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PubMed2004

Species specificity in rodent pheromone receptor repertoires.

The mouse V1R putative pheromone receptor gene family consists of at least 137 intact genes clustered at multiple chromosomal locations in the genome. Species-specific pheromone receptor repertoires may partly explain species-specific social behavior. We conducted a genomic analysis of an orthologous pair of mouse and rat V1R gene clusters to test for species specificity in rodent pheromone systems. Mouse and rat have lineage-specific V1R repertoires in each of three major subfamilies at these loci as a result of postspeciation duplications, gene loss, and gene conversions. The onset of this diversification roughly coincides with a wave of Line1 (L1) retrotranspositions into the two loci. We propose that L1 activity has facilitated postspeciation V1R duplications and gene conversions. In addition, we find extensive homology among putative V1R promoter regions in both species. We propose a regulatory model in which promoter homogenization could ensure that V1R genes are equally competitive for a limiting transcriptional structure to account for mutually exclusive V1R expression in vomeronasal neurons.

Animals↗
PubMed1996

Species-specific identification of Candida krusei by hybridization with the CkF1,2 DNA probe.

The species specificity of the Candida krusei DNA fingerprinting probe CkF1,2 has been investigated. A total of 149 pathogenic and nonpathogenic fungal and bacterial DNAs were screened with CkF1,2. The probe was cold labeled with peroxidase, and its specificity was assessed by using Southern blot, dot blot, and colony blot hybridization. Its sensitivity was determined by dot blot hybridization. The CkF1,2 probe proved to be species specific. It hybridized with DNA for the 112 C. krusei strains studied, whereas it failed to hybridize under low-stringency conditions to 37 DNAs from 27 different yeast species, including Candida albicans, Candida glabrata, Candida norvegensis, Candida inconspicua, Candida tropicalis, Candida valida, Candida zeylanoides, and Yarrowia lipolytica, as well as DNAs from the filamentous fungi and bacteria tested. However, CkF1,2 hybridized strongly with DNA of the yeast species Issatchenkia orientalis, the putative ascogenous perfect state of C. krusei. Amounts as small as 60 to 120 ng of C. krusei target DNA were detected by dot blot hybridization with CkF1,2. It permitted the direct screening of colony blots for early identification. The CkF1,2 probe has potential value as a diagnostic reagent for identifying C. krusei.

Blotting, Southern↗
PubMed1997

Rapid identification of pathogenic Leptospira species (Leptospira interrogans, L. borgpetersenii, and L. kirschneri) with species-specific DNA probes produced by arbitrarily primed PCR.

Arbitrarily primed PCR (AP-PCR) assays can be used to discriminate between species of Leptospira. Comparative analysis of the fingerprints obtained from representative sets of serovar reference strains of Leptospira interrogans sensu stricto, L. borgpetersenii, and L. kirschneri and the reference strains of the other Leptospira spp. revealed species-specific DNA fragments. These species-specific sequences were reamplified in order to produce digoxigenin-11-dUTP-labeled genomic DNA probes that could be used to identify Leptospira species. Three probes (specific for L. interrogans sensu stricto, L. borgpetersenii, and L. kirschneri) were selected and tested with 72 representative serovar reference strains, all of which had previously been studied by DNA-DNA hybridization. The two techniques were in general agreement, and hybridization with AP-PCR-derived probes was shown to be a useful approach for rapid species determination of leptospires, without the prior need for DNA sequence information. These nonradioactive probes can be used to identify Leptospira species in nonspecialized laboratories, and this should contribute to a better knowledge of the molecular epidemiology of leptospirosis.

Bacterial Typing Techniques↗
PubMed1984

Species-specific rDNA transcription is due to promoter-specific binding factors.

