Validation of criteria for nosocomial use of amikacin in Brazil with the Delphi technique

The Delphi technique has been used since the 1950s to collect the opinions of experts; to gauge their indications, and in some instances, to develop a consensus. This systematic collection and aggregation of informed judgments from a group of experts on specific questions or issues is a highly efficient and cost-effective means to establish guidelines and policies, when compared to other strategies, such as committee meetings or personal interviews. OBJECTIVE: Examine the content validation process of the proposed criteria of the American Society of Health System Pharmacists (ASHP) for amikacin use in hospital settings. MATERIAL AND METHODS: The Delphi technique was applied using the proposed ASHP criteria questionnaire containing 102 specific questions related to the nosocomial use of amikacin by individual patients. The questionnaire contained six groups of questions: 1) Identification and basic demographic data, 2) Relevant data for the use of amikacin, 3) Justification of its usage, 4) Critical parameters of amikacin use, 5) Complications, 6) Measurement of results. Eight hospital specialist medical doctors were selected, including five in the area of infectious diseases, one surgeon, one nephrologist and one in critical care medicine. The questionnaire was e-mailed to the doctors and they were asked for their opinion about the appropriateness of the questions. They were to say whether the general concept seemed totally or partially adequate to the proposed process, what grade (0 to 10) they would give to each section, and if there were any perceived deficiencies, they could add, omit or modify individual questions. A second questionnaire containing the questions for which there had been no consensus based on the answers to the previous one was re-sent to the participants for consolidation. RESULTS: Feedback revealed an agreement of 75 percent concerning the utility and appropriateness of sections 1 and 2. The section about the justification of amikacin usage was agreed on by 50 percent. There was a total agreement of 62 percent for the critical parameters of amikacin use, and a partial agreement of 37 percent. The complication of usage of the questionnaire was agreed upon by 50 percent of the participants, and positive measurement of the results was totally agreed on by 62 percent, and partially by 37 percent. The overall score for the questionnaire was 8.77 ± 0.25. CONCLUSION: The usage criteria for amikacin recommended by ASHP were validated by the Delphi technique for utilization in Brazilian hospital settings. The Delphi technique applied to validate a questionnaire instrument for monitoring the correct use of a specific strategic antibiotic indicated for the treatment and prophylaxis of serious antibiotic-resistant Gram-negative bacteria, proved to be a reliable and simple tool for designing guidelines and a consensus document for hospital use of antibiotics.

Validation of criteria for nosocomial use of amikacin in Brazil with the Delphi technique.

UNLABELLED: The Delphi technique has been used since the 1950s to collect the opinions of experts; to gauge their indications, and in some instances, to develop a consensus. This systematic collection and aggregation of informed judgments from a group of experts on specific questions or issues is a highly efficient and cost-effective means to establish guidelines and policies, when compared to other strategies, such as committee meetings or personal interviews. OBJECTIVE: Examine the content validation process of the proposed criteria of the American Society of Health System Pharmacists (ASHP) for amikacin use in hospital settings. MATERIAL AND METHODS: The Delphi technique was applied using the proposed ASHP criteria questionnaire containing 102 specific questions related to the nosocomial use of amikacin by individual patients. The questionnaire contained six groups of questions: 1) Identification and basic demographic data, 2) Relevant data for the use of amikacin, 3) Justification of its usage, 4) Critical parameters of amikacin use, 5) Complications, 6) Measurement of results. Eight hospital specialist medical doctors were selected, including five in the area of infectious diseases, one surgeon, one nephrologist and one in critical care medicine. The questionnaire was e-mailed to the doctors and they were asked for their opinion about the appropriateness of the questions. They were to say whether the general concept seemed totally or partially adequate to the proposed process, what grade (0 to 10) they would give to each section, and if there were any perceived deficiencies, they could add, omit or modify individual questions. A second questionnaire containing the questions for which there had been no consensus based on the answers to the previous one was re-sent to the participants for consolidation. RESULTS: Feedback revealed an agreement of 75% concerning the utility and appropriateness of sections 1 and 2. The section about the justification of amikacin usage was agreed on by 50%. There was a total agreement of 62% for the critical parameters of amikacin use, and a partial agreement of 37%. The complication of usage of the questionnaire was agreed upon by 50% of the participants, and positive measurement of the results was totally agreed on by 62%, and partially by 37%. The overall score for the questionnaire was 8.77 +/- 0.25. CONCLUSION: The usage criteria for amikacin recommended by ASHP were validated by the Delphi technique for utilization in Brazilian hospital settings. The Delphi technique applied to validate a questionnaire instrument for monitoring the correct use of a specific strategic antibiotic indicated for the treatment and prophylaxis of serious antibiotic-resistant Gram-negative bacteria, proved to be a reliable and simple tool for designing guidelines and a consensus document for hospital use of antibiotics.

