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PubMed1989

Lack of antibodies to Legionella pneumophila and Legionella micdadei in hemodialysis patients.

In 231 adult hemodialysis patients and 134 healthy adults, we measured antibodies to Legionella pneumophila (serogroups 1-6) and Legionella micdadei by indirect immunofluorescent antibody tests to assess the risk of Legionnaires' disease. One of the patients had a titer of 1:512 to Legionella pneumophila but he had no history of Legionnaires' disease. Two had a titer of 1:64 to Legionella pneumophila, and none had a titer of greater than or equal to 1:64 to Legionella micdadei. By contrast, none of the control group had a titer greater than or equal to 1:64 to Legionella pneumophila and two had a titer of 1:64 to Legionella micdadei. Thus, our population of maintenance hemodialysis patients did not display increased prevalence of antibodies of Legionella pneumophila and Legionella micdadei, but prospective studies of pneumonia in hemodialysis patients might further evaluate possible risk of Legionnaires' disease.

Adult↗
PubMed2005

[Evaluation of Dipstick 'Eiken' Legionella for detection of Legionella pneumophila serogroup 1 urinary antigen using the immunochromatographical method].

Dipstick 'Eiken' Legionella is a reagent for detection of Legionella pneumophila serogroup 1 antigen in urine using the immunochromatographical method. The reagent was evaluated using the reference and clinical isolated strains and clinical specimens. BinaxNOW Legionella and Legionella antigen [Mitsubishi] were evaluated simultaneously. Using four of L. pneumophila serogroup 1 strains, the minimal detectable concentration of Dipstick 'Eiken' Legionella was 5.0 x 10(4) to 2.0 x 10(5) colony forming unit/ml, it was approximate four times high in comparison with other two kits. And no positive reaction was obtained from 45 of non-L. pneumophila serogroup 1 strains. When Dipstick 'Eiken' Legionella was compared with BinaxNOW Legionella among 50 urine samples obtained from patients with pneumonia and 50 urine from healthy adults, the sensitivity was 94.7%, the specificity was 100.0% and the agreement was 99.0%. Dipstick 'Eiken' Legionella is found to be useful diagnostic reagent for Legionella infection in clinical laboratories.

Antigens, Bacterial↗
PubMed2005

Rapid method for enumeration of viable Legionella pneumophila and other Legionella spp. in water.

A sensitive and specific method has been developed to enumerate viable L. pneumophila and other Legionella spp. in water by epifluorescence microscopy in a short period of time (a few hours). This method allows the quantification of L. pneumophila or other Legionella spp. as well as the discrimination between viable and nonviable Legionella. It simultaneously combines the specific detection of Legionella cells using antibodies and a bacterial viability marker (ChemChrome V6), the enumeration being achieved by epifluorescence microscopy. The performance of this immunological double-staining (IDS) method was investigated in 38 natural filterable water samples from different aquatic sources, and the viable Legionella counts were compared with those obtained by the standard culture method. The recovery rate of the IDS method is similar to, or higher than, that of the conventional culture method. Under our experimental conditions, the limit of detection of the IDS method was <176 Legionella cells per liter. The examination of several samples in duplicates for the presence of L. pneumophila and other Legionella spp. indicated that the IDS method exhibits an excellent intralaboratory reproducibility, better than that of the standard culture method. This immunological approach allows rapid measurements in emergency situations, such as monitoring the efficacy of disinfection shock treatments. Although its field of application is as yet limited to filterable waters, the double-staining method may be an interesting alternative (not equivalent) to the conventional standard culture methods for enumerating viable Legionella when rapid detection is required.

Antibodies, Bacterial↗
PubMed2001

Distribution of Legionella longbeachae and other legionellae in Japanese potting soils.

