Sadashi Shiga

Differentiation of Chlamydia species by combined use of polymerase chain reaction and restriction endonuclease analysis.

To differentiate Chlamydia spp., a primer pair designed to generate a genus‐specific region of the major outer membrane protein (MOMP) gene was used in a PCR to amplify a single DNA fragment of 245‐259 bp. In the PCR, the expected single DNA fragment was amplified from strains of Chlamydia trachomatis, C. psittaci, C. pneumoniae and C. pecorum, respectively. By restriction endonuclease analysis with AluI and PvuII, the amplified products exhibited four distinct patterns, corresponding to the four species. It is, therefore, concluded that one‐step PCR followed by restriction endonuclease analysis as described in this study could be a valuable method for the detection and differentiation of Chlamydia species.

In Vitro Inhibitory Effects of Hinokitiol on Proliferation of Chlamydia trachomatis

ABSTRACT The inhibitory effects of hinokitiol (β-thujaplicin) on Chlamydia trachomatis D/UW-3/Cx were shown by MIC, minimum lethal concentration (MLC), and preinoculation minimal microbicidal concentration assays using HeLa 229 cells. The MIC and the MLC were both 32 μg/ml. Further evaluation of hinokitiol as a topical agent against C. trachomatis is warranted.

In vitro analysis of the change in resistance of Chlamydia trachomatis under exposure to sub-MIC levofloxacin for a therapeutic term.

Background: While fluoroquinolone-resistant Chlamydia trachomatis strains have not been clinically isolated, they were isolated in an in vitro study recently. Methods: To determine whether C. trachomatis strains develop resistance under sub-MIC antibacterial exposure in a clinical therapeutic term, C. trachomatis strains were exposed to sub-MIC levofloxacin (LVFX) for about 2 weeks. The MIC of LVFX was measured and DNA fingerprinting was performed every 72 h by PCR using random primers. Results: There was almost no change in the MIC under exposure to 0.125 μg/ml LVFX. However, some mutational changes in DNA fingerprints developed. Conclusions: In clinical therapeutic terms, resistant strains of C. trachomatis will probably not develop, even if sub-MIC LVFX is employed.

Biosynthesized Tea Polyphenols Inactivate Chlamydia trachomatis In Vitro

ABSTRACT Biosynthesized tea polyphenols showed antichlamydial activity against Chlamydia trachomatis D/UW-3/Cx and L2/434/Bu using cell culture. The most active compounds were (−)-epigallocatechin gallate and (−)-epicatechin gallate, followed by (−)-epicatechin (EC). (+)-Epicatechin and (−)-epigallocatechin were intermediate. EC was the least toxic. These results warrant evaluation of tea polyphenols as topical antichlamydial agents.

A case of pediatric Chlamydia pneumoniae infection and properties of the isolate

Chlamydia pneumoniae was isolated from the throat of a 2-year-old girl with upper respiratory illness. The isolate, Shizuoka-37, was stained with C. pneumoniae specific monoclonal antibody (RR402), as well as the genus specific antibody (Cultureset), but not with C. trachomatis specific monoclonal antibody (Micro-Trak). C. pneumoniae genome was amplified by polymerase chain reaction in the isolate. Elementary bodies (EB) of the isolate was round shaped by electron micrograph.

Monoclonal Antibodies to Herpes Simplex Viruses Antigens-Specificity and Utility in Rapid Serotyping of Clinical Isolates

Sixteen hybridomas secreting antibodies to HSV-1 and 22 hybridomas secreting antibodies to HSV-2 were derived from fusion of SP2/0 myeloma cells with spleen cells from BALB/c mice immunized with each respective virus. Four of the former 16 hybridomas and seven of the latter 22 hybridomas were subcloned and injected into pristane-primed mice to obtain high titers of monoclonal antibodies. Antigen specificity of these monoclonal antibodies were determined by the Western blotting (WB) assay. Two out of four monoclonal antibodies that showed selective reactivity for HSV-1 in IFA, reacted with HSV-1 specific proteins; #1 reacted with 100 KD and 70 KD proteins and #4 with a 150 KD protein, respectively, while the remaining two antibodies reacted only with a 50 KD protein that is type-common antigen. On the other hand, two out of seven antibodies which showed selective reactivity for HSV-2 in IFA, reacted with HSV-2 specific proteins: #5 with a 100 KD protein and #10 with three proteins of 30, 25, and 20 KD, and the other two antibodies reacted with a 50 KD protein that is a type-common antigen. The remaining three antibodies, two of which were found to be immunoglobulin type IgM, reacted with neither HSV-1 nor HSV-2 antigens in WB assay. In order to determine their utility in serotyping, 11 monoclonal antibodies were examined by IFA test for reactivity to cells that were infected with 20 HSV-1 or 16 HSV-2 isolates which had been typed by neutralization test.(ABSTRACT TRUNCATED AT 250 WORDS)