Rapid detection of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) in wound specimens and blood cultures: multicenter preclinical evaluation of the Cepheid Xpert MRSA/SA skin and soft tissue and blood culture assays.

A multicenter preclinical evaluation was conducted to evaluate the performance of two Cepheid Xpert assays for detection of methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus. Sensitivity was 97.1% and 98.3% for MRSA in wound and blood culture specimens, respectively. Sensitivity was 100% for S. aureus from both specimen types.

Drug-resistant tuberculosis in Castilla-León, Spain, 1996-2000.

SETTING: During 1996-2000, a regional anti-tuberculosis drug resistance survey was conducted in Castilla-León, Spain. OBJECTIVE: To determine the incidence of drug-resistant tuberculosis (TB) in newly treated human immunodeficiency virus (HIV) negative and HIV-positive TB patients. DESIGN: Nine hundred and eighty-five Mycobacterium tuberculosis strains isolated from HIV-negative (926) and HIV-positive (59) patients were studied (one strain per patient). Univariate and multivariate analyses were used to determine the prevalence of drug resistance in high-risk groups. RESULTS: Thirty-eight isolates (3.8%) showed resistance to one of the following drugs: streptomycin (S), isoniazid (H), rifampicin (R) or ethambutol (E). Of these, 36 (3.9%) were from HIV-negative and 2 (3.4%) from HIV-positive patients. The rate of drug resistance among HIV-negative patients was 1.2%, 2.0%, 0.3% and 0.8%, respectively, for S, H, R and E, and for HIV-positive patients it was 3.4%, 0%, 0% and 1.7%. Among the HIV-negative patients, monoresistance was observed in 32 (3.4%) strains and resistance to both H and R (multi-drug resistance) was detected in one. CONCLUSION: The incidence of primary drug resistance in the surveyed area was low and increased resistance was not observed in the HIV-positive group (P = 0.99). Routine surveillance of drug resistance is recommended by the TB control programme in representative patient populations to optimise treatment regimens.

In vitro establishment of AIDS-related lymphoma cell lines: phenotypic characterization, oncogene and tumor suppressor gene lesions, and heterogeneity in Epstein-Barr virus infection.

Lymphoma represents a major source of morbidity and mortality among AIDS patients. AIDS-associated non-Hodgkin lymphomas (AIDS-NHL) are almost invariably B-cell derived, are classified as high or intermediate grade lymphomas, and display three main histologic types: namely, small non-cleaved cell lymphoma (SNCCL), large cell immunoblastic plasmacytoid lymphoma (LC-IBPL), and large cell lymphoma (LCL). Here we report the in vitro establishment of three new AIDS-NHL cell lines (termed HBL-1, HBL-2, and HBL-3) derived from three AIDS-SNCCL patients differing in primary tumor sites and risk factors for HIV infection. The derivation of the cell lines from the original tumor clones was established by immunophenotypic and molecular genetic analysis. These cell lines display clonal immunoglobulin gene rearrangement, express surface immunoglobulin and B-cell restricted markers, and exhibit a phenotype consistent with SNCCL. Monoclonal Epstein-Barr virus infection was found in only one of the cell lines (HBL-1). Cytogenetic analysis demonstrated the presence of a chromosomal translocation involving the c-myc proto-oncogene and an immunoglobulin locus in all three cell lines. The pattern of genetic lesions detected in HBL-1, HBL-2, and HBL-3 reflects that found in primary AIDS-SNCCL and includes activation of the c-myc oncogene as well as inactivation of the p53 tumor suppressor gene. These cell lines should prove useful in studies of the biological, immunological, and viral factors involved in AIDS-associated lymphomagenesis.

An outbreak of Legionella micdadei pneumonia in transplant patients: evaluation, molecular epidemiology, and control.

