Analysis of two sequential SARS-CoV-2 outbreaks on a haematology-oncology ward and the role of infection prevention.

Two SARS-CoV-2 nosocomial outbreaks occurred on the haematology ward of our hospital. Patients on the ward were at high risk for severe infection because of their immunocompromised status. Whole Genome Sequencing proved transmission of a particular SARS-CoV-2 variant in each outbreak. The first outbreak (20 patients/31 healthcare workers (HCW)) occurred in November 2020 and was caused by a variant belonging to lineage B.1.221. At that time, there were still uncertainties on mode of transmission of SARS-CoV-2, and vaccines nor therapy were available. Despite HCW wearing II-R masks in all patient contacts and FFP-2 masks during aerosol generating procedures (AGP), the outbreak continued. Therefore, extra measures were introduced. Firstly, regular PCR-screening of asymptomatic patients and HCW; positive patients were isolated and positive HCW were excluded from work as a rule and they were only allowed to resume their work if a follow-up PCR CT-value was ≥30 and were asymptomatic or having only mild symptoms. Secondly, the use of FFP-2 masks was expanded to some long-lasting, close-contact, non-AGPs. After implementing these measures, the incidence of new cases declined gradually. Thirty-seven percent of patients died due to COVID-19. The second outbreak (10 patients/2 HCW) was caused by the highly transmissible omicron BA.1 variant and occurred in February 2022, where transmission occurred on shared rooms despite the extra infection control measures. It was controlled much faster, and the clinical impact was low as the majority of patients was vaccinated; no patients died and symptoms were relatively mild in both patients and HCW.

Controlling a human parainfluenza virus-3 outbreak in a haematology ward in a tertiary hospital: the importance of screening strategy and molecular diagnostics in relation to clinical symptoms.

BACKGROUND: The human parainfluenza virus 3 (HPIV-3) outbreak at the haemato-oncology ward of the Maastricht University Medical Centre in the summer of 2016. AIM: To describe an effective strategy to control the largest reported HPIV-3 outbreak at an adult haematology-oncology ward in the Netherlands by implementing infection control measures and molecular epidemiology investigation. METHODS: Clinical, patient and diagnostic data were both pro- and retrospectively collected. HPIV-3 real-time polymerase chain reaction (HPIV-3 RT-PCR) was validated using oropharyngeal rinse samples. Screening of all new and admitted patients was implemented to identify asymptomatic infection or prolonged shedding of HPIV-3 allowing cohort isolation. FINDINGS: The HPIV-3 outbreak occurred between 9 July and 28 September 2016 and affected 53 patients. HPIV-3 RT-PCR on oropharyngeal rinse samples demonstrated an up to 10-fold higher sensitivity compared with pharyngeal swabs. Monitoring showed that at first positive PCR, 20 patients (38%) were asymptomatic (of which 11 remained asymptomatic) and the average duration of shedding was 14 days (range 1-58). Asymptomatic patients had lower viral load, shorter period of viral shedding (≤14 days) and were mostly immune-competent oncology patients. The outbreak was under control five weeks after implementation of screening of asymptomatic patients. CONCLUSION: Implementation of a sensitive screening method identified both symptomatic and asymptomatic patients which had lower viral loads and allowed early cohort isolation. This is especially important in a ward that combines patients with varying immune status, because both immunocompromised and immune-competent patients are likely to spread the HPIV-3 virus, either through prolonged shedding or through asymptomatic course of disease.

Role of the environment in transmission of Gram-negative bacteria in two consecutive outbreaks in a haematology-oncology department.

In 2019-2020, two subsequent outbreaks caused by phenotypically identical ESBL-producing Enterobacter cloacae and multi-drug-resistant (MDR) Pseudomonas putida were detected in respectively 15 and 9 patients of the haematology-oncology department. Both bacterial species were resistant to piperacillin-tazobactam, used empirically in (neutropenic) sepsis in our hospital, and ciprofloxacin, used prophylactically in selective digestive decontamination for haematology patients. The E. cloacae outbreak was identified in clinical cultures of blood and urine. Despite intensified infection control measures, new cases were found in weekly point-prevalence screening cultures. Environmental samples of sinks and shower drains appeared positive in 18.1%. To diminish the environmental contamination burden, all siphons of sinks were replaced, and disinfection of sinks and shower drains was intensified using chlorine and soda on a daily basis. Replacement of shower drains was not possible. The outbreak of P. putida remained limited to rectal cultures only, and disappeared spontaneously without interventions. During both outbreaks, multiple strains of the incriminated bacterium were found simultaneously (demonstrated by Amplified-Fragment Length Polymorphism and/or Whole-Genome Multi-locus Sequencing Typing) in patients as well as the environment. It was experimentally shown that a biofilm on the toilet edge may act as a source for nosocomial transmission of Gram-negative bacteria. In conclusion, the drainage system of the hospital is an important reservoir of MDR bacteria, threatening the admitted patients. In existing hospitals, biofilms in the drainage systems cannot be removed. Therefore, it is important that in (re)building plans for hospitals a plan for prevention of nosocomial transmission from environment to patients is incorporated.

