Clonal dissemination of vancomycin-resistant Enterococci and comparison of susceptibility testing methods.

One hundred fifty clinical isolates of Enterococcus faecalis (88 isolates) and Enterococcus faecium (62 isolates) were tested in vitro for their susceptibility to vancomycin and high-level aminoglycosides (HLA). Remel's Synergy Quad Plates (RSQ) were used as the reference method and compared to Kirby-Bauer disc diffusion test, Vitek GPS-TA card, MicroScan Panel (GP-6), and Etest. Streptomycin susceptibility results for MicroScan GP-6 and RSQ were recorded at 24 and 48 h and all other methods and antibiotics were read at 24 h or less. When compared with the agar screen method, all of the methods demonstrated > 99% agreement. One isolate was falsely sensitive to gentamicin at 24 h, but resistant at 48 h, when tested on both MicroScan and RSQ agar screen. Thirty-nine isolates showed resistance to vancomycin with all methods. These isolates were from three different local hospitals and were identified as E. faecium. Pulse-field gel electrophoresis demonstrated that all of the vancomycin-resistant isolates were derived from the same clone. Of interest is the observation that high-level resistance to aminoglycosides varied between the clonally related isolates.

Are serum levels of vancomycin useful in the first week of therapy?

Controversy exists regarding the need to monitor serum concentrations of vancomycin with some investigators recommending measurement of peak and trough concentrations in the first week of therapy and regularly thereafter, whereas others contend that empiric dosing produces safe and effective drug concentrations so that testing is unnecessary. Since vancomycin concentrations are measured, routinely in our hospital in the first week of therapy, we conducted a 12 month study to assess their clinical value in patients who were treated when gram positive cocci were detected in blood culture smears. One-hundred-five patients had gram positive cocci on blood culture smears. These bacteria were pathogens in 15 patients with Staphylococcus aureus and in 18 with coagulase negative staphylococci based on microbiologic criteria and a chart review confirming their clinical significance. Ten patients with S. aureus and 8 patients with coagulase negative staphylococci that were pathogens and 10 patients with coagulase negative staphylococci that were contaminants were treated with vancomycin. Serum peak and trough concentrations of vancomycin obtained within the first 5 days of therapy in these 28 patients were 14 to 40 micrograms/ml and 4.8 to 20 micrograms/ml. These concentrations were much above the MIC's of the microorganisms (< 4 micrograms/ml). Five patients had increases of serum creatinine of more than 0.6 mg% and in each patient the increases were attributable to other causes-shock, heart failure, and preexisting renal failure. Fifty five peak and trough concentrations 19 of which were drawn in patients with contaminated cultures were measured at a cost of $2,475.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of two culture approaches, blind passage and dual observation, for detecting Chlamydia trachomatis in various prevalence populations.

Chlamydia trachomatis diagnosis in our laboratory consisted of dual inoculation of shell vials and detection of inclusions by using fluorescein-conjugated monoclonal antiserum; the second culture vial was conventionally used for blind passage when the first vial was negative. We compared the increase in positivity using blind passage with that of a strategy utilizing observation of two stained monolayers (dual observation) without blind passage, in an effort to reduce the reporting time and labor associated with the conventional approach. A total of 4,329 specimens were obtained from an obstetrics and gynecology (OB-GYN) clinic (2,563 specimens) and the sexually transmitted disease clinic (1,766 specimens). These specimens were used to compare the two strategies. Blind passage of 1,269 initially culture-negative specimens from the OB-GYN clinic resulted in an additional 6 positive chlamydial diagnoses. In comparison, a similar number of specimens (1,294) from the OB-GYN clinic collected subsequently to the first group were tested by dual observation. There were five additional positive findings. A similar evaluation of specimens from the sexually transmitted disease clinic was performed. Blind passage of 313 initially culture-negative specimens yielded 3 additional positive diagnoses, whereas dual observation of 1,435 similar specimens resulted in 9 positive diagnoses. On the basis of analysis of 4,332 specimens, sensitivity of dual observation is comparable to that of blind passage; labor, cost, and reporting time of dual observation are reduced in comparison to those of blind passage.

Evaluation of a visual, rapid, membrane enzyme immunoassay for the detection of herpes simplex virus antigen.