PURPOSE: To describe a nosocomial outbreak of Legionella micdadei pneumonia in transplant patients and to characterize the source of the outbreak and the control measures utilized. SUBJECTS AND METHODS: We performed retrospective Legionella micdadei serologic testing to enhance case finding in transplant patients with pneumonia that lacked a documented microbial etiology, as well as prospective environmental surveillance of water sites and testing for Legionella in clinical specimens. RESULTS: During a 3-month period, 12 cases of Legionella micdadei pneumonia were identified either by culture or serologic testing among 38 renal and cardiac transplant patients. Legionella micdadei isolates from hot water sources were found by pulsed-field gel electrophoresis to have a DNA banding pattern that was identical to the isolates from the first 3 culture-positive cases and from 2 cases that occurred 16 months later. CONCLUSIONS: Hospitals caring for organ transplant recipients and other immunosuppressed patients must be aware of the possibility of environmental sources of outbreaks of Legionella infection. A first-line screen with the Legionella urine antigen test will identify Legionella pneumophila serogroup 1. However, specific cultures in outbreak situations should be considered to identify other Legionella pneumophila serotypes and the nonpneumophila Legionella species.

Induction of anti-GM1 ganglioside antibodies by Campylobacter jejuni lipopolysaccharides.

A frequent association exists between acute motor neuropathy, antecedent Campylobacter jejuni (CJ) and anti-GM ganglioside antibodies. Despite the chemical and immunological similarity between CJ lipopolysaccharides (LPS) and GM1, the mechanism of induction of anti-GM1 antibodies is still unclear. We used CJ LPS to immunize rats, mice and immunodeficient mice lacking in NK, CD8+ or T-cell populations. None of these animals developed significant anti-GM1 titers. However, rats immunized with keyhole limpet hemocyanin which contains the cross-reactive sugar epitope Gal(beta1-3)GalNAc developed high titers of IgM anti-GM1 antibodies. This occurred only after these rats were given an intraperitoneal injection of CJ LPS. These results suggest that a glycoprotein antigenic stimulus can induce B-cells which are autoreactive to ganglioside but which remain anergic. A second stimulus with a cross-reactive LPS can then overcome the anergy to induce autoantibody production. A similar mechanism may explain the occurrence of GM1 antibodies in patients after CJ enteritis.

Plesiomonas shigelloides sepsis and splenic abscess in an adolescent with sickle-cell disease.

Plesiomonas shigelloides is a rare cause of self-limiting gastroenteritis. We report a case of extraintestinal P. shigelloides infection in an adolescent with sickle-cell disease who presented with bacteremia complicated by a splenic abscess. Despite the high mortality rate reported in extraintestinal P. shigelloides infection, the patient survived after drainage of the abscess and treatment with antibiotics.

Molecular epidemiology of staphylococcal scalded skin syndrome in premature infants.

BACKGROUND: Outbreaks of nosocomial staphylococcal scalded skin syndrome (SSSS) in infants have been well-described associated with the well baby nursery or delivery room. We describe two cases of SSSS in very low birth weight infants in a neonatal intensive care unit (NICU) and the success of infection control strategies used to prevent an outbreak. METHODS: Staphylococcal scalded skin syndrome was diagnosed in two infants in the NICU: Case I (a 47-day-old, formerly 530-g female); and Case II diagnosed 48 h later (a 41-day old, formerly 706-g female). Multiple infection control measures were implemented: (1) isolation and intravenous antibiotic treatment of cases; (2) placement of exposed infants into a cohort; (3) prophylactic mupirocin treatment of the anterior nares of all infants in the NICU and staff colonized with Staphylococcus aureus; and (4) personnel hand washing with hexachlorophene. Detection of exfoliative toxin A and studies to determine the genetic relatedness of S. aureus strains isolated from patients and staff were performed. RESULTS: In addition to the two SSSS cases, S. aureus was isolated from 2 of 12 (17%) exposed asymptomatic infants, 2 of 20 (10%) ancillary staff, 8 of 30 (27%) nurses and 6 of 24 (25%) physicians. Exfoliative toxin A-producing strains were isolated from both cases and one asymptomatic infant. No toxin was expressed by strains isolated from staff. Pulse field gel electrophoresis demonstrated genetically identical strains of S. aureus from the two SSSS cases and the asymptomatic infant, whereas three staff members harbored strains genetically related to the case strain. Unexpectedly two additional unique clusters of genetically related S. aureus strains were identified from the surveillance cultures. CONCLUSIONS: This report documents the rare occurrence of nosocomial SSSS attributed to transmission in the NICU among extremely low birth weight infants. Multiple infection control strategies were effective in limiting the outbreak. Molecular epidemiology investigation supported a unique S. aureus strain responsible for this event and the presence of bidirectional spread between staff and patients of non-toxin-producing strains.