Despite Excellent Test Characteristics of the cobas 4800 CT/NG Assay, Detection of Oropharyngeal Chlamydia trachomatis and Neisseria gonorrhoeae Remains Challenging.

Oropharyngeal Chlamydia trachomatis (CT) infections and, especially, Neisseria gonorrhoeae (NG) infections are common, but few commercial nucleic acid amplification tests (NAATs) specify extragenital samples for intended use. The test characteristics of the cobas 4800 CT/NG assay were evaluated for oropharyngeal swabs. The technical validation included analysis of the specificity, sensitivity, dynamic range, linearity, efficiency, and precision. The probability of detection curve combined with historical data enabled the estimation of potentially missed diagnoses. A clinical evaluation was performed on a subset of 2,798 clinical samples available from routine diagnostics. Results of the cobas 4800 were compared with those from in-house C. trachomatis/N. gonorrhoeae PCR assays. Discrepant samples were tested with resolver assays, and these results were considered decisive. No cross-reactivity was seen in the analytical specificity analysis. High linearity (R2 ≥ 0.983), efficiency (89% to 99%), and precision (cycle threshold [CT ] value of 0.1 to 0.9) were seen for both C. trachomatis and N. gonorrhoeae The limit of detection in oropharyngeal samples was 3.2 × 102 inclusion-forming units (IFU)/ml for C. trachomatis and 6.7 × 102 CFU/ml for N. gonorrhoeae Estimates on potentially missed diagnoses were up to 7.2% for C. trachomatis and up to 24.7% for N. gonorrhoeae Clinical sensitivity and specificity were evaluated with 25 C. trachomatis-positive, 86 N. gonorrhoeae-positive, and 264 negative samples, resulting in 100% and 99.6% for C. trachomatis and 100% and 96.7% for N. gonorrhoeae, respectively. The findings in this study demonstrate the utility of the cobas 4800 CT/NG assay for oropharyngeal samples. Despite its being a highly accurate test, the range of reported CT values, especially for N. gonorrhoeae, suggests relatively low oropharyngeal loads. Hence, consistent detection over the full range of oropharyngeal loads could be impaired.

Multidisciplinary investigation of a multicountry outbreak of Salmonella Stanley infections associated with turkey meat in the European Union, August 2011 to January 2013.

Between August 2011 and January 2013, an outbreak of Salmonella enterica serovar Stanley (S. Stanley) infections affected 10 European Union (EU) countries, with a total of 710 cases recorded. Following an urgent inquiry in the Epidemic Intelligence Information System for food- and waterborne diseases (EPIS-FWD) on 29 June 2012, an international investigation was initiated including EU and national agencies for public health, veterinary health and food safety. Two of three local outbreak investigations undertaken by affected countries in 2012 identified turkey meat as a vehicle of infection. Furthermore, routine EU monitoring of animal sources showed that over 95% (n=298) of the 311 S. Stanley isolates reported from animal sampling in 2011 originated from the turkey food production chain. In 2004­10, none had this origin. Pulsed-field gel electrophoresis (PFGE) profile analysis of outbreak isolates and historical S. Stanley human isolates revealed that the outbreak isolates had a novel PFGE profile that emerged in Europe in 2011. An indistinguishable PFGE profile was identified in 346 of 464 human, food, feed, environmental and animal isolates from 16 EU countries: 102 of 112 non-human isolates tested were from the turkey production chain. On the basis of epidemiological and microbiological evidence, turkey meat was considered the primary source of human infection, following contamination early in the animal production chain.

Social integration of people with intellectual disability: insights from a social psychological research programme.