Gatifloxacin in the treatment of community-acquired pneumonias: a comparative trial of ceftriaxone, with or without macrolides, in hospitalized adult patients with mild to moderately severe pneumonia.

Community-acquired pneumonia is very common, but some of the cases do require hospitalization for treatment, particularly when older patients and/or co-morbidities are involved; both "typical" and "atypical" respiratory pathogens take part etiologically, and there is increasing concern about the emergence of resistance. There is interest in therapeutic options that can: a) comprehend such a spectrum of bacteria and resistance; b) allow parenteral to oral sequential treatment. We made a multicenter, prospective and randomized trial to compare the "standard" treatment of ceftriaxone IV alone or in combination with erythromycin IV, followed by clarithromycin PO (ceftriaxone treatment arm), with gatifloxacin IV, followed by oral administration (gatifloxacin treatment arm). The need for hospitalization was based on clinical criteria as judged by the investigators. Standardized criteria for diagnosis and follow-up were employed. Fifty-six patients were enrolled, with 48% over 65 years old, and there were frequent co-morbidities. Of these, 51 were clinically evaluable, 26 in the gatifloxacin and 25 in the ceftriaxone arm, with comparable success rates, 92% and 88%, respectively, even when major prognostic factors were considered. There were no serious adverse events or significant laboratory value changes attributable to the study drugs. Gatifloxacin as monotherapy (initially IV then orally until completion of treatment) was shown to be effective and safe, comparable to ceftriaxone IV alone or in combination with a macrolide (initially IV then orally until completion of treatment), in empirical therapy for community-acquired pneumonias, for patients that, at the physician s discretion, require initial treatment as inpatients.

Gatifloxacin in the treatment of community-acquired pneumonias: a comparative trial of ceftriaxone, with or without macrolides, in hospitalized adult patients with mild to moderately severe pneumonia

Community-acquired pneumonia is very common, but some of the cases do require hospitalization for treatment, particularly when older patients and/or co-morbidities are involved; both "typical" and "atypical" respiratory pathogens take part etiologically, and there is increasing concern about the emergence of resistance. There is interest in therapeutic options that can: a) comprehend such a spectrum of bacteria and resistance; b) allow parenteral to oral sequential treatment. We made a multicenter, prospective and randomized trial to compare the "standard" treatment of ceftriaxone IV alone or in combination with erythromycin IV, followed by clarithromycin PO (ceftriaxone treatment arm), with gatifloxacin IV, followed by oral administration (gatifloxacin treatment arm). The need for hospitalization was based on clinical criteria as judged by the investigators. Standardized criteria for diagnosis and follow-up were employed. Fifty-six patients were enrolled, with 48 percent over 65 years old, and there were frequent co-morbidities. Of these, 51 were clinically evaluable, 26 in the gatifloxacin and 25 in the ceftriaxone arm, with comparable success rates, 92 percent and 88 percent, respectively, even when major prognostic factors were considered. There were no serious adverse events or significant laboratory value changes attributable to the study drugs. Gatifloxacin as monotherapy (initially IV then orally until completion of treatment) was shown to be effective and safe, comparable to ceftriaxone IV alone or in combination with a macrolide (initially IV then orally until completion of treatment), in empirical therapy for community-acquired pneumonias, for patients that, at the physician s discretion, require initial treatment as inpatients.

The pathogenesis of Leishmania/HIV co-infection: cellular and immunological mechanisms.

