Isolation of Klebsiella ozaenae from a corneal abscess.

In this report, a case of bacterial keratitis caused by Klebsiella ozaenae is presented. The patient was an otherwise healthy 65-year-old woman with a history of chronic ocular inflammation and a corneal transplant two years prior to the appearance of the corneal ulcer. K. ozaenae is uncommon as a cause of keratitis and is recognized as a cause of atrophic rhinitis and as an opportunistic pathogen in patients with various underlying diseases. The organism was not identified with the computer-assisted API 20E for identification of the Enterobacteriaceae, and conventional methods were required to demonstrate its unique properties. The clinical spectrum of disease, the characteristics of the organism, and its susceptibility to antimicrobial agents are discussed.

Brevibacterium casei bacteremia and line sepsis in a patient with AIDS.

Brevibacteria are obligately aerobic gram-positive bacilli that are associated with milk products and are also found on human skin. Strains of Brevibacterium casei have been found to correspond to Centers for Disease Control coryneform groups B-1 and B-3 and have been isolated from a variety of human clinical specimens. In this report, we describe a case of B. casei bacteremia and sepsis in a patient with AIDS associated with a contaminated Hickmann catheter and review the microbiology and characteristics of these emerging opportunistic pathogens.

Negative images of mycobacteria in aspiration biopsy smears from the lymph node of a patient with acquired immunodeficiency syndrome (AIDS): report of a case and a review of the literature.

Negative images of acid-fast bacilli were observed in Diff-Quik-stained smears of lymph node aspirates from a patient with acquired immunodeficiency syndrome (AIDS) and disseminated Mycobacterium avium-intracellulare infection. The significance of this finding in relation to the diagnosis and treatment of this infection is discussed and the literature pertaining to this observation is reviewed.

Confirming positive results of nucleic acid amplification tests (NAATs) for Chlamydia trachomatis: all NAATs are not created equal.

The Centers for Disease Control and Prevention recommended confirming positive screening tests for Chlamydia trachomatis when positive predictive values are <90%. It is accepted that less sensitive tests (i.e., culture and immunoassays) should not be used to confirm the results of more sensitive nucleic acid amplification tests (NAATs). We show that the same principle applies when NAATs are used for confirmation.

Production of extracellular and cell-associated glucosyltransferase activity by Streptococcus mutans during growth on various carbon sources.

The production of extracellular and cell-associated glucosyltransferase activity by Streptococcus mutans strain GS-5 was examined during growth on various carbon sources in a chemically defined medium. S. mutans cells produced glucosyltransferase activity only during logarithmic growth when glucose, fructose, mannitol, or sorbitol was the sole carbon source. Cells growing on mannitol or sorbitol produced approximately half as much extracellular glucosyltransferase activity as cells growing on glucose, although the proportions of the glucosyltransferase activity capable of synthesizing insoluble glucans were similar. Cells growing on fructose produced slightly more extracellular glucosyltransferase activity than cells grown on glucose, yet the proportion of the glucosyltransferase activity capable of synthesizing insoluble glucans was again similar to glucose cultures. S. mutans cells growing in the presence of both glucose and mannitol displayed diauxic growth and initial preferential utilization of glucose. Glucosyltransferase enzyme production occurred only during the phases of cell growth in the presence of the two carbon sources. The cell-associated glucosyltransferase activities of glucose-, fructose-, mannitol-, and sorbitol-grown cells were relatively low, yet all the cells were capable of adherence to glass in the presence of sucrose. When glucose-containing cultures of S. mutans were supplemented with sucrose, extracellular glucosyltransferase activity first became cell associated and then appeared to become inactivated, presumably due to the accumulation of insoluble glucans.

Enhanced glucosyltransferase activity in penicillin-treated cultures of Streptococcus mutans.

Penicillin-treated cultures of Streptococcus mutants GS-5 produced elevated levels of extracellular glucosyltransferase activity.

Evaluation of Gonodecten for the presumptive diagnosis of gonococcal urethritis in men.