BACKGROUND: Social integration of people with intellectual disability (ID) moving into regular neighbourhoods tends to be studied and evaluated without detailed knowledge about the social psychological aspects of everyday interaction between neighbours with and without ID. The goal of the present paper is to show how the authors' social psychological research programme may contribute to this field of inquiry. METHODS: The different ways in which societies respond to features and behaviours that may be perceived as deviant are theoretically analysed. Results of empirical studies are reported to clarify how social responses to people with ID are special in terms of perceptions, emotions and interaction desires of people with and without ID during a pre-contact and contact phase. RESULTS: On the basis of the theoretical analysis, it is concluded that regular neighbouring in modern Western society often takes the form of benevolent tolerance, rather than stigmatisation and prejudice. However, empirical studies reveal that, prior to getting people with ID as new neighbours, prospective neighbours without ID experience a specific pattern of emotions that are associated with specific desires (e.g. with respect to information supply or a caring relationship). These anticipatory reactions are dependent on the expected size of the group moving in and on the severity of ID. Furthermore, while actually engaging in neighbouring, neighbours with and without ID appear to have experiences related to behaviour of residents, staff and features of housing facilities that are perceived as (in)congruent with regular neighbouring. CONCLUSIONS: It is concluded that interpersonal relationships between neighbours with and without ID should not be simplified in terms of attitudes that would be primarily prejudiced/stigmatising versus entirely accepting. Rather, our studies paint a more complex picture of sometimes ambivalent thoughts, feelings and interaction needs that all should be taken into account to make social integration a success.

The significance of neighbours: views and experiences of people with intellectual disability on neighbouring.

BACKGROUND: People with intellectual disability (ID) who live in regular neighbourhoods have experiences with their neighbours, which are important to understand when studying social integration. METHOD: This study describes and analyses the opinions on, and experiences with, neighbour relationships of 39 people with ID living in neighbourhood housing facilities. RESULTS: We found that, while the views of people with ID on 'good neighbouring' were consistent with 'neighbouring' described in sociological literature, their experiences may be influenced by an organisational context, the tendency to formalise relationships and apprehension towards meeting unfamiliar people. CONCLUSION: Understanding influential factors to neighbouring for people with ID may shed light on the processes involved in social integration of people with ID at a neighbourhood level. This paper contributes to understanding the opinions of people with ID on satisfactory neighbourhood relationships, and explores opportunities to improve them.

Incorporation of LpxL1, a detoxified lipopolysaccharide adjuvant, in influenza H5N1 virosomes increases vaccine immunogenicity.

The increasing number of human influenza H5N1 infections accentuates the need for the development of H5N1 vaccine candidates to prevent a potential influenza pandemic. The use of adjuvants in such vaccines can contribute significantly to antigen dose-sparing. In this study, we evaluated the capacity of the non-toxic Neisseria meningitidis lipopolysaccharide analog LpxL1 to function as an adjuvant for an influenza H5N1 virosomal vaccine. Inactivated influenza H5N1 virus (NIBRG-14) was used to construct virosomes (reconstituted virus envelopes) with LpxL1 incorporated in the virosomal membrane thus combining the influenza hemagglutinin (HA) antigen and the adjuvant in the same particle. Mice were immunized in a one- or two-dose immunization regimen with H5N1 virosomes with or without incorporated LpxL1. After a single immunization, H5N1 virosomes with incorporated LpxL1 induced significantly enhanced H5N1-specific total IgG titers as compared to non-adjuvanted virosomes but hemagglutination inhibition (HI) titers remained low. In the two-dose immunization regimen, LpxL1-modified H5N1 virosomes induced HI titers above 40 which were significantly higher than those obtained with non-adjuvanted virosomes. Incorporation of LpxL1 had little effect on virosome-induced IgG1 levels, but significantly increased IgG2a levels in both the one- and two-dose immunization regimen. Compared to non-adjuvanted virosomes, LpxL1-modified virosomes induced similar numbers of IFNgamma-producing T cells but decreased numbers of IL-4-producing T cells irrespective of the number of immunizations. We conclude that LpxL1 incorporated in H5N1 influenza virosomes has the capacity to function as a potent adjuvant particularly stimulating Th1-type immune reactions.

Enhanced decision support for policy makers using a web interface to health-economic models--illustrated with a cost-effectiveness analysis of nation-wide infant vaccination with the 7-valent pneumococcal conjugate vaccine in the Netherlands.

