Showing papers in "Journal of Clinical Microbiology in 1986"

Sorbitol-MacConkey medium for detection of Escherichia coli O157:H7 associated with hemorrhagic colitis

Abstract: Escherichia coli serotype O157:H7 is a recently recognized human pathogen associated with hemorrhagic colitis. Unlike most E. coli strains, E. coli O157:H7 does not ferment sorbitol. Therefore, the efficacy of MacConkey agar containing sorbitol (SMAC medium) instead of lactose as a differential medium for the detection of E. coli O157:H7 in stool cultures was determined in comparison with MacConkey agar. The relative frequency of non-sorbitol-fermenting (NSF) organisms other than E. coli O157:H7 in feces was low at 10 to 20% (95% confidence limits), and NSF organisms also occurred mostly in small numbers. In a field trial involving over 1,000 diarrheal stools, E. coli O157:H7 was isolated from 18 stools, all of which were from patients with bloody diarrhea. In every instance, the growth of E. coli O157:H7 on SMAC medium was heavy and occurred in almost pure culture as colorless NSF colonies in contrast to fecal flora, which are mostly sorbitol fermenting and hence appear pink on this medium, whereas on MacConkey agar cultures, the growth of E. coli O157:H7 was indistinguishable from fecal flora. SMAC medium permitted ready recognition of E. coli O157:H7 in stool cultures. Detection of E. coli O157:H7 on SMAC medium had a sensitivity of 100%, a specificity of 85%, and an accuracy of 86%. SMAC medium stool culture is a simple, inexpensive, rapid, and reliable means of detecting E. coli O157:H7, and we recommend routine use of SMAC medium especially for culturing bloody stools.

Serum and nasal wash antibodies associated with resistance to experimental challenge with influenza A wild-type virus.

Abstract: To identify immunological predictors of resistance to influenza A infection and illness, the immunological status of live and inactivated virus vaccines subsequently challenged with H1N1 or H3N2 wild-type virus was examined. We refer to prechallenge antibodies of vaccinees receiving live attenuated virus as infection induced and those receiving inactivated virus as inactivated vaccine induced. Inactivated vaccine-induced protection against wild-type virus infection or illness correlated with the level of neuraminidase-inhibiting antibody in serum, local hemagglutinin immunoglobulin G (IgG) (but not IgA) enzyme-linked immunosorbent assay antibody, and hemagglutination-inhibiting antibody in serum. In contrast, infection-induced resistance to wild-type virus infection correlated with local hemagglutinin IgA antibody and neuraminidase-inhibiting antibody in serum, but not with hemagglutination-inhibiting antibody in serum. These observations suggest that live vaccine virus infection-induced and inactivated vaccine-induced immunity may involve different compartments of the immune system; sufficient antibody in either serum or nasal secretions is capable of conferring resistance.

The role of beta-lactamase in staphylococcal resistance to penicillinase-resistant penicillins and cephalosporins.

Abstract: We showed that most Staphylococcus aureus strains that have borderline or intermediate susceptibility to the penicillinase-resistant penicillins (PRPs) react this way because of the activity of their beta-lactamase on these antimicrobial agents These strains produced large amounts of staphylococcal beta-lactamase that rapidly hydrolyzed penicillin and partially hydrolyzed the PRPs Susceptibility to hydrolysis was penicillin greater than oxacillin greater than cephalothin greater than methicillin The borderline results and the hydrolysis could be prevented by the beta-lactamase inhibitors clavulanic acid and sulbactam For intrinsically methicillin-resistant (heteroresistant) S aureus, the inhibitors reduced the penicillin MICs, but the strains remained resistant to all the beta-lactam antimicrobial agents, including penicillin We conclude that the borderline in vitro susceptibility or resistance to PRPs in most of these S aureus strains is mediated by beta-lactamase and they are not heteroresistant or intrinsically resistant We do not know whether this in vitro resistance is expressed clinically

Dissociation between serum neutralizing and glycoprotein antibody responses of infants and children who received inactivated respiratory syncytial virus vaccine.

