Numerical analysis of electrophoretic protein patterns of 'Achromobacter'group B, E and F strains from human blood

Thirty-two clinical strains representing ' Achromobacter 'groups B, E and F were characterized by one-dimensional SDS-PAGE of cellular proteins. All the strains were isolated from blood samples from hospital patients in the United Kingdom. The protein patterns, which contained 40 to 45 discrete bands, were highly reproducible and were used as the basis for a numerical analysis which included all the protein bands. The 32 ' Achromobacter ' strains formed two clusters at the 77% S level. The first, phenon 1, included the 28 group B and the two group E strains and the second, phenon 2, contained the two strains of group F. The strains in each phenon were characterized by a clearly distinct pattern of protein bands. Phenon 1 could be further divided at the 87% S level into three subphenons which correlated with differences in the principal bands found between 40.0 and 42.5 kD. Strains of group E clustered with group B strains from which they could not be distinguished by protein patterns. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides a useful method for the classification of this group of bacteria. Reference strains of each of the PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia rustigianii strains from human diarrhoea and other sources

Twenty strains of Providencia rustigianii (including the type strain of Prov. friedericiana ) have been characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 12 strains (almost exclusively associated with the intestinal tract) from humans, plus eight largely from the intestinal tract of pig, penguin and environmental sources. The protein patterns, which contained 45–50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 20 Prov. rustigianii strains formed six clusters at the 88% S level. One of these clusters included the type strains of both Prov. friedericiana and Prov. rustigianii , thereby confirming the synonymy of these two species. In the second analysis, the principal protein bands were excluded. At the 86% S level the 20 Prov. rustigianii strains formed a single cluster, whilst a field strain of Morganella morganii and the respective type strains of three other Providencia species remained unclustered. The total protein pattern of the type strain of Prov. alcalifaciens was very similar to that of Prov. rustigianii phenon 3 and the M. morganii field strain, which indicates that careful biochemical characterization may be necessary to ascribe strains to a species before typing by the PAGE technique. Alternatively, a selective analysis of the protein bands may be used to confirm the identity of the strains, as shown in this study.     

Numerical analysis of electrophoretic protein patterns of Providencia rustigianii strains from human diarrhoea and other sources

Twenty strains of Providencia rustigianii (including the type strain of Prov. friedericiana) have been characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 12 strains (almost exclusively associated with the intestinal tract) from humans, plus eight largely from the intestinal tract of pig, penguin and environmental sources. The protein patterns, which contained 45-50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 20 Prov. rustigianii strains formed six clusters at the 88% S level. One of these clusters included the type strains of both Prov. friedericiana and Prov. rustigianii, thereby confirming the synonymy of these two species. In the second analysis, the principal protein bands were excluded. At the 86% S level the 20 Prov. rustigianii strains formed a single cluster, whilst a field strain of Morganella morganii and the respective type strains of three other Providencia species remained unclustered. The total protein pattern of the type strain of Prov. alcalifaciens was very similar to that of Prov. rustigianii phenon 3 and the M. morganii field strain, which indicates that careful biochemical characterization may be necessary to ascribe strains to a species before typing by the PAGE technique. Alternatively, a selective analysis of the protein bands may be used to confirm the identity of the strains, as shown in this study.     

Numerical analysis of electrophoretic protein patterns of Providencia alcalifaciens strains from human faeces and veterinary specimens

Twenty-five strains of Providencia alcalifaciens from various countries have been characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 15 from human faeces, one from duck faeces, one from a guinea-pig eye and eight from unknown sources. Also included, for reference purposes, were the type strains of three other Providencia species. The protein patterns, which contained 45-50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, in which the principal protein bands (in the 33-40 kD range) were excluded, the 25 Prov. alcalifaciens strains formed, at the 83% S level, a single cluster whilst the three Providencia reference strains remained unclustered. In the second, which included all the protein bands, the 25 Prov. alcalifaciens strains formed 10 clusters at the 85% S level. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov. alcalifaciens. Reference strains of each of the 10 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia alcalifaciens strains from human faeces and veterinary specimens

