In 2000, >400 cases of disease caused by Neisseria meningitidis serogroup W135 (MenW135), the largest MenW135 outbreak reported to date, occurred worldwide among Hajj pilgrims and their contacts. To elucidate the origin of the outbreak strains and to investigate their relatedness to major clonal groups, genotypic and phenotypic subtyping was performed on 26 MenW135 outbreak-associated isolates and 50 MenW135 isolates collected worldwide from 1970 through 2000. All outbreak-associated isolates were members of a single clone of the hypervirulent electrophoretic type (ET)-37 complex, designated the "(W)ET-37 clone"; 19 additional MenW135 strains were also members of this clone, and the remaining 31 MenW135 strains were clearly distinct. The 2000 MenW135 outbreak was not caused by emergence of a new MenW135 strain but rather by expansion of the (W)ET-37 clone that has been in circulation at least since 1970; the strains most closely related to those causing the 2000 outbreak have been isolated in Algeria, Mali, and The Gambia in the 1990s.

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Listeria bacteremia occurred in 2 pregnant women whose only common exposure was attendance at a party. The incubation period, the possibility of mild disease due to Listeria infection, and foods associated with risk of disease were evaluated. Ten (28%) of 36 party attenders met a case definition, which included isolation of Listeria monocytogenes from blood or stool or two of the following: fever, musculoskeletal symptoms, nausea, vomiting, diarrhea. One of 25 stool cultures was positive. The 2 blood isolates and 1 stool isolate were serotype 4b and identical by enzyme typing. The incubation periods for illness in the 2 pregnant women were 19 and 23 days. Consumption of large amounts of shrimp, nonalcoholic beverages, Camembert cheese, and cauliflower was significantly associated with illness. Eating shrimp remained a significant risk factor for illness after controlling for consumption of other foods. This study suggests a milder illness may exist in healthy persons who consume foods contaminated with L. monocytogenes and demonstrates a prolonged incubation period for disease.

Streptococcus pneumoniae isolates resistant to penicillin, chloramphenicol, tetracycline and sulfamethoxazole-trimethroprim are being recovered with increasing frequency in the United States. We analyzed the penicillin-binding proteins (PBPs), multilocus enzyme electrophoresis (MLEE) genotypes, and ribotpes of 22 multiresistant serotype 23F isolates of S. pneumoniae from the United States and 1 isolate each from Spain and South Africa. Also included were seven multiresistant isolates of other serotypes, three penicillin-resistant but chloramphenicol-susceptible serotype 23F isolates, and two penicillin-susceptible isolates (one penicillinsusceptible isolate was serotype 23F). Fifteen of the 22 multiresistant isolates from the United States and the isolates from Spain and South Africa had identical PBP patterns, MLEE profiles, and ribotypes. Six of the remaining seven multiresistant isolates were related by PBP pattern, but demonstrated slightly different MLEE and/or ribotype profiles, possibly because of acquisition of additional resistance markers (four of the six isolates were also resistant to erythromycin). The remaining multiresistant serotype 23F isolate had a unique PBP pattern and ribotype and was only distantly related to the other pneumococcal isolates by MLEE analysis. The PBP patterns, MLEE profiles, and ribotypes of the multiresistant serotpe 23F isolates were easily distinguished from those of six multiresistant isolates of other serotypes; three other penicillin-resistant, chloramphenicol-susceptible, serotype 23F isolates; and two penicillin-susceptible isolates. One exception was a multiresistant serotype 19A isolate that was highly related to the clonal group by PBP pattern and MLEE analysis and that had a ribotype similar to those of the other erythromycin-resistant serotype 23F isolates. MLEE analysis and ribotyping were more discriminating than were the PBP patterns in discerning strain differences. These data strongly suggest that a multiresistant clone ofS. pneumoniae serotype 23F that is related to multiresistant isolates from Spain and South Africa has become disseminated in the United States. Clinicians should be alerted to the spread of these multiresistant strains in the United States.

Since 1990, the frequency of Neisseria meningitidis serogroup C (NMSC) outbreaks in the United States has increased. Based on multilocus enzyme electrophoresis (MEE), the current molecular subtyping standard, most of the NMSC outbreaks have been caused by isolates of several closely related electrophoretic types (ETs) within the ET-37 complex. We chose 66 isolates from four well-described NMSC outbreaks that occurred in the United States from 1993 to 1995 to evaluate the potential of pulsed-field gel electrophoresis (PFGE) to identify outbreak-related isolates specific for each of the four outbreaks and to differentiate between them and 50 sporadic isolates collected during the outbreak investigations or through active laboratory-based surveillance from 1989 to 1996. We tested all isolates collected during the outbreak investigations by four other molecular subtyping methods: MEE, ribotyping (ClaI), random amplified polymorphic DNA assay (two primers), and serotyping and serosubtyping. Among the 116 isolates, we observed 11 clusters of 39 NheI PFGE patterns. Excellent correlation between the PFGE and the epidemiological data was observed, with an overall sensitivity of 85% and specificity of 71% at the 95% pattern relatedness breakpoint using either 1.5 or 1.0% tolerance. For all four analyzed outbreaks, PFGE would have given public health officials additional support in declaring an outbreak and making appropriate public health decisions.

