Benzylpenicillin plus an aminoglycoside versus meropenem in neutropenic lymphoma and leukaemia patients with a suspected bacterial infection: a randomized, controlled trial.

OBJECTIVES: In Norway, initial treatment of febrile neutropenia (FN) has traditionally been benzylpenicillin plus an aminoglycoside. Internationally, FN is often treated with a broad-spectrum ß-lactam antibiotic. We aimed to compare these two regimens in a prospective, randomized, trial in patients with lymphoma or leukaemia with an expected period of neutropenia ≥7 days, and a suspected bacterial infection. METHODS: Adult neutropenic patients with lymphoma or leukaemia, and a suspected bacterial infection, were randomized for treatment with benzylpenicillin plus an aminoglycoside or meropenem. The primary endpoint was clinical success, defined as no modification of antibiotics and clinical stability 72 h after randomization. RESULTS: Among 322 randomized patients, 297 proved evaluable for analyses. Fifty-nine per cent (95% CI 51%-66%), (87/148) of the patients given benzylpenicillin plus an aminoglycoside were clinically stable, and had no antibiotic modifications 72 h after randomization, compared with 82% (95% CI 75%-87%), (122/149) of the patients given meropenem (p <0.001). When the antibiotic therapy was stopped, 24% (95% CI 18%-32%), (36/148) of the patients given benzylpenicillin plus an aminoglycoside, compared with 52% (95% CI 44%-60%), (78/149) of the patients given meropenem, had no modifications of their regimens (p <0.001). In the benzylpenicillin plus an aminoglycoside arm, the all-cause fatality within 30 days of randomization was 3.4% (95% CI 1.2%-7.9%), (5/148) of the patients, compared with 0% (95% CI 0.0%-3.0%), (0/149) of the patients in the meropenem arm (p 0.03). CONCLUSION: Clinical success was more common in FN patients randomized to meropenem compared with the patients randomized to benzylpenicillin plus an aminoglycoside. The all-cause fatality was higher among the patients given benzylpenicillin plus an aminoglycoside.

The mild inflammatory response in febrile neutropenic lymphoma patients with low risk of complications is more pronounced in patients receiving tobramycin once daily compared with three times daily.

We evaluated inflammatory markers in febrile neutropenic lymphoma patients undergoing high-dose chemotherapy with autologous stem cell support. Based on MASCC scores, our patients had a low risk of serious complications and a perspective of a benign initial clinical course of the febrile neutropenia. We also studied the impact of tobramycin given once versus three times daily on these immune markers. Sixty-one patients participating in a Norwegian multicentre prospective randomized clinical trial, comparing tobramycin once daily versus three times daily, given with penicillin G to febrile neutropenic patients, constituted a clinically homogenous group. Four patients had bacteraemia, all isolates being Gram-positive. Thirty-two patients received tobramycin once daily, and 29 patients received tobramycin three times daily. Blood samples were taken at the onset of febrile neutropenia and 1-2 days later. All samples were frozen at -70 °C and analysed at the end of the clinical trial for C-reactive protein (CRP), procalcitonin (PCT), complement activation products, mannose-binding lectin (MBL) and 17 cytokines. We found a mild proinflammatory response in this series of patients. CRP was non-specifically elevated. Ten patients with decreased MBL levels showed the same mild clinical and proinflammatory response. Patients receiving tobramycin once daily showed a more pronounced proinflammatory response compared with patients receiving tobramycin three times daily. Overall, febrile neutropenic cancer patients with a benign clinical course show a mild proinflammatory immune response.

Colonization by Candida in children with cancer, children with cystic fibrosis, and healthy controls.

