Penicillin-resistant (penr) clinical isolates of Neisseria meningitidis, which do not produce beta-lactamase, were first identified in Spain in 1985; the frequency of their recovery, which has been increasing in the past few years, reached 20% in 1989. Serogrouping, determination of serotypes and subtypes, and multilocus enzyme electrophoresis of the penr strains showed an extensive diversity. Resistance is due, at least in part, to a decreased affinity of penicillin-binding protein (PBP) 2 for penicillin. Similar low-affinity forms of PBP 2 are also found in penr isolates of Neisseria lactamica, Neisseria polysaccharea, and Neisseria gonorrhoeae. Genetic transformation of an N. meningitidis type strain to low-level penicillin resistance with DNA from resistant meningococci and other Neisseria species resulted in transformants that possessed low-affinity forms of PBP 2. These altered forms of PBP 2 have been shown to arise from recombinational events that replace parts of the PBP 2 gene with the corresponding regions from the PBP 2 genes of commensal Neisseria species.

We analysed a collection of 95 multiply antibiotic-resistant pneumococci, recovered since 1988 from 14 Spanish hospitals, that have MlCs 3 0-25 pg benzylpenicillin ml-l. The majority of the isolates were of serogroups 14, 23, 6, 19 and 15, which are currently the serogroups mainly associated with multiresistance in Spain. All of the serogroup 23 isolates were members of the major Spanish serotype 23F multiresistant clone. Similarly, most of the serogroup 6 isolates were members of the major multiresistant serotype 6B clone, or variants of this clone. Eighteen of the 24 isolates of serogroup 1 9 were members of a highly penicillin-resistant clone that appears to be a serotype 19F variant of the major Spanish serotype 23F multiresistant clone. Eighteen of the 25 isolates of serotype 14 were members of a previously uncharacterized highly penicillin-resistant clone. Thirteen of the 16 isolates of serogroup 1 5 were members of a single previously unreported clone of serotype 15F that had moderate levels of resistance to penicillin. Approximately 65 O/ O of the multiresistant pneumococci that are currently circulating in Spain were members of the three new clones of serotype 14, 15F and 19F that we describe here, or the previously described serotype 6B and 23F clones. The other 35% of isolates were minor variants of the major clones, unrelated minor clones, and unique isolates, many of which appeared to have arisen b y horizontal gene transfer events.

Multiply-antibiotic-resistant isolates of serogroup 19 Streptococcus pneumoniae, possessing altered penicillin-binding protein (PBP) 1A, 2B, and 2X genes that are indistinguishable from those of the Spanish multiresistant serogroup 23F clone, are now commonly encountered in Spain. Those isolates that have been serotyped express type 19F capsular polysaccharide. Serotyping of further isolates, and hybridization using a serotype 19F-specific probe, has shown that some of them are serotype 19A, rather than 19F. The Spanish multiresistant serotype 19A, 19F, and 23F multiresistant strains were all shown to be very closely related in overall genotype, as they were indistinguishable by REP-PCR and by the sequencing of internal fragments of three house-keeping genes. The serotype 19A multiresistant strains, like the serotype 19F multiresistant strains, therefore appear to be a serotype variant of the Spanish multiresistant serotype 23F clone, which presumably has arisen by recombination at the capsular locus.

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The increase in antibiotic resistance and the possible changes in serotype prevalence as a consequence of a new conjugated vaccine have contributed to renewed interest in the study of pneumococcal serotypes and their antibiotic resistances. Spain still has one of the highest penicillin resistance rates, but in the past 4-5 years a slight decrease has been observed. The level of resistance has not increased either, 12.7% of the 11 165 isolates studied showed high-level penicillin resistance but 94% of these had an MIC of only 2 mg/L. Serotypes 6, 9, 14, 19 and 23 included 83% of the penicillin-resistant pneumococci; the remaining 17% belonged to 18 different serotypes. We analysed these minor penicillin-resistant serotypes in view of their potential increase following a possible child vaccination programme. Four of these serotypes (11, 15, 21 and 35) were the most prevalent, and among them serotype 15 was particularly frequent with >50% of its strains resistant. The effective control of these minor penicillin-resistant serotypes should be based on continuous surveillance of pneumococcal epidemiology.

