Shizuko Kagawa

Bio: Shizuko Kagawa is an academic researcher from Laboratory of Molecular Biology. The author has contributed to research in topics: Taylorella equigenitalis & Campylobacter lari. The author has an hindex of 5, co-authored 13 publications receiving 106 citations. Previous affiliations of Shizuko Kagawa include Azabu University.

Homogeneity of the 16S rDNA sequence among geographically disparate isolates of Taylorella equigenitalis.

Abstract: Background At present, six accessible sequences of 16S rDNA from Taylorella equigenitalis (T. equigenitalis) are available, whose sequence differences occur at a few nucleotide positions. Thus it is important to determine these sequences from additional strains in other countries, if possible, in order to clarify any anomalies regarding 16S rDNA sequence heterogeneity. Here, we clone and sequence the approximate full-length 16S rDNA from additional strains of T. equigenitalis isolated in Japan, Australia and France and compare these sequences to the existing published sequences.

Cloning and sequencing of 16S rDNA and 16S-23S rDNA internal spacer region (ISR) from urease-positive thermophilic Campylobacter (UPTC).

Abstract: Aims: To clone and sequence the 16S rDNA and 16S-23S rDNA internal spacer region (ISR) from urease-positive thermophilic Campylobacter (UPTC). Methods and Results: The primer sets for 16S rDNA and 16S-23S rDNA ISR amplified almost the full length of 16S rDNA and 16S-23S rDNA ISR. About 1500 bp for 16S rDNA and about 720 bp for 16S-23S rDNA ISR of the rrn operon of four strains of UPTC were identified after molecular cloning and sequencing. Conclusions: The four strains and CCUG18267 of UPTC showed approximately 99% sequence homology of 16S rDNA to each other, 96–97% to Camp. coli, 97–98% to Camp. jejuni and 97–98% to Camp. lari. Significance and Impact of the Study: For the first time, the nucleotide sequence of 16S-23S rDNA ISR of UPTC has been analysed. The sequence of ISR was almost identical among the four strains of UPTC. It is interesting that the UPTC intercistronic tRNAs demonstrated an order of tRNA of 5′-16S-tRNAAla-tRNAIle-23S-3′ in the organisms.

Nucleotide sequencing and analysis of 16S rDNA and 16S-23S rDNA internal spacer region (ISR) of Taylorella equigenitalis, as an important pathogen for contagious equine metritis (CEM).

Abstract: The primer set for 16S rDNA amplified an amplicon of about 1500 bp in length for three strains of Taylorella equigenitalis (NCTC11184T, Kentucky188 and EQ59). Sequence differences of the 16S rDNA among the six sequences, including three reference sequences, occurred at only a few nucleotide positions and thus, an extremely high sequence similarity of the 16S rDNA was first demonstrated among the six sequences. In addition, the primer set for 16S-23S rDNA internal spacer region (ISR) amplified two amplicons about 1300 bp and 1200 bp in length for the three strains. The ISRs were estimated to be about 920 bp in length for large ISR-A and about 830 bp for small ISR-B. Sequence alignment of the ISR-A and ISR-B demonstrated about 10 base differences between NCTC11184T and EQ59 and between Kentucky188 and EQ59. However, only minor sequence differences were demonstrated between the ISR-A and ISR-B from NCTC11184T and Kentucky188, respectively. A typical order of the intercistronic tRNAs with the 29 nucleotide spacer of 5'-16S rDNA-tRNAIle-tRNAAla-23S rDNA-3' was demonstrated in the all ISRs. The ISRs may be useful for the discrimination amongst isolates of T. equigenitalis if sequencing is employed.

Demonstration of heterogeneous genotypes of Taylorella equigenitalis isolated from horses in six European countries by pulsed-field gel electrophoresis.

Abstract: Forty-six isolates of Taylorella equigenitalis were analysed by pulsed-field gel electrophoresis (PFGE) after separate digestion of the genomic DNA with ApaI and with NotI. The isolates had been obtained from horses in six European countries and were classified into 18 genotypes. In Belgium, 2 genotypes were detected in 2 isolates, in England 9 among 15, in Finland 2 in 2, in France 2 among 10, in Sweden 3 among 5, and in Switzerland 3 among 12. Two English isolates and 4 French isolates gave identical PFGE profiles to those of Kentucky 188 from the United States. A common genotype was found in 5 isolates from Belgium and England and also in 10 isolates from France and Switzerland. The analysis of genomic DNA from 12 isolates of T. equigenitalis obtained from male horses in France, Sweden and Switzerland gave no evidence of a sex-related difference in the genomic DNA. Genomic DNA from 11 streptomycin (STM)-susceptible isolates obtained in Sweden and Switzerland were classified into four genotypes by PFGE. Each of the six genotypes determined among the 17 isolates from these two countries had single phenotypes for resistance or susceptibility to STM.