The distribution of Legionella longbeachae and Legionella spp. in Japanese potting soils was examined. Thirty samples were collected: 13 were composted wood products, 11 were potting mixes (containing composted wood products, sand, mineral fertilizer, and manure), 2 were peatmoss, 3 were peatmoss-sand mixes (containing peatmoss and sand), and 1 was a sample of hydroponic clay balls. A suspension of each sample was made in sterile distilled water and acidified, and 100 microl was plated on buffered charcoal yeast extract alpha (BCYEalpha) agar containing modified wadowsky yee (MWY) supplement and pimaricin (direct method). In parallel, each suspension was incubated at 33 degrees C for several months to allow for amebic enrichment, if present; the suspensions were then plated onto culture media as described above (enrichment method). A total of 46 strains of legionellae were isolated from 22 of the 24 samples (13 composted wood products, 11 potting mixes). L. longbeachae was isolated from 9 samples. The most predominant species of legionellae in potting soils was Legionella bozemanii, which was isolated from 13 samples. Legionella spp. and L. micdadei were isolated from 8 and 7 samples each. Compared with findings in potting soils in Australia (26/45; 58%), Japanese potting soils had an only 8.3% (2/24) isolation rate for L. longbeachae by the direct method. The components of composted wood products were broadleaves such as oak and Japanese oak, in contrast to the pine and eucalypt used in Australia, which may account for the different isolation rates. However, the amebic enrichment method was useful in increasing the recovery of legionellae in potting soils. Legionellae were not isolated from the peatmoss samples, a result identical to findings in surveys of similar material in Europe.

Bacteriological Techniques↗
PubMed1999

Legionella taurinensis sp. nov., a new species antigenically similar to Legionella spiritensis.

A group of 42 Legionella-like organisms reacting specifically with Legionella spiritensis serogroup 1 antisera were collected throughout Europe by the Centre National de Référence (French National Reference Centre) for Legionella. This group of isolates differed somewhat from L. spiritensis in terms of biochemical reactions, ubiquinone content and protein profile. The latter two analyses revealed that one of these L. spiritensis-like isolates, Turin I no. 1T, was highly related, but not identical to any of the red autofluorescent species of Legionella. In fact, this strain was the first of these particular isolates recognized to emit a red autofluorescence when exposed to UV light. Profile analysis of randomly amplified polymorphic DNA established that the red autofluorescent L. spiritensis-like isolates constituted a homogeneous group distinct from Legionella rubrilucens and Legionella erythra. DNA-DNA hybridization studies involving the use of S1 nuclease confirmed that the indicated group of isolates are a new species of Legionella, for which the name Legionella taurinensis is proposed with strain Turin I no. 1T (deposited as ATCC 700508T) as the type strain.

Antigens, Bacterial↗
PubMed1984

Legionella sainthelensi: a new species of Legionella isolated from water near Mt. St. Helens.

Six strains of a new species, Legionella sainthelensi, were isolated from freshwater in areas affected by the volcanic eruptions of Mt. St. Helens in the state of Washington. Strains of L. sainthelensi are culturally and biochemically similar to other legionellae. They grow on buffered charcoal yeast agar but not on media that lack cysteine. They are gram-negative, nonsporeforming, motile rods that are positive in reactions for catalase, oxidase, gelatin liquefaction, and beta-lactamase. They are negative in reactions for urease, hydrolysis of hippurate, reduction of nitrates, fermentation of glucose, and blue-white autofluorescence. Their cell wall fatty acid composition is qualitatively similar to those of other legionellae, with 50 to 62% branched-chain fatty acids. They contain the isobranched-chain 14- and 16-carbon acids and anteisobranched-chain 15- and 17-carbon acids and relatively large amounts of straight-chain 16-carbon acid. All strains of L. sainthelensi contain approximately equal amounts of ubiquinones Q9, Q10, Q11, and Q12, a pattern similar to those of Legionella bozemanii, Legionella dumoffi, and Legionella longbeachae. Serological cross-reactions were observed between L. sainthelensi, both serogroups of L. longbeachae, and Legionella oakridgensis. Three strains of L. sainthelensi were greater than 90% related by DNA hybridization. The type strain of L. sainthelensi, Mt. St. Helens 4, was 36% related to the type strain of L. longbeachae and 3 to 14% related to the other nine described Legionella species.

Disasters↗
PubMed1989

Evaluation of the Merifluor-Legionella immunofluorescent reagent for identifying and detecting 21 Legionella species.