Gonodecten (Gd; U.S. Packaging Corp., LaPorte, Ind.) is a commercially available test for the presumptive diagnosis of gonococcal urethritis in men. With this test, urethral discharge is collected on a swab, placed in a plastic tube, and moistened with oxidase reagent. Development of a purple color on the swab within 3 min indicates the presence of gonococci. This system was compared with culture and Gram-stained smear for the diagnosis of gonorrhea. Of 240 specimens tested, 174 were positive with culture, Gram stain, and the Gd test and 48 were negative with all tests. For eight specimens, cultures and smears were positive, but the Gd test was negative. Nine specimens produced positive Gd tests with negative smears and cultures. Only one specimen was positive on culture and with the Gd test and negative on Gram-stained smear. Haemophilus species were isolated from 14 specimens; 5 produced positive Gd test reactions, with gonococci being coisolated from 4. The sensitivity and specificity of the Gd test compared with culture were 95.6 and 84.2%, respectively, whereas the Gram stain was 99.5% sensitive and 100% specific compared with culture. The Gd test may be a useful screening test for the diagnosis of gonorrhea in men with urethral discharge, particularly in settings where a microscope, Gram stain reagents, or technological expertise is not readily available.

Properties of a variant of Streptococcus mutans altered in its ability to interact with glucans.

After continuous subculture of Streptococcus mutans GS-5, the resultant organism, GS-5var, was found to be altered in its ability to undergo dextran- and sucrose-induced cell agglutination and to adhere to smooth surfaces in the presence of sucrose. Both GS-5 and GS-5var cells possessed the serotype c polysaccharide antigen, displayed similar growth kinetics, appeared to have similar metabolic and biosynthetic capabilities, and produced similar amounts of cell-associated and extracellular glucosyltransferase activity in chemically defined medium. GS-5 cells which had bound exogenous glucosyltransferase enzymes from either cell type were able to undergo sucrose-dependent adherence to smooth surfaces, whereas GS-5var cells which had bound comparable levels of glucosyltransferase activity were unable to adhere. In contrast to GS-5var cells, GS-5 cells were also able to adhere to preformed glucan deposits. In addition, [(14)C]glucan binding studies demonstrated that GS-5 cells were able to bind fourfold-greater levels of [(14)C]glucan relative to GS-5var cells having similar cell-associated glucosyltransferase activity. Further study of GS-5 cells also suggested the existence of a glucan binding site, distinct from the cell-associated glucosyltransferase activity, which is important in dextran-induced agglutination and sucrose-dependent adherence to smooth surfaces. These results are discussed in terms of the alteration of the GS-5var cells and their possible relationship to the putative receptors for glucosyltransferase enzymes and glucans on the cell surface of S. mutans.

Evaluation of a ten-minute chromogenic substrate test for identification of pathogenic Neisseria species and Branhamella catarrhalis.

A ten-minute chromogenic substrate test was evaluated for its ability to rapidly identify pathogenic Neisseria spp. and Branhamella catarrhalis. Identifications obtained with this system were compared to those obtained using conventional procedures. The test correctly identified 98.9% of 90 Neisseria gonorrhoeae, 98.3% of 60 Neisseria meningitidis, 96.2% of 26 Neisseria lactamica, and 100% of 36 Branhamella catarrhalis strains. Eight Neisseria subflava strains that grew on modified Thayer-Martin agar were prolyl aminopeptidase positive and were misidentified as Neisseria gonorrhoeae. Other strains of saprophytic Neisseria spp. also reacted with the chromogenic substrates. The system was accurate and reliable for identifying the commonly encountered pathogenic species. In light of recent reports describing new species and atypical Neisseria strains, however, careful attention to the salient features of both common and atypical organisms is necessary for proper use of rapid enzymatic identification tests.

Regulation and extracellular glucosyltransferase production and the relationship between extracellular and cell-associated activities in Streptococcus mutans.

The regulation of extracellular glucosyltransferase production in Streptococcus mutans GS-5 has been studied using a chemically defined medium. Most of the glucosyltransferase activity produced by cells grown in the chemically defined medium was extracellular, in contrast with the distribution between cell-associated and extracellular glucosyltransferase activity when cells were grown in complex medium. The production of extracellular glucosyltransferase activity coincided with the logarithmic growth phase, and further accumulation ceased when glucose was exhausted from the medium. Accumulation of extracellular glucosyltransferase activity was inhibited immediately by chloramphenicol and rifamycin, added either at the beginning of growth or during mid-logarithmic growth. Low concentrations of chloramphenicol inhibited both cellular protein synthesis and the accumulation of extracellular glucosyltransferase activity to the same extent, indicating a close coupling between glucosyltransferase synthesis and secretion. Experiments using cell lysates showed that no intracellular accumulation of glucosyltransferase activity occurred in the presence of the inhibitors and that the intracellular activity is very low relative to the cell-surface activity. The utilization of cells depleted of cell-associated glucosyltransferase activity indicated that most of the cell-associated glucosyltransferase activity does not act as a precursor for the extracellular enzyme. Sugar analogue inhibitors of glycoprotein synthesis did not have any specific effects on the synthesis or secretion of extracellular glucosyltransferase activity.