A longitudinal, prospective study was conducted intermittently in Norway, from 1999 to 2008, to investigate the Candida colonization rates and species distributions in the tonsillopharyngeal and faecal flora in: (i) children with cancer; (ii) children with cystic fibrosis (CF); and (iii) healthy children. The effect of antibiotic treatment on Candida colonization was also studied, and we looked for changes in antifungal susceptibility over time within each child and between the different groups of children. In total, 566 tonsillopharyngeal swabs and 545 faecal samples were collected from 45 children with cancer, 37 children with CF, and 71 healthy, age-matched controls. The overall colonization rate with Candida was not significantly higher in the two groups of children undergoing extensive treatment with broad-spectrum antibiotics than in healthy controls. Approximately one-third of the cancer patients had a total lack of Candida colonization or had only one Candida-positive sample, despite multiple samples being taken, treatment with broad-spectrum antibiotics, long hospital stays, and periods with neutropenia. Children with CF had the highest prevalence of Candida albicans. Amoxycillin, azithromycin, third-generation cephalosporins and oral vancomycin resulted in a significantly increased Candida colonization rate. Phenoxymethylpenicillin, second-generation cephalosporins, metronidazole, trimethoprim-sulphamethoxazole, ciprofloxacin, penicillinase-resistant penicillins and inhaled tobramycin or colistin showed minimal effects on the Candida colonization rate. We found no evidence of development of antifungal resistance over time.

Molecular characteristics of pharyngeal and invasive emm3 Streptococcus pyogenes strains from Norway, 1988-2003.

A major virulence factor of group A streptococci (GAS) is the M protein. Strains with the M3 type are more often associated with necrotizing fasciitis (NF) and streptococcal toxic shock syndrome, and have a higher case fatality rate than strains of other M types. To better understand the epidemiology of M3 GAS strains in Norway, we analyzed 59 invasive and 69 pharyngeal isolates with respect to prophage content, allelic variation in emm3, mtsR encoding the metal transporter of Streptococcus repressor (mtsR), and sclB coding for streptococcal collagen-like protein B. The Norwegian emm3 strains were very homogeneous, mainly harboring the emm allele 3.1 and prophage profile PhiG3.01. Other prophage profiles were transient. The mutation in mtsR known to truncate the protein and result in decreased capacity to cause NF was not found in our isolates. The sclB gene usually harbored five or eight contiguous repeats of a CAAAA pentanucleotide sequence and a highly modular and variable collagen structural motif (CSM) region with 9 and 12 amino acid M3-specific conserved motif repeats distributed across the entire CSM region. Strains with 5 CAAAA repeats emerged in 1993 and these strains were associated with the increase in invasive M3 cases in the period 1993-2003.

Carriage of Streptococcus pneumoniae in healthy Norwegian children attending day-care centres.

An observational study to examine Streptococcus pneumoniae carriage in Norwegian children was initiated after two cases of pneumococcal meningitis, caused by the England(14)-9 clone, occurred in one day-care centre in Oslo. All children recruited from the day-care centre where the cases occurred were vaccinated with a seven-valent pneumococcal conjugate vaccine; the other participants who attended three other day-care centres nearby were not. The children were followed for 9 months, and three samplings took place. At the first visit, 45.7% of the children were colonised by pneumococci in the nasopharynx. The children harboured a variety of serotypes, with serotypes 6A, 23F, 6B and 19F being the most frequent. The numbers of children carrying vaccine serotypes decreased in both the vaccinated and the non-vaccinated groups. Thus, no significant effect of vaccine on carriage was detected in this relatively small study.

High case-fatality rates of meningococcal disease in Western Norway caused by serogroup C strains belonging to both sequence type (ST)-32 and ST-11 complexes, 1985-2002.