Abstract: The serum antibody response of infants and children immunized with Formalin-inactivated respiratory syncytial virus (RSV) vaccine 20 years ago was determined by using an enzyme-linked immunosorbent assay specific for the RSV fusion (F) and large (G) glycoproteins and a neutralization assay. Twenty-one young infants (2 to 6 months of age) developed a high titer of antibodies to the F glycoprotein but had a poor response to the G glycoprotein. Fifteen older individuals (7 to 40 months of age) developed titers of F and G antibodies comparable to those in children who were infected with RSV. However, both immunized infants and children developed a lower level of neutralizing antibodies than did individuals of comparable age with natural RSV infections. Thus, the treatment of RSV with Formalin appears to have altered the epitopes of the F or G glycoproteins or both that stimulate neutralizing antibodies, with the result that the immune response consisted largely of "nonfunctional" (i.e., nonneutralizing) antibodies. Subsequent natural infection of the vaccinees with wild-type RSV resulted in enhanced pulmonary disease. Despite this potentiation of illness, the infected vaccinees developed relatively poor G, F, and neutralizing antibody responses. Any or all of three factors may have contributed to the enhancement of disease in the RSV-infected vaccinees. First, nonfunctional antibodies induced by the inactivated RSV vaccine may have participated in a pulmonary Arthus reaction during RSV infection. Second, the poor antibody response of infants to the G glycoprotein present in the Formalin-inactivated vaccine may have been inadequate to provide effective resistance to subsequent wild-type virus infection. Third, the relatively reduced neutralizing antibody response of the infant vaccinees to wild-type RSV infection may have contributed to their enhanced disease by delaying the clearance of virus from their lungs.

Identification of Campylobacter pyloridis isolates by restriction endonuclease DNA analysis.

Abstract: Campylobacter pyloridis isolates recovered from gastric biopsy specimens of 16 patients were examined by restriction endonuclease DNA analysis with HindIII. For 8 of these 16 patients two different isolates were compared to study the persistence of the colonizing strains and the stability of their DNA digest patterns during a period of 2 years (two patients), the identity or nonidentity of different colony types within one culture (two patients), and the nature of the relapses after apparently successful antibacterial therapy (four patients). The isolates from the 16 patients all produced different DNA digest patterns. Comparison of the two different isolates recovered from the same patients showed that these isolates were identical in all eight cases. Laboratory subculturing of a C. pyloridis strain (10 times) did not change its DNA digest pattern. These results indicate the stability of the DNA digest patterns and a marked variability of these patterns among isolates from different patients. Using restriction endonuclease DNA analysis, we found the persistence in the stomach of the same C. pyloridis strain during a period of 2 years and the identity of different colony types within one culture. The relapses after apparently successful antibacterial treatment could be attributed to recrudescence rather than reinfection. Restriction endonuclease DNA analysis is a sensitive and useful method for identifying C. pyloridis isolates.

Detection of antibodies and antigens of human parvovirus B19 by enzyme-linked immunosorbent assay.

Abstract: Acute-phase serum from a patient with aplastic crisis provided sufficient human parvovirus B19 to make a monoclonal antibody against B19 and to develop antigen and immunoglobulin M (IgM) and IgG antibody detection enzyme-linked immunosorbent assays (ELISAs). The indirect capture antibody method was used for all three assays. Antigen was detected in 8 of 29 sera drawn within 2 days of onset of illness from patients with aplastic crisis. These sera had high titers of virus by electron microscopy and DNA hybridization and had no detectable B19 antibody. Antigen was not detected in serum specimens that had low titers of B19 DNA and had B19 antibody. With the IgM ELISA, we detected B19 IgM in over 85% of clinical cases of aplastic crisis and fifth disease and less than 2% of controls. The prevalence of B19 IgG antibodies increased with age. Approximately 2% of children less than 5 years of age and 49% of adults greater than 20 years of age had B19 IgG antibodies. The B19 antibody ELISAs are sensitive and specific tests to detect B19 infections.

Development and persistence of local and systemic antibody responses in adults given live attenuated or inactivated influenza A virus vaccine.

Abstract: An enzyme-linked immunosorbent assay was used to measure nasal-wash and serum isotype-specific hemagglutinin antibody responses in 109 seronegative (hemagglutination-inhibiting titer less than or equal to 1:8) adults vaccinated intranasally with live attenuated A/Washington/897/80 (H3N2) or A/California/10/78 (H1N1) cold-adapted (ca) virus or with licensed subvirion vaccine subcutaneously Live and inactivated virus elicited serum immunoglobulin A (IgA) responses in 83 and 96% of vaccinees, respectively, and elicited serum IgG responses in 72 and 100% of vaccinees Inactivated virus induced higher titers of serum antibodies than did live virus and stimulated a nasal-wash IgG response more often than did live virus (94 versus 59%, P less than 001) In contrast, only 38% of inactivated virus vaccinees had local IgA responses compared with 83% of live virus vaccinees Serum IgA and IgG and nasal IgG antibody titers remained elevated above prevaccination levels for at least 6 months in most of the live and inactivated vaccine responders, but the mean level of local IgA antibody induced by infection with live virus vaccine, in particular, decreased substantially Considered in the context of previous work, the finding that live virus vaccine induced relatively long-lasting antibody in both local and serum compartments suggested that this vaccine may be a suitable alternative to inactivated vaccine for use in healthy persons

