Molecular characterization of chromosomal class C beta-lactamase and its regulatory gene in Ochrobactrum anthropi.

Ochrobactrum anthropi, formerly known as CDC group Vd, is an oxidase-producing, gram-negative, obligately aerobic, non-lactose-fermenting bacillus of low virulence that occasionally causes human infections. It is highly resistant to all beta-lactams except imipenem. A clinical isolate, SLO74, and six reference strains were tested. MICs of penicillins, aztreonam, and most cephalosporins tested, including cefotaxime and ceftazidime, were >128 microg/ml and of cefepime were 64 to >128 microg/ml. Clavulanic acid was ineffective and tazobactam had a weak effect in association with piperacillin. Two genes, ampR and ampC, were cloned by inserting restriction fragments of genomic DNA from the clinical strain O. anthropi SLO74 into pBK-CMV to give the recombinant plasmid pBK-OA1. The pattern of resistance to beta-lactams of this clone was similar to that of the parental strain, except for its resistance to cefepime (MIC, 0.5 ,micro/ml). The deduced amino acid sequence of the AmpC beta-lactamase (pI, 8.9) was only 41 to 52% identical to the sequence of other chromosomally encoded and plasmid-encoded class C beta-lactamases. The kinetic properties of this beta-lactamase were typical for this class of beta-lactamases. Upstream from the ampC gene, the ampR gene encodes a protein with a sequence that is 46 to 62% identical to those of other AmpR proteins and with an amino-terminal DNA-binding domain typical of transcriptional activators of the Lys-R family. The deduced amino acid sequences of the ampC genes of the six reference strains were 96 to 99% identical to the sequence of the clinical strain. The beta-lactamase characterized from strain SLO74 was named OCH-1 (gene, bla(OCH-I)).

Inhibitor-resistant OXY-2-derived beta-lactamase produced by Klebsiella oxytoca.

Klebsiella oxytoca strains are generally moderately resistant to amoxicillin and ticarcillin due to the activities of the chromosomally encoded OXY-1 and OXY-2 class A beta-lactamase families. These enzymes have the ability to hydrolyze not only penicillins but also cephalosporins, including cefuroxime, ceftriaxone, and aztreonam, and are inhibited by clavulanic acid. A Klebsiella oxytoca strain was isolated from a culture of blood from a patient who had been treated with amoxicillin-clavulanate (3 g/day) for 10 days 1 month earlier. This strain harbored an unusual phenotype characterized by resistance to amoxicillin-clavulanate. It produced an OXY-2-type beta-lactamase (pI 6.3), as confirmed by PCR amplification with primers specific for the OXY-2-encoding gene. Gene sequencing revealed a point mutation (A-->G) corresponding to the amino acid substitution Ser-->Gly at position 130. This mutant enzyme was poorly inhibited by inhibitors, and its kinetic constants compared to those of the parent enzyme were characterized by an increased Km value for ticarcillin, with a drastically reduced activity against cephalosporins, as is observed with inhibitor-resistant TEM enzymes. The substitution Ser-->Gly-130 was previously described in the inhibitor-resistant beta-lactamase SHV-10 derived from an SHV-5 variant, but this is the first report of such a mutant in OXY enzymes from K. oxytoca.

Metabolites from the sponge-associated bacterium Micrococcus luteus.

In an ongoing survey of the bioactive potential of microorganisms associated with marine invertebrates, the extract of the sponge-associated bacterial strain Micrococcus luteus was found to exhibit potent antimicrobial activity. The previously known synthetic 2,4,4'-trichloro-2'-hydroxydiphenylether was found to be responsible for the antimicrobial activity. The major metabolite isolated was a new acyl-1-(acyl-6'-mannobiosyl)-3-glycerol.

Effects of N-alkylation and n-acylation on tobramycin activity.

The activities of tobramycin derivatives acetylated and ethylated on the 6'-N,2'-N and 3-N positions were examined. The MICs of these derivatives against tobramycin sensitive strains indicated that 2'-N-ethylated and 6'-N-ethylated derivatives have a fairly good activity, and confirmed that the 3-N position is the most important one for antibiotic activity since 3-N derivatives were less active. The MICs of these derivatives against tobramycin resistant strains, and their inactivation by tobramycin modifying enzymes were examined. These results showed that 2'-N or 6'-N ethylation protects the drug against inactivation by AAC(2') or AAC(6'), respectively, and 2'-N-ethyltobramycin and 6'-N-ethyltobramycin were active against strains containing these modifying enzymes. On the other hand, 3-N ethylation protects the drug against inactivation by AAC(3) but 3-N-ethyl tobramycin does not inhibit strains containing this enzyme.

Plasmid-mediated pristinamycin resistance. PAC IIA: a new enzyme which modifies pristinamycin IIA.

A wild strain of Staphylococcus aureus which inactivates a wide variety of antibiotics has been found to inactivate pristinamycin IIA, an antistaphylococcal antibiotic. This phenomenon has been demonstrated to be plasmid mediated. The plasmid directs the biosynthesis of an acetyltransferase which is able to O-acetylate the drug. We propose to call the new enzyme PAC (IIA): Pristinamycin acetyltransferase.