DNA fragments of 8.5 kb containing the gyrA gene were cloned from Escherichia coli KL-16 and from four spontaneous gyrA mutants which showed various levels of resistance to quinolones. The gyrA gene was situated at about 4 kb in front of the nrdA gene and transcribed counterclockwise on the E. coli chromosome. It encoded a polypeptide of 875 amino acids with a molecular weight of about 97,000. The four gyrA mutations were located strikingly close to one another within a small region near the N-terminus of the gyrA polypeptide, i.e., nucleotide changes from C to T, from C to G, from G to T and from G to T at nucleotides 248, 248, 318 and 199, respectively, resulting in amino acid changes from Ser to Leu, from Ser to Trp, from Gln to His and from Ala to Ser at amino acids 83, 83, 106 and 67, respectively. These mutations were situated in the relatively hydrophilic regions of the GyrA polypeptide and close to Tyr at amino acid 122 which has been shown to be the site covalently bound to DNA.

The proportion of gyrA and gyrB mutations in quinolone-resistant Escherichia coli strains was examined by introducing cloned wild-type gyrA and gyrB genes. In 25 spontaneous mutants of strain KL16, 13 had gyrA and 12 had gyrB mutations. In eight clinical isolates, five had gyrA mutations and one had a gyrB mutation; mutations in two isolates remained unidentified.

A 4.2-kb DNA fragment conferring quinolone resistance was cloned from a quinolone-resistant clinical isolate of Staphylococcus aureus and was shown to possess a part of the grlB gene and a mutated grlA gene. S-80-->F and E-84-->K mutations in the grlA gene product were responsible for the quinolone resistance. The mutated grlA genes responsible for quinolone resistance were dominant over the wild-type allele, irrespective of gene dosage in a transformation experiment with the grlA gene alone. However, dominance by mutated grlA genes depended on gene dosage when bacteria were transformed with the grlA and grlB genes in combination. Quinolone-resistant gyrA mutants were easily isolated from a strain, S. aureus RN4220, carrying a plasmid with the mutated grlA gene, though this was not the case for other S. aureus strains lacking the plasmid. The elimination of this plasmid from such quinolone-resistant gyrA mutants resulted in marked increases in quinolone susceptibility. These results suggest that both DNA gyrase and DNA topoisomerase IV may be targets of quinolones and that the quinolone susceptibility of organisms may be determined by which of these enzymes is most quinolone sensitive.

AT-4140, 5-amino-1-cyclopropyl-6,8-difluoro-1,4-dihydro-7-(cis-3,5-dimethyl-1-piperazinyl)-4-oxoquinoline-3-carboxylic acid, showed broad and potent antibacterial activity. Its MICs for 90% of the strains tested were 0.1 to 0.78 ,ug/mi against gram-positive organisms, such as members of the genera Staphylococcus, Streptococcus, and Enterococcus, and 0.0125 to 1.56 ,ug/ml against gram-negative organisms, such as members of the family Enterobacteriaceae and the genera Pseudomonas, Branhamella, Campylobacter, Haemophilus, and Neisseria. Its MICs were 0.025 to 0.78 ,ig/ml against glucose nonfermenters, such as members of the genera Xanthomonas, Acinetobacter, Alcaligenes, Moraxella, Flavobacterium, and Brucella; 0.2 to 0.78 ,ug/mi against anaerobes, such as Clostridium perfringens and Bacteroides fragilis; 0.0125 to 0.05 p.g/ml against Legionella spp.; 0.0125 to 0.2 ,ug/ml against Mycoplasma spp.; 0.031 to 0.063 ,u.g/ml against Chlamydia spp.; and 0.1 to 0.3 ,ig/ml against Mycobacterium spp. The potencies of AT-4140 against gram-negative organisms were comparable to those of ciprofloxacin and higher than those of ofloxacin, enoxacin, and norfloxacin. The potencies of AT-4140 against gram-positive organisms, glucose nonfermenters, anaerobes, Mycoplasma spp., Chlamydia spp., and Mycobacterium spp. were generally higher than those of the quinolones with which AT-4140 was compared. AT-4140 showed good oral efficacy against systemic infections with Staphylococcus aureus, Streptococcus pyogenes, Streptococcus pneumoniae, Escherichia coli, and Pseudomonas aeruginosa in mice. Its efficacy was better when a daily dose was given once than when it was given in two doses. Good efficacies of the orally administered drug were also observed in pulmonary, dermal, and urinary tract infection models in mice. The in vivo efficacies of AT-4140 were equal to or better than those of ciprofloxacin, ofloxacin, enoxacin, and norfloxacin.New quinolones used clinically these days have a broad spectrum of activity against both gram-positive and gramnegative organisms. However, some important pathogens, such as streptococci, enterococci, Mycoplasma spp., Chlamydia spp., and Mycobacterium spp., are not sufficiently susceptible to the quinolones. To improve this point, we screened of this group of compounds and found a new compound called AT-4140 (Fig. 1) with a broader antibacterial spectrum. This paper describes the in vitro and in vivo antibacterial activities of AT-4140 compared with those of ciprofloxacin, ofloxacin, enoxacin, and norfloxacin.(

A good prognosis can be expected for most, but not all, cases of thyroid papillary cancer. Numerous molecular studies have demonstrated beneficial treatment and prognostic factors in various molecular markers. Whereas most previous reports have focused on genomics and proteomics, few have focused on lipidomics. With the advent of mass spectrometry (MS), it has become possible to identify many types of molecules, and this analytical tool has become critical in the field of omics. Recently, imaging mass spectrometry (IMS) was developed. After a simple pretreatment process, IMS can be used to examine tissue sections on glass slides with location information.Here, we conducted an IMS analysis of seven cases of thyroid papillary cancer by comparison of cancerous with normal tissues, focusing on the distribution of phospholipids. We identified that phosphatidylcholine (16:0/18:1) and (16:0/18:2) and sphingomyelin (d18:0/16:1) are significantly higher in thyroid papillary cancer than in normal thyroid tissue as determined by tandem mass (MS/MS) analysis. These distributional differences may be associated with the biological behavior of thyroid papillary cancer.