Twenty-five strains of Providencia alcalifaciens from various countries have been characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 15 from human faeces, one from duck faeces, one from a guinea-pig eye and eight from unknown sources. Also included, for reference purposes, were the type strains of three other Providencia species. The protein patterns, which contained 45–50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, in which the principal protein bands (in the 33–40 kD range) were excluded, the 25 Prov. alcalifaciens strains formed, at the 83% S level, a single cluster whilst the three Providencia reference strains remained unclustered. In the second, which included all the protein bands, the 25 Prov. alcalifaciens strains formed 10 clusters at the 85% S level. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov, alcalifaciens . Reference strains of each of the 10 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia rettgeri strains from human faeces, urine and other specimens

Thirty-one strains of Providencia rettgeri (mainly from humans) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries and comprised 14 from urine, eight from faeces, two from bile (plus one from the liver of a sheep), two from sputum, one from an insect pupa and three the sources of which were unknown. Also included, for reference purposes, were the type strains of the four other Providencia species. The protein patterns, which contained 45-50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 31 Prov. rettgeri strains formed 13 clusters at the 84% S level. In the second analysis, in which the principal protein bands (in the 33.3-41.3 kD range) were excluded, 29 of the 31 Prov. rettgeri strains formed a single cluster at the 81% S level, whilst the four Providencia reference strains remained unclustered. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov. rettgeri. Reference strains of each of the 13 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia rettgeri strains from human faeces, urine and other specimens

Thirty-one strains of Providencia rettgeri (mainly from humans) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries and comprised 14 from urine, eight from faeces, two from bile (plus one from the liver of a sheep), two from sputum, one from an insect pupa and three the sources of which were unknown. Also included, for reference purposes, were the type strains of the four other Providencia species. The protein patterns, which contained 45–50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 31 Prov. rettgeri strains formed 13 clusters at the 84% S level. In the second analysis, in which the principal protein bands (in the 33.3–41.3 kD range) were excluded, 29 of the 31 Prov. rettgeri strains formed a single cluster at the 81% S level, whilst the four Providencia reference strains remained unclustered. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov. rettgeri. Reference strains of each of the 13 PAGE types identified are available from NCTC for inclusion in future studies.     

Typing of Proteus mirabilis from clinical sources by computerized analysis of electrophoretic protein patterns

B. HOLMES, M. COSTAS AND A.C. WOOD. 1991. Seventy-five strains of Proteus mirabilis (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 24 were from urine, 17 from faeces (including one from a dog and one from a monkey), 12 from the respiratory tract (including nine from sputum), four from a cerebral abscess, 12 from miscellaneous sources and six from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of the 75 P. mirabilis cultures plus those of the type strains of six Proteus and Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the P. mirabilis cultures formed nine clusters at the 85% S level. In the second analysis, in which the principal protein bands (in the 34.0–44.6 kDa range) were excluded, 74 of the 75 cultures of P. mirabilis formed a single cluster at the 77% S level distinct from the six Proteus and Providencia reference strains. The P. mirabilis strain which failed to cluster with the others had a background band pattern typical of the species in the lower molecular weight region but appeared to be less typical in the heavier bands. It is concluded that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of P. mirabilis. Reference strains of each of the nine PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Morganella morganii strains from faeces, wound, urine and other clinical sources

Sixty-five strains of Morganella morganii (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 13 were from stools (including one from a toucan), 13 from wounds, 11 from urine, five from blood (including one from a snake), five from the respiratory tract (four sputum, one lung), 12 from miscellaneous sources and six from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of 67 M. morganii cultures plus those of the type strains of seven Proteus and Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the M. morganii strains formed 21 clusters at the 91% S level. In the second analysis, in which the principal protein bands (in the 31.6—43.2 kDa range) were excluded, the 67 M. morganii cultures formed a single cluster at the 80% S level distinct from the seven Proteus and Providencia reference strains. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of M. morganii . Reference strains of each of the 21 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia stuartii strains from urine, wound and other clinical sources

