Mycobacterium pseudoshottsii sp. nov., a slowly growing chromogenic species isolated from Chesapeake Bay striped bass (Morone saxatilis) A group of slowly growing photochromogenic mycobacteria was isolated from Chesapeake Bay striped bass (Morone saxatilis) during an epizootic of mycobacteriosis. Growth characteristics, acid-fastness and 16S rRNA gene sequencing results were consistent with those of the genus Mycobacterium. Biochemical reactions, growth characteristics and mycolic acid profiles (HPLC) resembled those of Mycobacterium shottsii, a non-pigmented mycobacterium also isolated during the same epizootic. Sequencing of the 16S rRNA genes, the gene encoding the exported repeated protein (erp) and the gene encoding the 65 kDa heat-shock protein (hsp65) and restriction enzyme analysis of the hsp65 gene demonstrated that this group of isolates is unique. Insertion sequences associated with Mycobacterium ulcerans, IS2404 and IS2606, were detected by PCR. These isolates could be differentiated from other slowly growing pigmented mycobacteria by their inability to grow at 37 6C, production of niacin and urease, absence of nitrate reductase, negative Tween 80 hydrolysis and resistance to isoniazid (1 mg ml "1 ), p-nitrobenzoic acid, thiacetazone and thiophene-2-carboxylic hydrazide. On the basis of this polyphasic study, it is proposed that these isolates represent a novel species, Mycobacterium pseudoshottsii sp. nov. The type strain, L15 T , has been deposited in the American Type Culture Collection as ATCC BAA-883 T and the National Collection of Type Cultures (UK) as NCTC 13318 T .

An automated cellular fatty acid (CFA) bacterial identification system, Microbial Identification System (MIS; Microbial ID, Newark, Del.), was compared with a conventional system for the identification of 573 strains of gram-negative nonfermentative bacteria. MIS identifications were based exclusively on the CFA composition following 22 to 26 h of growth at 28°C on Trypticase soy agar. MIS identifications were listed with a confidence measurement (similarity index [SI]) on a scale of 0 to 1.0. A value of .0.5 was considered a good match. The MIS correctly listed as the first choice 478 of 532 (90%) strains contained in the data base. However, only 314 (59%) had SI values of 20.5. Of the 54 strains in which there was not agreement, 37 belonged to the genera Acinetobacter, Moraxella, or Alcaligenes or were Pseudomonas pickettii. Reproducibility studies suggest that SI variation is most likely a function of a difference in culture age at the time of analysis, which is due to the relatively low temperature and time of incubation. Other discrepancies were attributable to insufficiently characterized library entries or an inability to differentiate chemotaxonomically closely related species. The MIS, as the first automated CFA identification system, is an accurate, efficient, and relatively rapid method for the identification of gram-negative nonfermentative bacteria. The development of a CFA library with the media and incubation conditions routinely used for the isolation of clinical pathogens could further decrease the identification time and provide an increase in accuracy.

One hundred and ninety‐four strains of aerobically growing Gram‐positive rods of the genera Corynebacterium, Actinomyces, Arcanobacterium, Erysipelothrix, Listeria, Oerskovia, Nocardia, Rhodococcus, and of unnamed Center for Disease Control (CDC) groups were checked for cellular fatty acid profiles with the Microbial Identification System (Microbial ID, Newark, Del., USA). In order to obtain unified data usable for the clinical laboratory, 24 or 48 h sheep blood agar cultures were used. It was thought that grouping and perhaps identification could be aided by this approach. With the aid of numerical analysis, four groups (two consisting of two subgroups each) were established. The discriminatory ability of the scheme, however, was only 77.2%, indicating that grouping of an unknown isolate could be done with some accuracy, but that speciation would still require biochemical testing.