Development of a standardized subgrouping scheme for Legionella pneumophila serogroup 1 using monoclonal antibodies.

Abstract: A panel of monoclonal antibodies to Legionella pneumophila serogroup 1 and a subclassification scheme were developed in a collaborative project among three laboratories. The seven most useful monoclonal antibodies were selected from three previously developed panels on the basis of indirect fluorescent antibody patterns with 83 strains of L. pneumophila serogroup 1 that were obtained from widely distributed geographic locations. The isolates were divided into 10 major subgroups on the basis of reactivity patterns that can be readily reproduced in any laboratory and are not subject to major inconsistencies of interpretation of staining intensity. A standard protocol for the indirect fluorescent antibody procedure was also developed.

Rapid microprocedure for isolating detergent-insoluble outer membrane proteins from Haemophilus species.

Abstract: A rapid microprocedure for isolating detergent (sodium N-lauroyl sarcosinate)-insoluble major outer membrane proteins from Haemophilus species produced results qualitatively identical to those obtained with a commonly used preparative isolation procedure. Proteins isolated by both procedures were compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis after staining with Coomassie brilliant blue R-250. The time for outer membrane protein isolation was substantially reduced with the rapid procedure, allowing a larger number of membrane preparations to be obtained rapidly for routine analysis. Images

Spheroplastic phase of mycobacteria isolated from patients with Crohn's disease.

Abstract: Two strains of an unclassified Mycobacterium species were isolated after 18 and 30 months of incubation of media inoculated with resected intestinal tissues from patients with Crohn's disease. These strains represented the third and fourth isolates of this organism from Crohn's disease patients. Ultrastructural examination of this strain and two previously isolated strains revealed the presence of spheroplasts which eventually transformed into the bacillary form of a previously unrecognized Mycobacterium species. These cell wall-deficient forms did not stain with conventional dyes and failed to grow on hypertonic media. Restriction polymorphism of the ribosomal DNA genes was used to determine the relationship between the cell wall-deficient and bacillary forms. Identical restriction patterns of the ribosomal DNA genes were found between the spheroplasts and Mycobacterium sp. isolates with EcoRI, BamHI, and XhoI restriction endonucleases, thus providing definitive evidence of their origin. Unidentified spheroplasts were isolated from an additional 12 patients with Crohn's disease, of which 7 of 10 seroagglutinated with antiserum prepared against the Mycobacterium sp. Spheroplasts were isolated from 16 of 26 (61%) patients with Crohn's disease but not from tissues of 13 patients with ulcerative colitis or 13 patients with other diseases of the bowel. These findings support the role of mycobacteria as etiologic agents in some cases of Crohn's disease.

Susceptibility testing of slowly growing mycobacteria by a microdilution MIC method with 7H9 broth.

Abstract: Based on previous success with rapidly growing mycobacteria, a microdilution MIC system was devised for slowly growing mycobacterial species using 7H9 broth Test drugs included isoniazid, rifampin, ethambutol, streptomycin, clofazamine, and sulfamethoxazole Sixty isolates of four mycobacterial species, including Mycobacterium tuberculosis, from patients who had never received drug therapy were evaluated in the system, as well as 25 drug-resistant isolates and 11 control strains MICs were read when good macroscopic control growth was evident, a period which varied with each species Most species exhibited a narrow range of MICs with easily discernible growth endpoints The aminoglycosides, ethambutol, clofazamine, and sulfamethoxazole were the only drugs with activity against all species at clinically achievable levels in serum Correlation between susceptibilities by the proportion method in agar with single drug concentrations and the broth method were excellent for M tuberculosis, M kansasii, and M marinum for isoniazid, rifampin, and ethambutol Isolates of the M avium complex were much more susceptible in broth than in agar for rifampin, ethambutol, and streptomycin Given the successful transition of most microbiology laboratories to MIC plates for other bacterial species, this method would allow for testing of multiple drugs at multiple concentrations and has good potential for evaluation of drug combinations and drug-resistant isolates

Emerging agents of phaeohyphomycosis: pathogenic species of Bipolaris and Exserohilum.