Diagnosis of pediatric tuberculosis in the modern era.

BACKGROUND: Correctly diagnosing tuberculosis (TB) in children is critical to provide appropriate treatment and to detect undiagnosed source cases. However, diagnosing TB in children may be difficult. OBJECTIVE: We sought to determine whether Amplicor, a Food and Drug Administration-approved polymerase chain reaction (PCR) assay used to detect Mycobacterium tuberculosis in sputum and computerized tomography (CT) would facilitate the diagnosis of TB in children. We also examined the applicability of the Centers for Disease Control and Prevention clinical case definition for TB. SETTING: A university-affiliated pediatric hospital in New York City. SUBJECTS: From March, 1995, to November, 1997, 27 children < 15 years of age (mean age, 3.9 years) were evaluated for suspected TB. RESULTS: M. tuberculosis was cultured from 5 of 76 (6.6%) gastric aspirate specimens, and PCR detected M. tuberculosis DNA in 3 (4.1%) of these specimens. There was poor correlation between culture and PCR because 6 specimens were discordant. CT scans were diagnostic of mediastinal or hilar adenopathy in 6 children with equivocal or negative chest radiographs and confirmed adenopathy in 8 others. Six children received alternative diagnoses. CONCLUSIONS: We conclude that the commercially available PCR technology had very limited utility in detecting M. tuberculosis from gastric aspirates, but CT scans were useful in assessing pediatric patients with suspected TB.

Nonsusceptible Streptococcus pneumoniae in children with chronic otitis media with effusion and recurrent otitis media undergoing ventilating tube placement.

CONTEXT: Children with chronic otitis media are at risk for nonsusceptible Streptococcus pneumoniae (NSP) infection. If these children undergo ventilating tube placement, there is an opportunity to culture middle ear fluid and the nasopharynx to determine carriage of NSP. OBJECTIVE: To determine the incidence of NSP carriage, NSP antibiotic susceptibility and risk factors for NSP carriage in children with chronic otitis media undergoing tube placement. DESIGN AND SETTING: Prospective cohort study in an academic medical center with recruitment of patients from an otolaryngology private practice and clinic. PATIENTS: Children < 18 years of age undergoing tube placement for chronic otitis media. INTERVENTIONS: Myringotomy and tube placement, with culture of middle ear fluid and nasopharynx. MAIN OUTCOME MEASURES: The incidence of NSP cultured from the middle ears and nasopharynx of recruited subjects with the use of the minimum inhibitory concentration break points for penicillin susceptibility recommended by the National Committee for Clinical Laboratory Standards. RESULTS: S. pneumoniae was identified in at least 1 site from 23 of 300 study subjects (7.6%); of these 23, 12 case subjects (52.2%) harbored NSP. Of the risk factors assessed by preoperative questionnaire, only younger age was associated with NSP colonization (P < 0.0001). Of the six oral cephalosporins studied, cefpodoxime and cefuroxime showed good in vitro activity against S. pneumoniae isolates with intermediate penicillin resistance. CONCLUSIONS: Children with chronic otitis media undergoing tube placement may carry NSP and provide a means of monitoring the incidence of NSP and antibiotic susceptibilities for children with ear infections in their communities. Younger age is a risk factor for NSP carriage in this population.

Persistence of immunity to varicella-zoster virus after vaccination of healthcare workers.