We evaluated a 33-valent polyclonal indirect immunofluorescent-reagent kit (Merifluor-Legionella; Meridian Diagnostics Inc., Cincinnati, Ohio) made for the detection of Legionella species by testing bacterial isolates, seeded sputum, and negative sputum samples. Use of the reagent according to the directions of the manufacturer gave false-negative staining of homologous culture isolates due to a prozone phenomenon; this was solved by diluting test strain suspensions. After this change in testing protocol was made, the reagent gave bright fluorescent staining with 31 of the 33 Legionella strains with which it supposedly reacts. Strongly reacting Legionella strains included the type strains of L. pneumophila serogroups 1 to 10, L. longbeachae serogroups 1 and 2, and serogroup 1 of L. anisa, L. bozemanii, L. cherrii, L. dumoffii, L. gormanii, L. hackeliae, L. jamestowniensis, L. jordanis, L. maceachernii, L. micadedi, L. oakridgensis, L. rubrilucens, L. sainthelensi, L. spiritensis, L. steigerwaltii, and L. wadsworthii. Type strains of L. erythra and L. feeleii fluoresced only dimly with the reagent. Of 10 non-Legionella bacteria known to cross-stain with other polyvalent antisera, 5 also cross-reacted with the Merifluor reagent; these included 3 Bacteroides fragilis and 2 Pseudomonas fluorescens strains. The lower limit of detection of L. pneumophila serogroup 1 in seeded sputum was about 5 x 10(4) to 5 x 10(5) cells per ml. None of 21 randomly collected sputum specimens tested contained fluorescing legionellalike organisms, but 6 specimens did contain brightly fluorescing bacteria atypical in morphology for Legionella species. The Merifluor-Legionella kit appears to perform as well as other polyclonal immunofluorescent reagents used for detection of Legionella species. Because of the cross-reactions observed, which are common to all polyclonal reagents, utilization of this reagent for either bacterial identification or detection must be performed in combination with culture.

Cross Reactions↗
PubMed1989

[Legionella dumoffii and Legionella pneumophila serogroup 5 isolated from 2 cases of fulminant pneumonia].

We encountered two cases of legionella pneumonia which ran a dramatic course and isolated Legionella dumoffii from one patient and Legionella pneumophila serogroup 5 from the other patient. The patient from whom L. dumoffii was isolated was a 59-year-old male with no basic disease. He presented chill, fever, coughing and other symptoms, starting on July 3, 1986, his disease was diagnosed as pneumonia at the clinic of his company. The patient was then introduced and admitted to our hospital. On admission chest radiography disclosed zonal pneumonia with an unclear border in the right superior lobe of the lung; a beta-lactam preparation was administered, but no effect was obtained and the lung lesion showed a rapid advance. From this condition, we suspected legionella pneumonia and changed the therapy to treatment with erythromycin and rifampicillin. Despite this, no improvement occurred and the patient died on the 26th hospital day. Colonies like Legionella colonies were separated from a total of seven specimens of biopsy aspirated matter from the airway and autopsy collected lung abscess and tracheal secretions, and the bacterium was identified L. dumoffii based on the biochemical and serological properties. In addition, the patient's serum was found to have an increased antibody titer against L. dumoffii. Based on these findings, the patient's disease was diagnosed as pneumonia as caused by L. dumoffii, a relatively rare bacterium as a member of the genus Legionella. The patient from whom Legionella pneumophila serogroup 5 was isolated was an 81-year-old man with basic diseases such as heart failure, anemia and hypothyroidism. He presented fever, general fatigue, anorexia and other symptoms, starting around June 2, 1987; pneumonia was suspected and the patient was urgently admitted to our hospital. The patient died of pneumonia of unknown cause on the second hospital day. To clarify the cause, autopsy was conducted; a large number of colonies like Legionella colonies were noted in the lung tissue. Identification test was then conducted and the bacterium was identified as L. pneumophila; we concluded that the patient's pneumonia had been caused by the identified bacterium L. pneumophila. The isolate was further subjected to slide agglutination test and identified as L. pneumophila serogroup 5.

Aged↗
PubMedDate not supplied

Legionella moravica sp. nov. and Legionella brunensis sp. nov. isolated from cooling-tower water.