The intracellular protozoan parasites of the genus Leishmania have been recognized as opportunistic pathogens in immunosuppressed individuals, including those infected with human immunodeficiency virus type-1 (HIV-1). Leishmaniasis and AIDS overlap in several sub-tropical and tropical regions around the world, including the Mediterranean area. In 1994, 3%-7% of HIV-1-infected individuals in southern Europe developed visceral leishmaniasis. In humans, interestingly, both HIV-1 and Leishmania interact with, invade, and multiply within cells of myeloid or lymphoid origin. The combined modulation of Leishmania - and HIV-1-related pathogenesis in the co-infected cases is therefore probably a realistic goal. In the light of the recent demonstration that L. donovani can up-regulate HIV-1 replication, both in monocytoid and lymphoid cells in vitro and in co-infected individuals, it is clear from the epidemiological data available that Leishmania can probably act as a powerful co-factor in the pathogenesis of HIV-1 infection. In those who are co-infected, complex mechanisms involving cytokine secretion and cellular-signalling events play pivotal roles in the Leishmania-mediated activation and pathogenesis of HIV-1. An overview of the recent findings concerning this Leishmania/HIV-1 interaction is presented here.

Evaluation of viral resistance to reverse transcriptase inhibitors (RTI) in HIV-1-infected patients before and after 6 months of single or double antiretroviral therapy.

We evaluated samples of peripheral blood mononuclear (PBMC) cells from 46 AIDS patients, before starting therapy with HIV-1 reverse transcriptase inhibitors (RTI), and after 6 months of drug use. PBMC were stored and tested by a Line Probe Assay (LiPA), in order to assess the frequency of RT mutations in this population. Six patients were taking AZT before initial blood collection (1 to 16 weeks of drug use) and 40 patients had no prior therapy. After baseline evaluation, 19 patients received AZT, 23 AZT plus DDI, 3 started AZT only with DDI added after 3 months, and 3 received a combination of AZT plus 3TC. Detection of at least one mutation was found in 33% (15/46) of patients at baseline, and 83% (38/46) had at least 1 mutation after 6 months of therapy. In the majority of cases, samples presented with the wild type and variants of HIV, simultaneously. Patients receiving monotherapy had a higher frequency of mutations (L41 and F214, Y215) than did patients receiving double-drug therapy (19 vs. 10). No specific mutation associated with DDI was identified in 26 patients so treated. Despite the finding of a mean increase in CD4 count and a mild decrease in viral load, patients tended to have an inverse correlation between the CD4 variation and number of mutations detected after 6 months, suggesting potential loss of drug efficacy in the presence of these genotypic changes.

Successful use of a defined antigen/GM-CSF adjuvant vaccine to treat mucosal Leishmaniasis refractory to antimony: A case report.

Immunotherapy has been proposed as a method to treat mucosal leishmaniasis for many years, but the approach has been hampered by poor definition and variability of antigens used, and results have been inconclusive. We report here a case of antimonial-refractory mucosal leishmaniasis in a 45 year old male who was treated with three single injections (one per month) with a cocktail of four Leishmania recombinant antigens selected after documented hypo-responsiveness of the patient to these antigens, plus 50 microg of GM-CSF as vaccine adjuvant. Three months after treatment, all lesions had resolved completely and the patient remains without relapse after two years. Side effects of the treatment included only moderate erythema and induration at the injection site after the second and third injections. We conclude that carefully selected microbial antigens and cytokine adjuvant can be successful as immunotherapy for patients with antimonial-refractory mucosal leishmaniasis.

Co-infection with HTLV-1 is associated with a shorter survival time for HIV-1-infected patients in Bahia, Brazil.

Co-infection with HTLV-1 reaches 20% among patients infected by HIV-1 in Bahia, Brazil. To evaluate its impact on survival, we conducted a retrospective, case-control study involving 198 patients (63 cases). Co-infection was associated with parenteral exposure (P = 0.0001) and female sex (P = 0.02). Co-infected patients had a shorter mean survival (1849 days) than controls (2430 days, P = 0.001), regardless of sex or baseline CD4 cell count. In Bahia, Brazil, co-infection with HIV-1 and HTLV-1 is associated with a shorter survival time.

Transcriptional complementarity in breast cancer: application to detection of circulating tumor cells.