A total of 293 meningococcal disease (McD) patients from Western Norway hospitalized during 1985-2002 were examined for risk factors related to death. The case-fatality rate (CFR) increased from 4% during 1985-1993 to 17% during 1994-2002. We analysed the phenotypic and genotypic characteristics of the meningococcal patient isolates, with the aim of identifying whether highly virulent meningococcal strains contributed to the increased CFR. The Norwegian epidemic strain B:15:P1.7,16/ST-32 complex was overall the most common phenotype/genotype (n=75) and caused most deaths (n=9; CFR 12.0%). However, fatality was significantly associated with disease caused by serogroup C meningococcal strains; C:15:P1.7,16/ST-32 and C:2a/ST-11 complex strains, which had the highest CFRs of 21.1% and 18.2% respectively. Serogroup B strains of the ST-32 complex differing by serotype and/or serosubtype from the epidemic strain had a CFR of 5.1%, while the CFR of disease caused by other strains (all phenotypes and genotypes pooled) was 2.2%. The distribution of phenotypes/clonal complexes varied significantly between 1985-1993 and 1994-2002 (P<0.001); B:15/ST-32 complex strains decreased whereas both C:15:P1.7,16/ST-32 complex strains and strains with other phenotypes/clonal complexes increased. Our results indicate that C:15:P1.7,16/ST-32 and C:2a/ST-11 complex strains were highly virulent strains and contributed to the high CFR of McD in patients from Western Norway. To reduce fatality, rapid identification of such virulent strains is necessary. In addition, early and specific measures should include public information, vaccination of populations at risk of disease and carriage eradication, when clustering of patients occurs.

Macrolide-resistant Streptococcus pyogenes in Norway: population structure and resistance determinants.

A 2.7% prevalence of macrolide resistance in 1,657 Norwegian clinical Streptococcus pyogenes isolates was primarily due to erm(TR) (59%) and mef(A) (20%). Four clonal complexes comprised 75% of the strains. Macrolide resistance in S. pyogenes in Norway is imported as resistant strains or locally selected in internationally disseminated susceptible clones.

Molecular epidemiology of macrolide-resistant isolates of Streptococcus pneumoniae collected from blood and respiratory specimens in Norway.

Norway has a low prevalence of antimicrobial resistance, including macrolide-resistant Streptococcus pneumoniae (MRSP). In a nationwide surveillance program, a total of 2,200 S. pneumoniae isolates were collected from blood cultures and respiratory tract specimens. Macrolide resistance was detected in 2.7%. M-type macrolide resistance was found in 60% of resistant isolates, and these were mainly mef(A)-positive, serotype-14 invasive isolates. The erm(B)-encoded macrolide-lincosamide-streptogramin B (MLS(B)) type dominated among the noninvasive isolates. One strain had an A2058G mutation in the 23S rRNA gene. Coresistance to other antibiotics was seen in 96% of the MLS(B)-type isolates, whereas 92% of the M-type isolates were susceptible to other commonly used antimicrobial agents. Serotypes 14, 6B, and 19F accounted for 84% of the macrolide-resistant isolates, with serotype 14 alone accounting for 67% of the invasive isolates. A total of 29 different sequence types (STs) were detected by multilocus sequence typing. Twelve STs were previously reported international resistant clones, and 75% of the macrolide-resistant isolates had STs identical or closely related to these clones. Eleven isolates displayed 10 novel STs, and 7/11 of these "Norwegian strains" coexpressed MLS(B) and tetracycline resistance, indicating the presence of Tn1545. The invasive serotype-14 isolates were all classified as ST9 or single-locus variants of this clone. ST9 is a mef-positive M-type clone, commonly known as England(14)-9, reported from several European countries. These observations suggest that the import of major international MRSP clones and the local spread of Tn1545 are the major mechanisms involved in the evolution and dissemination of MRSP in Norway.

Systemic pneumococcal disease in Norway 1995-2001: capsular serotypes and antimicrobial resistance.

A total of 4624 pneumococcal isolates from episodes of systemic pneumococcal disease were received at the Norwegian Institute of Public Health during the period 1995-2001. All isolates were serotyped and tested for susceptibility to benzylpenicillin, lincomycin, erythromycin, tetracycline and trimethroprim sulphamethoxazole. The proportion of strains resistant to these antimicrobial agents remained stable at a low level, ranging from 0.1% for benzylpenicillin to 2.5% for erythromycin. The distribution of serotypes was also stable over the 7 years: serotypes 1, 4, 9, 14, 7, 6 and 23 were the most frequent, representing 70.5% of isolates. Overall, 95.8% of the isolates were of serotypes/groups included in the current 23-valent polysaccharide vaccine, 52.2% were of serotypes/groups included in the 7-valent conjugated vaccine and 85.5% were of serotypes/groups included in the 11-valent conjugated vaccine.