Abstract: Study of numerous living isolates of Bipolaris, Drechslera, Exserohilum, and Helminthosporium spp., as well as a mycological assessment of published case reports of phaeohyphomycosis attributed to these fungi, showed that Bipolaris australiensis, B. hawaiiensis, B. spicifera, Exserohilum longirostratum, E. mcginnisii, and E. rostratum are well-documented pathogens. Conidial shape, septation, and size, hilar characteristics, the origin of the germ tube from the basal cell and, to a lesser extent, from other conidial cells, and the sequence and location of the conidial septa are useful criteria for distinguishing these taxa. Images

Effect of age and preexisting antibody on serum antibody response of infants and children to the F and G glycoproteins during respiratory syncytial virus infection.

Abstract: The serum antibody response of 50 infants and children infected with respiratory syncytial virus (RSV) was determined by a glycoprotein-specific enzyme-linked immunosorbent assay, and the effects of age and preexisting antibody titer at the time of RSV infection on response to the G and F glycoproteins of RSV were examined The immune response to the G and F glycoproteins was assessed with anti-human immunoglobulin A to permit measurement of the response of young infants in the presence of maternally derived immunoglobulin G The findings suggested that age primarily affects the response to the F glycoprotein and that preexisting antibody titer affects the response to the G glycoprotein

Saliva, breast milk, and serum antibody responses as indirect measures of intestinal immunity after oral cholera vaccination or natural disease.

Abstract: The possibility that antibody responses in serum, saliva, or breast milk samples to oral vaccines or enteric infections may reflect the intestinal immune response was evaluated in Bangladeshi volunteers orally immunized with a cholera B subunit-whole-cell vaccine (B + WCV) and in patients convalescing from enterotoxin-induced diarrheal disease. Two peroral doses of B + WCV induced antitoxin and antibacterial antibody responses in the intestinal fluids of 76 and 92%, respectively, of the volunteers and in serum samples in 90 and 69% of those tested. These responses were comparable to those obtained after cholera or enterotoxigenic Escherichia coli disease. Whereas immunoglobulin A (IgA) antitoxin titer increases in saliva (44%) and breast milk (29%) specimens after vaccination were less frequent than in intestinal fluid (76%), antitoxin responses in saliva and breast milk occurred in 80 to 90% of the patients after disease. Also, antilipopolysaccharide (anti-LPS) titer increases in extraintestinal body fluids were found more frequently after disease than after vaccination. A comparison of the frequency and magnitude of antibody response in different body fluids with those in intestinal lavage fluid revealed no extraintestinal antibody that directly reflected the intestinal immunity. However, comparison of vibriocidal and IgG antitoxin antibodies in serum specimens with antitoxin and anti-LPS IgA responses in intestinal fluids after the vaccination of volunteers showed a sensitivity of 70 to 90% and a predictive accuracy of about 80% for the serum analyses reflecting the intestinal immune responses. Furthermore, antitoxin and anti-LPS antibody responses in saliva and breast milk samples seemed to be useful proxy indicators of a gut mucosal response of these antibodies after enterotoxin-induced diarrheal disease showing sensitivity vales of 70 to 90% and predictive accuracy vales of 70 to 100%.

Characterization of an organism that produces type E botulinal toxin but which resembles Clostridium butyricum from the feces of an infant with type E botulism.

Abstract: The apparent causative organism from the only reported case of type E infant botulism was isolated and characterized. Except for its ability to produce type E botulinal toxin, this organism (strain 5262) would be unquestionably identified as Clostridium butyricum. This is the second time an organism resembling a defined Clostridium species other than a member of the C. botulinum group has been implicated in infant botulism.

Identification of a new group of Chlamydia psittaci strains called TWAR.