RNA polymerase I transcription factors were purified from HeLa and mouse L cell extracts by phosphocellulose chromatography. Three fractions from each species were found to be required for transcription. One of these fractions, virtually devoid of RNA polymerase I activity, was found to form a stable preinitiation complex with small DNA fragments containing promoter sequences from the homologous but not the heterologous species. These species-specific DNA-binding factors can explain nucleolar dominance in vivo in mouse-human hybrid somatic cells and species specificity in cell-free, RNA polymerase I-dependent transcription systems. The evolution of species-specific transcriptional control signals may be the natural outcome of a special relationship that exists between the RNA polymerase I transcription machinery and the multigene family coding for rRNA.

Animals↗
PubMed1986

Isolation and characterization of proteolytic fragments of the sea urchin sperm receptor that retain species specificity.

The sea urchin sperm receptor isolated from the eggs of Strongylocentrotus purpuratus is a high molecular weight proteoglycan-like molecule. Previous studies in our laboratory suggested that the sperm receptor has two functional components, glycosaminoglycan chains that are responsible for sperm binding and polypeptide chains that control species specificity in the binding process. We have investigated this idea further by generating fragments of the receptor by limited proteolytic digestion of the egg cell surface. The results of experiments with these receptor preparations support the hypothesis that the species specificity of inhibition of fertilization observed in a competitive bioassay is conferred by the polypeptide portion of the receptor molecule. Studies with various receptor preparations reveal that the presence of at least 30% of the polypeptide by weight is required to inhibit fertilization species specifically. Receptor preparations containing less than 10% protein lack species specificity and inhibit fertilization in both S. purpuratus and Arbacia punctulata.

Animals↗
PubMed1990

Inheritance of species-specific behaviors in the paradise fish (Macropodus opercularis): a diallel study.

Species-specific elements of the paradise fish's ethogram were recorded in one familiar and three different unfamiliar environments, which were designed to model certain features of this species' natural habitat: (1) a densely vegetated home range, (2) a novel open field, (3) a small novel place, and (4) a small novel place with a predator. The inheritance of the behavioral elements was investigated employing a five-times-replicated diallel cross among three inbred strains. A detailed Hayman analysis of variance and a variance-covariance analysis were performed to uncover the genetic architectures of these phenotypes. Additive genetic effects and/or ambidirectional dominance was found to be characteristic of most species-specific behavioral elements studied, suggesting an evolutionary history of stabilizing selection.

Animals↗
PubMed1965

Studies on tissue thromboplastin. II. Species specificity.

Purified brain tissue thromboplastins of human, bovine and chicken origin have been tested on homologous and heterologous plasmas. Purified brain tissue thromboplastin is species specific. Lipid extracts from purified brain tissue thromboplastin prepared with pyridin show a negligable residual species specificity, probably caused by a slight contamination with brain tissue thromboplastin. The activity curves of our lipid extracts differ from typical curves expected for lipid activators by a less distinct inhibition in high concentrations probably caused by a different composition of petrol-ether and pyridin extracts. The protein part of tissue thromboplastin does not activate prothrombin in any system. The protein part of tissue thromboplastin of one species could be combined with the lipid part of another species to form an active tissue thromboplastin. The species specificity of these combinations was determined by the source of protein used.

Animals↗
PubMed1994

Species-specific binding of sperm proteins to the extracellular matrix (zona pellucida) of the egg.