BACKGROUND: We used a combination of genetic subtraction, silicon DNA microarray analysis, and quantitative PCR to identify tissue- and tumor-specific genes as diagnostic targets for breast cancer. METHODS AND RESULTS: From a large number of candidate antigens, several specific subsets of genes were identified that showed concordant and complementary expression profiles. Whereas transcriptional profiling of mammaglobin resulted in the detection of 70% of tumors in a panel of 46 primary and metastatic breast cancers, the inclusion of three additional markers resulted in detection of all 46 specimens. Immunomagnetic epithelial cell enrichment of circulating tumor cells from the peripheral blood of patients with metastatic breast cancer, coupled with RT-PCR-based amplification of breast tumor-specific transcripts, resulted in the detection of anchorage-independent tumor cells in the majority of patients with breast cancer with known metastatic disease. CONCLUSION: Complementation of mammaglobin with three additional genes in RT-PCR increases the detection of breast cancers in tissue and circulating tumor cells.

Serological expression cloning and immunological evaluation of MTB48, a novel Mycobacterium tuberculosis antigen.

Improved diagnostics are needed for the detection of Mycobacterium tuberculosis, especially for patients with smear-negative disease. To address this problem, we have screened M. tuberculosis (H37Rv and Erdman strains) genomic expression libraries with pooled sera from patients with extrapulmonary disease and with sera from patients with elevated reactivity with M. tuberculosis lysate. Both serum pools were reactive with clones expressing a recombinant protein referred to here as MTB48. The genomic sequence of the resulting clones was identical to that of the M. tuberculosis H37Rv isolate and showed 99% identity to the Mycobacterium bovis and M. bovis BCG isolate sequences. The genomic location of this sequence is 826 bp upstream of a region containing the esat-6 gene that is deleted in the M. bovis BCG isolate. The mtb48 1,380-bp open reading frame encodes a predicted 47.6-kDa polypeptide with no known function. Southern and Western blot analyses indicate that this sequence is present in a single copy and is conserved in the M. tuberculosis and M. bovis isolates tested but not in other mycobacterial species tested, including Mycobacterium leprae and Mycobacterium avium. In addition, the native protein was detected in the cytoplasm, as was a processed form that was also shed into the medium during culture. Serological analysis of recombinant MTB48 and the M. tuberculosis 38-kDa antigen with a panel of patient and control sera indicates that the inclusion of recombinant MTB48 in a prototype serodiagnostic test increases assay sensitivity for M. tuberculosis infection when it is combined with other known immunodominant antigens, such as the 38-kDa antigen.

Assessment of the serodiagnostic potential of nine novel proteins from Mycobacterium tuberculosis.

To identify antigens that would improve the accuracy of serological diagnosis of active tuberculosis, we cloned the genes encoding nine potentially immunogenic secreted or surface-associated proteins of Mycobacterium tuberculosis. Recombinant proteins were reacted with sera from HIV-negative individuals with extrapulmonary tuberculosis (EP-TB) or HIV-positive individuals with pulmonary tuberculosis (TBH). Specific and high level antibody responses were obtained for four recombinant proteins, of which antigen GST-822 was recognized by 60% of EP-TB and 42% of TBH and antigen MBP-506 was recognized by 45% of EP-TB and 61% of TBH. These results suggest that these proteins are strong candidates as subunits in a polyvalent serodiagnostic test.

Identification and characterization of prostein, a novel prostate-specific protein.

In this report, we describe the application of a systematic, genome-based approach to identify prostein, a novel prostate-specific protein expressed in normal and malignant prostate tissues. Characterization of the prostein gene shows that prostein cDNA encodes a 553-amino acid protein. The protein is predicted to be a type IIIa plasma membrane protein with a cleavable signal peptide and 11 transmembrane-spanning regions. The prostein gene is located on chromosome 1 at the WI-9641 locus between q32 and q42. Prostein mRNA is shown to be uniquely expressed in normal and cancerous prostate tissues using Northern blot, eDNA microarray, and real-time PCR analyses. Furthermore, prostein mRNA expression does not appear to be prostate tumor grade related and is restricted exclusively to prostate cell lines. Immunohistochemical staining using a mouse monoclonal antibody generated against prostein demonstrates that this protein is specifically detected in prostate tissues both at the plasma membrane and in the cytoplasm. Prostein expression is androgen responsive because treatment of LNCaP cells with androgen up-regulates prostein message and protein expression levels. These results validate prostein as a prostate-specific marker with potential utility in the diagnosis and treatment of prostate cancer.

Identification and characterization of T cell-stimulating antigens from Leishmania by CD4 T cell expression cloning.