We have developed a web-based user-interface (web interface) to enhance the usefulness of health-economic evaluations to support decision making (http://pcv.healtheconomics.nl). It allows the user to interact with a health-economic model to evaluate predefined and customized scenarios and perform sensitivity analysis. To explore its usefulness, it was applied to an evaluation of cost-effectiveness of nation-wide infant vaccination with the 7-valent pneumococcal conjugate vaccine (PCV7), that was used to support a policy decision on the inclusion of PCV7 in the national vaccination program (NVP) of the Netherlands. We used a decision-tree analytic model to project the impact of infant vaccination with four doses of PCV7 on an annual cohort of infants born in the Netherlands. The base-case analysis includes the beneficial effects on unvaccinated individuals (herd protection). Additional scenarios varying the number of doses, discount rate for effects and the number of serotypes in the vaccine were evaluated and can be analysed on the web. Our model projects a base-case incremental cost-effectiveness ratio (iCER) of euro14,000 (95% uncertainty interval (UI): 9,800-20,200) per quality adjusted life year (QALY) or euro15,600 (95% UI: 11,100-23,900) per life year gained (LYG).

Economic evaluation of meningococcal serogroup C conjugate vaccination programmes in The Netherlands and its impact on decision-making.

The cost-effectiveness of one time vaccination of all persons aged 14 months to 18 years (catch-up programme) and of routine childhood immunisation at either ages 2 + 3 + 4 months, 5 + 6 months, or 14 months with a meningococcal C conjugate vaccine was estimated for The Netherlands, from a societal and a health care payer perspective. A decision analysis cohort model was employed (time horizon 77 years), direct and indirect costs (friction cost method) were considered and future costs and effects were discounted at 4%. The results showed that all vaccination options yield a substantial health gain and that the catch-up programme and routine vaccination at 14 months render favourable cost-effectiveness ratios: between about 13,200 and 17,700 per life year gained for the catch-up programme and between about 2200 and 2400 per life year gained for routine childhood vaccination at 14 months, depending on the perspective. In comparison to vaccination at 14 months, routine childhood vaccination during the first year of life is much less cost-effective: each additional life year gained costs approximately 147,000 (2 + 3 + 4 months) or 102,000 (5 + 6 months), from both perspectives. Additionally, inclusion of the likely herd immunity effect of the catch-up programme increases these incremental cost-effectiveness ratios. These results played a major role in the decision to add meningococcal C vaccination to the routine childhood immunisation schedule at 14 months and to implement a catch-up vaccination programme in The Netherlands in 2002.

PorA-specific differences in antibody avidity after vaccination with a hexavalent Men B outer membrane vesicle vaccine in toddlers and school children.

A clinical phase II trial with an experimental hexavalent outer membrane vesicle (OMV) vaccine (HexaMen) containing six different porin A (PorAs) was carried out in toddlers (2-3 years) and schoolchildren (7-8 years) in The Netherlands. HexaMen exists of two OMVs each containing three different PorA types. The serum bactericidal activity (SBA) after vaccination against the six PorAs was significantly different and was higher in toddlers than in schoolchildren. After vaccination the SBA against P1.5-2,10 was 4-6 times higher than against P1.7-2,4. The aim of this study was to test whether the differences in SBA could be explained by a difference in subtype-specific antibody avidity maturation. The avidity index (AI) of antibodies against three subtypes (PorA types P1.5-2,10; P1.12-1,13 and P1.7-2,4) was measured by ELISA and evaluated in relation to SBA. A significant avidity maturation for the 3 PorA subtypes was found. This maturation was most pronounced for P1.5-2,10 (mean AI = 72%), correlating with the highest SBA titres. Generally, the avidity titre correlated best with SBA. No differences in avidity indices against the three tested PorAs were found between toddlers and school children indicating that avidity maturation induced by this vaccine is not age-dependent.

Functional activity of antibodies against the recombinant OpaJ protein from Neisseria meningitidis.

The opacity proteins belong to the major outer membrane proteins of the pathogenic Neisseria and are involved in adhesion and invasion. We studied the functional activity of antibodies raised against the OpaJ protein from strain H44/76. Recombinant OpaJ protein was obtained from Escherichia coli in two different ways: cytoplasmic expression in the form of inclusion bodies followed by purification and refolding and cell surface expression followed by isolation of outer membrane complexes (OMCs). Immunization with purified protein and Quillaja saponin A (QuilA) induced high levels of Opa-specific antibodies, whereas the E. coli OMC preparations generally induced lower levels of antibodies. Two chimeric Opa proteins, hybrids between OpaB and OpaJ, were generated to demonstrate that the hypervariable region 2 is immunodominant. Denatured OpaJ with QuilA induced high levels of immunoglobulin G2a (IgG2a) in addition to IgG1, whereas refolded OpaJ with QuilA induced IgG1 exclusively. These sera did not induce significant complement-mediated killing. However, all sera blocked the interaction of OpaJ-expressing bacteria to CEACAM1-transfected cells. In addition, cross-reactive blocking of OpaB-expressing bacteria to both CEACAM1- and CEA-transfected cells was found for all sera. Sera raised against purified OpaJ and against OpaJ-containing meningococcal OMCs also blocked the nonopsonic interaction of Opa-expressing meningococci with human polymorphonuclear leukocytes.