The zona pellucida is an extracellular matrix surrounding the mammalian egg where species-specific gamete recognition and signaling occur. Pig zona pellucida were isolated in large amounts and used as an affinity matrix for detergent-solubilized, biotinylated membrane proteins of pig spermatozoa. On non-reducing SDS-polyacrylamide gel electrophoresis, specifically bound sperm proteins migrated with M(r) 170,000, 150,000, 130,000, 56,000, and 50,000 (p50). Disulfide bond reduction separated each of the M(r) 130,000-170,000 proteins into M(r) 105,000 (p105) and M(r) 45,000 (p45) subunits, indicating that these high M(r) proteins are related. Based on two-dimensional electrophoresis, the M(r) 56,000 band was composed of three to four proteins that migrated with M(r) 56,000-62,000 (p56-62) in the second (reducing) dimension. p50 bound to heterologous zona pellucida (murine, bovine) and to Xenopus laevis oocyte envelopes, demonstrating a lack of species specificity to its binding and was identified as proacrosin/acrosin based on amino acid sequences of two tryptic peptides and its interaction with monospecific antibodies to proacrosin. In contrast, p105/p45 and one or more of the p56-62 proteins bound to pig zona pellucida but not to the egg extracellular matrices of the other species; these proteins therefore exhibited the species-specific binding to the zona pellucida expected for molecules involved in specific gamete adhesion. Amino acid sequences of nine tryptic peptides derived from p105/p45 did not match peptide sequences in existing databases, establishing it as a unique protein. These (p105/p45 and at least one p56-62 protein) are the first sperm membrane proteins to be identified that bind in a species-specific manner to the egg extracellular matrix.

Acrosin↗
PubMedDate not supplied

[Detection of humoral antibodies to species-specific and group ornithosis antigens in the dynamics of the infection].

The time course of antibody production to group- and species-specific antigens of ornithosis agent was studied by complement fixation (CFT) and hemagglutination-inhibition (HI) tests. In rabbits after a single intravenous inoculation of the ornithosis agent, antibody to the homologous species-specific antigen appeared in the peripheral blood 3--5 days after inoculation and reached maximum levels during the 1-st week, and to the group-specific antigen could be detected 3--5 days after inoculation but reached maximum titers only 3--4 weeks later. In practical serological diagnosis of ornithosis and other chlamydia infections, the group antigen is used which gives retrospective diagnosis at 10--14 days or later. The results of the study suggest that the use of CFT with the species-specific antigen will not only differentiate ornithosis from other chlamydial infections but also will permit early serological diagnosis of ornithosis.

Animals↗
PubMed1976

Schistoma mansoni soluble egg antigens: determination of the stage and species specificity of their serologic reactivity by radioimmunoassay.

Three highly purified Schistosoma mansoni egg antigens reactivity with sera from mice with chronic schistosomiasis mansoni (major serologic antigens-MSA1, MSA2, and MSA3) were utilized in radioimmunoassays for determining their stage and species specificity. Inhibition studies revealed that MSA1 was completely stage specific, since its binding to S. mansoni chronic infection serum (CIS) could not be inhibited either with cercarial or adult worm antigen. MAS2 and MSA3 were specific with regard to adult worm antigen but their binding to CIS was slightly inhibited by cercarial antigen. Immature eggs in comparison with mature eggs contained only negligible amounts of MSA1 whereas MSA2 and MSA3 were present in large quantities. Fluid from eggs hatched in spring water (hatch fluid-HF) contained considerable amounts of MSA1 as well as MSA2 and MSA3. With regard to species specificity, MSA1 essentially did not cross-react with crude soluble egg antigen (SEA) from Schistosoma japonicum and Schistosoma haematobium whereas MSA2 and MSA3 showed partial cross-reactivity. Antibody dilution studies with sera from animals and humans infected with heterologous schistosome species (i.e., S. japonicum and S. haematobium) indirectly confirmed the high degree of species specificity of MSA1. The degree of serologic specificity of MSA1 as demonstrated in this study parallels that of the sensitizing activity of S. mansoni eggs and SEA for granuloma formation, as demonstrated in previous investigations. This study, therefore, indicates that MSA1 may be a major immunopathologically active egg antigen. In addition, the high degree of species specificity of MSA1 suggests its use in the serodiagnosis of schistosomiasis mansoni.

Animals↗
PubMed2004

Rapid, species-specific detection of uropathogen 16S rDNA and rRNA at ambient temperature by dot-blot hybridization and an electrochemical sensor array.