Eighty-six strains of Providencia stuartii (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 52 were from urine, 11 from wounds, five from blood (one of these also from urine), four from ear infections, two each from faeces and sputum, one from 'alimentation'and nine from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of 46 Prov, stuartii strains (selected to represent the full range of protein pattern diversity) plus those of the type strains of the four other Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the Prov. stuartii strains formed 13 clusters at the 88% S level. In the second analysis, in which the principal protein bands (in the 33.8–40.7 kDa range) were excluded, 45 of the 46 Prov. stuartii strains formed a single cluster at the 82% S level, whilst the four Providencia reference strains remained unclustered. The 40 strains of Prov. stuartii not included in the cluster analysis were assigned to a protein type by calculating their similarity with the strains in the database used for the cluster analysis. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov. stuartii . Reference strains of each of the 13 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Providencia stuartii strains from urine, wound and other clinical sources

Eighty-six strains of Providencia stuartii (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 52 were from urine, 11 from wounds, five from blood (one of these also from urine), four from ear infections, two each from faeces and sputum, one from 'alimentation' and nine from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of 46 Prov. stuartii strains (selected to represent the full range of protein pattern diversity) plus those of the type strains of the four other Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the Prov. stuartii strains formed 13 clusters at the 88% S level. In the second analysis, in which the principal protein bands (in the 33.8-40.7 kDa range) were excluded, 45 of the 46 Prov. stuartii strains formed a single cluster at the 82% S level, whilst the four Providencia reference strains remained unclustered. The 40 strains of Prov. stuartii not included in the cluster analysis were assigned to a protein type by calculating their similarity with the strains in the database used for the cluster analysis. We conclude that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of Prov. stuartii. Reference strains of each of the 13 PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Group EF-4 bacteria, predominantly from dog-bite wounds of humans

Thirty-seven strains of Group EF-4 bacteria (from various countries) were characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 21 from dog-bite wounds of humans, three from cat-bite wounds of humans and five from human limb wounds which may have been inflicted by dogs or cats; there was also one each from a pet monkey, a tiger lung (fatal), a dog tonsil, a mouse, a cat liver, a wallaby mandible, a human vagina and one from a human limb wound which was apparently not inflicted by an animal. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible and were used as the basis for three numerical analyses. In the first, in which the principal protein bands (in the 34.8 to 41.3 kD range) were excluded, the 37 Group EF-4 strains formed, at the 62% S level, two major clusters corresponding to strains producing a dihydrolase for arginine and those not doing so. In the second analysis, which included all the protein bands and which was performed only on the 22 arginine-positive strains, two phenons formed (one of which could be further divided into two sub-phenons) at the 56% S level. The third analysis, also based on all the protein bands, divided the 15 arginine-negative strains into three clusters at the 56% S level. We conclude that high resolution PAGE combined with computerised analysis of protein patterns correlates exactly with the separation of Group EF-4 into two biovars (also with the distinction of the biovars on the basis of G + C content). Reference strains of each of the PAGE types identified are available from the NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Group EF-4 bacteria, predominantly from dog-bite wounds of humans

Thirty-seven strains of Group EF-4 bacteria (from various countries) were characterized by one-dimensional SDS-PAGE of cellular proteins. They comprised 21 from dog-bite wounds of humans, three from cat-bite wounds of humans and five from human limb wounds which may have been inflicted by dogs or cats; there was also one each from a pet monkey, a tiger lung (fatal), a dog tonsil, a mouse, a cat liver, a wallaby mandible, a human vagina and one from a human limb wound which was apparently not inflicted by an animal. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible and were used as the basis for three numerical analyses. In the first, in which the principal protein bands (in the 34.8 to 41.3 kD range) were excluded, the 37 Group EF-4 strains formed, at the 62% S level, two major clusters corresponding to strains producing a dihydrolase for arginine and those not doing so. In the second analysis, which included all the protein bands and which was performed only on the 22 arginine-positive strains, two phenons formed (one of which could be further divided into two sub-phenons) at the 56% S level. The third analysis, also based on all the protein bands, divided the 15 arginine-negative strains into three clusters at the 56% S level. We conclude that high resolution PAGE combined with computerised analysis of protein patterns correlates exactly with the separation of Group EF-4 into two biovars (also with the distinction of the biovars on the basis of G+C content). Reference strains of each of the PAGE types identified are available from the NCTC for inclusion in future studies.     

