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.

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.

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.

Citrobacter sedlakii meningitis and brain abscess in a premature infant.

Citrobacter sedlakii was isolated from blood and cerebrospinal fluid cultures of a 5-day-old premature infant with sepsis, meningitis, and brain abscess. This newly described organism was difficult to identify due to discrepancies between the Vitek and API 20E identification systems. To our knowledge, this is the first report of the isolation of C. sedlakii from cerebrospinal fluid.

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.

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.

In vitro study of amniotic fluid gram stain: effect of centrifugation.

OBJECTIVE: Gram stain of amniotic fluid (AF) is used to detect intraamniotic infection. The purpose of this study was to determine if centrifugation improved the ability of AF Gram stain to detect bacteria. METHODS: AF obtained by amniocentesis from patients with preterm labor (PTL) or preterm premature rupture of membranes (PPROM) was pooled. Individual AF samples as well as the pooled sample had a negative Gram stain for microorganisms or white blood cells (WBCs) and negative cultures. With pure bacterial cultures, a suspension equivalent to a 0.5 McFarland turbidity standard was prepared and then serially diluted in the AF to either 10(6), 10(5), 10(4), or 10(3) colony forming units (cfu)/ml. Each sample was divided into 2 equal portions, with 1 undergoing centrifugation. The Gram stains were interpreted by technologists in the clinical microbiology laboratory in a blinded fashion. Fisher's exact test was used to compare the bacterial detection rate in centrifuged vs. uncentrifuged AF samples at each concentration. RESULTS: Centrifugation of AF significantly improved the ability of the Gram stain to detect bacteria at bacterial concentrations < or =10(4) cfu/ml (P < 0.01). At concentrations > or =10(5) cfu/ml, centrifugation did not improve the ability of the Gram stain to dtect bacteria. CONCLUSIONS: At low bacterial concentrations, centrifugation of AF increases the bacterial detection rate of AF Gram stain.

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.

Comparison of monoclonal antibody methods and a ribosomal ribonucleic acid probe test for Neisseria gonorrhoeae culture confirmation.

Recently, a chemiluminescent nucleic acid probe test that specifically detects the ribosomal ribonucleic acid of Neisseria gonorrhoeae has been released for clinical laboratory use (AccuProbe Neisseria gonorrhoeae). In this study, three coagglutination tests (GonoGen I, Meritec GC, and GC Omni), the GonoGen II immunofiltration method and the Micro Trak Neisseria gonorrhoeae fluorescent monoclonal antibody test were compared with AccuProbe for identification of gonococci. Strains tested (n = 376) included 194 Neisseria gonorrhoeae, 82 Neisseria meningitidis, 32 Neisseria lactamica, 32 Neisseria species, 32 Moraxella catarrhalis, 2 Moraxella spp. and 2 Kingella denitrificans. The GonoGen I, Meritec GC and GC Omni coagglutination tests produced clearly positive results for 93.8%, 92.3% and 95.9% of the gonococci, respectively. The GonoGen II unequivocally identified 91.8% and the MicroTrak fluorescent antibody test identified 90.7% with 2+ or greater fluorescence. AccuProbe identified 100% of the gonococci tested. GonoGen I and GonoGen II were 98% specific, Meritec GC was 99% specific and the specificity of the GC Omni, MicroTrak fluorescent antibody and AccuProbe tests was 100%. While antibody-based tests were reliable when results were clearly interpretable, the AccuProbe was the only confirmatory test that was 100% accurate. Serotyping studies indicate that an array of beta-lactamase positive and negative gonococcal serotypes fail to react with the monoclonal antibody-based tests in general and with the fluorescent antibody test in particular.

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.

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.

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.

Osteomyelitis caused by Rhodococcus equi in a renal transplant recipient.

We report the first case of osteomyelitis due to Rhodococcus equi, which occurred in a renal transplant patient. Infection with this organism is rare and usually causes a distinct clinical syndrome resembling pulmonary tuberculosis. We investigated by time-kill curve analysis various antimicrobial combinations for in vitro efficacy. The literature is briefly reviewed, and aspects of diagnosis and therapy are discussed.

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.

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.

Comparison of the Gono-Pak system with the candle extinction jar for recovery of Neisseria gonorrhoeae.

Maximum CO2 levels in the Gono-Pak system were obtained 2 h after activation, with a mean value of 1.5%. Although this is less than the 2 to 3% CO2 level obtained with the candle jar, Gono-Pak produced comparable recovery of Neisseria gonorrhoeae in 357 clinical specimens and stock cultures. Using adjustable CO2 incubators, we found the recovery at 1% CO2 to be comparable to that at 5%, whereas 10% CO2 was inhibitory.

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.

Prevalence of enteric parasites in homosexual patients attending an outpatient clinic.

A total of 372 pooled stool specimens from 274 homosexual men with diarrhea were submitted for parasitologic examination over a 2.5-year period. Each two-vial pooled specimen set contained portions of stool from 3 consecutive days in Formalin and polyvinyl alcohol. Of the 274 patients, 133 (48.5%) harbored one or more intestinal protozoa, with 161 (43.3%) of the 372 specimens submitted being positive for one or more organisms. The parasites identified included Entamoeba histolytica (71 patients), Giardia lamblia (22 patients), Endolimax nana (106 patients), Entamoeba coli (39 patients), Entamoeba hartmanni (25 patients), Dientamoeba fragilis (3 patients), Iodamoeba bütschlii (2 patients), and Chilomastix mesnili (2 patients). Cryptosporidium sp. (2 patients) and Isospora belli (1 patient) were also detected. Results of this study support the experience of other workers regarding high rates of infection with intestinal parasites in the homosexual population and also indicate that symptomatic individuals belonging to this acquired immunodeficiency syndrome risk group be screened for both common and uncommon intestinal pathogens.