Comparison of the value of pulsed-field gel electrophoresis, random amplified polymorphic DNA and amplified rDNA restriction analysis for subtyping Taylorella equigenitalis

Abstract: Eight strains of Taylorella equigenitalis were identified by a polymerase chain reaction using a primer pair specific to the 16S rDNA of T. equigenitalis. These eight strains were chosen because they had previously been shown to represent eight distinct genotypes by pulsed-field gel electrophoresis analysis after separate digestion of the genomic DNA with ApaI or NotI. The eight strains could be classified into six or seven types by random amplified polymorphic DNA analysis using different kinds of primers. Amplified rDNA restriction analysis after separate digestion with five restriction enzymes, including AluI and MboI, of the 1500 bp fragments of rDNA amplified by polymerase chain reaction did not discriminate the genomic variations among the eight strains of T. equigenitalis. Thus, pulsed-field gel electrophoresis was shown to discriminate these eight organisms better than random amplified polymorphic DNA analysis, while amplified rDNA restriction analysis was found to be unsuitable for subtyping T. equigenitalis.

High-throughput analysis of the satellitome illuminates satellite DNA evolution

Abstract: Satellite DNA (satDNA) is a major component yet the great unknown of eukaryote genomes and clearly underrepresented in genome sequencing projects. Here we show the high-throughput analysis of satellite DNA content in the migratory locust by means of the bioinformatic analysis of Illumina reads with the RepeatExplorer and RepeatMasker programs. This unveiled 62 satDNA families and we propose the term “satellitome” for the whole collection of different satDNA families in a genome. The finding that satDNAs were present in many contigs of the migratory locust draft genome indicates that they show many genomic locations invisible by fluorescent in situ hybridization (FISH). The cytological pattern of five satellites showing common descent (belonging to the SF3 superfamily) suggests that non-clustered satDNAs can become into clustered through local amplification at any of the many genomic loci resulting from previous dissemination of short satDNA arrays. The fact that all kinds of satDNA (micro- mini- and satellites) can show the non-clustered and clustered states suggests that all these elements are mostly similar, except for repeat length. Finally, the presence of VNTRs in bacteria, showing similar properties to non-clustered satDNAs in eukaryotes, suggests that this kind of tandem repeats show common properties in all living beings.

Adoption of the transiently non-culturable state. A bacterial survival strategy?

Abstract: Microbial culturability can be ephemeral. Cells are not merely either dead or alive but can adopt physiological states in which they appear to be (transiently) non-culturable under conditions in which they are known normally to be able to grow and divide. The reacquisition of culturability from such states is referred to as resuscitation. We here develop the idea that this "transient non-culturability" is a consequence of a special survival strategy, and summarise the morphological, physiological and genetic evidence underpinning such behaviour and its adaptive significance.

Molecular approaches : advantages and artifacts in assessing bacterial diversity

Abstract: Bacteria account for a major proportion of Earth's biological diversity They play essential roles in quite diverse environments and there has been an increasing interest in bacterial biodiversity Research using novel and efficient tools to identify and characterize bacterial communities has been the key for elucidating biological activities with potential for industrial application The current approach used for defining bacterial species is based on phenotypic and genomic properties Traditional and novel DNA-based molecular methods are improving our knowledge of bacterial diversity in nature Advances in molecular biology have been important for studies of diversity, considerably improving our knowledge of morphological, physiological, and ecological features of bacterial taxa DNA-DNA hybridization, which has been used for many years, is still considered the golden standard for bacteria species identification PCR-based methods investigating 16S rRNA gene sequences, and other approaches, such as the metagenome, have been used to study the physiology and diversity of bacteria and to identify novel genes with potential pharmaceutical and other biotechnological applications We examined the advantages and limitations of molecular methods currently used to analyze bacterial diversity; these are mainly based on the 16S rRNA gene These methods have allowed us to examine microorganisms that cannot be cultivated by routine methods and have also been useful for phylogenetic studies We also considered the importance of improvements in microbe culture techniques and how we can combine different methods to allow a more appropriate assessment of bacterial diversity and to determine their real potential for industrial applications

Nucleotide sequence of the gyrA gene of Arcobacter species and characterization of human ciprofloxacin‐resistant clinical isolates

Abstract: The nucleotide sequence of the gyrA gene of Arcobacter butzleri, Arcobacter cryaerophilus , Arcobacter cibarius , and Arcobacter skirrowii was determined. The deduced GyrA proteins are closely related to those of Wolinella succinogenes and Helicobacter pullorum , whereas those of Campylobacter species showed less sequence identity. The phylogenetic analysis of GyrA sequences provides a result similar to 16S rRNA gene sequence phylogenetic analysis and allows the discrimination among A. butzleri species. In addition, a Thr→Ile mutation at amino acid 85 in the quinolone resistance-determining region was associated with ciprofloxacin resistance for two A. butzleri and one A. cryaerophilus ciprofloxacin-resistant strains.