The characterization of a novel Mycobacterium sp. isolated from granulomatous skin lesions of moray eels is reported. Analysis of the hsp65 gene, small-subunit rRNA gene, rRNA spacer region, and phenotypic characteristics demonstrate that this organism is distinct from its closest genetic match, Mycobacterium triplex, and it has been named M. montefiorense sp. nov.Granulomatous skin disease has occurred sporadically and persistently within captive exhibit populations of moray eels, including green moray (Gymnothorax funebris) and spotted moray (G. moringa) eels (5), where it has led to eel death, reducing sizable aquarium collections of these fish. We have identified a mycobacterium as the etiologic agent of granulomatous skin disease in moray eels (5). On sequence analysis, the 16S rRNA gene of the eel Mycobacterium sp. was most closely related to that of Mycobacterium triplex, an opportunistic pathogen of humans (3, 4). The present report describes the phenotypic and genotypic characterization of this novel, slowgrowing, acid-fast bacterium and clarifies the taxonomy of this isolate in the genus Mycobacterium. On the basis of previous recommendations for the description of new mycobacteria (10) and the data presented here, we propose this as a new mycobacterium species called M. montefiorense.M. triplex (ATCC 700071) was obtained from the American Type Culture Collection (ATCC; Manassas, Va.). The Mycobacterium sp. previously described by Herbst et al. was stored at Ϫ70 o C (5). All phenotypic and genetic analyses were performed with biomass obtained from growth of this isolate on either Trypticase soy agar with 5% sheep blood or Middlebrook 7H10 agar. Type strains have been deposited at the American Type Culture Collection (ATCC BAA-256) and the Deutsche Sammlung von Mikroorganismen und Zellkulturen, Braunschweig, Germany (DSM-44602).As 16S ribosomal typing indicated that the organism isolated from moray eels was a Mycobacterium sp., acid-fast stains and biochemical tests were performed. Both the coccus form from blood agar and the rod form from Middlebrook medium were acid fast when stained with Kinyoun stain. The coccobacillary morphology of M. montefiorense on blood agar was consistent with the "streptococcus-like" organisms observed in previous studies of the pathology of eel skin lesions (5). Mycobacterial biochemical tests were performed by using standard methods as previously described (7). M. triplex has previously been demonstrated to be similar to M. simiae and M. avium complex (3) by conventional biochemical tests. Several differences in biochemical reactions were demonstrated for M. triplex and M. montefiorense (Table 1); e.g., M. triplex was urease positive and M. montefiorense was urease negative. There was a clear difference in temperature requirements and colony morphology. M. montefiorense would not grow at temperatures of Ͼ30°C. Growth from Middlebrook-based medium at 25°C demonstrated beaded acid-fast rods that were slow growing and nonchromogenic. The colonies were small and transparent. M...

Electrophoretic karyotype (EK) patterns, determined by using contour-clamped homogeneous pulsed-field electrophoresis, and isoenzyme (IZ) profiles were evaluated as methods for strain delineation among 35 isolates of Candida lusitaniae recovered from 15 patients. All isolates were identified to the species level by using conventional morphologic and physiologic criteria, and the identification was confirmed by gas-liquid chromatography analysis of the cellular fatty acids. The isolates were then typed without knowledge of the patient source. The IZ profiles showed all isolates to be closely related. Fifteen EK patterns were found; each pattern was restricted to isolates recovered from a single patient. In contrast, on the basis of heterogeneity in phosphatases, I-glucosidases, esterases, and catalases, 10 IZ profiles were found; 4 were shared by isolates recovered from more than one patient. Multiple isolates from six patients were analyzed, and for each patient, a single EK-and IZ-defined type was found. The types of isolates obtained from two patients, after the emergence of resistance to amphotericin B, remained the same as the types of isolates obtained earlier. The data suggest that a patient becomes colonized by a single strain of C. lusitaniae which may disseminate to multiple sites, that the colonizing strain can persist during the patient's hospitalization, and that it may develop resistance to amphotericin B. Both EK patterns and IZ profiles can be used to delineate strains of C. lusitaniae, but the EK pattern provides more discriminatory power.

