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Background A psychosocial evaluation is an important component of the preoperative assessment process for people seeking metabolic and bariatric surgery (MBS), and is required for accreditation of MBS programs. Recently, independent companies without affiliations with MBS programs have been marketing remotely administered, unaffiliated psychosocial evaluations for MBS (RUS), and American Society for Metabolic and Bariatric Surgery (ASMBS) members have raised concerns about these evaluations. Objectives To explore ASMBS members’ beliefs about RUS. Setting Online survey. Methods We developed a survey to evaluate ASMBS members’ opinions, experiences, and/or concerns about in-person and RUS psychosocial evaluations for MBS. Results In total, 635 ASMBS members responded to the online survey and 156 responded to an open-ended question on RUS. Responses were coded based on a manual developed for this study, yielding themes of concerns about the quality of RUS, lack of ongoing relationships in RUS, and conditions under which/reasons why RUS evaluations could be acceptable. Conclusion Respondents expressed both interest in and concerns about RUS in pre-MBS psychosocial evaluations. Use of RUS has the potential to improve access to MBS by providing a convenient and efficient means of completing the psychosocial evaluation. Conversely, respondents expressed concerns about the background and training of RUS providers, the quality of the reports, and the limited relationships between the RUS provider and both the MBS patient and the MBS team. We discuss the clinical and research implications of response themes, particularly for patients in rural areas or those who have other barriers to care.

Summary Background Antiretroviral therapy (ART) management is challenging for individuals in resource-limited settings presenting for third-line treatment because of complex resistance patterns, partly due to reduced access to viral load monitoring. We aimed to evaluate use of newer antiretroviral drugs and contemporary management approaches, including population-based sequencing, to select appropriate antiretrovirals, plasma viral load monitoring, and interventions to improve adherence in individuals presenting with second-line viral failure. Methods A5288 was a phase 4, third-line ART strategy study done at 19 urban sites in ten countries that enrolled adult participants with confirmed plasma HIV-1 RNA (viral load) of 1000 copies per mL or more after more than 24 weeks of protease inhibitor-based second-line ART. The primary objective was to use antiretrovirals (raltegravir, etravirine, and ritonavir-boosted darunavir) and diagnostic monitoring technologies, including viral load, genotyping, and adherence support to achieve viral load suppression (defined as ≤200 copies per mL) in 65% or more of participants. ART history and real-time drug resistance genotypes were used to assign participants to one of four cohorts: cohort A (no lopinavir resistance) stayed on second-line ART and cohorts B (B1, best available nucleoside reverse transcriptase inhibitors [NRTIs] plus ritonavir-boosted darunavir plus raltegravir; B2, ritonavir-boosted darunavir plus raltegravir plus etravirine; B3, ritonavir-boosted darunavir, raltegravir, and either tenofovir plus emtricitabine or tenofovir plus lamivudine), C (ritonavir-boosted darunavir plus raltegravir plus tenofovir-emtricitabine or tenofovir plus lamivudine), and D (best available NRTIs plus ritonavir-boosted darunavir plus raltegravir) were defined by increasing levels of resistance and received appropriate regimens, including new antiretrovirals. Participants in Cohort B without detectable hepatitis B surface antigen were assigned by blocked randomisation to cohorts B1 and B2, and those with detectable hepatitis B surface antigen were assigned to cohort B3. The trial is registered with ClinicalTrials.gov , number NCT01641367 . Findings From Jan 10, 2013, to Sept 10, 2015, 545 participants were enrolled. 287 (53%) were assigned to cohort A, 74 (14%) to B1, 72 (13%) to B2, eight (1%) to B3, 70 (13%) to C, and 34 (6%) to D. Overall, 349 (64%, 95% CI 60–68) participants achieved viral suppression at week 48, with proportions varying from 125 (44%) of 287 in cohort A to 65 (88%) of 74 in cohort B1, 63 (88%) of 72 in B2, eight (100%) of eight in B3, 63 (90%) of 70 in C, and 25 (74%) of 34 in D. Participants in cohort A remained on their second-line protease inhibitor, and had the most participants with grade 3 or higher adverse events (147 [51%]). Interpretation Targeted real-time genotyping to select third-line ART can appropriately allocate more costly antiretrovirals to those with greater levels of HIV drug resistance. Funding National Institutes of Health.

Serious Candida glabrata infections, which can be difficult to treat, are often treated with echinocandins. We compared in vitro the effects of high and low concentrations of 3 echinocandins (micafungin [MCF], caspofungin [CAS], and anidulafungin [ANF]), voriconazole (VRC), and amphotericin B (AmB), singly and VRC in combination with MCF, CAS, and ANF, on the production of cytokines/chemokines by human monocyte-derived macrophages (MDM). MDM were activated by infection with C. glabrata or lipopolysaccharide (LPS). Luminex multi-analyte microsphere technology was used for cytokine/chemokine analysis. Concentrations of cytokines/chemokines were significantly elevated following activation by infection or LPS. Treatment with high concentrations of echinocandins, singly or in combination with VRC, was most effective in lowering the elevated cytokine/chemokine levels. This effect occurred only with MDM activated by infection with C. glabrata and not with LPS. Treatment with VRC or AmB alone had little or no effect on cytokine/chemokine levels. In severe C. glabrata infection associated with very high concentrations of dysregulated cytokines/chemokines, echinocandins, singly or in combination with VRC, may decrease cytokine/chemokine concentrations and thus may improve host survival.

Background:Candida glabrata causes infections associated with severe sepsis, production of high concentrations of cytokines/chemokines, and high mortality. This study describes the effects of anidulafungin (ANF) and voriconazole (VRC), singly and in combination, on the production of eight cytokines/chemokines by human monocyte-derived macrophages (MDM) infected with C. glabrata or activated by lipopolysaccharide (LPS). Methods: MDM monolayers were established, infected with C. glabrata or activated with LPS, and then treated with high or low concentrations of ANF, VRC, or both. Cytokine/chemokine levels in MDM supernatants were determined. Results: Levels of cytokines/chemokines were significantly elevated in supernatants of infected or LPS-activated MDM. Except for interleukin-10, all significant decreases in cytokine/chemokine concentrations (p < 0.01) occurred in supernatants of infected MDM treated with high concentrations of ANF or ANF + VRC. Conclusions: Decreases in cytokine/chemokine levels in supernatants of infected MDM treated with high concentrations of ANF or ANF + VRC suggest that similar treatment could improve survival in patients with severe, invasive C. glabrata infections and markedly elevated levels of serum cytokines/chemokines.

The purpose of this study was to determine by time-kill methodology the anticandidal effectiveness and durability of micafungin (MCF) and voriconazole (VRC), singly and in combination, against Candida glabrata (Cgl), intracellularly in human monocyte-derived macrophages and extracellularly in RPMI-MOPS broth with and without fetal calf serum (FCS) or pooled human serum (PHS). The anticandidal activity of MCF was concentration-dependent and durable. Combinations of MCF + VRC both intra- and extracellularly were more effective than single drugs. However, in extracellular experiments, FCS, and even more PHS, significantly decreased the anticandidal activity of MCF and VRC. Our in vitro studies suggest that MCF alone may be adequate where protein concentration is low (intracellular or extravascular sites), while MCF + VRC combinations may be preferred where protein concentration is high such as in the intravascular space.

The activity of tigecycline against Legionellae, which are intracellular pathogens, was evaluated intracellularly in human phagocytes and extracellularly, and compared to the activities of erythromycin and levofloxacin. Clinical isolates of L. pneumophila serogroups 1, 5, and 6 and L. micdadei were tested in time-kill experiments. Extracellular experiments were done using buffered yeast extract broth. For intracellular assays, monolayers of human monocyte-derived macrophages (MDM) were infected with L. pneumophila or L. micdadei. Antibiotics (0.05-2.5 × MIC) were then added. MDM were lysed at 0, 24, 48, and 72 h and viable bacteria in the lysates were enumerated. Based on multiples of the MICs, tigecycline was less active extracellularly than levofloxacin or erythromycin. However, intracellular killing of both L. pneumophila and L. micdadei by tigecycline at 72 h was greater than for erythromycin or levofloxacin. Currently, evidence does not support the use of tigecycline as a first-line drug for treatment of Legionella infections. However, since Legionellae are intracellular pathogens, these results suggest that tigecycline should be effective for treatment of infections caused by these bacteria.