We evaluated a 12-min, direct, monoclonal antibody-based enzyme immunoassay (EIA) (SureCell; Kodak, Rochester, N.Y.) which aids in the detection of herpes simplex virus infection; the assay system is also approved for culture confirmation. The test was evaluated from direct clinical samples and compared with conventional culture methodology by using a single swab. A total of 265 specimens from 180 female cervical-urogenital sites, 62 male urogenital sites, 4 rectal sites, 3 skin sites, 6 oral sites, and 10 colposcopy sites were collected on Dacron or cotton swabs and placed in viral transport medium (VTM). Within 6 h of receipt, 0.2 ml of the vortexed VTM was inoculated into each of two replicate cell cultures. Cell monolayers were observed daily for ten days, and cytopathic effect was confirmed by using an indirect immunoperoxidase reagent. The procedure for the SureCell assay conformed to the manufacturer's recommendations. When conventional culture was compared with EIA results, the overall sensitivity, specificity, positive predictive value, negative predictive value, and agreement were 64.4, 98.9, 96.7, 84.4, and 87.2%, respectively. Variables affecting the EIA sensitivity are the stage of the lesion and conventional culture methodologies. A review of culture results for 32 EIA false-negative tests indicated that 15 were detected after 48 h of incubation. Cytopathic effect observed at 48-, 72-, and 96-h cutoffs altered the sensitivity for the EIA. To ensure detection of SureCell herpes simplex virus-negative specimens, it is recommended that an unused aliquot of VTM be tested in cell culture.

Case report and seeded blood culture study of Brucella bacteremia.

Diagnosis of brucellosis requires prompt detection and identification of the coccobacillus for appropriate patient management, as the organism is associated with a potentially severe outcome. In a recent experience, an 18-year-old migrant farm worker presented at a local hospital with nonspecific symptoms. A significant Brucella titer of 2,560 was followed by the recovery of a gram-negative coccobacillus, subsequently identified as Brucella abortus, from subcultured 5-day-old BACTEC NR730 negative blood cultures. The organism proved to be susceptible to a variety of antimicrobial agents and resistant to nitrofurantoin. The patient was administered antimicrobial therapy for Brucella spp. consisting of tetracycline and streptomycin for 21 days. During the course of therapy the patient experienced defervescence and was discharged with the recommendation for periodic follow-up examinations. Seeded culture studies of this isolate with fresh human blood and target inocula of 5 and 500 CFU/ml indicated that the larger (500-CFU/ml) inoculum produced positive instrument detection within 2 days, whereas the smaller (5-CFU/ml) inoculum required 5.5 to 7.5 days for detection, depending on the medium used. These findings underscore the potential for Brucella bacteremia to escape instrument detection given a low bacterial inoculum.

Complement components and receptors: deficiencies and disease associations.

The complement system, accessory to many immunological functions, consists of a number of interdependent components and receptors. Numerous in vitro approaches have elucidated the biological role of these components and receptors. However, it is the in vivo "natural" experiments that underscore their importance. The phagocytosis and subsequent digestion of pyogenic bacteria is significantly enhanced by the fixation of the third complement component to the bacterial cell wall. Equally important is the intact expression of a receptor (CR3) for the C3b cleavage fragment. Breakdown in this ligand-receptor interaction due to either C3 or CR3 deficiency leads to pyogenic infection. Interestingly, C3-deficient individuals do not demonstrate leukocytic infiltration at the site of infection. Undoubtedly, this is due to the lack of C5 convertase and failure to produce C5a. CR3-deficient individuals, on the other hand, do demonstrate leukocytosis since the third complement component is functional. C3 deficiency is not necessarily a primary lesion and may be secondary to factor I deficiency. In this case, the C3b fragment, along with factor B, acts as a C3 convertase. Inefficient inactivation of C3b, due to factor I deficiency, leads to the uncontrolled consumption of the third component, resulting in C3 deprivation. It appears that phagocytosis by neutrophils and monocytes followed by enzyme-interaction is not sufficient for destruction of the Neisseria organisms. In addition to this leukocyte activity, an intact membrane attack complex, composed of the late complement components C5, 6, 7, 8, and 9, is required for the lysis of these bacteria. This is supported by findings that individuals deficient in late components are highly susceptible to systemic Neisseria infections. Diseases of an autoimmune nature are frequently associated with a deficiency of one of the early complement components C1, C2, or C4 and a deficiency of erythrocytic CR1 receptors as well. This may suggest that proper interaction between a complement fragment of the immune complex with the complement receptor expressed on the erythrocyte is important for proper management and clearance of the complex. Deficiency of the early complement components would prevent the activation of C3 and the fixation of a resulting C3 cleavage product. In this case, erythrocytes would be unable to participate in the transport of the immune complex to the reticuloendothelial system. Instead, tissue deposition of the complex would occur more readily, contributing to the pathologic process. Provided that the early complement cascade were intact, deficiency of erythrocytic CR1 receptors would contribute to the pathologic response for the same reason.(ABSTRACT TRUNCATED AT 400 WORDS)

Erythrocyte sedimentation rate and C-reactive protein compared in the elderly.

The erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) concentrations were studied in 101 elderly individuals (mean age 72 y) to determine their utility as diagnostic aids in subjects with underlying infection/inflammation. Whereas ESR and CRP were both significantly increased in patients with infection or inflammation, or both, analysis of variance indicated that those subjects still alive six months later had significantly lower ESR values. Analysis of sensitivity, specificity, and positive predictive values indicated that neither test satisfactorily discriminated between patients with and those without ongoing active or chronic disease. Receiver-operating characteristic curve analysis confirmed the low true-positive/false-positive ratios of both ESR and CRP. In the elderly, neither CRP nor ESR has distinct advantages over the other, and both tests evidently have limited utility.

Neisseria meningitidis and Moraxella osloensis: dual infection in blood and peritoneal fluid.

The clinical course of a malnourished alcoholic in which Neisseria meningitidis was isolated from the blood and Moraxella osloensis from the peritoneal fluid is described. Following bacteriologic diagnosis, the patient was treated and responded to a course of penicillin therapy. To our knowledge, this represents the first case of peritonitis associated with M. osloensis. Clinical reports of the isolation of this organism are rare; its pathogenicity is not clearly established, and the presence of the organism may often be unrecognized.

Long-term evaluation of the hepatitis B vaccine (Heptavax-B) in hemodialysis patients.

Hemodialysis patients were screened for hepatitis B surface antibody (anti-HBs) prior to immunization at two teaching hospitals. Thirty-one of 111 patients (28%) had baseline sera positive for anti-HBs, while anti-HBs was found in 30 of 420 (7.1%) health care employees (P less than 0.001). A total of 72 hemodialysis patients (mean age, 55.7), received the hepatitis B vaccine (Heptavax-B, Merck Sharp & Dohme, West Point, PA). The responder rates (34 of 72; 47%) and nonresponder (38 of 72; 53%) rates were similar to previous reports. Neither age (P greater than 0.05) nor injection site (P greater than 0.05) appeared to influence results. Nonresponders (16 of 17; 94%) who were given a fourth vaccine dose also failed to mount an antibody response. Of the 34 responders, 18 were followed by serial anti-HBs determinations. Seven transient responders (7 of 18; 39%) were identified, and anti-HBs fell below 10 S/N (sample/control counts per minute) within 12 to 15 months of the first vaccine dose. A fourth dose was administered to this group and it extended the presence of serum anti-HBs (S/N greater than or equal to 10) in four of six patients for another 2, 8, 10, and 15 months, respectively. Antibody persisted but declined over the study period in the remainder of responders followed serially (11 of 18; 61%). When compared with those responders who lost anti-HBs, those with persistent antibody had higher anti-HBs values at 7 (P less than 0.02) and 12 months (P less than 0.005) after the first injection, and were younger (P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Mycobacterium haemophilum infection in a patient with acquired immune deficiency syndrome.

Mycobacterium haemophilum was isolated from wrist and ankle aspirates as the organism responsible for tenosynovitis in a patient with acquired immune deficiency syndrome. Mycobacterium isolates recovered from synovial fluid were identified as hemin requiring by their failure to grow on subculture unless the medium was supplemented with hemin. M. haemophilum is of low virulence and rarely associated with infections in humans. This is the first documented case of M. haemophilum infection in a patient with acquired immune deficiency syndrome.

Pathophysiology of primary meningococcal pericarditis associated with Neisseria meningitidis group C. A case report and review of the literature.

Pericarditis associated with Neisseria meningitidis in the absence of meningitis or meningococcemia is an extremely rare event. We report herein a case of a 59-yr-old woman with primary meningococcal pericarditis caused by Neisseria meningitidis group C. The patient responded to a course of penicillin therapy and recovery was uncomplicated. The pathophysiologic features underlying or contributing to the disease are discussed and the pertinent literature is reviewed.