Development of rapid molecular approaches for pathogen detection is key to improving treatment of infectious diseases. For this study, the kinetics and temperature-dependence of DNA probe hybridization to uropathogen species-specific sequences were examined. A set of oligonucleotide probes were designed based on variable regions of the 16S gene of the Escherichia coli, Proteus mirabilis, Klebsiella oxytoca, and Pseudomonas aeruginosa. A universal bacterial probe and probes-specific for gram-positive and gram-negative organisms were also included. The oligonucleotide probes discriminated among 16S genes derived from 11 different species of uropathogenic bacteria applied to nylon membranes in a dot-blot format. Significant binding of oligonucleotide probes to target DNA and removal of nonspecific binding by membrane washing could both be achieved rapidly, requiring as little as 10 min. An oligonucleotide probe from the same species-specific region of the E. coli 16S gene was used as a capture probe in a novel electrochemical 16-sensor array based on microfabrication technology. Sequence-specific hybridization of target uropathogen 16S rDNA was detected through horseradish peroxidase acting as an electrochemical transducer via a second, detector probe. The sensor array demonstrated rapid, species-specific hybridization in a time course consistent with the rapid kinetics of the dot-blot hybridization studies. As in the dot-blot hybridization studies, species-specific detection of bacterial nucleic acids using the sensor array approach was demonstrated both at 65 degrees C and at room temperature. These results demonstrate that molecular hybridization approaches can be adapted to rapid, room temperature conditions ideal for an electrochemical sensor array platform.

Bacteria↗
PubMed2004

Determination of methionine and selenomethionine in yeast by species-specific isotope dilution GC/MS.

A method for the simultaneous determination of methionine (Met) and selenomethionine (SeMet) in yeast using species-specific isotope dilution (ID) gas chromatography/mass spectrometry (GC/MS) is described. Samples were digested by refluxing for 16 h with 4 M methanesulfonic acid. Analytes were derivatized with methyl chloroformate and extracted into chloroform for GC/MS analysis. In addition to use of commercially available 13C-enriched Met and SeMet spikes for species specific ID analysis, a 74Se-enriched SeMet spike was also available for comparison of results. In selective ion monitoring mode, the intensities of ions at m/z 221, 222, 269, 270, and 263 were used to calculate the 221/222, 269/270, and 269/263 ion ratios for quantification of Met and SeMet. Concentrations of 5959 +/- 33 and 3404 +/- 12 microg g(-1) (one standard deviation, n = 6) with relative standard deviations of 0.55 and 0.36% for Met and SeMet, respectively, were obtained using 13C-enriched spikes. A concentration of 3417 +/- 8 microg g(-1) (one standard deviation, n = 6) was obtained using the 74Se-enriched SeMet spike. The concentration of SeMet measured in the yeast is equivalent to 66.43 +/- 0.24% of total Se and 30.31 +/- 0.11% of total Met is in the form of SeMet. Method detection limits (three times the standard deviation) of 3.4 and 1.0 microg g(-1) were estimated for Met and SeMet, respectively, based on a 0.25-g subsample of yeast with 1 mL of extract used for derivatization. A similar concentration of 5930 +/- 29 microg g(-1) (one standard deviation, n = 4) for Met and a lower concentration of 2787 +/- 49 microg g(-1) (one standard deviation, n = 4) for SeMet were obtained for this yeast sample using species-specific ID analysis based on GC/MS with 13C-enriched Met and SeMet spikes when a 2-h open microwave digestion approach using 8 M methanesulfonic acid was used.

Carbon Isotopes↗
PubMed1996

Species-specific measurement of the second group of Dermatophagoides mite allergens, Der p 2 and Der f 2, using a monoclonal antibody-based ELISA.