B.CAT CONFIRM, a rapid test for confirmation of Branhamella catarrhalis.

B.CAT CONFIRM (Scott Laboratories, Inc., Fiskeville, R.I.), a rapid test for detection of tributyrin hydrolysis, was evaluated for its ability to identify strains of Branhamella catarrhalis and to differentiate them from Neisseria species and related species. On initial testing, B.CAT CONFIRM was positive for 65 (96%) of 68 B. catarrhalis strains within 30 min after inoculation. Retesting of the remaining three strains resulted in their correct identification. B.CAT CONFIRM was negative for all Neisseria spp. (130 strains) and for Kingella spp. (3 strains). Two of the three Moraxella spp. were weakly positive in the B.CAT CONFIRM after 60 min, but these reactions were easily distinguishable from the strong reactions of B. catarrhalis strains. This test will be helpful in the clinical laboratory for the rapid identification of B. catarrhalis in clinical specimens.

Evaluation of RapiDEC Staph for identification of Staphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus saprophyticus.

RapiDEC Staph is a test for presumptive identification of the principal human staphylococcal species, Staphylococcus aureus, S. epidermidis, and S. saprophyticus. The test includes control and test cupules for fluorogenic detection of coagulase and chromogenic substrates for alkaline phosphatase and beta-galactosidase. These tests identify S. aureus, S. epidermidis, and S. saprophyticus, respectively. Positive results with both chromogenic substrates provide a presumptive identification of S. xylosus or S. intermedius (S. xylosus-S. intermedius). Test cupules are inoculated with an organism suspension, and reactions are read after a 2-h incubation. RapiDEC-Staph was evaluated with 303 clinical and stock staphylococcal strains. Identifications were compared with those obtained by the tube coagulase test, a latex slide coagulase test (StaphAUREX), another commercial identification system (Staph-TRAC), and additional conventional tests. RapiDEC-Staph correctly identified 100% of 130 S. aureus strains, 70.3% of 74 S. epidermidis strains, and 81.3% of 32 S. saprophyticus strains. Four of five S. xylosus isolates were called S. xylosus-S. intermedius. Unidentified S. epidermidis and S. saprophyticus strains were called "Staphylococcus spp." Among the 62 other coagulase-negative staphylococci, 4 were misidentified as S. epidermidis and 7 were misidentified as S. saprophyticus. While the sensitivity and specificity of the fluorogenic coagulase test for S. aureus were 100%, failure to detect alkaline phosphatase activity in several S. epidermidis isolates resulted in fewer correct identifications by the RapiDEC-Staph test for this species.

Evaluation of the BactiCard Neisseria for identification of pathogenic Neisseria species and Moraxella catarrhalis.

The BactiCard Neisseria (Remel, USA) is a chromogenic enzyme substrate system for identifying Neisseria gonorrhoeae, Neisseria meningitidis, Neisseria lactamica, and Moraxella catarrhalis. The identification system consists of a card with four test circles impregnated with chromogenic substrates for indoxyl butyrate esterase (IB), prolyl aminopeptidase (PRO), gamma-glutamyl aminopeptidase (GLUT), and ss-galactosidase (BGAL). These substrates permit the identification of Moraxella catarrhalis, Neisseria gonorrhoeae, Neisseria meningitidis, and Neisseria lactamica, respectively. After hydration of the circles with buffer, colonies from growth on selective media or a subculture are applied to the four circles. IB and BGAL reactions are read for a blue-green color after 2 and 15 min, respectively. PRO and GLUT reactions are read at 15 min for a red color after addition of a developer reagent. Identifications obtained with the BactiCard Neisseria were compared with those obtained using conventional procedures for 558 isolates in a blinded fashion. The BactiCard Neisseria identified 100% of 254 Neisseria gonorrhoeae, 100% of 125 Neisseria meningitidis, 53 (98.2%) of 54 Neisseria lactamica, and 123 (98.4%) of 125 Moraxella catarrhalis isolates. The BactiCard Neisseria is an accurate and rapid system for identification of these microorganisms in the clinical laboratory.