Numerical analysis of electrophoretic protein patterns of Campylobacter laridis and allied thermophilic campylobacters from the natural environment

Twenty-one strains comprising Campylobacter laridis (nine), nalidixic acid sensitive campylobacters (NASC) (four), and urease-positive thermophilic campylobacters (UPTC) (eight) were characterized by one-dimensional SDS-PAGE of cellular proteins. The UPTC and NASC strains included six from river water, two from mussels and four from sea water. The type strains of three other Campylobacter species were included for reference. The protein patterns, which contained 45-50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 21 strains formed nine clusters at the 80% similarity (S) level. The typical C. laridis strains were restricted to two phenons (2 and 5); the atypical strains being distributed among the remaining phenons. In the second analysis, which excluded the principal protein bands (40-48.5 kD range), the 21 strains formed five clusters at the 80% S level. The typical C. laridis strains were relatively homogeneous and fell into a single phenon (2) within which two subgroups were discernable. The atypical strains were more heterogeneous with respect to background protein pattern, with representatives appearing in all five phenons. An electropherotyping scheme comprising six electropherotypes, and based on both analyses is proposed. The high within-group S level and separation from reference strains of Campylobacter in the second analysis, suggested that UPTC and NASC strains belonged within C. laridis possibly as biovars.     

Numerical analysis of electrophoretic protein patterns of Campylobacter laridis and allied thermophilic campylobacters from the natural environment

Twenty-one strains comprising Campylobacter laridis (nine), nalidixic acid sensitive campylobacters (NASC) (four), and urease-positive thermophilic campylobacters (UPTC) (eight) were characterized by one-dimensional SDS-PAGE of cellular proteins. The UPTC and NASC strains included six from river water, two from mussels and four from sea water. The type strains of three other Campylobacter species were included for reference. The protein patterns, which contained 45–50 discrete bands, were highly reproducible and were used as the basis for two numerical analyses. In the first, which included all the protein bands, the 21 strains formed nine clusters at the 80% similarity (S) level. The typical C. laridis strains were restricted to two phenons (2 and 5); the atypical strains being distributed among the remaining phenons. In the second analysis, which excluded the principal protein bands (40–48.5 kD range), the 21 strains formed five clusters at the 80% S level. The typical C. laridis strains were relatively homogeneous and fell into a single phenon (2) within which two subgroups were discernable. The atypical strains were more heterogeneous with respect to background protein pattern, with representatives appearing in all five phenons. An electropherotyping scheme comprising six electropherotypes, and based on both analyses is proposed. The high within-group S level and separation from reference strains of Campylobacter in the second analysis, suggested that UPTC and NASC strains belonged within C. laridis possibly as biovars.     

Typing of Proteus mirabilis from clinical sources by computerized analysis of electrophoretic protein patterns.

Seventy-five strains of Proteus mirabilis (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 24 were from urine, 17 from faeces (including one from a dog and one from a monkey), 12 from the respiratory tract (including nine from sputum), four from a cerebral abscess, 12 from miscellaneous sources and six from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of the 75 P. mirabilis cultures plus those of the type strains of six Proteus and Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the P. mirabilis cultures formed nine clusters at the 85% S level. In the second analysis, in which the principal protein bands (in the 34.0-44.6 kDa range) were excluded, 74 of the 75 cultures of P. mirabilis formed a single cluster at the 77% S level distinct from the six Proteus and Providencia reference strains. The P. mirabilis strain which failed to cluster with the others had a background band pattern typical of the species in the lower molecular weight region but appeared to be less typical in the heavier bands. It is concluded that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of P. mirabilis. Reference strains of each of the nine PAGE types identified are available from NCTC for inclusion in future studies.     