Two Legionella-like organisms were isolated from cooling-tower water samples in Czechoslovakia. They were presumptively identified as legionellae by their growth on buffered charcoal-yeast extract agar (BCYE) containing L-cysteine and their absence of growth on BCYE without L-cysteine. Both strains contained predominately branch-chained cellular fatty acids and were therefore definitively placed in the genus Legionella. They were serologically distinct from other described Legionella species and were shown by DNA studies to constitute two new Legionella species, Legionella moravica (type strain 316-36; ATCC 43877) and Legionella brunensis (type strain 441-1; ATCC 43878).

Czechoslovakia↗
PubMed1981

Susceptibility of Pittsburgh pneumonia agent (Legionella micdadei) and other newly recognized members of the genus Legionella to nineteen antimicrobial agents.

The susceptibilities of 11 strains representing the five recognized species of Legionella were determined by agar dilution testing on buffered charcoal-yeast extract agar. All of the legionellae tested were susceptible to rifampin, erythromycin, rosaramycin, chloramphenicol, and the aminoglycosides and were resistant to clindamycin and vancomycin. Susceptibilities to penicillins and cephalosporins were variable. Legionella micdadei, Legionella bozemanii, and Legionella gormanii were susceptible to these agents, but minimal inhibitory concentrations for each species were different. Legionella dumoffii resembled Legionella pneumophila in being resistant to penicillin, cephalothin, and cephamandole and susceptible to moxalactam and cefoxitin. All species except L. micdadei produced beta-lactamase.

Anti-Bacterial Agents↗
PubMed1985

Legionella anisa: a new species of Legionella isolated from potable waters and a cooling tower.

Between March 1980 and June 1981, five strains of Legionella-like organisms were isolated from water. Four were recovered from potable water collected from hospitals in Chicago, Ill., and Los Angeles, Calif., during outbreaks of nosocomial legionellosis. The fifth strain was isolated from water collected from an industrial cooling tower in Jamestown, N.Y. The strains exhibited biochemical reactions typical of Legionella species and were gram-negative motile rods which grew on buffered charcoal-yeast extract agar but not on blood agar, required cysteine, and were catalase positive, urease negative, nitrate negative, hippurate negative, and nonfermentative. All strains were positive for oxidase and beta-lactamase and produced a brown, diffusible pigment. Of the five strains, four exhibited blue-white autofluorescence under long-wavelength UV light. The fatty-acid composition and ubiquinone content of these strains were consistent with those of other Legionella species. Direct fluorescent-antibody examination of the five strains with conjugates to previously described Legionella species demonstrated no cross-reactions except with the conjugates to L. longbeachae serogroup 2 and L. bozemanii serogroup 2. Four strains gave a 4+ reaction to the L. longbeachae serogroup 2 conjugate and the fifth strain gave a 1+ reaction. Each of the five strains gave a 4+ reaction with the conjugate to L. bozemanii serogroup 2. DNAs from the five strains were highly related (84 to 99%) and showed 5 to 57% relatedness to other Legionella species. These strains constitute a new species in the genus Legionella, and the name Legionella anisa sp. nov. is proposed. The type strain of L. anisa is WA-316-C3 (ATCC 35292).

Air Conditioning↗
PubMed1987

Purified 60-kilodalton Legionella protein antigen with Legionella-specific and nonspecific epitopes.

In a previous study, all convalescent-phase sera from patients with culture-confirmed legionellosis reacted on immunoblots with a Legionella genus-wide 58-kilodalton (kDa) protein antigen (J.S. Sampson, B.B. Plikaytis, and H.W. Wilkinson, J. Clin. Microbiol. 23:92-99, 1986). The present study was done to immunologically characterize and determine the diagnostic relevance of this purified antigen. The antigen was precipitated from enriched cell extracts with ammonium sulfate and purified by high-pressure liquid chromatography. High-titered rabbit antiserum produced to the purified protein was used to show its presence on immunoblots in the 60-kDa range in 38 Legionella serogroups, representing 23 species, and in 39 non-Legionella bacteria. The antiserum was made specific for Legionella strains by sequential absorptions with Bordetella pertussis, Pseudomonas aeruginosa, and Pseudomonas fluorescens whole cells. Serum from legionellosis patients reacted with both specific and nonspecific epitopes. Results of indirect immunofluorescence experiments showed that neither specific nor nonspecific epitopes of the 60-kDa protein were surface exposed on Legionella cells and that cross-reactive epitopes were variably exposed on non-Legionella bacteria. The 60-kDa protein antigen should be useful in diagnostic tests for legionellosis if care is taken to expose cryptic epitopes and if the tests use or measure only the Legionella-specific epitopes.