Application of bioluminescent urine screens in a tertiary care facility.

Two adenosine triphosphate (ATP)-detection systems for quantitating bacteriuria, the LUMAC (noncentrifugation method) and MONOLIGHT (centrifugation method) urine screens, were separately evaluated for their capacity to detect bacteriuria in specimens from patients at a tertiary care teaching hospital. Results of each study were compared with the findings of conventional culture. Indices of test efficacy, sensitivity/predictive value for a negative test, were as follows: at greater than or equal to 10(4) CFU/ml--LUMAC 88%/93% and MONOLIGHT 82%/88%; and at greater than or equal to 10(5) CFU/ml--LUMAC 99%/99% and MONOLIGHT 97%/99%. Both systems were satisfactory urine screens for catheterized and midstream urine specimens when used at the traditional level of significance (greater than or equal to 10(5) CFU/ml). An assessment of the MONOLIGHT noncentrifugation protocol demonstrated efficacy of the system to detect significant bacteriuria at greater than or equal to 10(5) CFU/ml. Decreased numbers of false-positive results compared to the centrifugation method were obtained with this assay. False-positive and false-negative results were attributable to threshold sensitivity of the instruments. The presence of somatic cells and yeasts were associated with false-positive results. False-positive results might stem from the inability of conventional culture to recover selected microorganisms. Time and cost analyses of the LUMAC system indicated that significant savings over conventional methodology were not effected.

Nodular glomerulosclerosis associated with multiple myeloma. Role of light chain isoelectric point.

A 68-year-old female patient with multiple myeloma exhibited advanced nodular glomerulosclerosis. Immunofluorescence of the kidney showed kappa light chain deposition in the mesangium and in glomerular and tubular basement membrane. Isoelectric focusing and immunofixation of urinary proteins revealed an isolated kappa light chain with an unusually high isoelectric point of 8.4. Most light chain proteins have isoelectric points in the 4.6 to 6.7 range. Since loss of fixed negative charges may precede experimental glomerulosclerosis, it is proposed that this cationic circulating kappa chain may have interacted with glomerular polyanion, thereby inducing a nodular sclerotic reaction leading to irreversible renal damage.

Leukocyte esterase-nitrite and bioluminescence assays as urine screens.

The 1-min leukocyte esterase (LE)-nitrite test (Chemstrip 9; Biodynamics, Division of Boehringer Mannheim Biochemicals, Indianapolis, Ind.) and a bioluminescence assay (Monolight centrifugation method; Analytical Luminescence Laboratory, Inc., San Diego, Calif.) were tested for their efficacy as urine screens among 453 patients at a tertiary-care teaching hospital. Both methods had the capacity to exclude significant bacteriuria (greater than or equal to 10(5) CFU/ml) when compared with the results of conventional culture methods, with predictive values of 99 and 93%, respectively, for a negative test. Bioluminescence was the more accurate nonculture method used. Sensitivity and specificity values were 97 and 71%, respectively, for bioluminescence, 82 and 60%, respectively, for LE with nitrite, and 72 and 64%, respectively, for LE without nitrite. At reduced levels of bacteriuria less than 10(5) CFU/ml), the sensitivities of LE-nitrite and bioluminescence were decreased but comparable. The addition of protein and blood test results in the Chemstrip 9, along with LE-nitrite as bacteriuria indicators, were unsatisfactory because of the large numbers of false-positive results attributed to protein and blood determinations. LE activity as detected by the LE test was a poor predictor of significant bacteriuria in both male and female patients. The sensitivity (71%) and specificity (57%) of the LE test in male patients were significantly lower than those previously reported and varied with the patient population studied.

Staphylococcus simulans septicemia in a patient with chronic osteomyelitis and pyarthrosis.

Staphylococcus simulans was identified as the etiological agent of osteomyelitis and septic arthritis in an adult male who had sustained a fracture of the fibula and syndesmosis separation which required the installation of orthopedic hardware. Identifying characteristics and antibiograms for this organism, recovered from blood, wound exudate, and deep tissue samples, were determined. Recent evidence has linked slime production (adherence to smooth surfaces) by coagulase-negative staphylococci to infections by these organisms at sites where foreign bodies had been inserted. Tests for adherence showed this S. simulans strain to be a strong slime producer. This is the first reported case of osteomyelitis and septicemia due to S. simulans.