CDC group IVc-2 is a gram-negative, oxidase-positive, nonfermentative bacillus that has been implicated in human infections, including septicemia and peritonitis. Biochemically it most closely resembles Bordetella bronchiseptica andAlcaligenes sp. Results of cellular fatty acid (CFA) and 16S rRNA gene analysis were combined with biochemical data to assist in identification and classification. The predominant CFAs were hexadecanoic acid (16:0), cis-9-hexadecanoic acid (16:1ω7c), cis-11-octadecanoic acid (18:1ω7c), and Δ-cis-9,10-methylenehexadecanoic acid (17:0cyc). Small amounts (2 to 5%) of 3-hydroxytetradecanoic acid (3-OH-14:0), tetradecanoic acid (14:0), 2-hydroxyhexadecanoic acid (2-OH-16:0), and Δ-cis-11,12-methyleneoctadecanoic acid (19:0cyc) were also consistently present. The highest 16S rRNA gene similarity was with Ralstonia eutropha and Ralstonia solanacearum. The CFA and 16S rRNA gene sequence data support the inclusion of CDC group IVc-2 in the recently created genusRalstonia, which includes R. eutropha, R. pickettii, and R. solanacearum.

Multidrug-resistant (MDR)Mycobacterium tuberculosis and extrensively drug-resistant (XDR) M. tuberculosis are emerging public health threats whose threats are compounded by the fact that current techniques for testing the susceptibility of M. tuberculosis require several days to weeks to complete. We investigated the use of high-performance liquid chromatography (HPLC)-based quantitation of mycolic acids as a means of rapidly determining drug resistance and susceptibility in M. tuberculosis. Standard susceptibility testing and determination of the MICs of drug-susceptible (n ‫؍‬ 26) and drug-resistant M. tuberculosis strains, including MDR M. tuberculosis strains (n ‫؍‬ 34), were performed by using the Bactec radiometric growth system as the reference method. The HPLC-based susceptibilities of the current first-line drugs, isoniazid (INH), rifampin (RIF), ethambutol (EMB), and pyrazinamide (PZA), were determined. The vials were incubated for 72 h, and aliquots were removed for HPLC analysis by using the Sherlock mycobacterial identification system. HPLC quantitation of total mycolic acid peaks (TMAPs) was performed with treated and untreated cultures. At 72 h, the levels of agreement of the HPLC method with the reference method were 99.5% for INH, EMB, and PZA and 98.7% for RIF. The inter-and intra-assay reproducibilities varied by drug, with an average precision of 13.4%. In summary, quantitation of TMAPs is a rapid, sensitive, and accurate method for antibiotic susceptibility testing of all first-line drugs currently used against M. tuberculosis and offers the potential of providing susceptibility testing results within hours, rather than days or weeks, for clinical M. tuberculosis isolates.Tuberculosis, one of the oldest diseases known to humans, remains a major public health threat worldwide. Two billion people, or one-third of the world's population, are infected with the causative agent, Mycobacterium tuberculosis (5). More than 9 million new cases of tuberculosis (TB) are reported each year, resulting in 2 million deaths (5). Drug-resistant M. tuberculosis strains are rapidly becoming the next global health emergency and are the harbingers of an impending pandemic (2,6,7,8,9,11,13,16,18,23). Much of the drug resistance worldwide is the result of inadequate or inappropriate therapy and is linked to the impossibly long and costly treatment courses, which have toxic side effects (8, 17). Multidrug resistance (MDR), defined as resistance to at least isoniazid (INH) and rifampin (RIF), is now common throughout the world, with average rates of resistance of 15% in high-burden countries (16, 23). Second-line drugs, which are generally more costly and more toxic than their first-line counterparts, have been the mainstay of therapy for MDR TB. However, this last line of defense is now failing. Recently, MDR M. tuberculosis strains with extensive resistance to second-line drugs have been identified throughout the world. These strains, known as extrensively drug-resistant (XDR) M. tuberculosis, threaten to move cont...

We report a case of a patient who developed cystitis caused by non-serogroup 01 Vibrio cholerae after swimming in the Chesapeake Bay. Treatment was empirical, with complete symptomatic resolution. Genitourinary tract infections by Vibrio spp. are uncommon but should be considered when cystitis occurs after saltwater exposure in appropriate geographic regions.