Antigens, Bacterial↗
PubMed2001

Legionella gresilensis sp. nov. and Legionella beliardensis sp. nov., isolated from water in France.

Novel Legionella-like isolates, strains Montbéliard A1T and Gréoux 11 D13T, isolated from two different French water sources, were studied taxonomically and phylogenetically. Morphological and biochemical characterization revealed that they were Gram-negative, aerobic, non-spore-forming bacilli with a cut-glass appearance that grew only on L-cysteine-supplemented buffered charcoal yeast extract agar. Phenotypic characterization using fatty acid and ubiquinone profiles and SDS-PAGE analysis confirmed that they were closely related, but distinct from, other species of the genus Legionella, since serotyping could not relate them to any existing serogroup. Genotypic profiles generated by randomly amplified polymorphic DNA and 16S-23S rDNA spacer region PCR analyses were unique for each of these isolates. DNA-DNA relatedness values of strains Montbéliard A1T and Gréoux 11 D13T to each other and to other Legionella type strains were less than 25%. Phylogenetic affiliation of these organisms obtained by 16S rDNA sequence comparisons confirmed that they were distinct from any other known Legionella species. All the above results confirm that these strains constitute two novel species for which the names Legionella gresilensis sp. nov. (type strain Gréoux 11 D13T = ATCC 700509T = CIP 106631T) and Legionella beliardensis sp. nov. (type strain Montbéliard A1T = ATCC 700512T = CIP 106632T) are proposed.

Bacterial Proteins↗
PubMed2005

Legionella waterline colonization: detection of Legionella species in domestic, hotel and hospital hot water systems.

AIMS: An evaluation was made of the prevalence of Legionella species in hot water distribution systems in the city of Bologna (Italy) and their possible association with bacterial contamination (total counts and Pseudomonadaceae) and the chemical characteristics of the water (pH, Ca, Mg, Fe, Mn, Cu, Zn and Total Organic Carbon, TOC). METHODS AND RESULTS: A total of 137 hot water samples were analysed: 59 from the same number of private apartments, 46 from 11 hotels and 32 from five hospitals, all using the same water supply. Legionella species were detected in 40.0% of the distribution systems, L. pneumophila in 33.3%. The highest colonization was found in the hot water systems of hospitals (93.7% of samples positive for L. pneumophila, geometric mean: 2.4 x 10(3) CFU l(-1)), followed by the hotels (60.9%, geometric mean: 127.3 CFU l(-1)) and the apartments with centralized heating (41.9%, geometric mean: 30.5 CFU l(-1)). The apartments with independent heating systems showed a lower level of colonization (3.6% for Legionella species), with no evidence of L. pneumophila. Correlation analysis suggests that copper exerts an inhibiting action, while the TOC tends to favour the development of L. pneumophila. No statistically significant association was seen with Pseudomonadaceae, which were found at lower water temperatures than legionellae and in individual distribution points rather than in the whole network. CONCLUSIONS: The water recirculation system used by centralized boilers enhances the spreading of legionellae throughout the whole network, both in terms of the number of colonized sites and in terms of CFU count. SIGNIFICANCE AND IMPACT OF THE STUDY: Differences in Legionella colonization between types of buildings are not due to a variation in water supply but to other factors. Besides the importance of water recirculation, the study demonstrates the inhibiting action of copper and the favourable action of TOC on the development of L. pneumophila.

Environmental Monitoring↗
PubMed2004

Phylogenetic characterization of Legionella-like endosymbiotic X-bacteria in Amoeba proteus: a proposal for 'Candidatus Legionella jeonii' sp. nov.