More than 50% of the nontypeable (NT) pneumococcal strains received in our laboratory for reference purposes are isolated in sporadic cases of conjunctivitis. To determine the genetic structure of the population of these NT conjunctival strains, we analyzed 75 pneumococci (40 NT and 35 typeable) isolated from conjunctivas and 30 (15 NT and 15 typeable) isolated from other sources. The NT and typeable conjunctival strains grouped in separate clusters, whereas NT and typeable pneumococci isolated from other sources were similarly distributed. NT conjunctival strains belonged to two well-differentiated clonal lineages. The first, represented by three newly described sequence types, featured fully antibiotic susceptible strains and appeared to be characteristic of conjunctival tissue; the second, represented by the previously described ST344, had a pattern of multiresistance to penicillin, tetracycline, and erythromycin and shared a genetic background with some NT strains isolated from other sources.Streptococcus pneumoniae is an important pathogenic bacterium associated with pneumonia, septicemia, meningitis, and otitis. It is also a common cause of acute conjunctivitis, particularly in children, but also in adults (11,19). For reference purposes, our laboratory receives many pneumococcus samples from all origins that have been isolated in Spanish hospitals (8). Around one-third of the strains isolated from children below the age of 6 months that were studied in our laboratory between 1990 and 1999 caused acute conjunctivitis (9).Pneumococcal serotyping usually fails to detect a small number of strains that do not react with antipneumococcal typing sera. Nontypeable (NT) strains are infrequently isolated from sterile clinical specimens (2.2%), in which case they are rarely implicated as causes of invasive disease (2, 12), since they are otherwise relatively common in nonsterile samples (10%). The identification of these NT pneumococci is dubious (14, 17), particularly in nonsterile specimens, and they may be confused with other Streptococcus species. The association between the presence of NT isolates and the occurrence of conjunctivitis was first suggested in 1977 (10) in a retrospective study of the incidence of capsular types in a Boston hospital between 1935 and 1974.Further studies associated NT strains with outbreak and sporadic cases of conjunctivitis (1,6,18,20), and a recent report has confirmed NT S. pneumoniae-like strains isolated from an outbreak of epidemic conjunctivitis as being S. pneumoniae (3).In the last 10 years, approximately 50% of the NT pneumococcal strains received in our laboratory have been isolated from cases of conjunctivitis, and the frequency of these noncapsular strains was five times that found in other pathologies (laboratory data).In general, the NT strains have been characterized in cases related to outbreaks, but as yet we have little information concerning those strains isolated from sporadic cases. The purpose of this study was to characterize NT pneumococcal strains isolated...

Summary.A scheme is proposed for biotyping Gardnerella vaginalis, based on detection of hippurate hydrolysis, 0-galactosidase (ONPG) and lipase, and fermentation of arabinose, galactose and xylose. Seventeen biotypes were found among 197 strains from asymptomatic women and patients with bacterial vaginosis (non-specific vaginitis). The distribution of biotypes was similar in both populations but some biotypes were found more frequently in patients. The proposed scheme is compared with those previously described. Introductionsexually transmitted diseases and family planning centres. The strains were isolated from 84 asymptomatic women and from 13 Several simple schemes have been described for the rapid identification of isolates of Gardnerella vaginaliswith bacteria] vaginosis.

We studied the susceptibilities of relatively penicillin G-resistant and -susceptible strains of Neisseria meningitidis, as well as Neisseria lactamica and Neisseria polysaccharea, to penicillin, ampicillin, and several cephalosporins. The MICs of penicillin, ampicillin, cephalothin, and cefuroxime for moderately resistant meningococci have increased two- to sixfold in relation to MICs for susceptible strains. For these strains of meningococci, N. lactamica, and N. polysaccharea, penicillin, ampicillin, cephalothin, and cefuroxime MICs for 50 and 90% of strains were similar. By genetic transformation of a penicillin-susceptible strain of N. meningitidis to low-level penicillin resistance with DNA from penicillin-resistant strains of N. meningitidis, N. lactamica, N. polysaccharea, and N. gonorrhoeae, isogenic strains with the same pattern of resistance to beta-lactams were obtained, suggesting that these commensal Neisseria spp. could be the source of meningococcal resistance genes.