The four mouse IgG isotypes differ extensively in bactericidal and opsonophagocytic activity when reacting with the P1.16 epitope on the outer membrane PorA protein of Neisseria meningitidis.

Mouse monoclonal antibodies (MoAbs) of the four IgG isotypes, all specific for the P1.16 epitope on the meningcoccal PorA protein, were tested for functional activities. The avidities of the antibodies, measured by NH4SCN elution in enzyme-linked immunosorbent assay, showed similar values for all the MoAbs. The serum bactericidal activity (SBA) defined as the lowest concentration of antibodies giving 50% reduction in the number of meningococcal colony-forming units using human serum as complement, showed a hierarchy of IgG3 >> IgG2b > IgG2a >> IgG1. For the opsonophagocytosis (OP), the hierarchy was IgG3 > IgG2b = IgG2a >> IgG1. OP was measured in flow cytometry using log-phase live meningococci as target cells, normal human peripheral blood polymorphonuclear cells (PMNs) as effector cells and human serum as a complement source. The mouse MoAbs were negative in OP when using human PMNs in the absence of complement. The results demonstrate the importance of choosing the right isotype of mouse MoAbs when using them to judge the potential vaccine importance of their corresponding antigen. If such MoAbs should be used for passive vaccination against infectious diseases, the isotype would presumably play an important role for their anticipated clinical effects.

Comparison of functional immune responses in humans after intranasal and intramuscular immunisations with outer membrane vesicle vaccines against group B meningococcal disease.

A serogroup B meningococcal outer membrane vesicle (OMV) vaccine was delivered either intranasally or intramuscularly to 12 and 10 volunteers, respectively. The mucosal vaccine was given as four weekly doses followed by a fifth dose after 5 months; each dose consisted of OMVs equivalent to 250 microg of protein. The intramuscular (i.m.) vaccine, consisting of the same OMVs but adsorbed to Al(OH)(3), was administered as three doses each of 25 microg of protein, with 6 weeks interval between first and second doses and the third dose after 10 months. Both groups of vaccinees demonstrated significant immune responses when measured as specific IgG antibodies against live meningococci, as serum bactericidal activity (SBA) and as opsonophagocytic activity. Two weeks after the last dose, the anti-meningococcal IgG concentrations were significantly higher in the i.m. group (median IgG concentration: 43.1 microg/ml) than in the intranasal group (10.6 microg/ml) (P=0.001). The corresponding opsonophagocytic activity was 7.0 and 3.0 (median log(2) titre) (P=0.001), and the SBA was 5.0 and 2.0 (median log(2) titre) (P=0.005), for the i.m. and intranasal groups, respectively. The last immunisation induced an enhanced immune response in the i.m. group, whereas the intranasal group showed no significant booster response. Accordingly, affinity maturation of anti-OMV-specific IgG antibodies was seen only after i.m. vaccination. The IgG1 subclass dominated the responses in both groups, whereas the significant IgG3 responses observed in the i.m. group were absent in the intranasal group. Although the intranasal OMV vaccination schedule used here induced functional immune responses relevant to protection, an improved vaccine formulation and/or a modified mucosal immunisation regimen may be needed to achieve a systemic effect comparable to that seen after three doses of intramuscular vaccination.

Serum bactericidal activity correlates with the vaccine efficacy of outer membrane vesicle vaccines against Neisseria meningitidis serogroup B disease.