Persistent immunity against Leishmania: infections in humans is mediated predominantly by CD4(+) T cells of the Th1 phenotype. Herein we report the expression cloning of eight Leishmania: Ags using parasite-specific T cell lines derived from an immune donor. The Ags identified by this technique include the flagellar proteins alpha- and beta-tubulin, histone H2b, ribosomal protein S4, malate dehydrogenase, and elongation factor 2, as well as two novel parasite proteins. None of these proteins have been previously reported as T cell-stimulating Ags from Leishmania: beta-tubulin-specific T cell clones generated against Leishmania: major amastigotes responded to Leishmania:-infected macrophages and dendritic cells. IFN-gamma enzyme-linked immunospot analysis demonstrated the presence of T cells specific for several of these Ags in PBMC from self-healing cutaneous leishmaniasis patients infected with either Leishmania: tropica or L. major. The responses elicited by Leishmania: histone H2b were particularly striking in terms of frequency of histone-specific T cells in PBMC (1 T cell of 6000 PBMC) as well as the percentage of responding donors (86%, 6 of 7). Ags identified by T cells from immune donors might constitute potential vaccine candidates for leishmaniasis.

High prevalence of giardiasis and stronglyloidiasis among HIV-infected patients in Bahia, Brazil.

Diarrhea due to intestinal microbial infections is a frequent manifestation among HIV-infected patients. It has been postulated that HIV-infected patients may have special types of intestinal infections, and that immune activation from such parasites may affect the progression of HIV disease. To evaluate these associations, the frequency of infections was examined in HIV-infected patients in Bahia, Brazil. To determine the potential impact of the presence of intestinal parasitic infections on HIV disease progression, a retrospective study approach was used. The medical charts of 365 HIV-infected patients who had been treated at the AIDS Clinic of the Federal University of Bahia Hospital were reviewed, and the prevalence of parasites was compared with 5,243 HIV-negative patients who had attended the hospital during the same period of time. Among HIV-infected subjects, CD(4) count, RNA plasma viral load (VL), and number of eosinophils were compared according to their stool examination results. The overall prevalence of each parasite was similar for HIV-positive and HIV-negative patients. However, the prevalence of S. stercoralis (p<10(-7)) and G. lamblia (p=0.005) was greater for HIV-infected subjects. The mean CD(4) count and viral load of HIV patients in our clinic who had stool examinations was 350 cells +/- 340 and 4.4 +/- 1.4 log RNA viral load, respectively. In this patient group there was no clear association between the level of the absolute CD(4) count or the viral load and a specific parasitic infection. The presence of an intestinal parasitic infection was not associated with faster progression of the HIV disease among HIV-infected patients. We conclude that strongyloidiasis and giardiasis are more frequent in HIV-infected patients in Bahia, Brazil. If this association is due to immune dysregulation, as has been proposed elsewhere, it must occur in patients after only minor shifts in CD(4) count from normal levels, or as a result of immune dysfunction not represented by CD(4) count. These infections do not appear to alter the progression of HIV disease.

Evaluation of DPPD, a single recombinant Mycobacterium tuberculosis protein as an alternative antigen for the Mantoux test.

Although the tuberculin test has aided in the diagnosis of tuberculosis for more than 85 years, its interpretation is difficult particularly because sensitization with non-tuberculous mycobacteria leads to false positive tests. Using the guinea pig model of tuberculosis, we have recently described a recombinant antigen (DPPD) that could circumvent this problem. The DPPD gene is unique to the M. tuberculosis complex organisms and is absent in the organisms representative of all other members of the Mycobacterium genus. Moreover, DPPD induced strong DTH in 100% of the guinea pigs infected with M. tuberculosis and in none of the guinea pigs immunized with nine different species of Mycobacterium. Here we present results of a clinical investigation using DPPD. Mantoux test using both PPD and DPPD was initially performed in 26 patients with confirmed pulmonary tuberculosis and in 25 healthy PPD negative individuals. The results indicated that both PPD and DPPD elicited DTH in 24 out of the 26 patients. No DTH was observed in any of the PPD negative individuals. In addition, a small clinical trial was performed in a population of 270 clinically healthy and randomly selected individuals. DPPD produced a bimodal histogram of skin reaction size and PPD produced a skewed histogram. Because the DPPD gene is not present in non-tuberculous bacilli, these results suggest that this molecule can be an additional tool for a more specific diagnosis of tuberculosis.

