Utilization of protein profiles for the characterization of halophilic bacteria

Proteins from 24 halophilic bacteria, including Haloarcula marismortui, Haloarcula vallismortis, Haloferax mediteranei, Haloferax gibbonsii, Halobacterium salinarium, as well as unknown isolates from Enid, Oklahoma; Jefferson Island, Louisiana; and the Salado Formation-New Mexico, were analyzed by one-dimensional SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Their protein profiles were compared, and the bacteria were grouped according to the Statistical Analysis System (SAS Institute, Cary, North Carolina) on an IBM 4316 computer. The groupings made from protein profiles showed agreement with groupings made from DNA reassociation data. The various known halophiles were easily separated into the three main halobacterial genera. The data show that one-dimensional SDS-PAGE can be easily used to rapidly screen large numbers of unknown strains to group them into related clusters. This technique offers a way to reduce the total number of halophilic isolates being studied in large taxonomic research programs.     

Electrophoretic characterization of Amaranthus L. seed proteins and its systematic implications

The seed protein profiles of 11 Amaranthus taxa (Amaranthaceae) from Spain were studied. These profiles were evaluated as a chemical character to clarify the taxonomic complexity in the genus. Tricine-sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) profiles of the Amaranthus seed proteins studied showed a range of peptides varying from 64 to 12 kDa, with a larger number of protein bands observed between 25.1 and 12 kDa. For the taxonomic study, 14 bands, some of them subdivided into several isoforms, were considered. The similarity analysis based on the SDS-PAGE profile is a useful character for the discrimination of species in Amaranthus , except for A. cruentus and A. hypochondriacus , for which a hybrid population was found. © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 57–63.     

Differentiation of Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus delbrueckii subsp. lactis by SDS-PAGE of cell-wall proteins

AIMS: In the present study, a method based on SDS-PAGE fingerprinting of surface layer proteins was developed to identify Lactobacillus delbrueckii subsp. bulgaricus and subsp. lactis dairy isolates. METHODS AND RESULTS: The two subspecies, identified by species-specific PCR, were characterized by different SDS-PAGE cell-wall protein profiles; subspecies bulgaricus showed one band of about 31 kDa which, in some cases, was observed at a doublet, and subspecies lactis showed one band of about 21 kDa or 18 kDa. CONCLUSION: The sensitivity of this procedure for discriminating between the two subspecies was very high. The different types of SDS-PAGE profile for cell-wall proteins of the strains studied in this work did not seem to be correlated to the different dairies of origin. SIGNIFICANCE AND IMPACT OF THE STUDY: The method appears to be an efficient taxonomic tool. It has the advantage of easy gel interpretation over fingerprinting of whole-cell protein extracts, and may be used as an alternative to established PCR-based techniques which, though rapid and safe, require expensive instruments and reagents.     

A polyphasic taxonomic study of Chryseobacterium strains isolated from dairy sources

A polyphasic taxonomic study, employing protein electrophoresis (SDS-PAGE), gas chromatographic analysis of cellular fatty acids (FAME), mol% G+C determination and DNA-DNA hybridizations, was undertaken on 103 dairy isolates shown to belong to Chryseobacterium. Reference strains of the Chryseobacterium species, CDC group IIb and Embedobacter brevis were included. SDS-PAGE analysis yielded good differentiation between the investigated species. About half of the strains could be clustered into nine major groups while the other half occupied a separate position. With FAME analysis no clear differentiation of the Chryseobacterium species (except C. meningosepticum) and SDS-PAGE groups could be achieved. FAME analysis, however, gave good differentiation between the Chryseobacterium and Empedobacter strains. The mol% G+C of the isolates tested, ranged between 36.4 and 39.0. The combination of SDS-PAGE and DNA-DNA hybridization identified a large group of dairy isolates as C. indologenes, one isolate as C. gleum and two new genotypic groups, comprising five and 15 dairy isolates respectively, emerged from the polyphasic study. Another large part of strains have a separate or uncertain position in Chryseobacterium and remained classified as Chryseobacterium species CDC group IIb.     