We investigated the antistaphylococcal activity of daptomycin, vancomycin, oxacillin, gentamicin, and rifampin in human monocyte-derived macrophages. Compared with vancomycin and oxacillin, daptomycin had the most rapid and greatest antibacterial activity, but that of oxacillin was most sustained. The combination of daptomycin, gentamicin, and rifampin was most effective intracellularly, while daptomycin plus gentamicin and the three-drug combination were most effective extracellularly, completely eliminating viable Staphylococcus aureus .

Infections caused byCandidaspecies other thanCandida albicansare increasingly common, and decreased susceptibility to azoles has made them more difficult to treat. Since phagocytic killing is important in elimination ofCandidainfections, intracellular killing of fluconazole-resistantCandida glabrata,Candida kruseiandCandida parapsilosis(four strains each) by voriconazole was investigated in human monocyte-derived macrophages (MDMs). MDMs were infected withCandida, and voriconazole was then added. MDMs were lysed at 0, 24 or 48 h after infection, and viableCandidain the lysates enumerated. Compared to the starting inoculum, the number of viable intracellularC. parapsilosisandC. glabratain untreated MDMs increased to 28 121 and 351 %, respectively, in 48 h. In contrast, the number ofC. kruseidecreased to 42 %. In MDMs treated with voriconazole, the decrease in viable count was dependent upon drug concentration. At 48 h,C. glabratawas killed only at 5× MIC (PC. kruseiwas killed at all voriconazole concentrations, whileC. parapsilosiswas inhibited at 0.5 and 1× MIC and killed at ⩾2.5× MIC (PCandidaspecies in the absence or presence of voriconazole vary markedly. The activity of voriconazole depends on the concentration of the drug and the time of exposure. For the 12Candidastrains studied, regression curves show that the maximum intracellular anticandidal activity of voriconazole was reached at 3.5–5× MIC.

Nonsurgical weight management strategies have been largely unsuccessful in treating and reducing the prevalence of morbid obesity. Bariatric surgery has developed into a viable and successful long-term option for the morbidly obese patient. Unfortunately, bariatric surgery does not address the psychosocial, emotional, behavioral, and lifestyle challenges that often await many patients as they move through their weight loss. Patients may expect that their weight loss journey will be hurdle-free. The prebariatric surgery psychological evaluation can help identify the risk factors that might become barriers to optimal weight loss postsurgery. Components of the assessment may include developmental history, degree of social integration, willingness to access meaningful support, past and present psychiatric history, the presence of unusual life stressors, coping skills, and psychological resources. Other variables to be assessed include the presence of significant psychological eating, the onset of obesity, mai...

Infections caused by fluconazole-resistant Candida glabrata and Candida krusei are increasingly common causes of morbidity and mortality. We investigated the intracellular killing of fluconazole-resistant C. glabrata and C. krusei by cytokine-activated human monocyte-derived macrophages (MDM) in the presence and absence of voriconazole. For C. glabrata , MDM were activated with either granulocyte-macrophage colony-stimulating factor (GM-CSF) or interferon γ (IFN-γ) before infection, after infection, or both before and after infection, whereas for C. krusei MDM were activated with cytokines both before and after infection. Activated MDM were infected, treated with voriconazole, and then lysed, and viable yeast in the lysates enumerated at 0, 24, or 48 h after infection. In the presence of voriconazole (2.5 × MIC), the best activity against C. glabrata occurred when MDM were activated with GM-CSF for 24 h before infection as well as after infection or when they were activated for 24 h before infection alone. A lesser effect was observed when MDM were activated for at least 1 h before infection or when they were treated with cytokines only after infection. IFN-γ activation had a significant but lesser effect than GM-CSF. Activity against C. krusei in the presence of voriconazole was greatest when MDM were activated with IFN-γ rather than GM-CSF. Our results suggest that cytokines increase the intracellular anticandidal effect of voriconazole and may be useful as therapeutic adjuvants to voriconazole for treatment of infections caused by fluconazole-resistant C. glabrata and C. krusei .

Although antibiotics are known to affect the intracellular growth of Chlamydia pneumoniae in acute infections, their efficacy in therapy for chronic infections, including atherosclerosis, remains debatable. Human monocyte-derived macrophages (MDM) obtained from monocytes of healthy donors were infected with C. pneumoniae AR-39 and treated with levofloxacin (8 microg/mL) immediately after infection (0 hours) or 24 hours after infection. Levofloxacin treatment at 24 hours, but not at 0 hours, resulted in a significant decrease in the number of C. pneumoniae inclusions within the MDM (p < 0.05). Also decreased were concentrations of proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-6, and IL-8 in the extracellular medium (p < 0.01). Viable counts in titrations remained similar to those in untreated controls. In summary, levofloxacin administered to MDM at serum-attainable levels 24 hours after C. pneumoniae infection significantly decreased inclusion counts and proinflammatory cytokine production, but did not eliminate the C. pneumoniae infection.

Background:Chlamydia pneumoniae is known to cause acute respiratory infection and more recently it has been studied as a pathogen causing inflammatory changes in chronic diseases such as atherosclerosis. This study addresses the antichlamydial effect of levofloxacin and its role in modulation of a proinflammatory cytokine IL-6 production by uninfected and infected HEp-2 cells. Methods: HEp-2 cell monolayers were infected with previously prepared and frozen aliquots of C. pneumoniae [1 × 103 inclusion-forming units (IFU)/ml] by centrifugation for 30 min and incubation at 37°C for 1 h. Infected monolayers were treated with levofloxacin (3 or 8 µg/ml) immediately after infection (0 h) or 24 h after infection. Monolayers were examined daily for 96 h after infection by counting inclusions with fluorescently labeled antichlamydial monoclonal antibody. Aliquots of disrupted monolayers were titrated to determine the numbers of viable C. pneumoniae IFU/ml. IL-6 concentrations in cell supernatants were determined by ELISA assays. Results: Infected HEp-2 cells produced IL–6. Noninfected HEp-2 cells demonstrated modulation of IL-6 production by levofloxacin. No viable C. pneumoniae were detected in infected HEp-2 cells when the monolayer was treated with levofloxacin immediately after infection (0 h). In contrast, when cells were treated 24 h after infection, a gradual decline in the number of viable C. pneumoniae occurred; by 96 h into the assay ≧98% of C. pneumoniae were killed. IL-6 concentrations were similar in the supernatants of levofloxacin-treated and nontreated HEp-2 cells. Conclusions: (1) Levofloxacin is effective in eliminating C. pneumoniae from infected HEp-2 cells; (2) although levofloxacin modulates the production of IL-6 in untreated HEp-2 cells, no evidence for such modulation was observed in HEp-2 cells infected with C. pneumoniae. (3) Presence of viable C. pneumoniae may not be necessary for IL-6 production by infected and treated HEp-2 cells.

Altersschatzungen mussen immer haufiger an Individuen durchgefuhrt werden, die in ihrer biologischen und soziokulturellen Herkunft nicht der Population entsprechen, an dem das Referenzmaterial erhoben wurde. Bei der biochemischen Altersschatzung aufgrund des Razemisierungsgrades von Asparaginsaure in Dentin waren genetische bzw. soziokulturell bedingte Einflusse als Fehlerquellen theoretisch denkbar, wenn dadurch Variationen der Proteinkomposition des Dentins bedingt wurden. Zur Klarung der Frage, ob solche Einflusse tatsachlich zu berucksichtigen sind, wurden Zahne von turkischen und deutschen Individuen mit bekanntem Alter im unmittelbaren Vergleich untersucht. Die Beziehung zwischen dem Razemisierungsgrad von Asparaginsaure in Dentin und dem Dentinalter war in den untersuchten Kollektiven gleich. Auch unter Berucksichtigung der einschlagigen Literatur ergibt sich derzeit kein Anhaltspunkt dafur, dass genetische bzw. soziokulturelle Einflusse auf die Dentinproteinkomposition bei der biochemischen Altersschatzung berucksichtigt werden mussen.