API QuadFERM+ with rapid DNase for identification of Neisseria spp. and Branhamella catarrhalis.

The QuadFERM+ system (Analytab Products, Plainview, N.Y.), a 2-h carbohydrate degradation method for the identification of Neisseria spp., was evaluated along with a rapid DNase test for confirmation of Branhamella catarrhalis. QuadFERM+ identified 100% of 82 N. gonorrhoeae and 96% of 54 N. meningitidis strains. The two misidentified meningococcal strains were biochemically atypical and were also misidentified by the conventional method. Of 26 N. lactamica strains, 25 (96%) were correctly identified. Of 21 Neisseria spp., 14 (67%) produced carbohydrate reactions in agreement with the conventional procedure, and 7 strains produced detectable acid in the QuadFERM+ from maltose and sucrose but not glucose. All 9 N. cinerea and 30 B. catarrhalis strains were asaccharolytic by QuadFERM+. The rapid DNase test was positive for all B. catarrhalis strains and negative for all other organisms. Two beta-lactamase-positive N. gonorrhoeae strains and 25 (93%) of 27 beta-lactamase-positive B. catarrhalis strains were detected by the 2-h acidometric beta-lactamase test on the strip. QuadFERM+ with rapid DNase is a simple and easily interpretable method for identification of these organisms in the clinical laboratory.

Clinical evaluation of the Vitek Neisseria-Haemophilus Identification card.

A clinical evaluation of the Vitek Neisseria-Haemophilus Identification (NHI) card (Vitek Systems, Inc., Hazelwood, Mo.) was performed with 480 clinical isolates and stock strains of Neisseria spp., Haemophilus spp., and other fastidious microorganisms included in the data base of the system. Identifications obtained with the NHI card were compared with those determined by conventional methods. The card identified 83.2% of 244 Neisseria spp. and Branhamella catarrhalis, 54.9% of 164 Haemophilus spp., and 84.7% of 72 fastidious gram-negative species with no further testing required. Some isolates produced good confidence-marginal separation identifications, in which the correct identification was listed with one or two other possible identifications and extra tests were required and suggested. When isolates producing good confidence-marginal separation identifications were included, correct identifications of these organism groups increased to 97.1, 92.7, and 94.4%, respectively. Among the commonly isolated microorganisms, the NHI card identified 99.1% of 110 N. gonorrhoeae, 98.5% of 68 N. meningitidis, 93.9% of 98 H. influenzae, and 95.6% of 46 H. parainfluenzae strains. All of these organisms produced excellent to very good confidence level identifications except for H. influenzae biotypes II, III, and VII, for which hemolytic reactions were required for differentiation from H. haemolyticus. The NHI card reliably identified other fastidious gram-negative species, including H. aphrophilus, Eikenella corrodens, Gardnerella vaginalis, and Kingella denitrificans.

Clinical evaluation of the Vitek ANI card for identification of anaerobic bacteria.

An evaluation of the Vitek Anaerobe Identification (ANI) card was performed with 341 bacterial isolates, including 313 clinical isolates and 28 stock strains of anaerobic microorganisms. Identifications obtained with the ANI card were compared with those determined by conventional methods. The card identified 73.2% of 149 anaerobic gram-negative bacilli, 63.6% of 44 Clostridium spp., 65.8% of 38 anaerobic nonsporeforming gram-positive bacilli, and 69.1% of 110 anaerobic cocci, with no further testing required. When genus-level identifications were included, 83.9% of the anaerobic gram-negative bacilli, 70.5% of Clostridium spp., 73.7% of the anaerobic nonsporeforming gram-positive bacilli, and 73.6% of the anaerobic cocci were identified. Nineteen isolates (5.6%) produced identifications of good confidence but marginal separation or questionable biotype, in which the correct identification was listed with one or two other possible choices and extra tests were required and suggested. A total of 28 (8.2%) were not identified and 29 isolates (8.5%) were misidentified by the ANI card. Among the commonly isolated clinically significant anaerobes, the ANI card identified 100% of 55 Bacteroides fragilis and 100% of 8 Clostridium perfringens. Use of supplemental tests and expansion of the data base to include additional strains of organisms that are difficult to separate even with conventional methods may improve the accuracy of the ANI card as a method for identification of anaerobic bacteria in the clinical laboratory.