Differentiation of Lactobacillus isolates from infant faeces by SDS-PAGE and rRNA-targeted oligonucleotide probes

Isolates of lactobacilli from infant faeces phenotypically characterized as Lactobacillus paracasei subsp. paracasei (six strains), Lact. rhamnosus (six strains), Lact. gasseri (three strains), Lact. acidophilus (one strain) and Lact. fermentum/reuteri (three strains) according to recent classification systems were subjected to SDS-PAGE of whole cell proteins and rRNA-targeted oligonucleotide probe hybridization, in order to confirm the phenotypic characterization and elucidate the exact taxonomic position of the three strains that had properties between fermentum and reuteri. Results suggested a good agreement between the phenotypic characterization, SDS-PAGE and rRNA-targeted oligonucleotide probe hybridization for strains of all species except for the Lact. fermentum/reuteri strains. Results obtained by rRNA probes suggested a possible phylogenetic relatedness of the strains to Lact. reuteri. Isolates from infant faeces with interesting probiotic properties could be used as components of fermented milk products.     

Identification of Paenibacillus larvae to the subspecies level: an obstacle for AFB diagnosis

This study was initially aimed at developing a PCR-test to differentiate between the pathogenic agent of American foulbrood (Paenibacillus larvae subsp. larvae) and powdery-scale disease (P. larvae subsp. pulvifaciens) of the honeybee. The test was based on the "insert of clone 9" (iC9), referring to a cloned 1.9 kB HaeIII fragment that occurs only in the P. larvae subsp. larvae reference strains and possibly correlates with American foulbrood virulence. It was shown that an iC9-based PCR-test discriminates between the BCCM/LMG reference strains of both subspecies. However, the screening of 179 Belgian field strains revealed five isolates that gave no iC9-based amplicon, thus rather resembling to P. larvae subsp. pulvifaciens. In addition, they all produced acid from mannitol, a characteristic previously assigned to the pulvifaciens subspecies. Because the reference strains gave conflicting data, this carbohydrate acidification was not conclusive. Therefore, the exact taxonomic position of the five retained strains was determined by a polyphasic approach using SDS-PAGE, AFLP, and ERIC-based PCR. Four iC9-negative field strains could be identified as P. larvae subsp. larvae; the taxonomic position of the fifth field strain remained ambiguous. The latter was provisionally classified as a subspecies pulvifaciens strain on the basis of SDS-PAGE. The present paper demonstrates the existence of field strains that do not fit well in the subdivision of the species P. larvae into two subspecies. Knowing that only one of both subspecies represents the pathogenic agent of AFB, this is a serious obstacle for the diagnosis of this honeybee disease.     

Description and partial characterization of a new chlamydia-like microorganism

Abstract A new obligate intracellular bacterium which we called 'Z' was isolated as a cell culture contaminant of unknown origin. The organism grew in a variety of cultured cells with a 5–7-day developmental cycle, within cytoplasmic phagosomes, similarly to Chlamydia and some Rickettsia spp. Two alternating developmental forms (elementary bodies and reticulate bodies) were observed by electron microscopy. SDS-PAGE electrophoresis, immunoblotting with chlamydia-specific antibodies, and polymerase chain reaction using chlamydial genus specific primers provided evidence that our bacterium differs significantly from chlamydiae. Further characterization of 'Z' including determination of 16S ribosomal RNA sequences will allow its taxonomic position to be established.     

五种啮齿类动物核型和血清蛋白质的SDS-聚丙烯酰胺凝胶电泳分析

本文对5种啮齿类动物(岩松鼠、花鼠、五趾跳鼠、棕背(鼠平)和岢岚绒鼠)的核型及其血清蛋白质的SDS-聚丙烯酰胺凝胶电泳进行了分析。通过分析结果,探讨了它们的核型及血清蛋白在其分类中的意义。     

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.     

Evaluation of biochemical and serological methods to identify and clustering yeast cells of oral Candida species by CHROMagar test, SDS-PAGE and ELISA.

The purpose of this work was to evaluate biochemical and serological methods to characterize and identify Candida species from the oral cavity. The strains used were five Candida species previously identified: C. albicans, C. guilliermondii, C. parapsilosis, C. krusei, C. tropicalis, and Kluyveromyces marxianus, as a negative control. The analyses were conducted through the SDS-PAGE associated with statistical analysis using software, chromogenic medium, and CHROMagar Candida (CA), as a differential medium for the isolation and presumptive identification of clinically important yeasts and an enzyme-linked immunoabsorbent assay (ELISA), using antisera produced against antigens from two C. albicans strains. This method enabled the screening of the three Candida species: C. albicans, C. tropicalis, and C. krusei, with 100% of specificity. The ELISA using purified immunoglobulin G showed a high level of cross-reaction against protein extracts of Candida species. The SDS-PAGE method allowed the clustering of species-specific isolates using the Simple Matching coefficient, S(SM) = 1.0. The protein profile analysis by SDS-PAGE increases what is known about the taxonomic relationships among oral yeasts. This methodology showed good reproducibility and allows collection of useful information for numerical analysis on information relevant to clinical application, and epidemiological and systematical studies.     