BACKGROUND: The group 2 Dermatophagoides mite allergens, Der p 2 and Der f 2, were known to be highly crossreactive, and previous assays to measure Der p 2 and Der f 2 were not species-specific. OBJECTIVE: The aim of this study was to develop a monoclonal antibody-based ELISA (MoAb-ELISA) to species-specifically measure Der p 2 and Der f 2. METHODS: The MoAb-ELISA for Der p 2 and Der f 2 was performed using species-specific MoAbs for Der p 2 and Der f 2 and a biotinylated second MoAb which recognized a common epitope on both Der p 2 and Der f 2. RESULTS: The assay was highly species-specific, reproducible and sensitive. Thirty-two house dust samples were assayed by the MoAb-ELISA for Der p 2 and Der f 2 and by a previously reported radioimmunoassay for Der 2 with rabbit anti-Der 2 antibodies. The summed values for Der p 2 and Der f 2 by the MoAb-ELISA demonstrated a good correlation with the Der 2 values using the radioimmunoassay (r = 0.978). Furthermore, the proportion of the Der p 2 level in the total Der 2 level (Der p 2 divided by Der p 2 plus Der f 2) correlated well with that of the D. pteronyssinus mite number to the total Dermatophagoides mite number identified by species (r = 0.970). CONCLUSION: The MoAb-ELISA for Der p 2 and Der f 2, as well as that of Der p 1 and Der f 1, will be useful for the standardization of mite extracts and for the assessments of mite allergen exposure.

Animals↗
PubMed1996

Multiplex PCR using conserved and species-specific 16S rRNA gene primers for simultaneous detection of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis.

Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis are strongly associated with periodontitis. However, little is known about their distribution in periodontally healthy individuals, because culturing techniques are not sufficiently sensitive. A modified multiplex PCR was developed to address that question. Our method uses two species-specific forward primers in combination with a single reverse primer. These primers target variable and conserved regions of the 16S rRNA gene. Sensitivity was determined by testing serial dilutions of A. actinomycetemcomitans and P. gingivalis cells. Primer specificity was tested against (i) six A. actinomycetemcomitans strains and four P. gingivalis strains, (ii) seven different species of oral bacteria, and (iii) supra- and subgingival plaque from 20 subjects. The multiplex PCR had a lower limit of detection of 2 A. actinomycetemcomitans and 30 P. gingivalis cells. Species-specific amplicons were obtained for all A. actinomycetemcomitans and P. gingivalis strains tested and did not occur with seven other bacterial species unless A. actinomycetemcomitans and P. gingivalis were added. Neither target species was detected in supragingival plaque; A. actinomycetemcomitans was detected in one subgingival specimen, and P. gingivalis was detected in another. When plaque samples were spiked with 10 A. actinomycetemcomitans cells and 100 P. gingivalis cells, species-specific amplicons were detected. These findings show our multiplex PCR to be highly sensitive and specific while allowing simultaneous detection of A. actinomycetemcomitans and P. gingivalis. This assay has potential applications in epidemiological studies, diagnosis, treatment planning, and monitoring of periodontal pathogens.

Aggregatibacter actinomycetemcomitans↗
PubMed1994

Characterization of a species-specific DNA fragment originating from the Candida albicans mitochondrial genome.

A species-specific DNA fragment EO3 isolated from Candida albicans was characterized. EO3 was shown to originate from a duplicated region of C. albicans mitochondrial (mt) DNA. Nucleotide sequence of a 0.9 kbp fragment (EO3S) which confers the species specificity of EO3 was determined. EO3S region in some strains contained two parts of deletions (or insertions) of 50-60 bp with short inverted repeats at both ends. The EO3S region was maintained stably in mtDNA of individual C. albicans strains and may be useful as target DNA for the diagnosis and epidemiological studies of candidiasis.

Base Sequence↗
PubMed2004

Detection of four Plasmodium species in blood from humans by 18S rRNA gene subunit-based and species-specific real-time PCR assays.