This study evaluates the effects of cytokines, used singly and in combination, on the microbicidal activity of human monocyte-derived macrophages (MDM) against intracellular Candida albicans in the presence and absence of fluconazole. In the absence of fluconazole, the addition of tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), gamma interferon (IFN-γ), or IL-4 had no effect on the growth of C. albicans . In contrast, the addition of granulocyte-macrophage colony-stimulating factor (GM-CSF) resulted in decreased growth ( P < 0.05), while the addition of IL-10 resulted in increased growth ( P < 0.01). In the presence of fluconazole, only the addition of IFN-γ resulted in an increase in the growth of C. albicans . In the presence or absence of fluconazole, all cytokine combinations except IFN-γ plus GM-CSF caused significant decreases in growth ( P < 0.01). IL-10 and IL-4 did not influence the activity of TNF-α or IL-1β. In the absence or presence of C. albicans the addition of fluconazole, all of the cytokines studied, and combinations of fluconazole and selected cytokines caused increases in nitric oxide (NO) production ( P < 0.01). Similar observations were made for superoxide (O 2 − ) only in the presence of C. albicans . The greatest concentrations of NO and O 2 − were produced when C. albicans alone was present in the assays. Our results demonstrate that in the presence of low concentrations of fluconazole (0.1 times the MIC), selected cytokines and their combinations significantly increase the microbicidal activity of MDM against intracellular C. albicans .

BACKGROUND Because breast-conserving surgery (BCS), mastectomy alone, and mastectomy with reconstruction are equally effective for the treatment of early stage breast carcinoma, women's choice among them often focuses on quality-of-life (QOL) issues. Information regarding QOL after these surgical treatments could help women with this decision. METHODS Participants in this prospective study were women, age 30–85 years, with newly diagnosed breast carcinoma who underwent BCS (n = 103), mastectomy alone (n = 55), or mastectomy with reconstruction (n = 40). Quality of life was assessed after diagnosis (baseline) and at 1, 3, 6, 12, 18, and 24 months after baseline by using the Mischel Uncertainty in Illness Scale, Profile of Mood States, and Functional Assessment of Cancer Therapy for Breast Cancer. RESULTS In multivariate regression analyses controlling for the QOL score obtained at baseline, age, and type of nonsurgical treatment, women who underwent mastectomy with reconstruction had greater mood disturbance (P = 0.002) and poorer well-being (P = 0.002) after baseline than women who had mastectomy alone; these differences remained 18 months after surgery. Although similar analyses also showed that women who underwent BCS had more mood disturbance than women who had mastectomy alone, this difference was significant only at 12 months after baseline. The BCS and mastectomy-only group did not differ significantly regarding well-being. CONCLUSIONS Aspects of QOL other than body image are not better in women who undergo BCS or mastectomy with reconstruction than in women who have mastectomy alone. In fact, mastectomy with reconstruction is associated with greater mood disturbance and poorer well-being. Cancer 2001;91:1238–46. © 2001 American Cancer Society.

Background: Vancomycin resistance among enterococci is an emerging nosocomial problem. Consequently, it is important to understand the distri-bution of vancomycin-resistant enterococci (VRE) within and between hospitals to implement appropriate infection control measures. Methods: In this study, 116 VRE isolates obtained from patients in 6 New York State hospitals were analyzed by antibiotic susceptibility testing, pulsed-field gel electrophoresis (PFGE) fingerprinting, plasmid profile analysis, vanA and vanB polymerase chain reaction, and DNA:DNA hybridization with vanA and vanB probes. Results: PFGE and plasmid typing generally agreed, but plasmid profiles were more variable. These analyses revealed that genetic heterogeneity among isolates from within each of the 6 hospitals varied considerably. Among 23 Enterococcus faecium isolates from one hospital, there were only 3 PFGE types, and 20 isolates had the same type. However, in another hospital, each isolate was genetically distinct. Closely related strains were not found in separate hospitals. VRE strains with vanA genes and strains with vanB genes were found in 3 hospitals. Both plasmid and chromosomal carriage of these genes was detected. Conclusions: PFGE typing showed that nosocomial VRE transmission had occurred in some hospitals. However, there was no evidence for it in others. Neither was there evidence for intrahospital transmission or for emergence of an endemic strain. These observations demonstrate that it is important to evaluate genetic heterogeneity among VRE before implementation of infection control measures. PFGE is the method of choice for epidemiologic typing, but polymerase chain reaction, plasmid, and hybridization studies can provide important information concerning the presence and potential for transfer of vancomycin resistance genes. (AJIC Am J Infect Control 1999;27:411-7)

One hundred ninety-five individual vancomycin-resistant Enterococcus faecium (VRE) isolates from five upstate New York hospitals were studied for antimicrobial susceptibilities to LY333328, quinupristin-dalfopristin, teicoplanin, ampicillin, and gentamicin. LY333328 was the most active antibiotic against VRE. The effect of media and methods on the antibacterial activity of LY333328, its synergy with ampicillin, and the postantibiotic effects (PAE) of LY333328 and ampicillin were evaluated. In microdilution tests, the MIC of LY333328 at which 90% of the isolates were inhibited (MIC 90 ) was 2 μg/ml in Mueller-Hinton II (MH II) broth and 1 μg/ml in brain heart infusion (BHI) broth. In contrast, on MH II agar the MIC 90 was 4 μg/ml and on BHI agar it was >16 μg/ml. Bactericidal activity was observed for most strains at concentrations from 8 to ≥133 times the MIC of the tube macrodilution in MH II broth. A bactericidal effect of LY333328 plus ampicillin was demonstrated in time-kill studies, but there was great strain-to-strain variability. By the MH II agar dilution method, bacteristatic synergy (defined as a fractional inhibitory concentration of

Study objective: To identify the oncology patient population presenting to the emergency department and examine the causes and clinical management of oncology symptoms in the ED. Design: Retrospective review of 5,640 adult patients, with the following variables identified for oncology patients presenting to the ED: age, sex, cancer type, cancer stage, previous cancer treatment, previous hospitalization, presenting symptoms, treatment provided in the ED, admission and discharge data, and day, time, and length of ED visit. Setting: Community teaching hospital with annual ED census of 31,000. Participants: All adult oncology patients who presented to the ED during the study period. We identified these patients by cross-referencing ED logs and tumor registry records. Results: Cancer history was identified for 284 of the 5,640 adult ED admissions (5%). Forty-three percent (n=122) of the 284 patients with cancer history had an oncology-related ED visit. The most common symptoms of these patients were gastrointestinal (48%), pain (40%), neurologic (38%), cardiac (25%), and pulmonary (23%). Ten percent of patients with oncology-related ED visits died during the admission, and 48% died within 1 year of the ED visit. Conclusion: Oncology patients present to the ED with symptoms of undiagnosed malignancy, complications of cancer treatment, and acute disease-related symptoms. Knowledge of an individual's cancer history and ability to recognize oncologic symptoms are important to the management of oncology patients. [Swenson KK, Rose MA, Ritz L, Murray CL, Adlis SA: Recognition and evaluation of oncology-related symptoms in the emergency department. Ann Emerg Med July 1995;26:12-17.]

In systemic candidiasis, endogenously produced tumor necrosis factor (TNF)-alpha prolongs survival of the infected host. To determine whether endogenously produced TNF-alpha has a beneficial effect beyond that provided by antifungal therapy, survival was assessed in infected mice that received fluconazole or amphotericin B alone and in combination with anti-TNF-alpha antibody. Neutralization of serum TNF-alpha did not affect survival in fluconazole recipients; however, for amphotericin B recipients, it significantly shortened mean survival. For both fluconazole and amphotericin B recipients, colony counts in organs were significantly higher in animals that also received anti-TNF-alpha antibody. Administration of anti-TNF-alpha antibody with amphotericin B or fluconazole did not affect the morphology of fungi or the inflammatory response in kidneys. This study suggests that exogenous TNF-alpha and drugs that increase the endogenous production of TNF-alpha by the host may be useful adjuncts to fluconazole and amphotericin B for the treatment of systemic candidiasis.