OBJECTIVE: Varicella-zoster virus (VZV) vaccine is recommended to protect susceptible healthcare workers (HCWs) from serious disease and to prevent nosocomial spread of VZV. We evaluated clinical outcomes and serological responses in HCWs after immunization with live attenuated VZV vaccine. DESIGN: Vaccinees were immunized from 1979 to 1998 during VZV vaccine trials, as well as after licensure, and followed prospectively for 1 month to 20.6 (mean 4.6) years after vaccination. Sera were tested by fluorescent antibody to membrane antigen (FAMA), latex agglutination (LA), and enzyme-linked immunoassay (EIA) to detect VZV-specific antibodies. STUDY PARTICIPANTS: The median age of the 120 HCWs was 26 years; 51 (42%) were males. INTERVENTIONS: Ninety eight (82%) of these study subjects received vaccine prepared by Merck and 22 (18%) by SmithKline Beecham; 25, 81, and 14 vaccinees received one dose, two doses, and three doses, respectively. RESULTS: The crude attack rate was 10%; 12 of 120 HCWs developed chickenpox 6 months to 8.4 years after vaccination. The attack rates following household and hospital exposures were 18% (4/22) and 8% (6/72), respectively. All resulting illness was mild to moderate (mean 40 vesicles). Seroconversion after vaccination was documented by FAMA in 96% of HCWs, although 31% lost detectable antibodies. Compared with FAMA, LA and EIA were 82% and 74% sensitive and 94% and 89% specific, respectively. CONCLUSIONS: The VZV vaccine effectively protected HCWs from varicella, particularly from serious disease. Currently available serological tests are not optimal, and improved assays are needed.

Nosocomial rotavirus in a pediatric hospital.

We describe a nosocomial rotavirus outbreak among pediatric cardiology patients and the impact of a prospective, laboratory-based surveillance program for rotavirus in our university-affiliated, quartenary-care pediatric hospital in New York City. Improved compliance with infection control and case-finding among patients and healthcare workers halted the outbreak.

Comprehensive evaluation of performance, laboratory application, and clinical usefulness of two direct amplification technologies for the detection of Mycobacterium tuberculosis complex.

The rapid detection of Mycobacterium tuberculosis from respiratory specimens is critical for optimal treatment of patients. Several nucleic acid amplification-based systems designed to detect Mycobacterium tuberculosis complex directly from specimens have been developed, and 2 are commercially available. We studied the performance characteristics of these 2 systems (Gen-Probe Amplified Mycobacterium Tuberculosis Direct (MTD), Gen-Probe, San Diego, Calif; AMPLICOR, Roche Molecular Systems, Branchburg, NJ). Each uses a different amplification strategy, detection modality, and approach to inhibition of amplicon contamination. When compared with culture, the respective sensitivities and specificities were as follows: 92.2% and 98. 7% (study 1) and 88. 7% and 95.3% (study 2); AMPLICOR, 87.5% and 99.7%. Resolution of discordant results was accomplished by incorporating clinical data and multiple specimen analysis. An increased rate of false-positive results was encountered during 1 phase of the study. The conditions under which the test was performed were modified and the "contamination" issue was resolved. This report discusses the benefits and limitations of each assay, proposes cost-effective algorithms for their incorporation into routine laboratory work flow, and discusses the clinical usefulness of these molecular technologies.

In vitro activity of recombinant lysostaphin-antibiotic combinations toward methicillin-resistant Staphylococcus aureus.

In vitro bactericidal experiments were performed with combinations of recombinant lysostaphin and various antibiotics against five strains of Staphylococcus aureus, including clinical isolates of methicillin-resistant S. aureus (MRSA). Sublethal concentrations of lysostaphin combined with beta-lactam antibiotics (including benzylpenicillin, methicillin, and cephalosporin B), bacitracin, or polymyxin B, exhibited rapid synergistic bactericidal activity against all S. aureus strains tested. Lysostaphin activity was additive when combined with either vancomycin, gentamicin, tetracycline, or erythromycin. Synergistic combinations of recombinant lysostaphin and antibiotics might be applicable to controlling cutaneous staphylococcal infections and MRSA carriage.

Differentiation of Nocardia from rapidly growing Mycobacterium species by PCR-RFLP analysis.