We investigated the diversity of the primary sequences of 16S rRNA genes among Neisseria meningitidis strains (Men) and evaluated the use of this approach as a molecular subtyping tool. We aligned and compared a 1,417-bp fragment of the 16S rRNA gene from 264 Men strains of serogroups A, B, C, and Y (MenA, MenB, MenC, and MenY, respectively) isolated throughout the world over a 30-year period. Thirty-one positions of difference were found among 49 16S types: differences between types ranged from 1 to 14 positions (0.07 to 0.95%). 16S types and serogroups were highly associated; only 3 out 49 16S types were shared by two or more serogroups. We have identified 16S types that are exclusively associated with strains of certain hypervirulent clones: 16S type 5 with MenA subgroup III, 16S type 4 with the MenB electrophoretic type 5 (ET-5) complex, and 16S types 12 and 13 with MenC of the ET-37 complex. For MenC strains, 16S sequencing provided the highest sensitivity and specificity and the best overall association with the outbreak-related versus sporadic isolates when compared with pulsed-field gel electrophoresis, multilocus enzyme electrophoresis, and multilocus sequence typing. We demonstrated for the first time an unexpected diversity among 16S rRNA genes of Men strains, identified 16S types associated with well-defined hypervirulent clones, and showed the potential of this approach to rapidly identify virulent strains associated with outbreaks and/or an increased incidence of sporadic disease.rRNA is essential for the survival of all cells, and the genes encoding the rRNA are highly conserved in the bacteria and other kingdoms. The sequences of the rRNA and proteins comprising the ribosome are highly conserved throughout evolution, because they require complex inter-and intramolecular interactions to maintain the protein-synthesizing machinery (13,38). Consequently, determination of differences in the sequence of the 16S rRNA gene is well established as a standard method for the identification and phylogenetic classification of prokaryotic species, genera, and families (3, 38) and infers the evolution of the organisms. The sequences of rRNA genes in species with multiple rRNA operons are identical or nearly identical. A few studies have clearly shown the presence of sequence heterogeneity between rRNA operons within single genomes (16,19). In most organisms, 16S rRNA gene sequences from only a few strains of any particular species have been studied; therefore, variation among operons and its significance have not been fully evaluated (2,

The amino acids required for growth and as energy sources by 10 strains of Legionella pneumophila were determined by using a chemically defined medium. All strains required arginine, cysteine, isoleucine, leucine, threonine, valine, methionine, and phenylalanine or tyrosine. Most strains (7 of 10) required serine, and two strains had to be supplied proline before growth could be established. All 10 strains used serine and, to a lesser extent, threonine as the sole sources of

Brazilian purpuric fever (BPF) is a recently recognized fulminant pediatric disease characterized by fever, with rapid progression to purpura, hypotensive shock, and death. BPF is usually preceded by purulent conjunctivitis that has resolved before the onset of fever. Both the conjunctivitis and RPF are caused by Haemophilus influenzae biogroup aegyptius (formerly called H. aegyptius). Isolates from 15 BPF cases, mainly from blood or hemorrhagic cerebrospinal fluid, case-associated isolates from 42 persons in towns where BPF cases occurred, and control strains from 32 persons in towns without BPF cases were characterized biochemically, genetically, and epidemiologically. Results indicated that a single clone was responsible for all BPF cases identified in six Brazilian towns from 1984 through 1986. All of 15 (100%) case strains were the same clone as was 1 of 32 (3%) control strains (P = <10-8). Isolates of the clone were preferentially intrarelated by DNA hybridization (99% relatedness, hydroxyapatite method at 60 and 75°C) and were separable from other H. influenzae biogroup aegyptius strains (approximately 90% relatedness at 60°C and 82% relatedness at 75°C). All isolates of the BPF clone and no other strains contained a 24-megadalton plasmid of restriction endonuclease type 3031, were of a single multilocus enzyme mobility type, were of a single sodium dodecyl sulfate-polyacrylamide gel electrophoresis type, and were in one of two ribosomal DNA restriction patterns. Ail BPF clone isolates reacted with monoclonal antibodies produced from a case strain; only 3 of 62 (5%) other strains reacted with this monoclonal antibody. Ninety percent of BPF clone strains and 27% of other strains were relatively resistant to sulfamethoxazole-trimethoprim.