Abstract: A new group of Chlamydia psittaci strains has been identified. They are called TWAR after the laboratory designation of the first two isolates. Twelve strains were isolated from pharyngeal swabs of different persons with acute respiratory disease in Seattle, Wash., during 1983 to 1986. One strain was obtained from the eye of a child during the trachoma vaccine study in Taiwan in 1965. Nine strains were characterized in this study. TWAR organisms formed intracytoplasmic inclusions in HeLa cells which were morphologically typical of C. psittaci and iodine stain negative (contained no glycogen). Immunological analysis with various chlamydia-specific monoclonal antibodies revealed that TWAR strains belong to the genus Chlamydia, are distinct from C. trachomatis, and are serologically unique among C. psittaci. All TWAR strains so far isolated appear identical serologically. TWAR organisms grew poorly in egg and cell cultures and demonstrated low virulence to mice by intracerebral, intranasal, and intravenous inoculation. Available data suggest that the TWAR strain is a primary human pathogen.

Monoclonal antibodies against Escherichia coli heat-stable toxin (STa) and their use in a diagnostic ST ganglioside GM1-enzyme-linked immunosorbent assay.

Abstract: Seven monoclonal antibodies (MAbs) against heat-stable enterotoxin (ST) from a human Escherichia coli isolate were prepared and evaluated for their usefulness in an ST immunodetection assay, the ST ganglioside GM1-enzyme-linked immunosorbent assay (ELISA). This assay is based on the ability of STa, as present in, for example, culture filtrates from ST-producing E. coli, to inhibit specific anti-ST antibody from binding to solid-phase-bound ST ganglioside (GM1-bound ST-cholera B subunit). Four of the MAbs were of immunoglobulin G1 (IgG1), one was of IgG2b, and two were of IgM isotype. All the IgG1 MAbs could be completely inhibited by addition of free ST; 0.2 to 0.4 ng of purified ST inhibited binding of these MAbs by 50%. The non-IgG1 MAbs were, in contrast, not inhibited by 200-fold-higher amounts of purified ST, probably because they were directed against linkage epitopes or were of low affinity or both. When the IgG1 MAbs were tested in the ST GM1-ELISA, ST could be detected in culture filtrates from stock human E. coli isolates with 100% sensitivity and specificity. ST in filtrates from fresh stool cultures was demonstrated with higher sensitivity with the MAbs ST GM1-ELISA than with the conventional infant mouse test. Both subtypes of STa, STaI and STaII, could be detected by the ST GM1-ELISA by using either IgG1 MAb in the immunodetection step, whereas infant-mouse-active ST from Yersinia enterocolitica failed to react.

Is semiquantitative culture of central vein catheter tips useful in the diagnosis of catheter-associated bacteremia?

Abstract: Semiquantitative culturing of catheter tips has been used as an index of catheter-related bacteremia. As the sensitivity and predictive values of this test have not been determined, we studied 780 tips from central vein catheters inserted into 440 critically ill patients in an intensive care unit. The results were correlated with clinical data for 30 bacteremic episodes which occurred in these patients, 14 of which were catheter related. When five or more colonies per plate were taken as a positive result, the sensitivity of the method was 92%, and the specificity was 83%. Although the predictive value of a negative result was excellent (99.8%), the predictive value of a positive result was low (8.8%) in our patient population, which had a relatively low incidence of catheter-related bacteremia (2%). We conclude that a semiquantitative culture technique is useful in the diagnosis of bacteremia associated with central vein catheters.

Serum and nasal-wash immunoglobulin G and A antibody response of infants and children to respiratory syncytial virus F and G glycoproteins following primary infection.

Abstract: An enzyme-linked immunosorbent assay (ELISA) with immunoaffinity-purified fusion (F) or attachment (G) glycoprotein was used to measure the serum and secretory immune responses of 18 infants and children, 4 to 21 months of age, who underwent primary infection with respiratory syncytial virus (RSV). Most of the 10 older individuals (9 to 21 months of age) developed moderate levels of serum and nasal-wash immunoglobin A (IgA) and IgG F and G antibodies. These individuals developed a moderate level of serum or nasal-wash antibodies that neutralized virus infectivity. One of the eight younger individuals (4 to 8 months of age) failed to develop an F antibody response, while three failed to develop a G antibody response. The most notable difference in the responses of the two age groups involved the titer in convalescent sera of G, F, and neutralizing antibodies which were 8- to 10-fold lower in younger individuals. Most of the younger infants failed to develop a rise in serum or nasal-wash neutralizing antibody. It is possible that the presence of maternally derived antibody in the younger infants suppressed the immune response to RSV infection, and that this accounted, in part, for the low level of postinfection antibody titer in this group. This low level and the irregular response of the infants less than 8 months of age may contribute to the severity of their initial infection and may also be responsible, in part, for their failure to develop effective resistance to subsequent reinfection by RSV.