For evaluation of serum bactericidal activity (SBA) as surrogate for the efficacy of outer membrane vesicle (OMV) vaccines against Neisseria meningitidis serogroup B disease, we have reanalyzed data from a randomized double blind placebo-controlled efficacy trial involving 172000 secondary school students (aged 13-14 years) in Norway (1988-1991). A cohort of the efficacy trial consisting of 880 individuals was selected for immunogenicity studies. An efficacy of 87% was calculated for a 10-month observation period. However, after an observation period of 29 months, the estimated efficacy against group B disease induced by vaccination was 57%. The immunogenicity study showed that the SBA geometric mean titer (GMT) for the vaccinees was 2.4 before vaccination and 19.0 six weeks after the second vaccine dose. One year after vaccination the GMT was reduced to 2.8. A separate three-dose study with 304 adolescents showed that with a third dose at 10 months after the second dose (i.e. when cases of disease started to appear) a strong booster response was induced. Ten months after the second dose the SBA was reduced to near pre-immunization level. Following the third dose the SBA geometric mean titer of 2.7 increased to 62.3. One year after the third dose, the GMT was markedly higher than 6 weeks after the second dose (12.6 versus 8.8). Thus, protection after vaccination corresponds with the level of SBA. In order to reach lasting protective levels of SBA in a population, three vaccine doses are probably required. Measurements of SBA are likely to be useful for evaluating various upcoming formulations and improvements of immunization regimens for OMV vaccines.

Occurrence and clinical presentation of systemic pneumococcal infections in an unselected population in Oslo, Norway, between 1993 and 1997.

In order to describe the clinical and microbiological manifestations of systemic pneumococcal infection in an unselected urban population, 147 cases that occurred in the period 1993-1997 were retrospectively reviewed. An unexpected finding was that gastrointestinal symptoms were remarkably common. All pneumococcal isolates were fully susceptible to penicillin. The 7-valent conjugated vaccine covered 71% of those under 2 years of age, but only 21% of those 15-65 years of age were covered. Although the case fatality rate was 17%, the rate of early fatality due to systemic pneumococcal infection was unchanged compared with data published in the era before antibiotics. This study emphasizes the importance of continuing efforts to prevent systemic pneumococcal infections.

Molecular characteristics of Neisseria meningitidis strains isolated in Burkina Faso in 2001.

In the course of an epidemic of meningitis in Burkina Faso in 2001, 27 cerebrospinal fluid samples from patients in 7 districts were forwarded to Norway for isolation and characterization of the causative agents. Neisseria meningitidis was isolated from 13 (48%) samples. The isolates were analysed using serological and genetic methods. Of the 13 strains, 4 were serogroup A, serotype 21:P1.9, sequence type (ST)-5 and belonged to clonal subgroup III, while the remaining 9 strains were serogroup W135, serotype 2a:P1.5,2, ST-11 and belonged to the electrophoretic type-37 complex. PCR analyses revealed meningococcal DNA in 13/14 culture-negative samples. Sequence analysis of the PCR products demonstrated that at least 3 different meningococcal strains were responsible for these 13 cases. Our results show that the W135 strain associated with the 2000 hajj (Muslim pilgrimage) outbreak was a significant cause of disease in Burkina Faso in 2001. Further studies are warranted to determine whether W135 is about to replace serogroup A in sub-Saharan Africa.

Do HIV-seropositive patients become colonised with drug-resistant microorganisms?

The aim of the present study was to investigate whether HIV-infected patients, a group that is supposedly at risk for infection with antibiotic-resistant microbes, really does so, and to assess possible risk factors for acquiring these organisms. During the period from January 1998 to July 1999, samples of normal flora were obtained from 107 HIV-infected patients attending an outpatient clinic in Oslo, Norway. The samples were cultured for Streptococcus pneumoniae, Haemophilus influenzae, Escherichia coli, coagulase-negative staphylococci and Candida spp., and the resulting isolates were tested for antimicrobial susceptibility. The patients studied represented all stages of HIV infection, from recently infected to severely immunocompromised. Samples were taken at one, two or three time-points to determine whether antimicrobial resistance in colonising microorganisms increases over time. Antimicrobial resistance was linked primarily to antimicrobial prophylaxis, but it did not increase during the observation period. The level of a patient's immunodeficiency and the consequently intensified medical care was also of some importance. Even though about 50% of the patients were receiving antimicrobial agents at the time of sampling, the level of resistance found in these patients was very similar to that found in other patient groups in Norway; except for Candida albicans isolates, which were less susceptible to fluconazole. Overall, antimicrobial resistance was uncommon in the HIV-seropositive patients studied, a finding that is probably related to the overall low prevalence of antimicrobial resistance in the general population in Norway.