The role of tumor necrosis factor alpha (TNF-alpha) in host defense against systemic Candida albicans infection was evaluated in a murine model of systemic candidiasis in which uniform death occurred between 5 and 6 days after infection. TNF-alpha was first detected at 16 h postinfection and progressively increased thereafter. Peak levels (700 to 900 pg/ml) were measured in mice near death. Administration of 0.5 to 1.0 mg of polyclonal immunoglobulin G (IgG) TNF-alpha antibody (TNF-alpha Ab) to mice 2 h preinfection neutralized serum TNF-alpha for up to 30 h. However, this regimen shortened survival from a mean of 5.5 days for IgG controls to 3.4 days (P = 1.9 x 10(-12)). Semiquantitative cultures of spleen, lung, liver, and kidney conducted at 1, 2, and 3 days postinfection found colony counts of spleen and kidney to be significantly higher for TNF-alpha Ab recipients but only for the first 48 h. Administration of 1.5 and 1.0 mg of TNF-alpha Ab at 2 h before and 48 h after fungal injection, respectively, shortened the mean survival from 4.9 to 2.3 days (P = 5.2 x 10(-8)). This regimen neutralized serum TNF-alpha throughout infection. With this regimen, colony counts of all organs were significantly higher in TNF-alpha Ab recipients at 1, 2, and 3 days postinfection. Histopathologic studies showed an increase in the number and size of C. albicans foci in tissues. Peripheral leukocyte counts and inflammatory response in tissue were similar for TNF-alpha Ab and IgG sham recipients. In vitro, incubation of C. albicans with four to eight times the peak serum levels of TNF-alpha for up to 24 h did not inhibit the rate of germ tube or pseudohypha formation. Thus, TNF-alpha that was produced during infection with C. albicans augmented host resistance against this organism and prolonged survival. The protective effect of TNF-alpha was not mediated by increased leukocytes in blood or tissues nor by a direct anticandidal effect of TNF-alpha. This study suggests that the administration of exogenous TNF-alpha may enhance host resistance against systemic C. albicans infection and may improve host survival.

The activities of FK037, cefpirome, ceftazidime and cefepime against 71 aminoglycoside-resistant, 35 aminoglycoside-sensitive, 29 cystic fibrosis Pseudomonas aeruginosa isolates, and 31 Pseudomonas spp. strains were studied using the agar dilution technique (final inoculum 10(4) c.f.u./spot). The MIC90 for aminoglycoside-sensitive P. aeruginosa against FK037, cefpirome, ceftazidime and cefepime was 32, 16, 8 and 16 mg/l, respectively. The MIC90 for P. aeruginosa strains resistant to one or more aminoglycosides was similar for FK037, cefpirome and ceftazidime (128 mg/l) and two dilutions lower for cefepime (32 mg/l). The MIC90 for P. aeruginosa isolates highly resistant to all three aminoglycosides (MIC > or = 128 mg/l) was 64 mg/l for FK037 and cefpirome, and 32 mg/l for ceftazidime and cefepime. The MIC90 for P. aeruginosa from patients with cystic fibrosis was 32 mg/l for all four cephalosporins tested, and 8, 32 and 64 for tobramycin, gentamicin and amikacin, respectively. Xanthomonas maltophilia was resistant to all four cephalosporins and three aminoglycosides. The activity of ceftazidime and cefepime was one to two dilutions greater against P. cepacia and P. picketti than of FK037 and cefpirome. The activity of ceftazidime was two dilutions greater than the other three cephalosporins against P. fluorescens. In kinetic time kill curves against P. aeruginosa, all four cephalosporins demonstrated similar activity at 6 and 24 h when tested at 1 x MIC. At 2 x MIC, regrowth was less at 24 h for cefepime, cefpirome and FK037 than for ceftazidime. In time kill curves for P. aeruginosa, synergy was clearly demonstrated at 1/4 MIC and 1/2 MIC concentrations for FK037 and tobramycin.

Candida glabrata (Cgl) and Candida parapsilosis (Cpa) can cause serious infections and can be resistant to some antifungal drugs. In treating infections caused by these organisms, killing rates and post-antifungal effects (PAFE) are important factors in both dose interval choice and outcome. Two strains each of Cgl and Cpa were studied. For PAFE studies, each organism was exposed to micafungin (MCF), anidulafungin (ANF), or caspofungin (CAS) for 1 h at concentrations ranging from 0.25 to 16×MIC. Cell suspensions were then washed 3 times and resuspended in fresh broth. Time 0 was immediately after resuspension of the yeast. Time-kill experiments were done using similar drug concentrations. Samples were removed at each time point (0-120 h) and viable counts determined. PAFE of ANF and CAS were generally very long, were markedly longer than those of MCF, and increased with increased drug concentration. For ANF and CAS, PAFE for Cgl were greater than those for Cpa only at 0.5 to 2 × MIC. Time-kill experiments showed that ANF, CAS, and MCF were fungicidal at 8 to 16 × MIC up to 120 h. CAS had the greatest activity against Cgl, while ANF and MCF were more active than CAS against Cpa. Because of the prolonged PAFE of these echinocandins, especially ANF and CAS, less frequent dosing during therapy of Cpa and Cgl infections could be considered. Further studies are needed to determine the clinical efficacy of longer dosing intervals.

Serial dilution susceptibility testing of imipenem against 59 clinical isolates of Pseudomonas aeruginosa, conducted simultaneously on single lots of Difco and BBL Mueller-Hinton agar (MHA), resulted in MICs for 90% of strains tested of 8 and 16 micrograms/ml, respectively. MICs for Escherichia coli, Klebsiella pneumoniae, and Pseudomonas spp. were also higher on BBL MHA. Quantification of the cation content of the two MHAs by atomic absorption spectroscopy demonstrated that the zinc concentration in BBL MHA was 15 times greater than that measured in Difco MHA (2.61 and 0.17 micrograms/ml, respectively). Concentrations of calcium, magnesium, iron, manganese, and copper in the two agars were similar. Addition of zinc to Difco MHA resulted in increases in MICs of imipenem for P. aeruginosa but not in the MICs of ceftazidime or cefpirome for P. aeruginosa (P < 0.01). A lesser zinc effect was seen on the activity of imipenem against E. coli, K. pneumoniae, and Pseudomonas spp. The activities of ceftazidime and cefpirome were similar on both MHAs when tested against all gram-negative organisms in this study. Thus, the effect of zinc in MHA was clearly demonstrated by a significant increase in the MICs of imipenem for P. aeruginosa, and, to a lesser extent, for other gram-negative bacilli.

The in vitro inhibitory and bactericidal activities of daptomycin and reference antibiotics were determined by serial macrobroth dilution for 23 gentamicin-susceptible (MIC:2,000 mg/l) and 21 high-level-gentamicin-resistant (HLGR) Enterococcus faecalis, Enterococcus faecium, and Enterococcus avium isolates. The activities of daptomycin, vancomycin, and teicoplanin were independent of the gentamicin susceptibility profile and species tested. For all the isolates, the inhibitory activity of daptomycin (MIC90: 2 mg/l) was comparable to vancomycin (MIC90: 2 mg/l), but less than that of teicoplanin (MIC90: 0.5 mg/l). Daptomycin demonstrated excellent bactericidal activity against all enterococci tested (MBC90: 8 mg/l). In contrast, all microorganisms were tolerant to vancomycin and teicoplanin. Ampicillin and ciprofloxacin MICs and MBCs were dependent on enterococcal gentamicin resistance profile and species, with MICs and MBCs that ranged between 1 and64 mg/l. By time-kill curves, the combination of daptomycin plus ampicillin demonstrated synergistic bactericidal activity against gentamicin-susceptible and HLGR E. faecalis. Daptomycin, singly and in combination, may be useful in treating enterococcal infections, including those caused by HLGR microorganisms.

The in vitro activity of sparfloxacin, a new fluoroquinolone, was assessed against 234 gram-positive bacterial isolates by agar dilution (10(4) CFU/spot). Sparfloxacin activity was compared with that of ciprofloxacin and five other antibiotics. Sparfloxacin was the most active drug tested against methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MRSA) and coagulase-negative staphylococci (MIC90, 0.125-0.25 mg/l). Sparfloxacin was also the most active drug tested against Enterococcus faecalis (MIC90, 1 mg/l) and showed equal activity against gentamicin-susceptible and gentamicin-resistant (MIC greater than 2,000 mg/l) enterococci. Sparfloxacin was the most active quinolone tested against Streptococcus pneumoniae and S. pyogenes (MIC90, 1 mg/l). Most Corynebacterium jeikeium showed exquisite susceptibility to sparfloxacin (MIC, 0.06-0.25 mg/l). For MRSA, time-kill curves showed sparfloxacin to be rapidly bactericidal at the MIC of the organism. Sparfloxacin showed greater and more sustained bactericidal activity than ciprofloxacin and vancomycin at 1x and 2x the MIC. Reduction in the activity of sparfloxacin occurred with decreased agar pH (from 7.0 to 6.0) and increased bacterial inoculum. Sparfloxacin showed superior activity compared to reference drugs against most gram-positive bacteria.