Phenotypic comparison of Pseudomonas aeruginosa strains isolated from a variety of clinical sites.

Abstract: Pseudomonas aeruginosa elaborates a number of extracellular products which have been shown to play a role in the pathogenesis of disease caused by this organism. In this study, we showed that the host environment markedly affects the levels of exoproducts produced. We compared the phenotypes of a number of P. aeruginosa strains obtained from a variety of clinical sources, including burn wounds, skin wounds, urine, cystic fibrosis sputum, acute pneumonia sputum, and blood. The clinical isolates were examined quantitatively for levels of total protease, elastase, phospholipase C, exotoxin A, and exoenzyme S produced in vitro under defined conditions. The exoproduct levels varied significantly, depending on the site of isolation. Elevated levels of elastase were demonstrated in strains isolated from acute lung infections, phospholipase C levels were elevated in urinary tract and blood isolates, exotoxin A levels were elevated in blood isolates, and exoenzyme S levels were increased in acute pneumonia isolates. Isolates from cystic fibrosis sputum produced low amounts of virtually all of the tested exoproducts, particularly as compared with sputum isolates from acute P. aeruginosa lung infections.

Association of Blastocystis hominis with signs and symptoms of human disease.

Abstract: Purged stools from 389 patients were evaluated microscopically for the presence of Blastocystis hominis. A total of five or more B. hominis cells per 40X field were observed in 43 patients (11%), and B. hominis was the only intestinal parasite present in 23 (6%) of these patients. Of the 23 patients, 19 had symptoms which included abdominal discomfort (15 patients), anorexia (10 patients), diarrhea (9 patients), and flatus (9 patients). The remaining four patients were asymptomatic. The proportion of eosinophils in the peripheral blood ranged from 4 to 12% in 11 (58%) of the symptomatic patients. Absolute eosinophil counts were greater than 250/microliter in 8 patients and greater than 400/microliter in 5 patients. Eosinophilia was not observed in the remaining symptomatic or asymptomatic patients. This study supports the emerging concept of the role of B. hominis as an intestinal parasite causative of human disease.

Autoclaved liquid medium for propagation of Treponema hyodysenteriae.

Abstract: Three liquid media that differ slightly in composition but not in the method of preparation were developed for the propagation of Treponema hyodysenteriae and Treponema innocens. The three media are unique in that all components are sterilized by autoclaving before use. These media supported better growth of T. hyodysenteriae than did previously used liquid media.

Comparison of 15 laboratory and patient-derived strains of Mycobacterium avium for ability to infect and multiply in cultured human macrophages.

Abstract: Mycobacterium avium is a cause of nontuberculous chronic granulomatous infections which is attracting increased attention as a frequent opportunistic pathogen in acquired immunodeficiency syndrome. Some important aspects of its human pathogenicity were investigated by using cultured human macrophages infected with it. The uptake and replication of various strains of M. avium in the macrophages could be measured by CFU counts of the bacteria in samples of lysed, sonicated macrophages. Microscopic counts of acid-fast bacilli were not useful because the bacteria multiplying in the macrophages were usually not acid fast. Electron microscopy showed the intracellular bacilli to multiply by transverse fission, to be surrounded in individual vacuoles by a broad electronlucent zone, and to have thinner cell walls than extracellularly grown M. avium. Fifteen strains, including examples of serovars 1, 2, 4, 8, and 9, were studied for uptake and rate of replication in cultured macrophages from three normal subjects. The strains were isolates from patients with nontuberculous granulomatous infection, acquired immunodeficiency syndrome, or unrelated problems, or they were laboratory reference cultures. There were no differences among them in phagocytosis, but there were differences in intracellular replication. Laboratory strains tended to be avirulent, that is, they did not replicate in the macrophages. Patient isolates usually were virulent and could be compared for virulence by intracellular replication rates. Virulence correlated with flat, transparent bacterial colony morphology on nutrient agar but not with serovar or kind of patient from whom the bacteria were isolated. However, among strains of transparent colony morphology there were wide differences in virulence. A virulent bacilli generally produced domed, opalescent colonies on nutrient agar. A virulent bacilli predominated in populations of M. avium conditioned to growth in bacteriologic culture medium. Bacilli of virulent colony morphology predominated in populations passaged through cultured macrophages. The model described here presents a new approach to the investigation of the pathogenicity of M. avium for human subjects and may be more patient relevant than animal models.