Antibiotic sensitivity still prevails in Norwegian blood culture isolates.

We describe the antimicrobial susceptibility of bacteraemia isolates from Norway. From March 1998 to February 1999, four university hospitals covering all parts of Norway collected their first 10 isolates each month. Minimal inhibitory concentrations were determined for: Enterobacteriaceae (n=192), staphylococci (n=89) and Streptococcus pneumoniae (n=69) using the Etest. NCCLS breakpoints were used. About 20% of all blood culture isolates in Norway in this period were investigated. Compared with countries outside Scandinavia antibiotic sensitivity still prevails. Only minor differences in resistance were found between participating hospitals, between hospital departments and between hospital- and community-acquired pathogens. The prudent use of antibiotics in Norway may contribute to the fact that antibiotic resistance still remains low in the most common bacterial pathogens causing bloodstream infections.

Meningococcal outer membrane vesicle vaccine given intranasally can induce immunological memory and booster responses without evidence of tolerance.

We have studied the ability of outer membrane vesicle (OMV) vaccines from Neisseria meningitidis serogroup B to induce vaccine-specific antibody and spleen cell proliferative responses in mice after being administered intranasally (i.n.) and/or subcutaneously (s.c.). A series of four weekly i.n. doses (25 microg) without adjuvant or a single s.c. dose (2.5 microg) with aluminum hydroxide was followed 2 months later by secondary i.n. or s.c. immunizations. After i.n. priming, both immunoglobulin G (IgG) antibody responses in serum, measured by enzyme-linked immunosorbent assay, and IgA antibodies in saliva and extracts of feces were significantly boosted by later i.n. immunizations. The IgG antibody responses in serum were also significantly augmented by secondary s.c. immunization after i.n. as well as s.c. priming. Sera from mice immunized i.n. reached the same level of bactericidal activity as after s.c. immunizations. The s.c. immunizations alone, however, had no effect on mucosal IgA antibody responses, but could prime for booster antibody responses in secretions to later i.n. immunizations. The i.n. immunizations also led to marked OMV-specific spleen cell proliferation in vitro. Both serum antibody responses and spleen cell proliferation were higher after i.n. priming and later s.c. immunizations than after s.c. immunizations alone. There was thus no evidence that i.n. priming had induced immunological tolerance within the B- or T-cell system. Our results indicate that a nonproliferating meningococcal OMV vaccine given i.n. can induce immunological memory and that it may be favorably combined with similar vaccines for injections.

Aminoethyl-isothiourea inhibits the increase in plasma endothelin-1 caused by serogroup A streptococci and prolongs survival in rat peritoneal sepsis.

To elucidate the possible roles of nitric oxide (NO), endothelin-1 (ET-1), and reactive oxygen species (ROS) in the pathophysiology of serogroup A streptococcal (GAS) peritoneal sepsis, we investigated the effects of aminoethylisothiourea (AE-ITU), an inducible NO synthase (iNOS) inhibitor, and a ROS scavenger, and the ET-1 receptor antagonist bosentan. In rats, live GAS inocula, 3 x 10(8) and 1 x 10(9) cfu/kg, entailed a 24-h mortality of 10% and 90%, respectively. GAS caused increases in tissue iNOS activity (9 h), in serum nitrite/nitrate (9-24 h), and in intracellular leukocyte ROS levels (3-6 h). These changes were all prevented by the pre-treatment with AE-ITU. A novel finding was that AE-ITU also prevented the GAS-induced marked increase in plasma ET-1 at 6 h. Short-term (7-h) survival was improved by both AE-ITU and by bosentan. The mechanism(s) for the beneficial effects of AE-ITU may possibly be a combined mode of action; iNOS inhibition, ROS scavenging, and inhibition of the increase in plasma ET-1 caused by GAS.