Cross-reactivity of antibodies against PorA after vaccination with a meningococcal B outer membrane vesicle vaccine.

The cross-reactivity of PorA-specific antibodies induced by a monovalent P1.7-2,4 (MonoMen) and/or a hexavalent (HexaMen) meningococcal B outer membrane vesicle vaccine (OMV) in toddlers and school children was studied by serum bactericidal assays (SBA). First, isogenic vaccine strains and PorA-identical patient isolates were compared as a target in SBA, to ensure that the vaccine strains are representative for patient isolates. Geometric mean titers (GMTs) in SBA against patient isolates with subtypes P1.5-2,10 and P1.5-1,2-2 after vaccination with HexaMen were generally lower than those against vaccine strains with the same subtype, although the percentage of vaccine responders (> or =4-fold increase in SBA after vaccination) was not affected. Using various P1.7-2,4 patient isolates, GMTs as well as the number of vaccine responders were higher than for the P1.7-2,4 vaccine strain, indicating that the use of the P1.7-2,4 vaccine strain may have underestimated the immunogenicity of this subtype in HexaMen. Secondly, the cross-reactivity of antibodies induced by MonoMen and HexaMen was studied using several patient isolates that differed from the vaccine subtypes by having minor antigenic variants of one variable region (VR), by having a completely different VR or by having a different combination of VRs. MonoMen induced P1.4-specific antibodies that were cross-reactive with P1.4 variants P1.4-1 and P1.4-3. HexaMen induced a broader cross-reactive antibody response against various patient isolates with one VR identical to a vaccine subtype or a combination of VRs included in HexaMen. Cross-reactivity, measured by a fourfold increase in SBA after vaccination, against these strains ranged from 23 to 92% depending on the subtype of the tested strain and was directed against both VR1 and VR2. The extended cross-reactivity of vaccinee sera induced by HexaMen against antigenic variants has important favorable implications for meningococcal B OMV vaccine coverage.

Antibody avidity and immunoglobulin G isotype distribution following immunization with a monovalent meningococcal B outer membrane vesicle vaccine.

The avidity maturation and immunoglobulin G (IgG) isotype distribution of antibodies after vaccination with a meningococcal B outer membrane vesicle (OMV) vaccine were evaluated as indicators of protective immunity. Pre- and postvaccination sera from 134 healthy toddlers (ages, 2 to 3 years) immunized with a monovalent meningococcal B OMV (serosubtype P1.7-2,4) vaccine adsorbed with AlPO(4) or Al(OH)(3) were analyzed by enzyme-linked immunosorbent assay (ELISA) methods. The children were vaccinated three times with intervals of 3 to 6 weeks between vaccinations or twice with an interval of 6 to 10 weeks between vaccinations. A booster was given after 20 to 40 weeks. The avidity index (AI) of antibodies increased significantly during the primary series of vaccinations and after the booster was given. No differences in AIs were found when the results obtained with the two vaccination schedules or with the two adjuvants were compared. After vaccination, IgG1 was the predominant IgG isotype, followed by IgG3. No IgG2 or IgG4 was detected. There was a strong correlation between serum bactericidal activity (SBA) and ELISA titers (r = 0.85 [P < 0.0001] for total IgG, r = 0.83 for IgG1 [P < 0.0001], r = 0.82 for IgG3 [P < 0.0001], and r = 0.84 [P < 0.0001] for the avidity titer). When two subgroups with similar anti-OMV IgG levels were compared before and after the booster vaccination, the higher AI after the booster vaccination was associated with significantly increased SBA. We concluded that avidity maturation occurs after vaccination with a monovalent meningococcal B OMV vaccine, especially after boosting, as indicated by a significant increase in the AI. Vaccination with the monovalent OMV vaccine induced mainly IgG1 and IgG3 isotypes, which are considered to be most important for protection against meningococcal disease. An increase in the AI of antibodies is associated with increased SBA, independent of the level of specific IgG and the IgG isotype distribution. Measuring the AI and IgG isotype distribution of antibodies after vaccination can be a supplementary method for predicting protective immunity for evaluation in future phase III trials with meningococcal serogroup B vaccines.