The highly leukotoxic JP2 clone of Aggregatibacter actinomycetemcomitans: evolutionary aspects, epidemiology and etiological role in aggressive periodontitis.

Abstract: For many years, attention has been given to the oral bacterium Aggregatibacter actinomycetemcomitans, as a species possibly implicated in the etiology of aggressive periodontitis in adolescents. One of the major virulence factors of A. actinomycetemcomitans is the leukotoxin which is able to kill important cells of the immune system. As demonstrated in population genetic analyses, the population structure of A. actinomycetemcomitans is mainly clonal with evolutionary lineages corresponding to the serotypes. A particular highly leukotoxic clone (JP2) of serotype b has been discovered. The JP2 clone, with an estimated origin some 2400 years ago, is found to be highly conserved, based on analyses of a collection of JP2 clone strains collected through more than 20 years from individuals of diverse origin and living geographically widespread. Despite demonstration of minor evolutionary changes within the genome of JP2 clone strains of A. actinomycetemcomitans, the JP2 clone strains constitute a unique clonal type, the characteristics of which include a 530 basepair deletion in the leukotoxin operon implicated in the enhanced leukotoxic activity of the clone. Mapping of the geographic occurrence of the JP2 clone of A. actinomycetemcomitans has revealed that its colonization is largely restricted to individuals of African descent. Characteristic mutations, which allow JP2 clone isolates from the Mediterranean region to be distinguished from isolates from West Africa, including the Cape Verde islands, suggest that the JP2 clone initially emerged as a distinct genotype in the Mediterranean region of Africa and subsequently spread to West Africa, from where it might have been transferred to the American continent during the transatlantic slave trade. The finding of a sustained selective colonization of individuals of African descent, despite geographical separation from the African continent for centuries, suggests that the JP2 clone might have a distinct host tropism. Further studies are needed to elucidate the reasons for the apparent selective colonization of the Mediterranean and Western African populations. The JP2 clone of A. actinomycetemcomitans appears to play a prominent role in the etiology of aggressive periodontitis compared to other clonal types of the species. While A. actinomycetemcomitans, in general, is considered an opportunistic pathogen of the resident oral microbiota, the JP2 clone has features similar to those of an exogenous pathogen. Clonal types other than JP2 can be isolated from healthy as well as periodontally diseased individuals, whereas the JP2 clone has been isolated primarily from periodontally diseased individuals. As demonstrated in a prospective cohort study in Morocco, where the JP2 clone is endemically present, the presence of this clone in dental plaque confers a remarkably increased risk for development of aggressive periodontitis, suggesting that the JP2 clone is an important etiological agent of aggressive periodontitis in adolescents. Support for association of clonal types other than JP2 of A. actinomycetemcomitans with aggressive periodontitis has also been provided, but the association is much weaker. Nearly half of the JP2 clone carriers were found to be persistently infected during a two-year follow-up period, which indicates a level of stability of colonization with the JP2 clone similar to that previously reported for non-JP2 clonal types of A. actinomycetemcomitans. The relative risk of aggressive periodontitis is highest for individuals with stable JP2 clone colonization. Although the method used is not quantitative, this finding adds to the evidence for a causal role of the JP2 clone in aggressive periodontitis. Longitudinal data shows that few individuals are colonized with the JP2 clone de novo after puberty. Patterns of parent-child carriage and shared colonization of JP2 clone strains among siblings have been demonstrated in other studies, altogether indicating that transmission of the JP2 clone, like other clonal types of A. actinomycetemcomitans, occurs vertically, and through close person to person contacts. In conclusion, a conserved and highly leukotoxic clone of A. actinomycetemcomitans with unique characteristics and with an apparent linkage to individuals of African descent has been identified. Being aware that the genetic constitution of the hosts has not been considered and that focus in our studies has been on A. actinomycetemcomitans only, and not on other members of the oral microbiota, we conclude that the JP2 clone of A. actinomycetemcomitans is a likely etiological agent of aggressive periodontitis in adolescents.