Objectives: The antifungal effects of voriconazole and caspofungin, singly and in combination, were determined against Candida glabrata in time–kill curves in broth, in human monocyte-derived macrophages (MDMs) and in MDMs activated by granulocyte-macrophage colony-stimulating factor (GM-CSF). Methods: Three strains of fluconazole-resistant C. glabrata were evaluated. For intracellular studies, MDM monolayers, with or without GM-CSF activation, were infected with C. glabrata and treated with voriconazole and caspofungin at 2.53 and 53 MIC, respectively, or at 13 MIC. Extracellular studies in broth were performed using drug concentrations from 0.1 to 103 MIC. Viable yeast were enumerated at 0, 24 and 48 h. Results: Significantly greater killing of C. glabrata occurred with the drug combination than with either single drug, both intracellularly and extracellularly (P < 0.01). For voriconazole, the antifungal activity in MDM activated by GM-CSF was greater than that in unactivated MDM, regardless of antibiotic concentration or time of exposure. However, for caspofungin and for the two-drug combination, enhanced activity in GM-CSF-activated MDM depended on the drug concentration and time of exposure. Conclusions: Our data suggest that combinations of voriconazole and caspofungin may be efficacious for the treatment of serious C. glabrata infections. With single-drug therapy, especially voriconazole, GM-CSF activation of monocytes could be considered.

We investigated the antistaphylococcal activities of daptomycin, gentamicin, and rifampin against two Staphylococcus aureus strains and their stable small-colony variants, singly and in combination, in human monocyte-derived macrophages and in broth. Intracellularly, the three-drug combination and two-drug combinations with rifampin were most effective. Extracellularly, daptomycin, daptomycin plus gentamicin, gentamicin plus rifampin, and the three-drug combination had similar activities.

Killing curves were used to characterize Staphylococcus haemolyticus isolates previously reported to contain subpopulations showing increased resistance to vancomycin. Results suggested that vancomycin and teicoplanin were ineffective at a concentration of 8 micrograms/ml and growth was seen between 24 and 48 h. Conversely, the lipopeptide antibiotic daptomycin at the same concentration rapidly killed tested strains by 6 h. Various staphylococcal strains were examined to determine if vancomycin resistance could be selected in all strains of staphylococci, was specie(s) restricted, or was unique to this patient's clinical isolates. About 1 x 10(8) colony-forming units were added to melted brain-heart infusion agar plates containing 12 micrograms/ml of vancomycin. Plates were examined after 48 h for presence of resistant clones. Results indicated that selection for vancomycin resistance was restricted to S. haemolyticus strains. Further, all S. haemolyticus isolates that displayed a double zone of growth around imipenem agar diffusion discs (Impdz) contained stably resistant subpopulations. Vancomycin resistance could not be selected in imipenem-sensitive derivative clones. Impdz isolates that were recovered from geographically distinct locations displayed nearly identical SDS-PAGE protein profiles. It appears that a characteristic susceptibility pattern displayed by clinical isolates of S. haemolyticus may provide a marker for those strains that contain subpopulations having increased resistance to vancomycin.

The in-vitro activity of cefpirome and ceftazidime when combined with aminoglycosides (gentamicin, amikacin, and tobramycin) in the presence and in the absence of rifampicin was evaluated against 32 isolates of Pseudomonas aeruginosa by two methods. Agar dilution susceptibilities demonstrated a marked reduction in synergy (FIC less than or equal to 0.5) when rifampicin was added to the combination. Synergy rates decreased from 59.4-84.4% without to 3.1-9.4% with the addition of rifampicin. In contrast, kill curve tests performed on two P. aeruginosa strains demonstrated synergy at 24 h when rifampicin was added to cefpirome, ceftazidime, gentamicin or a beta-lactam agent plus gentamicin combination. The addition of rifampicin to the combinations of cefpirome or ceftazidime plus gentamicin achieved a 2-log10 lower bacterial count at 24 h than that of the beta-lactam and gentamicin combination alone. When rifampicin was added to the combination cefpirome or ceftazidime plus gentamicin at different times during incubation, a greater bactericidal effect was observed when rifampicin was added at 0 and 1 h of incubation than when added later. No antagonism was observed with rifampicin when used in combination with beta-lactam agents and/or aminoglycosides.

To determine the effects of recombinant human activated protein C (rhAPC) on the antimicrobial activity and cytokine production of normal human monocyte-derived macrophages (MDMs) in the presence and absence of Escherichia coli infection, with and without treatment with levofloxacin or ampicillin.MDM monolayers were infected with E. coli ATCC 25922 and treated with levofloxacin or ampicillin in the presence or absence of rhAPC. Antimicrobial activity and cytokine (TNF-alpha, IL-1beta, IL-6 and IL-8) concentrations in the supernatants were measured.When low concentrations of levofloxacin were used, a therapeutic concentration of rhAPC enhanced intracellular antibacterial activity at all time points. With ampicillin, antibacterial activity increased, was unaffected or diminished depending upon the drug concentration and assay time. Without antibiotics, rhAPC had no antibacterial effect. E. coli caused cytokine production to increase many fold. This increase was significantly greater with antibiotics (P0.01). Without antibiotics, rhAPC decreased production of TNF-alpha, IL-1beta and IL-6, but not IL-8. At high levofloxacin concentrations, rhAPC was associated with further increases in the concentrations of these cytokines. Cytokine concentrations at 24 h were unaffected by rhAPC in the presence of ampicillin and E. coli.rhAPC can affect the bactericidal activity and cytokine production of human MDM in the presence of infection and antibiotic therapy. Importantly, factors such as type and concentration of antibiotics, presence of bacteria and timing must be taken into consideration when evaluating cytokine data from septic patients.

The prevalence of, and clinical risk factors associated with, vancomycin-resistant enterococcal colonization were investigated in patients suspected of having Clostridium difficile infection. Stools submitted for C difficile cytotoxin testing were screened for vancomycin-resistant enterococci (VRE). Isolates were speciated and characterized further by antibiotic susceptibility testing, DNA fingerprinting, and DNA:DNA hybridization for detection of specific vancomycin resistance genes. Of the 79 evaluable patients identified during a 3-month period, 16.5% were VRE-positive. The VRE isolates were genetically heterogeneous, although all carried the vanA gene. DNA fingerprinting data suggest that patient-to-patient transmission occurred, implicating colonized patients as potential reservoirs for VRE transmission. A positive C difficile cytotoxin assay and diabetes mellitus were the only identifiable risk factors associated with VRE colonization. Patients at risk for C difficile infection therefore may serve as reservoirs for VRE (Infect Control Hosp Epidemiol 1997;18:342-344). Vancomycin-resistant enterococci (VRE) were first identified in 1988.1 Since then, sporadic case reports and reports of nosocomial outbreaks of VRE infections have appeared in the literature.2,3 Enterococcus faecium and Enterococcus faecalis, the most common species of Enterococcus causing disease in humans, have been implicated in these outbreaks. From 1989 to 1993, the percentage of VRE among the enterococcal isolates reported by institutions participating in the National Nosocomial Infection Surveillance System increased from 0.3% to 7.9%.3 The greatest increase was seen in intensive-care-unit patients, for whom the percentage of enterococcal isolates resistant to vancomycin increased from 0.4% to 13.6%. Investigations of epidemic outbreaks have revealed that the patients at greatest risk are the critically ill, transplant and oncology patients, those who have undergone intra-abdominal or cardiothoracic surgical procedures, had prolonged hospital stays, have received multiple antibiotics, or have received vancomycin.4 It has been shown that patients with Clostridium difficile infection treated with oral vancomycin are at increased risk for colonization or infection with VRE.5 This 3-month prospective study was designed to investigate the possible correlation between oral vancomycin administration and VRE carriage, to define the clinical and epidemiological significance of asymptomatic VRE colonization, and to identify predictive risk factors for such colonization.