Enterobacter asburiae sp. nov., a new species found in clinical specimens, and reassignment of Erwinia dissolvens and Erwinia nimipressuralis to the genus Enterobacter as Enterobacter dissolvens comb. nov. and Enterobacter nimipressuralis comb. nov.

Abstract: Enterobacter asburiae sp. nov. is a new species that was formerly referred to as Enteric Group 17 and that consists of 71 strains, 70 of which were isolated from humans. Enterobacter asburiae sp. nov. strains gave positive reactions in tests for methyl red, citrate utilization (Simmons and Christensen's), urea hydrolysis, L-ornithine decarboxylase, growth in KCN, acid and gas production from D-glucose, and acid production from L-arabinose, cellobiose, glycerol (negative in 1 to 2 days, positive in 3 to 7 days), lactose, D-mannitol, alpha-methyl-D-glucoside, salicin, D-sorbitol, sucrose, trehalose, and D-xylose. They gave negative reactions in the Voges-Proskauer test and in tests for indole, H2S production, phenylalanine, L-lysine decarboxylase, motility, gelatin, utilization of malonate, lipase, DNase, tyrosine clearing, acid production from adonitol, D-arabitol, dulcitol, erythritol, i(myo)-inositol, melibiose, and L-rhamnose. They gave variable reactions in tests for L-arginine dihydrolase (25% positive after 2 days) and acid production from raffinose (69% positive after 2 days). Thirty-four Enterobacter asburiae sp. nov. strains were tested for DNA relatedness by the hydroxyapatite method with 32PO4-labeled DNA from the designated type strain (1497-78, ATCC 35953). The strains were 69 to 100% related in 60 degrees C reactions and 63 to 100% related in 75 degrees C reactions. Divergence within related sequences was 0 to 2.5%. Relatedness of Enterobacter asburiae sp. nov. to 84 strains of members of the Enterobacteriaceae was 5 to 63%, with closest relatedness to strains of Enterobacter cloacae, Erwinia dissolvens, Enterobacter taylorae, Enterobacter agglomerans, Erwinia nimipressuralis, and Enterobacter gergoviae. All strains tested were susceptible to gentamicin and sulfdiazine, and most were susceptible to chloramphenicol, colistin, kanamycin, nalidixic acid, carbenicillin and streptomycin. All strains were resistant to ampicillan, cephalothin, and penicillin, and most were resistant or moderately resistant to tetracycline. Enterobacter asburiae sp. nov strains were isolated from a variety of human sources, most prevalent of which were urine (16 strains), respiratory sources (15 strains), stools (12 strains), wounds (11 strains), and blood (7 strains). The clinical significance of Enterobacter aburiae is not known. As a result of this and previous studies, proposals are made to transfer Erwinia dissolvens and Erwinia nimipressuralis to the genus Enterobacter as Enterobacter dissolvens comb. nov. and Enterobacter nimipressuralis comb. nov., respectively.

Increased incidence of fungemia caused by Candida krusei.

Abstract: Candida krusei colonized 12.4% of 868 patients undergoing episodes of therapy-induced granulocytopenia over a 9-year period. The gastrointestinal tract was most frequently colonized, followed by the respiratory tract and urinary tract. Ten patients developed systemic infections with C. krusei; all 10 had two or more positive blood cultures. Nine of the 10 patients were colonized with C. krusei, and 6 were receiving systemic antifungal agents at the time of development of the infection. Seven patients died within 1 month of C. krusei sepsis; systemic candidiasis was seen in the autopsies of the four patients on whom autopsies were performed. Therefore, C. krusei should be recognized as an emerging pathogen in select patient populations.

Evaluation of a blood-free, charcoal-based, selective medium for the isolation of Campylobacter organisms from feces.

