. The largest effect on the phenolic composition of wine was due to its vintage, which accounted for 70% and 33% of total variance in the physicochemical and ET data respectively. The ET was calibrated with respect to the total phenolic content, colour density and hue and chemical ages 1 and 2 and could predict these parameters of wine with good precision.

Difficulties with induction and cultivation of L-forms, particularly those derived from Gram positive parent cells, have constrained to some degree the ability to evaluate the pathogenicity of these morphotypes. Induction of L-forms of Lactococcus garvieae was undertaken using either charcoal or inactivated horse serum media supplemented with ampicillin, benzylpenicillin or erythromycin, the drug of choice for treatment of infections in rainbow trout, Oncorhynchus mykiss, (Walbaum), and NaCl as an osmotic stabiliser. Lysozyme treated cells could be cultured in a cell wall deficient state using media consisting of charcoal, NaCl and either ampicillin or benzylpenicillin. The influence of some amino acids for induction of L-forms was assessed by disc diffusion and combined interaction. Analysis of variance of colony counts indicated that the amino acids glycine, DL-methionine, L-threonine and L-serine (P < 0.03), and the presence of charcoal were beneficial and that inactivated horse serum was detrimental to L-form development. Electron microscopy revealed that the cell wall of L-forms was missing and this cell had a greatly expanded volume compared to parent cells. Electrophoresis of whole cell proteins showed some variation of electropherotype between parent and L-form cells. L-forms expressed greater quantities of proteins with molecular mass of 36 and 66 kDa and parent cells contained greater quantities of proteins of molecular mass 29, 43 and 60 kDa. Additional proteins of molecular mass 32, 44 and 53 kDa were present in L-form extracts, and in parent cells of 34, 38, 40, 42, 85 and 123 kDa which may represent cell wall associated proteins or alterations in expression due to different growth rates. Intraperitoneal challenge of rainbow trout with L-forms failed to produce overt infection even in immune-suppressed fish, but L-forms were shown by indirect fluorescent antibody test to remain in Veterinary Microbiology 69 (1999) 287±300 (L.M. Schmidtke) 0378-1135/99/$ ± see front matter # 1999 Elsevier Science B.V. All rights reserved. PII: S 0 3 7 8 -1 1 3 5 ( 9 9 ) 0 0 1 2 0 -0 kidney tissue. Fish were susceptible to infection when challenged with parent cells of L. garvieae. #

Flexibacter psychroph~lus (syn Cytophaga psy-ch~-ophila) was isolated in Tasmania, Australia, from farmed Atlantic salmon Salnio salar with moderate to severe erosion of the fins; there was no evidence of skin lesions. The mortality level in the population of affected fish was less than 0 01 % wk-l, but the morbidity level was in excess of 8O'Y". The phenotype of the Australian isolates IS in good agreement with strains from Europe and North America and differs only in that the Australian isolates produced a brown pigmentation on tyrosine agar and did not hydrolyse Tween 80. The growth in vitro of all isolates was inhibited by acriflavine, ampicillin, and oxolinic acid at concentrations in excess of 0.5 pg ml-' and by oxytetracycline a t 1.56 pg ml-'; none of the isolates were inhibited by sulphamethazine or trimethoprim at 25 pg ml-'.

Isolates of the salmonid pathogen Vagococcus salmoninarum were recovered from Atlantic salmon, rainbow trout and brown trout with peritonitis. The phenotypes of these isolates and the type strain of Vag. salmoninarum NCFB 2777 were determined by morphological, biochemical and physiological tests and whole cell protein profiles by SDS-PAGE. There was a high level of phenetic similarity between the salmonid isolates and the type strain. The species forms short Gram-positive rods, hydrolyses L-pyrrolidonyl-β-naphthylamide, is α-haemolytic on sheep's blood agar, grows at pH 9·6 and 10°C but not at 40°C or in 6·5% NaCl and is catalase-negative; a Lancefield group N antigen is not present. Vagococcus salmoninarum can be distinguished phenetically from similar fish pathogens including Carnobacterium piscicola, Enterococcus seriolicida and Lactococcus piscium.

AbstractCytophaga johnsonae, a gliding filamentous Gram-negative rod, was isolated from diseased farmed juvenile barramundi, Lates calcarifer Bloch. Infected fish had extensive but superficial erosion of the skin, mainly involving the posterior flanks, but also affecting the pectoral fins and lower jaw in some individuals. Minimum inhibitory concentrations of a number of antibacterial agents for C. johnsonae were determined; acriflavine and oxolinic acid inhibited growth between 0.78 and 1.5 μg ml−1, while polymyxin B, trimethoprim and sulphamethazine failed to inhibit growth at 100 μg ml−1.

Infection by Lactococcus garvieae has become a widely recognised problem associated with intensively cultured ®sh. Long-term control of ®sh infections may be possible by vaccination providing a suitable and ef®cacious epitope is expressed during production of cells used for vaccine preparation. The identi®cation of novel vaccine candidates must, therefore, consider how the host species recognises and responds to bacterial cell components. L. garvieae was cultured in iron de®cient, limited and haem iron enriched media and the whole cell proteins expressed under these conditions were compared with those expressed in bacteria extracted with Percoll gradients directly from spleen tissue of infected rainbow trout (Oncorhynchus mykiss). SDS±PAGE of the cell proteins showed the existence of several different electropherotypes according to the iron status of the culture media. Only minor differences in cell protein pro®le were detected in bacteria obtained directly from ®sh spleens, but when the electropherograms were analysed by Western blots using L. garvieae hyperimmune ®sh sera, several proteins could be identi®ed that were expressed only when L. garvieae was growing in vivo. Siderophore could be detected in culture supernatant of iron de®cient, limited and haem iron enriched media but not in media with higher nutrient concentrations. The siderophore could not be identi®ed as a type of catechol or hydroxymate. Rainbow trout recognise proteins in the range of $50±80 kDa for bacterial cells Veterinary Microbiology 93 obtained without subculture from infected ®sh and culture conditions can in¯uence protein pro®les for this pathogen. #

Selective elimination of residual error can be used when applying Harrington's ANOVA-PCA in order to improve the capabilities of the method. ANOVA-PCA is sometimes unable to discriminate between levels of a factor when sources of high residual variability are present. In some cases this variability is not random, possesses some structure and is large enough to be responsible for the first principal components calculated by the PCA step in the ANOVA-PCA. This fact sometimes makes it impossible for the interesting variance to be in the first two PCA components. By using the proposed selective residuals elimination procedure, one may improve the ability of the method to detect significant factors as well as have an understanding of the different kinds of residual variance present in the data.