Numerical analysis of PAGE protein patterns and the taxonomic relationships within the 'Mycoplasma mycoides cluster'

Twenty-six isolates belonging to the 'Mycoplasma mycoides cluster' have been characterized by one-dimensional SDS-PAGE of their cellular proteins. A numerical classification based on the resulting patterns and using a correlation coefficient revealed four distinct phenons at a similarity (S) level of 70%, comprising: (a) bovine group 7 strains; (b) M. capricolum and F38-like strains; (c) M. mycoides subsp. capri and LC strains ('subsp. mycoides'); (d) M. mycoides subsp. mycoides (SC). At the 75% S level, they could be divided further to give eight phenons. The composition of the clusters at both levels was in good agreement with their previous classification, except for M. mycoides subsp. mycoides LC and M. mycoides subsp. capri, which were clustered in a single phenon at 70% S and could not be clearly separated at 75% S. We conclude that high-resolution SDS-PAGE, combined with computerized analysis of protein patterns, provides an extremely effective approach to the investigation of taxonomic relationships within this group of mycoplasmas.     

Analysis of the multiple forms of Gaucher spleen sphingolipid activator protein 2.

Gaucher spleen sphingolipid activator protein 2 was fractionated into concanavalin A binding- and non-binding fractions. These fractions each contained several bands on non-denaturing polyacrylamide gel electrophoresis (PAGE). The two fractions were further fractionated by electroblotting the proteins from preparative gels onto nitrocellulose, staining with Ponceau S to locate the bands of protein and then eluting the protein components from the nitrocellulose. A total of ten fractions, each containing only one or two major components, was collected. All of these subfractions activated beta-glucocerebrosidase and sphingomyelinase and most subfractions also activated beta-galactocerebrosidase. The structural relationship of the bands was investigated using endoglycosidase digestions. The results indicated that the two bands with the fastest mobility on non-denaturing PAGE did not contain any carbohydrate. The remaining bands showed only limited or partial digestion with endoglycosidase H and endoglycosidase D, but were readily hydrolysed with endoglycosidase F. The products of these digestions included bands with similar mobilities to the non-carbohydrate containing bands.     

Isolation and characterization of moderately halophilic bacteria from fully cured salted cod (bachalao)

A total of 128 Strains of moderately halophilic bacteria were isolated from wet, dry and desalted bachalao (salted codfish) as well as from fresh cod and curing salt. The viable count of these bacteria in fully cured wet and dry bachalao ranged from 10³ to 10⁷ per g. All strains were characterized with 40 phenotypic tests and clustered using the S _SM coefficient and UPGMA linking analysis. The strains clustered into five phena at 75% similarity, with 77 strains in phenon A and 37 in phenon E. Two main colony types, smooth and rough, were observed and correspond to phena A, B and C on one hand and phenon E on the other. These two types seem to represent the dominating bacterial flora in fully cured, wet and dry bachalao, respectively. Representative strains of the smooth colony type were characterized further and found to grow well in 0.1–4.5 mol I⁻¹ NaCl and at 15–37°C. They have not, at present, been assigned to any known bacterial species.     

Characterization of Staphylococcus species by SDS–PAGE of Whole-Cell and Extracellular Proteins

In this study, a total of fifteen staphylococcal strains belonging to different species were characterized by whole-cell and extracellular protein profiles using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE). The results are presented as dendrograms after quantitative analysis of the band patterns with a computer program. Visual inspection of protein bands and cluster analysis of protein patterns of to be used 15 strains, representing 10 Staphylococcus species, showed that whole-cell and extracellular protein profiles differed in several protein bands in Staphylococcus aureus, S. epidermidis, S. simulans, and other species of Staphylococcus; however, the differences were insufficient for reliable differentiation of Staphylococcus species by the SDS–PAGE method.