Objectives: The antibacterial activity of a new fluoroquinolone, gemifloxacin, was tested against intracellular Legionella pneumophila and Legionella micdadei and was compared with the activities of levofloxacin, gatifloxacin, moxifloxacin and erythromycin. Methods: For intracellular assays, bacteria were used to infect human monocyte-derived macrophages prepared from heparinized blood of healthy volunteers. Antibiotics were added following phagocytosis. Numbers of viable bacteria were determined at 0, 24, 48, 72 and 96 h. Results:Theintracellularantibacterialactivityofgemifloxacinwasconcentration-andtime-dependent.All of the quinolones had similar activities against L. pneumophila and L. micdadei at 10 · MIC, but there were minor differences: at 24 h moxifloxacin was significantly more active than the other quinolones against L. pneumophila, while gemifloxacin was more active against L. micdadei (P < 0.01). All of the quinolones were markedly more active than erythromycin (P < 0.01). The antibacterial effect of gemifloxacin against L. pneumophila following drug removal at 24 h persisted for 72 h at 20 · MIC but not at 10 · MIC, while for L. micdadei the antibacterial effect persisted for 24 h at 10 · MIC. Conclusions: All of the quinolones had similar activities against intracellular L. pneumophila and L. micdadei and were markedly more effective than erythromycin.

We studied 20 Chlamydia pneumoniae isolates obtained from respiratory sites and atheroma tissue of patients from various geographic areas to determine the susceptibilities of these isolates to a new des-fluoroquinolone, garenoxacin, and to levofloxacin. In addition, we assessed the cultures with these isolates by PCR for the presence or absence of Mycoplasma sp. DNA. Both the MIC at which 90% of isolates are inhibited (MIC 90 ) and the minimal bactericidal concentration at which 90% of isolates are killed (MBC 90 ) for garenoxacin were 0.06 μg/ml, and both the MIC 90 and the MBC 90 for levofloxacin were 2.0 μg/ml. The activity of garenoxacin against C. pneumoniae was 32-fold greater than that of levofloxacin. Mycoplasma sp. DNA was detected by PCR in 17 of 20 cultures. Mycoplasma amplicons from five Mycoplasma DNA-positive C. pneumoniae cultures were sequenced and found to represent four Mycoplasma species. Our data demonstrate that C. pneumoniae cultures frequently contain Mycoplasma DNA and that its presence in C. pneumoniae cultures does not appear to affect the susceptibility results for the two fluoroquinolones that we tested.

Using the standard Craig and Gudmundsson method (W. A. Craig and S. Gudmundsson, p. 296-329, in V. Lorian, ed., Antibiotics in Laboratory Medicine , 1996) as a guideline for determination of postantibiotic effects (PAE), we studied a large series of growth curves for two strains of Legionella pneumophila . We found that the intensity of the PAE was best determined by using a statistically fitted line over hours 3 to 9 following antibiotic removal. We further determined the PAE duration by using a series of observations of the assay interval from hours 3 to 24. We determined that inoculum reduction was not necessarily the only predictor of the PAE but that the PAE was subject to the type and dose of the drug used in the study. In addition, there was a variation between strains. Only levofloxacin at five and ten times the minimum inhibitory concentration (MIC) resulted in a PAE duration of 4 to 10 h for both strains of L. pneumophila tested. Ciprofloxacin at five and ten times the MIC and azithromycin at ten times the MIC caused a PAE for one strain only. No PAE could be demonstrated for either strain with erythromycin or doxycycline. Using the presently described method of measuring PAE for L. pneumophila , we were able to detect differences in PAE which were dependent upon the L. pneumophila strain, the antibiotic tested, and the antibiotic concentration. We suggest the use of mathematically fitted curves for comparison of bacterial growth in order to measure PAE for L. pneumophila .

The inhibitory and bactericidal effects of telithromycin (HMR 3647, RU 66647) were compared with those of gentamicin, ampicillin, erythromycin, azithromycin and vancomycin against 74 strains of enterococci (34 Enterococcus faecalis and 40 Enterococcus faecium) by agar dilution, broth dilution, time kill assays and postantibiotic effect (PAE). The telithromycin MIC90 for vancomycin-sensitive (VSE) E. faecalis strains tested using the agar dilution method was 8 µg/ml. For a different group of VSE E. faecalis strains tested using the broth dilution method it was 0.06 µg/ml The telithromycin MIC90s for vancomycin-resistant (VRE) and VSE E. faecium strains, determined using the agar dilution method, were 4 and 8 µg/ml, respectively, while for a different set of VRE and VSE E. faecium strains tested using the broth macrodilution method, they were 32 and 16 µg/ml, respectively. Telithromycin MBC90s for E. faecalis were 4–6 tubes higher and for E. faecium 3–5 tubes higher, respectively, than the MIC90s. In time kill assays, telithromycin had bactericidal activity against only 1 of 7 E. faecium strains; for all other E. faecium and E. faecalis strains, only inhibitory activity was demonstrated. Neither synergy nor drug interference was observed when telithromycin was used in combination with ampicillin, vancomycin or gentamicin. At 10 times the MIC, the PAE of telithromycin against E. faecalis was 2.8 h, while for E. faecium it was 1.6 h. Telithromycin should be evaluated for therapy of enterococcal infections, including those caused by VRE organisms. However, because of the strain-to-strain variability in susceptibility to telithromycin, MIC determinations are important, especially for erythromycin-resistant strains.

The activity of the ketolide telithromycin (HMR 3647) against intracellular Legionella pneumophila strain L-1033 was compared with the activities of erythromycin and levofloxacin. To assay intracellular antibacterial activity, human monocytes were allowed to adhere to wells in 24-well tissue culture plates and were then exposed to L. pneumophila cells for 1 h to allow phagocytosis to occur. Antibiotics were added to the wells after removal of unphagocytosed bacteria. Quantitative bacterial cell counts were made from lysed monocytes at 0, 24, 48, 72 and 96 h. The antibacterial effects of antibiotics against intracellular L. pneumophila L-1033 were concentration and time dependent; at 10 x MIC the activity of telithromycin was greater than that of erythromycin and was less than that of levofloxacin (P < 0.01); telithromycin-rifampicin combinations showed no synergy or interference; and removal of telithromycin from assays at 24 h did not affect its intracellular antibacterial activity. In conclusion, the ketolide telithromycin has excellent activity against intracellular L. pneumophila strain L-1033 and should be evaluated for therapy of legionnaires' disease.

Background: MDI-P (Medical Discoveries, Inc–Pharmaceutical, Layton, Utah) is a clear, colorless liquid generated by electrolysis of preservative-free and endotoxin-free, nonpyrogenic, sterile, injection saline (0.9% NaCl, wt/vol). It contains numerous highly reactive chlorine and oxygen species, including HOCl –1, OCl– 1 , Cl –1 , Cl 2 , O 2– 1 , and O 3 . This report presents data on the in vitro microbicidal activity of MDI-P against 4 clinically relevant microbial pathogens that are often difficult to eradicate. Methods: MDI-P was generated from injection saline by using a patented electrolysis instrument. It was then tested for microbicidal activity at concentrations ranging from 0.01% to 50% against Staphylococcus aureus, Pseudomonas aeruginosa, Legionella pneumophila, and Candida albicans (10 5 to 10 9 colony-forming units/mL). The effect of serum (50% and 90%) and pH on MDI-P activity were also tested. The morphologic effects of MDI-P on microbial cells were studied by light microscopy of cells stained by Gram's method and by transmission electron microscopy. Morbidity, mortality, and the effect of MDI-P on tissues were studied by using a mouse model. Results: The microbicidal activity of MDI-P occurred within the first minute of exposure for all the organisms tested. When 50% MDI-P was tested against cell titers of 10 5 or 10 7 colony-forming units/mL, all test organisms were killed within 1 minute; at lower MDI-P concentrations, C albicans was the most sensitive organism, and L pneumophila was the most resistant. Even with beginning cell titers of 10 9 colony-forming units/mL, killing by 50% MDI-P was >99.9% for all test strains. Furthermore, at the same beginning cell titer, killing of C albicans by MDI-P diluted to 50% with normal human serum rather than injection saline was only slightly reduced. No acute morbidity, mortality, or tissue damage was detected in mice that were intravenously given 17 mL/kg of undiluted MDI-P. Conclusions: MDI-P is a very fast-acting, broad-spectrum microbicidal material. The lack of evidence for acute morbidity, mortality, or tissue injury, ease of preparation, and low cost suggest that it may be useful for various sterilization and disinfection applications. (AJIC Am J Infect Control 2000;28:251-7)