Abstract: A blood-free, charcoal-based selective medium (CSM) consisting of a Columbia agar base, activated charcoal (4 g/liter), hematin (0.032 g/liter), sodium pyruvate (0.1 g/liter), cefoperazone (32 mg/liter), vancomycin (20 mg/liter), and cycloheximide (100 mg/liter) supported the growth of Campylobacter jejuni and C. coli with colony counts equivalent to those obtained on antibiotic-free horse blood agar. CSM was compared to Skirrow medium (SKM) for the recovery of C. jejuni and C. coli from stools of patients with diarrhea, the media being incubated for 2 days under reduced oxygen tension at 43 degrees C. These campylobacters were isolated from 35 (2.9%) of 1,227 stools tested (29 on both media, 5 on CSM alone, and one on SKM alone). Whenever C. jejuni and C. coli were recovered, growth was pure on 29 CSM cultures (85%), but on only 11 SKM cultures (37%). Complete suppression of "contaminating" flora occurred in 704 CSM cultures (57%) compared with 426 SKM cultures (35%). CSM more effectively suppressed contaminating pseudomonads, gram-positive organisms, and yeasts than did SKM; both media failed to suppress members of the family Enterobacteriaceae in about a quarter of the samples. Studies on 20 representative Enterobacteriaceae contaminants showed that susceptibility to cefoperazone and growth on CSM were markedly dependent on inoculum size; 12 strains were inhibited by cefoperazone (32 mg/liter) at inoculum sizes of 5 X 10(2) and 5 X 10(4) but not 5 X 10(6) organisms, indicating that the frequency of contaminants on CSM could probably be reduced further by ensuring that stools were not inoculated too heavily on CSM. Our findings confirm that charcoal is an effective substitute for blood in media for growing campylobacters, and that CSM is a highly effective blood-free selective medium for isolating C. jejuni and C. coli from stools.

Associated mortality and clinical characteristics of nosocomial Pseudomonas maltophilia in a university hospital.

Abstract: We studied the spectrum of clinical disease in 99 patients with nosocomial Pseudomonas maltophilia isolates at the University of Virginia Hospital from 1981 through 1984. The annual rate of isolation increased from 7.1 to 14.1 per 10,000 patient discharges. A crude mortality rate of 43% was documented in all patients from whom the organism was cultured, and the data include 12 patients with nosocomial bacteremia (four deaths). Risk factors associated with death for patients having a P. maltophilia isolate included the following: requirement for care in any intensive care unit during hospitalization (P = 0.0001), patient age over 40 years (P = 0.002), and a pulmonary source for the P. maltophilia isolate (P = 0.003). All P. maltophilia isolates were susceptible to trimethoprim-sulfamethoxazole, 60% of the isolates were resistant to all aminoglycosides (amikacin, tobramycin, and gentamicin), and more than 75% of the isolates were resistant to all beta-lactam antibiotics. The antibiotic susceptibility pattern allows for a niche exploitable in the hospital microbial environment by an organism with a marked associated mortality.

Seroepidemiology of Klebsiella bacteremic isolates and implications for vaccine development.

Abstract: The frequencies of capsular serotypes among 703 Klebsiella strains isolated from the blood of hospitalized patients were determined. More than 90% of the isolates were typeable, with 69 of the 77 known serotypes being identified. Serotypes 2, 21, and 55, representing 8.9, 7.8, and 4.8% of all the isolates, respectively, were observed at a frequency significantly higher (P less than 0.05) than that for other capsular serotypes. Approximately 43% of the serotypes appeared at a frequency of less than 0.5%. Differences were found when the seroepidemiology of North American and European isolates was compared. The current findings indicate that a capsular polysaccharide-based vaccine against Klebsiella organisms is feasible and should be multivalent, eliciting antibodies directed against the 25 serotypes which make up approximately 70% of all the bacteremic isolates.

Reevaluation of the yeast killer phenomenon.

Abstract: The killer effect of 36 Hansenula, Pichia, Saccharomyces, and Candida species on 26 hyphomycetes isolates, 1 isolate of the achlorophyllous microorganism Prototheca, 4 isolates of the lipophilic yeast Malassezia, 1 isolate of the aerobic actinomycete Nocardia, and 19 isolates of bacteria was studied. The killer phenomenon, which was previously considered to be restricted to yeasts, was found to occur among unrelated microorganisms.

Group C rotaviruses in humans.

Abstract: Atypical rotaviruses obtained from human feces from Australia, Brazil, and the United Kingdom were shown by a combination of techniques--immunoelectron microscopy, immunofluorescence, genome profile analysis, terminal fingerprint analysis of genome segments, and dot-blot hybridization--to be related to group C porcine rotaviruses. The prevalence of antibody to group C rotaviruses was found to be low in human sera and immunoglobulin pools from six countries. No signs of infection were obtained when one of the human viruses was inoculated into gnotobiotic piglets. We conclude that the atypical human viruses are the first examples of group C rotaviruses in humans.