Computer Program Designed to Follow Fluctuations in Microbial Populations and Its Application in a Study of Chesapeake Bay Microflora

A computer program has been developed which performs cluster analysis of microorganisms using methods of numerical taxonomy. The program is designed to group related strains, identify the groups by reference to known strains, and calculate a hypothetical median organism (HMO) for each group. The HMO serves to condense taxonomic information and provides a tag for each strain cluster. Every strain in a group is compared with the HMO established for that group. A representative strain for the group is obtained by selection of the strain showing highest similarity to the HMO. New data sets can be compared with data sets of previous analyses. Hence, the occurrence of the same taxonomic groups within separate data sets can be determined. Quantitative or qualitative differences in distribution of taxonomic groups within or between data sets can be measured. The output from the computer is a graphical display, using an on-line plotter; thus, the investigator is provided with visual comparison of data sets. Results obtained from a study applying the computer program in an analysis of taxonomic data obtained for 43 bacterial strains isolated from Chesapeake Bay indicate the usefulness of this method of taxonomic analysis in microbial ecology.     

A taxonomic analysis of seed proteins inPinus spp.(Pinaceae)

Seed storage proteins have proved to be a powerful biochemical marker for taxonomic research, but they have not been extensively employed in forest tree studies. In order to improve the understanding of the taxonomy of the genusPinus, total seed proteins of 12 pine species have been analyzed by means of SDS-PAGE (Sodium dodecylsulphate-polyacrylamide gel electrophoresis). The results showed the presence, in the genusPinus, of two main sub-taxa, corresponding to the subgeneraHaploxylon andDiploxylon. Differences and affinities between Mediterranean pine species were found in agreement with classification ofKlaus (1989).     

The oligomeric state of c rings from cyanobacterial F-ATP synthases varies from 13 to 15

We isolated the c rings of F-ATP synthases from eight cyanobacterial strains belonging to four different taxonomic classes (Chroococcales, Nostocales, Oscillatoriales, and Gloeobacteria). These c rings showed different mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), probably reflecting their molecular masses. This supposition was validated with the previously characterized c(11), c(14), and c(15) rings, which migrated on SDS-PAGE in proportion to their molecular masses. Hence, the masses of the cyanobacterial c rings can conveniently be deduced from their electrophoretic mobilities and, together with the masses of the c monomers, allow the calculation of the c ring stoichiometries. The method is a simple and fast way to determine stoichiometries of SDS-stable c rings and hence a convenient means to unambiguously determine the ion-to-ATP ratio, a parameter reflecting the bioenergetic efficacy of F-ATP synthases. AFM imaging was used to prove the accuracy of the method and confirmed that the c ring of Synechococcus elongatus SAG 89.79 is a tridecameric oligomer. Despite the high conservation of the c-subunit sequences from cyanobacterial strains from various environmental groups, the stoichiometries of their c rings varied between c(13) and c(15). This systematic study of the c-ring stoichiometries suggests that variability of c-ring sizes might represent an adaptation of the individual cyanobacterial species to their particular environmental and physiological conditions. Furthermore, the two new examples of c(15) rings underline once more that an F(1)/F(o) symmetry mismatch is not an obligatory feature of all F-ATP synthases.     

A reinvestigation into the taxonomic status of Riccia himalayensis and Riccia discolor based on scanning electron microscopic and chemotaxonomical studies

Riccia himalayensis St. (Ms.) Kashyap is a common species found in the Himalayas from the foot of the mountains to an altitude of 2743 m. Riccia discolor Lehm. et Lindb. on the other hand, is the common and most widely distributed monsoon species, that has been reported from both the western and eastern Himalayas, as well as from the plains of Assam, west Bengal, Madhya Pradesh, south India, etc. Dispute still exists regarding the taxonomic status of R. himalayensis , which is considered a synonym of R. discolor by some taxonomists, according to whom R. himalayensis has no specific status. This has led to a lot of errors in interpreting R. discolor . On the other hand many authors have considered R. himalayensis to be a composite species of R. discolor , R. billardieri and R. gangetica .
The present investigation was aimed to reinvestigate the taxonomic status of R. himalayensis and R. discolor based on morphological analysis of thallus and spore surface by light microscope and scanning electron microscopy (SEM) studies and biochemical analyses of the protein profile by SDS-PAGE. Results reveal that although the two species share certain common morphological features, they are two distinct species chemotaxonomically.     