Opsonophagocytic killing of some bacteria (Staphylococcus aureus, Pseudomonas aeruginosa) by phagocytes is enhanced by previous brief exposure of the organism to antibiotics. We studied the regrowth of Legionella pneumophila previously pretreated with levofloxacin, erythromycin and/or rifampicin in human monocytes. The MIC for the L. pneumophila isolate of levofloxacin, erythromycin and rifampicin was 0.03, 0.5 and 0.001 mg/L, respectively. Growth of L. pneumophila from buffered charcoal yeast extract (BCYE) agar for 24 h was subcultured into BYE broth containing from 0 to 4x MIC of levofloxacin, erythromycin or rifampicin. After incubation at 35 degrees C in 5% CO2 for 18 h, the organisms were washed and opsonized with 20% heat inactivated pooled normal human serum. Thereafter, L. pneumophila was exposed to human monocytes (5:1 ratio) previously adhered to wells in tissue culture plates containing RPMI and 10% fetal calf serum. After 0, 24, 48 and 72 h of incubation, quantitative cultures of lysed human monocytes were done on BCYE agar. Our results indicate effective inhibition on L. pneumophila at 0 h regardless of the antibiotic (levofloxacin, rifampicin or erythromycin) or their concentrations (1x, 2x or 4x MIC). At 24, 48 and 72 h, recovery and regrowth of L. pneumophila were both antibiotic- and concentration-dependent. In comparison with controls (no antibiotic pretreatment), peak regrowth of L. pneumophila pretreated with either 1x MIC of levofloxacin or erythromycin was delayed (48 versus 24 h) and reduced (30% of control peak regrowth). Regrowth of L. pneumophila pretreated with 1x MIC of rifampicin continued beyond 72 h. Pretreatment with levofloxacin at 4x MIC caused the greatest degree of growth inhibition (2 log10). In contrast, at 72 h, regrowth of organisms pretreated with 4x MIC of erythromycin or rifampicin was less than peak control (P < 0.01) but greater than that seen with levofloxacin (P < 0.01). The rate and degree of regrowth of L. pneumophila pretreated with combinations of levofloxacin or erythromycin with rifampicin, or levofloxacin with erythromycin (all at 1x MIC) was similar to that seen with single drugs. Thus, significant delay and reduction of regrowth in this phagocytic system occurred with levofloxacin only. Prolonged exposure of the organism at 4x MIC levofloxacin concentrations was effective in suppressing regrowth of pretreated L. pneumophila in human monocytes.

Pentoxifylline can decrease the production of tumour necrosis factor alpha (TNF alpha) by endotoxin-stimulated macrophages and may improve survival in animals with overwhelming bacterial sepsis. In this study various doses of pentoxifylline were administered to mice with systemic Candida albicans infection to determine its effect on serum TNF alpha levels, organ fungal burden, and host survival. Intraperitoneal injections of pentoxifylline at 20 mg/kg every 8 h did not affect these endpoints. However, fungal counts were significantly higher in kidneys of animals that received 30 and 60 mg/kg of pentoxifylline every 8 h when compared to controls. Injection of 60 mg/kg of pentoxifylline at 8 h intervals also significantly shortened mean survival from 5.8 to 3.8 days (P = 0.01). Pentoxifylline did not affect peripheral WBC counts, serum TNF alpha and interleukin-6 levels, or the density of neutrophils in tissues. In vitro, pentoxifylline decreased the production of TNF alpha by C. albicans-stimulated macrophages in a dose-dependent manner, but only at concentrations greater than 100 mg/L. In contrast, pentoxifylline suppressed TNF alpha production by endotoxin-stimulated macrophages at concentrations as low as 10 mg/L. Thus, higher doses of pentoxifylline are detrimental in systemic C. albicans infection. However, the detrimental effect is not mediated by alterations in serum TNF alpha or interleukin-6 levels or the aggregation of neutrophils in tissues.

Within a period of 13 years 39 patients underwent surgery for primary gastrointestinal non-Hodgkin's lymphoma. The stomach was the most frequent site of involvement (26 cases) followed by the small intestine (10 cases) and the large bowel (three cases). Operative procedures included: total gastrectomy (n = 19), subtotal gastrectomy (n = 5), partial gastrectomy (n = 2), small bowel resection (n = 7), right hemicolectomy (n = 5) and sigmoid resection (n = 1). According to Musshoff's modification of the Ann Arbor system, we found stage I in nine, stage II in 12, stage III in two and stage IV in 16 patients. Histological typing according to the Kiel classification showed low-grade malignancy in 11 cases and high-grade in 28. Twenty-eight patients received chemotherapy and/or radiation in accordance with the tumour staging and type of malignancy. The follow-up data were analysed by the method of Kaplan and Meier. Including five patients who died post-operatively from perforated or bleeding lymphomas, the overall 5-year survival rate was 53.8%.

The susceptibilities of 56 Legionella pneumophila isolates (43 clinical and 15 environmental isolates) to levofloxacin, ofloxacin, erythromycin, and rifampin were studied with buffered charcoal yeast extract (BCYE) agar (inoculum, 10(4) CFU per spot), and the susceptibilities of five isolates were studied with buffered yeast extract (BYE) broth (inoculum, 10(5) CFU/ml). The MICs inhibiting 90% of strains tested on BCYE agar were 0.125, 0.25, 1.0, and < or = 0.004 micrograms/ml for levofloxacin, ofloxacin, erythromycin, and rifampin, respectively. The MICs by the BYE broth dilution method were 1 to 3, 2, 1 to 2, and 1 tube lower than those by the agar dilution method for levofloxacin, ofloxacin, erythromycin, and rifampin, respectively. The MBCs were 1 to 2 tubes higher than the broth dilution MICs for levofloxacin, 1 to 3 tubes higher than the broth dilution MICs for ofloxacin, 1 to 3 tubes higher than the broth dilution MICs for erythromycin, and the same as the broth dilution MICs for rifampin. In kinetic time-kill curve studies, at drug concentrations of 1.0 and 2.0 times the MIC, the most active drugs were levofloxacin and rifampin. At 72 h, concentrations of levofloxacin and rifampin of 2.0 times the MIC demonstrated a bactericidal effect against L. pneumophila. In contrast, at concentrations of 1.0 and 2.0 times the MICs regrowth was observed with ofloxacin and only a gradual decrease in the numbers of CFU per milliliter was observed with erythromycin. Only a minor inhibitory effect was observed with 0.25 or 0.5 time the MICs of all drugs at 24 to 48 h, with regrowth occurring at 72 h. In contrast to erythromycin or ofloxacin plus rifampin at 0.25 time the MICs, only levofloxacin plus rifampin demonstrated synergy. Thus, levofloxacin demonstrated the best inhibitory and bactericidal effects against L. pneumophila when it was studied alone or in a combination with rifampin.