The X-bacteria which initiated organismic association with the D strain of Amoeba proteus in 1966 as parasites have changed to obligate endosymbionts on which the host depends for survival. Owing to the difficulty in cultivating the bacteria in vitro, the identity of X-bacteria has not been determined. As the life cycle of X-bacteria is similar to that of Legionella spp. in soil amoebae, we applied the polymerase chain reaction method with specific primers aimed at Legionella spp. for the detection and cloning of 16S rRNA gene. The identity and intracellular localization of the endosymbiont were confirmed by the application of a specific fluorescently labelled 16S rRNA-targeted probe. In addition we cloned RNA polymerase beta-subunit gene (rpoB) of X-bacteria by genomic library tagging. A phylogenetic analysis of the 16S rRNA gene placed the bacterium within a unique monophyletic group containing all other members of the genus Legionella. Phylogeny from rpoB and mip genes further confirmed the taxonomic context of X-bacteria to be a Legionella sp. In all three phylogenic analyses, X-bacterium was placed apart from Legionella-like amoebal pathogens present in soil amoebae. Thus, we propose the name 'Candidatus Legionella jeonii' sp. nov. for the endosymbiotic X-bacteria in Amoeba proteus.

Amino Acid Sequence↗
PubMed1990

Distribution of Legionella longbeachae serogroup 1 and other legionellae in potting soils in Australia.

Legionella longbeachae serogroup 1 and other Legionella spp. were isolated from 73% of 45 potting soils made in Australia by 13 manufacturers but were not detected in 19 potting soils made in Greece, Switzerland, and the United Kingdom examined between March 1989 and May 1990. Several Legionella species were isolated from a small number of samples of uncomposted pine sawdusts, but it is not known whether sawdust was the source of some of the legionellae found in potting soils. Legionella spp. persisted for periods ranging from 3 to 10 months in a potting soil held at temperatures between -20 and 35 degrees C. Isolates of L. longbeachae serogroup 1 from soil did not grow at 43 degrees C, a temperature which was also lethal for this species in soil. Most Legionella spp. isolated from potting and natural soils belonged to one distinct group according to analysis of ubiquinones and were serologically related to several known species in this group. A small number of potting soils contained L. pneumophila and L. micdadei.

Australia↗
PubMed2004

Mixed lung infection by Legionella pneumophila and Legionella gormanii detected by fluorescent in situ hybridization.

A mixed infection by Legionella pneumophila and a nonpneumophila Legionella species was detected in a lung biopsy specimen obtained from a patient with atypical pneumonia by fluorescent in situ hybridization (FISH). This result was confirmed by polymerase chain reaction (PCR). Sequencing of PCR products confirmed mixed infection by L. pneumophila and L. gormanii. Culture for Legionella spp. was negative and serology showed a rise only in IgG anti- Legionella pneumophila titer. To our knowledge, this is the first report of a mixed infection by L. pneumophila and a non-pneumophila Legionella species detected by FISH. Because FISH is a rapid and culture independent method that detects specific microorganisms in biopsy specimens it is recommended, in particular, for the detection of fastidious bacteria.

Aged↗
PubMed2000

Localization of Legionella bacteria within ribosome-studded phagosomes is not restricted to Legionella pneumophila.

In this report, we investigate the intracellular fate of selected members of the genus Legionella within the monocytic cell line Mono Mac 6 cells. By means of electron microscopy and immunocytochemistry, we could show that Legionella pneumophila as well as Legionella longbeachae are able to induce ribosome-studded phagosomes which associate with the rough endoplasmic reticulum (RER), whereas Legionella micdadei remains to be located within smooth phagosomes but also shows signs of RER association. In addition, we could demonstrate a remarkable correlation between the phagosome type and the morphological phenotype of intracellular bacteria: within ribosome-studded phagosomes, bacteria generally lacked the outer coat of low electron density whereas bacteria within the smooth phagosomes still possessed this outer coat. The virulence factors responsible for inhibition of phagosome maturation and their distribution within the genus Legionella as well as the biological significance of the morphological difference of bacteria within smooth and ER-associated phagosomes remain to be investigated.

Endoplasmic Reticulum↗