There have been reports of increasing numbers of cases of malaria among migrants and travelers. Although microscopic examination of blood smears remains the "gold standard" in diagnosis, this method suffers from insufficient sensitivity and requires considerable expertise. To improve diagnosis, a multiplex real-time PCR was developed. One set of generic primers targeting a highly conserved region of the 18S rRNA gene of the genus Plasmodium was designed; the primer set was polymorphic enough internally to design four species-specific probes for P. falciparum, P. vivax, P. malarie, and P. ovale. Real-time PCR with species-specific probes detected one plasmid copy of P. falciparum, P. vivax, P. malariae, and P. ovale specifically. The same sensitivity was achieved for all species with real-time PCR with the 18S screening probe. Ninety-seven blood samples were investigated. For 66 of them (60 patients), microscopy and real-time PCR results were compared and had a crude agreement of 86% for the detection of plasmodia. Discordant results were reevaluated with clinical, molecular, and sequencing data to resolve them. All nine discordances between 18S screening PCR and microscopy were resolved in favor of the molecular method, as were eight of nine discordances at the species level for the species-specific PCR among the 31 samples positive by both methods. The other 31 blood samples were tested to monitor the antimalaria treatment in seven patients. The number of parasites measured by real-time PCR fell rapidly for six out of seven patients in parallel to parasitemia determined microscopically. This suggests a role of quantitative PCR for the monitoring of patients receiving antimalaria therapy.

Animals↗
PubMed1993

Species-specific inhibition of fertilization by a peptide derived from the sperm protein bindin.

The sperm protein bindin is responsible for the species-specific adhesion of the sperm to the egg. The regions of the bindin molecule responsible for forming the contact between the sperm and the egg were investigated by measuring the ability of peptides representing various regions of the bindin sequence to inhibit fertilization. Twenty-four peptides were studied: 7 based on the Strongylocentrotus purpuratus bindin sequence, 11 based on the S. franciscanus bindin sequence, and 6 control peptides. Values for the concentration of peptide required to inhibit 50% of the productive sperm contacts (IC50) were extracted from experimental measurements of the extent of fertilization in the presence of various concentrations. of these peptides. The IC50 value averaged 220 microM for the control peptides. Active peptides representing certain specific subregions of the bindin sequence displayed IC50 values < 10% of the average value for control peptides, and the IC50 for the most potent of the peptides tested was only approximately 1% of the control peptide value (IC50 = 2.2 microM). Furthermore, we found that a peptide representing a particular region of the S. franciscanus bindin sequence that differs from the S. purpuratus bindin sequence inhibits fertilization species specifically. For the reaction of S. purpuratus sperm and eggs, the IC50 of this peptide was approximately 120 microM, whereas for the reaction of S. franciscanus sperm and eggs it was only 8.6 microM. These results demonstrate that a few specific regions of the bindin molecule are involved in the sperm-egg contact and that certain of these regions mediate the species specificity of the interaction in a sequence-specific manner.

Amino Acid Sequence↗
PubMed2004

Identification of Flavobacterium columnare by a species-specific polymerase chain reaction and renaming of ATCC43622 strain to Flavobacterium johnsoniae.

Species-specific polymerase chain reaction (PCR) primers have been designed to identify the causative agent of columnaris disease, Flavobacterium columnare. The 16S rRNA gene sequences of F. columnare (eight sequences representing the different genotypes of the species) and related species (18 sequences) were aligned and compared to choose specific regions that are unique to F. columnare and do not have significant intraspecies variability. The species-specific regions in the 16S rRNA gene were used to design a pair of species-specific PCR primers, ColF and ColR. The PCR technique produced a specific amplicon of about 675 base pairs (bp) in 27 isolates of F. columnare and there was no amplification in the closely related species. The specificity of the amplified product was confirmed by digesting with HhaI. The PCR primers did not produce a 675 bp product with F. columnare ATCC43622 strain. This ATCC43622 strain was characterized by biochemical and ribotyping methods and renamed Flavobacterium johnsoniae. The American Type Culture Collection has confirmed these findings and made the change.

Bacterial Typing Techniques↗