In vivo expression of Neisseria meningitidis proteins homologous to the Haemophilus influenzae Hap and Hia autotransporters.

The genome sequences of Neisseria meningitidis serogroup B strain MC58 and serogroup A strain Z2491 were systematically searched for open reading frames (ORFs) encoding autotransporters. Eight ORFs were identified, six of which were present in both genomes, whereas two were specific for MC58. Among the identified ORFs was the gene encoding the known autotransporter IgA1 protease. The deduced amino acid sequences of the other identified ORFs were homologous to known autotransporters and found to contain an N-terminal signal sequence and a C-terminal domain that could constitute a beta-barrel in the outer membrane. The ORFs NMB1985 and NMB0992, encoding homologs of the Hap (for Haemophilus adhesion and penetration protein) and Hia (for Haemophilus influenzae adherence protein) autotransporters of H. influenzae, were cloned from serogroup B strain H44/76 and expressed in Escherichia coli. Western blots revealed that all sera of patients (n=14) and healthy carriers (n=3) tested contained antibodies against at least one of the recombinant proteins. These results indicate that both genes are widely distributed among N. meningitidis isolates and expressed during colonization and infection.

Serum bactericidal activity and isotype distribution of antibodies in toddlers and schoolchildren after vaccination with RIVM hexavalent PorA vesicle vaccine.

A clinical phase II trial with the RIVM hexavalent OMV vaccine containing six different PorAs was carried out in toddlers (2-3 years) and schoolchildren (7-8 years) in The Netherlands. Children were vaccinated three times (0, 2, 8 months). Sera after two and three vaccinations were analysed for serum bactericidal activity (SBA) and isotype distribution in whole cell enzyme linked immunosorbent assay (ELISA). The SBA after vaccination against the six PorAs was significantly different. We investigated whether the age specific and PorA specific differences in SBA titers correlated with differences in PorA specific IgG isotype distribution. The SBA titers were higher in toddlers compared with schoolchildren. After vaccination, IgG1 antibodies dominated the response followed by IgG3 antibodies. IgG2 levels were low, whereas IgG4 was not detected. Irrespective of PorA, IgG total and isotype specific titers after two and three vaccinations were significantly higher in toddlers than in schoolchildren. A weak correlation was found between IgG total or IgG1 and SBA. Although the immunogenicity of the six PorAs is very different, the isotype distribution was similar for all six tested PorAs. We conclude that the RIVM hexavalent PorA vesicle vaccine induces bactericidal antibodies mainly of the IgG1 and IgG3 isotypes that are considered to be most important for protection against disease. The isotype distribution of the response is not age-dependent.

Modification of lipid A biosynthesis in Neisseria meningitidis lpxL mutants: influence on lipopolysaccharide structure, toxicity, and adjuvant activity.

Two genes homologous to lpxL and lpxM from Escherichia coli and other gram-negative bacteria, which are involved in lipid A acyloxyacylation, were identified in Neisseria meningitidis strain H44/76 and insertionally inactivated. Analysis by tandem mass spectrometry showed that one of the resulting mutants, termed lpxL1, makes lipopolysaccharide (LPS) with penta- instead of hexa-acylated lipid A, in which the secondary lauroyl chain is specifically missing from the nonreducing end of the GlcN disaccharide. Insertional inactivation of the other (lpxL2) gene was not possible in wild-type strain H44/76 expressing full-length immunotype L3 lipopolysaccharide (LPS) but could be readily achieved in a galE mutant expressing a truncated oligosaccharide chain. Structural analysis of lpxL2 mutant lipid A showed a major tetra-acylated species lacking both secondary lauroyl chains and a minor penta-acylated species. The lpxL1 mutant LPS has retained adjuvant activity similar to wild-type meningococcal LPS when used for immunization of mice in combination with LPS-deficient outer membrane complexes from N. meningitidis but has reduced toxicity as measured in a tumor necrosis factor alpha induction assay with whole bacteria. In contrast, both adjuvant activity and toxicity of the lpxL2 mutant LPS are strongly reduced. As the combination of reduced toxicity and retained adjuvant activity has not been reported before for either lpxL or lpxM mutants from other bacterial species, our results demonstrate that modification of meningococcal lipid A biosynthesis can lead to novel LPS species more suitable for inclusion in human vaccines.