The in vitro activity of cefdinir, an oral aminothiazolyl hydroxyimino cephalosporin was compared with that of cefixime, cefpodoxime, cefaclor, cephalexin, ciprofloxacin, ofloxacin, oxacillin, ampicillin, vancomycin and trimethoprim-sulfamethoxazole against 279 gram-positive and gram-negative recent clinical isolates from adult and pediatric patients. Cefdinir was the most active drug among the cephalosporins against oxacillin-sensitive Staphylococcus aureus and coagulase-negative staphylococci, Streptococcus pneumoniae, S. pyogenes, Escherichia coli and Moraxella catarrhalis (MIC90 0.015-2 mg/l). Cefixime was the most active agent against Hemophilus influenzae, Klebsiella pneumoniae, K. oxytoca, Proteus mirabilis and P. vulgaris (MIC900.015-0.125 mg/l). The activity of cefpodoxime was better than that of cefixime against S. pneumoniae and oxacillin-sensitive staphylococci (MIC90 0.25-8 vs. 0.5-32 mg/l), similar to cefixime against S. pyogenes (MIC90 0.06 mg/l) and not as good as cefixime against H. influenzae, M. catarrhalis, Klebsiella spp. and Proteus spp. (MIC900.015-0.25 vs. 0.125-0.5 mg/l). The activity of cefdinir was greater than that of the other cephalosporins against Enterococcus faecalis (MIC90 16-32 vs.64 mg/l). None of the cephalosporins were active against methicillin-resistant, coagulase-positive or -negative staphylococci or Pseudomonas aeruginosa (MIC9064 mg/l). Overall, the susceptibilities of adult and pediatric isolates were similar. Kinetic kill curves demonstrated rapid and similar killing at 6 h by cefdinir, cefixime, cefpodoxime and ofloxacin. At 24 h at 1 x MIC, the least regrowth was observed with cefdinir and cefpodoxime; at 2 x MIC, suppression of growth was similar with all four drugs.

The effect of Pseudomonas aeruginosa cytotoxin was assessed in leukopenic outbred Swiss male mice (20 g) using a high cytotoxin-producing strain (PA158) and its cytotoxin-deficient isogenic mutant (PA114F5) generated by Tn7::Tn5 transposon mutagenesis of PA158. Leukopenia was induced by intraperitoneal (i.p.) administration of cyclophosphamide (150 μg/g). Anesthetized mice were infected via a 4 mm incision on the shaved back with 300 CFU/mouse (9 ld50; expected death rate 85%). Precleared mouse cytotoxin-specific heat inactivated rabbit polyclonal antibody (RPA) was administered i.p. (0.2 ml) 24 hr before challenge. Controls received i.p. normal rabbit serum, RPA, cyclophosphamide alone, or a sham procedure. Challenge with the high cytotoxin-producing strain PA158 caused earlier and a significantly greater mortality than that observed with a cytotoxin-deficient strain PA114F5 (P < 0.01). Cytotoxin-specific polyclonal antibody was protective. Pretreatment with antibody decreased the mortality rate following challenge with PA158 from 88.9% to 27.8% (P < 0.01). Pretreatment with antibody decreased the mortality rate following challenge with PA114F5 from 27.8% to 5.6% (P < 0.05). These results demonstrate that P. aeruginosa cytotoxin contributes to the pathogenicity of the organism and that cytotoxin antibody is protective in a systemic P. aeruginosa infection in leukopenic mice.

The in-vitro activities of two new fluoroquinolones, clinafloxacin (CI-960, PD 127391, AM-1091) and PD 131628 (the active component of the pro-drug CI-990, PD 131112) and nine other antibiotics were tested against 107 clinical isolates of Pseudomonas aeruginosa, 12 isolates of Xanthomonas maltophilia, and 19 isolates of other Pseudomonas spp. Of the 107 P. aeruginosa isolates, 33 were resistant to gentamicin, tobramycin and amikacin, 17 were resistant to only one or two of these aminoglycosides and 24 were aminoglycoside sensitive. Thirty-three were isolates from cystic fibrosis patients. Susceptibility studies were performed using the agar dilution technique and kinetic time kill curves. With the exception of aminoglycoside-sensitive P. aeruginosa isolates where ciprofloxacin had similar activity to clinafloxacin and PD 131628, the two new fluoroquinolones were the most active agents against all isolates tested (MIC90 0.25-2.0 mg/L). Cross-resistance was identified with ciprofloxacin and ofloxacin-resistant strains, but the superior activity of clinafloxacin and PD 131628 resulted in 90% of the isolates having MICs < 2 mg/L. Kinetic kill curves with aminoglycoside-sensitive P. aeruginosa revealed ciprofloxacin to have the most rapid and sustained killing. However, with amino-glycoside-resistant P. aeruginosa isolates, clinafloxacin and PD 131628 were more rapidly bactericidal than ciprofloxacin.

The susceptibility of sparfloxacin, a new broad spectrum fluoroquinolone, was determined for 72 clinical isolates of Pseudomonas aeruginosa, 15 Xanthomonas maltophilia, and 19 Pseudomonas spp. The activity of sparfloxacin was compared with that of ciprofloxacin and other reference antibiotics. Sparfloxacin was the most active antibiotic tested against X. maltophilia (MIC90 1 mg/l) and the most active quinolone against P. cepacia and P. putrifaciens. Ciprofloxacin, however, demonstrated greater activity than sparfloxacin against P. fluorescens and P. stutzeri. P. aeruginosa was most susceptible to ciprofloxacin with an MIC90 of 2 mg/l, compared with an MIC90 of 8 mg/l for sparfloxacin and ofloxacin. Although cross-resistance between quinolones was noted, cross-resistance between antibiotic classes was not seen. Aminoglycoside-resistant and aminoglycoside-susceptible P. aeruginosa strains were equally susceptible to sparfloxacin. Kill curves showed sparfloxacin to be rapidly bactericidal against P. aeruginosa at 1 x MIC. Sparfloxacin demonstrated greater bactericidal activity than ciprofloxacin at 1 x and 2 x their MICs. Unlike ciprofloxacin and gentamicin, sparfloxacin showed sustained bactericidal activity at greater than or equal to 1 MIC for 24 h.

Treatment of disseminated Pseudomonas aeruginosa infection in leukopenic mice was evaluated using cefpirome alone and in combination with gentamicin and/or rifampin. Mice were made leukopenic with cyclophosphamide and infected through a skin incision with an inoculum of 1250 organisms (13 LD50). Antibiotics were administered subcutaneously for 48 h. Although the addition of cefpirome to gentamicin and/or rifampin improved survival significantly at 48 h compared with untreated controls (84.6%-100% vs. 38.5%), therapy with these combinations did not improve survival significantly from that achieved with cefpirome alone. Quantitative blood and tissue (liver, spleen, kidney, lung) cultures in mice treated with cefpirome alone or including rifampin were lower than in infected controls or groups receiving therapy that excluded cefpirome. Highest counts were observed in mice receiving cefpirome plus gentamicin. Except for the cefpirome plus gentamicin group, which demonstrated areas of acute tubular necrosis, the cefpirome group had less tissue pathology than infected controls.

Summary Using limited proteolysis by chymotrypsin or papain, we examined the effects of phosphorylation on the conformation of skeletal muscle myosin. In the absence of MgCl 2 , phosphorylation of the 19,000 dalton light chain (LC 2 ) inhibited digestion of LC 2 by chymotrypsin or papain. Phosphorylation also suppressed chymotryptic conversion of the heavy chain to subfragment 1 and increased its conversion to heavy meromyosin. These results indicate that phosphorylation alters the conformation of the N-terminal segment of LC 2 and suggest that it also affects the heavy chain. In the presence of MgCl 2 , phosphorylation inhibited the chymotryptic digestion of LC 2 but an effect on digestion of the heavy chain was not apparent. Thus, phosphorylation of LC 2 alters LC 2 conformation under physiological conditions.

The activation of the coupling factor-latent ATPase enzyme by tryptic proteolysis may resemble the activation of many proenzymes by limited proteolysis. The beta (53 000 dalton) subunit of solubilized coupling factor-latent ATPase from Mycobacterium phlei was selectively lost in some trypsin-treated samples. Since a concomitant loss of ATPase activity was not observed, the beta subunit may not be essential for ATPase catalytic activity. Treatment of solublized coupling factor with chymotrypsin rapidly produced an A′-type (61 000 dalton) species from the native alpha (64 000 dalton) subunits with partial activation of the ATPase enzyme. Secondary chymotryptic cleavage yielded an A″-type (58 000 dalton) species and a less-active enzyme. Storage of fresh coupling factor samples at −20°C in the presence of 4 mM MgCl 2 with several freeze-thaw cycles resulted in loss of ATPase activity without apparent change in alpha subunit structure. Storage at 4°C in the presence or absence of MgCl 2 both decreased ATPase activity and generated A′-type alpha subunit species. Since presence of phenylmethylsulfonyl fluoride prevented these changes, an unknown protease was suspected. The peptide bonds first cleaved by trypsin, chymotrypsin, and the unknown protease are all apparently located within the same small segment of alpha subunit polypeptide chain.