Identification of Neisseria spp., Haemophilus spp., and other fastidious gram-negative bacteria with the MicroScan Haemophilus-Neisseria identification panel.

The Haemophilus-Neisseria identification (HNID) panel (American MicroScan, Sacramento, Calif.) is a 4-h microdilution format system for identification of Haemophilus and Neisseria spp., Branhamella (Moraxella) catarrhalis, and Gardnerella vaginalis. The HNID panel was evaluated by using 423 clinical isolates and stock strains of these organisms, and HNID identifications were compared with those obtained by conventional methods. In addition, 32 isolates representing six genera not included in the HNID data base were tested to determine whether these organisms would produce unique biotype numbers for possible inclusion in the data base. The HNID panel correctly identified 95.3% of 86 Neisseria gonorrhoeae strains, 96% of 25 G. vaginalis strains, and 100% of 28 Neisseria lactamica strains and 48 B. catarrhalis strains. Only 64.7% of 68 Neisseria meningitidis isolates were identified correctly owing to false-negative or equivocal carbohydrate and/or aminopeptidase reactions. Among the Haemophilus spp., 98.8% of 83 H. influenzae strains, 97.1% of 34 H. parainfluenzae strains, and 80% of 15 H. aphrophilus and H. paraphrophilus strains were correctly identified. Eight strains of Neisseria cinerea, a species not included in the data base, produced profiles identical with those for B. catarrhalis and N. gonorrhoeae. Isolates of other species not included in the data base, including Eikenella corrodens, Kingella spp., and Cardiobacterium hominis, produced unique biochemical reaction patterns on the panel. Modification of interpretative criteria for certain tests, expansion of the data base to include other species, and suggestions for additional confirmatory tests will increase the accuracy and utility of the HNID panel.

Use of the API NeIdent system for identification of pathogenic Neisseria spp. and Branhamella catarrhalis.

The API NeIdent system (Analytab Products, Plainview, N.Y.) was evaluated for identifying Neisseria spp. and Branhamella catarrhalis commonly isolated from clinical specimens. The system identified 90% of 303 Neisseria gonorrhoeae isolates, 71% of 113 Neisseria meningitidis isolates, and 63% of 16 Neisseria lactamica isolates but failed to identify any of 22 B. catarrhalis isolates. Testing of gonococcal strains of various auxotypes revealed no relationship between nutritional requirements and NeIdent profile numbers. With the Neisseria species, interpretation of the cinnamaldehyde-coupled beta-naphthylamine reactions was difficult and resulted in profile numbers not listed in the Profile Register. Positive resazurin-glucose reactions resulted in unlisted numbers for all B. catarrhalis strains. Inconsistent results were also obtained when 62 N. gonorrhoeae isolates were tested more than once on the strip. In all cases, profile variability and failure to identify these organisms were related to the beta-naphthylamide substrate tests. Expansion of the data base and modification of the substrate formulations or their interpretive criteria may increase the reliability of the NeIdent system for identifying Neisseria spp. and B. catarrhalis.

Asymptomatic Neisseria subflava biovar perflava bacteriuria in a child with obstructive uropathy.

A case of asymptomatic urinary tract infection with Neisseria subflava biovar perflava in a 10-year-old male patient with congenital structural abnormalities of the urinary bladder is presented. The organism was recovered from three catheter urine specimens collected over a seven-month period. A brief review of the role of saprophytic Neisseria species in infectious processes is presented and the likely source of this unusual urinary tract isolate is discussed.

Unexpected isolation of Bordetella pertussis from a blood culture.

Bordetella pertussis was isolated from a culture of blood from a 31-year-old man with Wegener's granulomatosis. The organism was detected with the BACTEC 9240 system after 6 days of incubation and was confirmed as B. pertussis by the Centers for Disease Control and Prevention. To our knowledge, this is the first published report of the recovery of B. pertussis from blood.