A randomized, placebo-controlled trial of granulocyte-macrophage colony-stimulating factor and nucleoside analogue therapy in AIDS.

Preliminary preclinical and clinical data suggest that granulocyte-macrophage colony-stimulating factor (GM-CSF) may decrease viral replication. Therefore, 105 individuals with AIDS who were receiving nucleoside analogue therapy were enrolled in a placebo-controlled, double-blind study and were randomized to receive either 125 microgram/m(2) of yeast-derived, GM-CSF (sargramostim) or placebo subcutaneously twice weekly for 6 months. Subjects were evaluated for toxicity and disease progression. A significant decrease in mean virus load (VL) was observed for the GM-CSF treatment group at 6 months (-0.07 log(10) vs. -0.60 log(10); P=.02). More subjects achieved human immunodeficiency virus (HIV)-RNA levels <500 copies/mL at >/=2 evaluations (2% on placebo vs. 11% on GM-CSF; P=.04). Genotypic analysis of 46 subjects demonstrated a lower frequency of zidovudine-resistant mutations among those receiving GM-CSF (80% vs. 50%; P=.04). No difference was observed in the incidence of opportunistic infections (OIs) through 6 months or survival, despite a higher risk for OI among GM-CSF recipients. GM-CSF reduced VL and limited the evolution of zidovudine-resistant genotypes, potentially providing adjunctive therapy in HIV disease.

Molecular and immunological characterization of Mycobacterium tuberculosis CFP-10, an immunodiagnostic antigen missing in Mycobacterium bovis BCG.

In order to identify antigens that may be used in the serodiagnosis of active tuberculosis (TB), we screened a Mycobacterium tuberculosis genomic expression library with a pool of sera from patients diagnosed with active pulmonary TB. The sera used lacked reactivity with a recombinant form of the M. tuberculosis 38-kDa antigen (r38kDa), and the goal was to identify antigens that might complement r38kDa in a serodiagnostic assay. Utilizing this strategy, we identified a gene, previously designated lhp, which encodes a 100-amino-acid protein referred to as culture filtrate protein 10 (CFP-10). The lhp gene is located directly upstream of esat-6, within a region missing in M. bovis BCG. Immunoblot analysis demonstrated that CFP-10 is present in M. tuberculosis CFP, indicating that it is likely a secreted or shed antigen. Purified recombinant CFP-10 (rCFP-10) was shown to be capable of detecting specific antibody in a percentage of TB patients that lack reactivity with r38kDa, most notably in smear-negative cases, where sensitivity was increased from 21% for r38kDa alone to 40% with the inclusion of rCFP-10. In smear-positive patient sera, sensitivity was increased from 49% for r38kDa alone to 58% with the inclusion of rCFP-10. In addition, rCFP-10 was shown to be a potent T-cell antigen, eliciting proliferative responses and gamma interferon production from peripheral blood mononuclear cells in 70% of purified protein derivative-positive individuals without evident disease. The responses to this antigen argue for the inclusion of rCFP-10 in a polyvalent serodiagnostic test for detection of active TB infection. rCFP-10 could also contribute to the development of a recombinant T-cell diagnostic test capable of detecting exposure to M. tuberculosis.

HTLV-I associated myelopathy: clinical and epidemiological profile.

The prevalence of HTLV-I reaches 1.8% among blood donors in Salvador, and 40% among chronic myelopathy patients in the state of Bahia, Brazil. The present study shows the epidemiological and clinical picture of patients attending the HAM/TSP Outpatient Unit at the Foundation of Neurology and Neorusurgery (FNN). 114 patients had epidemiologic data collected and 51 of these patients, who had regularly attended the HAM/TSP Unit for at least 1 year, were evaluated for signs, symptoms and disease progression. Most of the 114 patients were female (70%), of African descent, and with a mean age of 51. Sexually transmitted diseases and blood transfusion were the most common risk factors. Paraparesis with spasticity was the predominant sign (85%), bladder dysfunction occurred in 75%, intestinal dysfunction was recorded in 48%. Sensory examination was normal in 50% of the cases studied. The patients' functional status, as measured by the Kurtzke Disability Scale, during the 1 year observation period changed only in early disease. Steroid therapy with prednisone was the most commonly used treatment in this group.