Biochemical investigation of the Rana esculenta complex in the Kis-Balaton Nature Reserve, Hungary

In this study it has been proved by PAGE of serum proteins that all the three members of the Rana esculenta complex occur in the Kis-Balaton Nature Reserve (Hungary). On the basis of the LDH isoenzyme pattern which is characteristic in green frogs we could distinguish all three variations of R. ridibunda and R. lessonae and one type of R. esculenta. The mobility of serum albumins on SDS-PAGE implies that the R. esculenta comes from hybridization of the two other species. The PAGE methods provide a reliable basis for the rapid taxonomic identification of both adults and immature speciments of the three forms of frogs.     

HICLAS: a taxonomic database system for displaying and comparing biological classification and phylogenetic trees

MOTIVATION: Numerous database management systems have been developed for processing various taxonomic data bases on biological classification or phylogenetic information. In this paper, we present an integrated system to deal with interacting classifications and phylogenies concerning particular taxonomic groups. RESULTS: An information-theoretic view (taxon view) has been applied to capture taxonomic concepts as taxonomic data entities. A data model which is suitable for supporting semantically interacting dynamic views of hierarchic classifications and a query method for interacting classifications have been developed. The concept of taxonomic view and the data model can also be expanded to carry phylogenetic information in phylogenetic trees. We have designed a prototype taxonomic database system called HICLAS (HIerarchical CLAssification System) based on the concept of taxon view, and the data models and query methods have been designed and implemented. This system can be effectively used in the taxonomic revisionary process, especially when databases are being constructed by specialists in particular groups, and the system can be used to compare classifications and phylogenetic trees. AVAILABILITY: Freely available at the WWW URL: http://aims.cps.msu.edu/hiclas/ CONTACT: pramanik@cps.msu.edu; lotus@wipm.whcnc.ac.cn     

The identification and classification ofAcinetobacter strains exhibiting variations in phosphate accumulation by SDS-PAGE and numerical analysis

The limitation of phosphate concentrations in effluents is of international concern because of the risk of eutrophication.Acinetobacter spp. have often been isolated from activated sludge plants exhibiting enhanced phosphate removal. The ability to remove phosphate from mixed liquor byAcinetobacter isolates and reference strains was, therefore, determined. The ability to remove phosphate was found to vary among theAcinetobacter strains. The taxonomic relationships among these various strains were elucidated by SDS-PAGE and numerical analysis, and the correlation between their taxonomy and phosphate uptake ability was determined. The ability to remove large amounts of phosphate was found to be strain specific rather than species specific. The classification ofA. haemolyticus andA. baumannii as separate species is questioned.     

An evaluation of electrophoresis as a taxonomic method using comparative data from the macropodidae (Marsupialia)

Comparative electrophoretic data collected for seven proteins from thirty-four species of marsupial have been examined to define the advantages and shortcomings of electrophoresis as a taxonomic method. The problems of relative rates of change, convergence and parallelism are examined and the range of taxonomic ranks which can usefully be studied with this technique is determined. The relative merits of this type of data, when combined with various methods of taxonomic analysis, are discussed.     

Studies on the pathophysiological mechanism of Campylobacter jejuni-induced fluid secretion in rat ileum

Abstract Lipopolysaccharide (LPS) patterns were obtained from fast-growing Rhizobium strains after silver staining of proteinase K treated cells lysates, run in SDS-PAGE. The rhizobia came from root nodules of Acacia senegal and Prosopis chilensis , collected in differents part of the Sudan. The LPS profiles of all strains were typical of rhizobia. Two different LPS region with lower and higher electrophoretic mobility (region I and region II, respectively) coulld be dinguished in the gels, and based on the profiles the strains were divided into three groups. Strains isolated from A. senegal showed a wider range of different profiles than strains isolated from P. chilensis , even though the plants belong to the same cross-infection group. Otherwise there was no clear correlation between the taxonomic relatedness or site of isolation of the strains and their LPS profiles.