The nucleotide sequences from a region of the groEL gene from one Nocardia asteroides and from several species of Mycobacterium were determined and found to be highly homologous. Based on these homologies, we developed a rapid method capable of differentiating between these two genera. The method is based on restriction fragment-length polymorphism (RFLP) analysis of DNA amplified from the groEL gene that is highly conserved between mycobacteria and nocardiae. When the groEL gene from species of these genera is enzymatically amplified by the polymerase chain reaction (PCR), a 422-bp fragment is generated. Correlation of the restriction endonuclease digestion patterns of the amplification products with reference and/or biochemically characterized clinical samples enabled us to establish RFLP profiles for ten species of Mycobacterium and five species of Nocardia. When a portion of the groEL gene from each of these organisms is digested with the restriction endonuclease Hae III, that organism is readily assigned to one of these two genera on the basis of the derived RFLP patterns. The utility of this approach was examined by testing 105 pure cultures from samples previously identified by routine culture techniques for the presence of groEL DNA sequences of mycobacterial or nocardial origin. This analysis correctly identified the organism in all samples tested. In summary, PCR-RFLP analysis provides a rapid and sensitive method for the differentiation of Nocardia species from rapidly growing Mycobacterium species.

A comparison of the bacteria found on the hands of 'homemakers' and neonatal intensive care unit nurses.

This prevalence study was conducted to compare the counts, types and antimicrobial resistance profiles of bacterial flora on the hands of individuals in the community to that of nurses at a nearby university teaching hospital, with an intense hand hygiene regimen. Hand cultures were obtained from 204 individuals during a home visit and 119 nurses in two neonatal intensive care units (NICUs). The mean total log counts of organisms were 5.73 and 5.24 for the homemakers [defined as the person (usually the mother) who is the primary person responsible for arranging childcare, cooking, cleaning etc] and nurse hands, respectively (P<0.0001). Significantly more homemakers had Acinetobacter lwoffii, Enterobacter cloacae, Klebsiella pneumoniae, Pseudomonas aeruginosa, P. fluorescens/putida, and Staphylococcus aureus on their hands compared with the nurses (all P<0.05). However, significantly more nurses had Enterococcus faecalis, S. epidermidis, and S. warneri on their hands (P<0.05). Of note, the hands of nurses harboured significantly more S. epidermidis strains resistant to amoxicillin/clavulanate, cefazolin, clindamycin, erythromycin, and oxacillin and S. warneri resistant to amoxicillin/clavulanate, cefazolin, clindamycin, and oxacillin (P<0.05). Surprisingly, significantly more trimethoprim/sulfamethoxazole-resistant S. epidermidis and ciprofloxacin-resistant S. warneri was recovered from the hands of homemakers (P<0.05). This study demonstrates differences in prevalence, bacterial composition and antimicrobial resistance of hand flora of hospital personnel compared with homemakers. Moreover, the hands of homemakers may serve as community reservoirs for antimicrobial resistant strains of clinical importance.

Infections in parenteral drug abusers. Further immunologic studies.

In our previous study of soft tissue infections in parenteral drug abusers, two thirds of the infections were polymicrobial. Oral and enteric organisms were frequently recovered. These patients and a group of uninfected addicts showed frequent cutaneous anergy, lymphopenia, and hypergammaglobulinemia. An additional group of uninfected addicts was studied. The mean levels of IgA, IgG, and IgM were higher in the uninfected addicts. In the addict and control groups, elevations in IgA (17 percent of total), IgG (65 percent), and IgM (19 percent) levels were found. Zinc levels were within normal limits. T-cell populations below 70 percent were seen in five of the seven addicts and two of the four control subjects. Reversed helper to suppressor cell ratios were found in three of the seven addicts and control subjects. No consistent pattern of immunologic abnormalities emerged. The interrelationship of the abnormalities in the addict and their relationship to AIDS is unclear.

Clinical mycobacteriology. Work flow and optional protocols for laboratories in industrialized countries.

The algorithm of the mycobacteriology laboratory of the 1990s relies heavily on growth-dependent procedures that are slow and labor intensive. The introduction of nucleic acid-based direct amplification tests portends future laboratories where molecular diagnostic assays will prevail. Financial constraints mandate an interdisciplinary approach to establish clinical pathways and rational tailoring of current protocols to curtail expensive overuse of dwindling resources.

Penicillin-nonsusceptible pneumococcus in acute otitis media in New York City.

OBJECTIVE: To determine the proportion of children with acute otitis media (AOM) presenting in our catchment area in New York City who were infected with nonsusceptible Streptococcus pneumoniae and to determine the susceptibility of these organisms to penicillins and other antibiotics commonly used to treat AOM. SETTING: Ambulatory clinics and the emergency department of a tertiary care, inner-city medical center. PATIENTS: During a 2-year period from 1993 to 1995, 115 children (aged 6 months to 12 years) with AOM underwent tympanocentesis. Patients did not receive antibiotics for at least 1 week before tympanocentesis. RESULTS: Thirty-one children were infected with S pneumoniae, and 83.9% of isolates were susceptible to penicillin. Of the 16.1% strains that were nonsusceptible, most (4 of 5 strains) were intermediately resistant, and only 1 exhibited high-level resistance to penicillin. Of all the cephalosporins tested, only cefotaxime had consistent activity against the intermediately resistant strains. Notably, all nonsusceptible pneumococci were inhibited by macrolides. CONCLUSIONS: This study provides unique reference data for nonsusceptible Streptococcus pneumoniae in children with AOM and documents that newer cephalosporin agents are not active against all of these strains.

Assessment of alternative hand hygiene regimens to improve skin health among neonatal intensive care unit nurses.

OBJECTIVE: The frequent handwashing and gloving required in high-risk, high-volume patient care areas such as critical care units damages skin of the hands. The purpose of this exploratory study was to compare 2 hand care regimens (traditional antiseptic wash with chlorhexidine-containing detergent versus mild soap wash with subsequent alcohol-based rinse for degerming as necessary) in a neonatal intensive care unit (NICU). DESIGN: Prospective, quasi-experimental, random assignment. SETTING: One NICU (47 beds) in a New York City children's hospital. SUBJECTS: Sixteen full-time NICU nurses. OUTCOME MEASURES: Microbial flora and skin condition of hands. INTERVENTION: Nurses were randomly assigned to one of the 2 hand care regimens. RESULTS: No significant differences in microbial counts or types of organisms from hands of staff were found, but after 2 weeks nurses in the mild soap and alcohol group had significant improvements in their skin condition (P =.005). CONCLUSIONS: Use of a mild soap for cleaning and an alcohol-based product for degerming may offer an acceptable alternative to the traditional antiseptic handwash and may reduce skin damage to health care professionals' hands.

Differences in skin flora between inpatients and chronically ill outpatients.

BACKGROUND: Changes in skin flora have been reported among hospitalized and critically ill patients, but little is known about whether these changes are associated with hospitalization or with chronic, serious illness. The purpose of this survey was to compare skin flora of chronically ill outpatients and inpatients. METHODS: Aerobic skin flora of forearm and midsternum of 250 patients in an intensive care unit and 251 outpatients was sampled by contact plates. RESULTS: Mean colony-forming units were 160.6, forearm; 229. 4, sternum (P <.000). In logistic regression analysis, patients in the medical intensive care unit were significantly more likely to have high counts on the arm (odds ratio, 2.48; 95% confidence interval: 1.34-4.43; P =.004), and blacks were significantly more likely to have higher counts on the sternum when compared with other ethnic groups (odds ratio, 1.92; confidence interval: 1.18-3.11; P =. 009). No differences were noted between inpatients or outpatients in prevalence of methicillin-sensitive Staphylococcus aureus, but inpatients were more likely to carry methicillin-resistant Staphylococcus aureus (arm, P =.007; sternum, P =.02). Outpatients had a higher prevalence of micrococci and gram-negative bacteria at both skin sites (all P <.01) and yeast at the sternal site (P =.007). CONCLUSIONS: This comparison provides data to differentiate between effects of hospitalization and effects of chronic illness on skin flora.