The morphological heterogeneity of lipopolysaccharides (LPSs) among salmonella mutants with different LPS chemotypes was analyzed in silver-stained polyacrylamide gels. The biochemical differences in the LPS chemotypes were reflected in the unique profiles of the purified LPSs. The LPS profiles in the whole-cell lysates were also unique for each chemotype. (Whole-cell lysates were assessed by a method which preferentially silver stains LPS and by a proteinase K digest of whole-cell lysates. The silver-stained LPS profiles of proteinase K-digested lysates were similar to the homologous purified LPS and could be used to preliminarily characterize the LPS chemotype before purification.) In summary, biochemical variation in LPS composition can be detected in silver-stained polyacrylamide gels.

Immunoglobulin M antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) is a rapid and versatile diagnostic method that readily permits the combination of multiple assays. Test consolidation is especially important for arthropod-borne viruses (arboviruses) which belong to at least three virus families: the Togaviridae,Flaviviridae, and Bunyaviridae. Using prototype viruses from each of these families and a panel of well-characterized human sera, we have evaluated and standardized a combined MAC-ELISA capable of identifying virus infections caused by members of each virus family. Furthermore, by grouping antigens geographically and utilizing known serological cross-reactivities, we have reduced the number of antigens necessary for testing, while maintaining adequate detection sensitivity. We have determined that a 1:400 serum dilution is most appropriate for screening antiviral antibody, using a positive-to-negative ratio of ≥2.0 as a positive cutoff value. With a blind-coded human serum panel, this combined MAC-ELISA was shown to have test sensitivity and specificity that correlated well with those of other serological techniques.

We report the development and evaluation of a Salmonella O-group-specific Bio-Plex assay to detect the six most common serogroups in the United States (B, C 1 , C 2 , D, E, and O13) plus serotype Paratyphi A. The assay is based on rfb gene targets directly involved in O-antigen biosynthesis; it can be completed 45 min post-PCR amplification. The assay correctly and specifically identified 362 of 384 (94.3%) isolates tested in comparison to traditional serotyping. Seventeen isolates (4.4%) produced results consistent with what is known about the molecular basis for serotypes but different from the results of traditional serotyping, and five isolates (1.3%) generated false-negative results. Molecular determination of the serogroup for rough isolates was consistent with a common serotype in most instances, indicating that this approach has the potential to provide O-group information for isolates that do not express an O antigen. We also report the sequence of the O-antigenencoding rfb gene cluster from Salmonella enterica serotype Poona (serogroup O13). Compared with other, previously characterized rfb regions, the O13 rfb gene cluster was most closely related to Escherichia coli O127 and O86. The O-group Bio-Plex assay described here provides an easy-to-use, high-throughput system for rapid detection of common Salmonella serogroups.Serotyping of salmonellae is a valuable phenotypic subtyping tool for understanding the epidemiology of this important food-borne pathogen. Salmonella isolates are serotyped using the Kauffmann-White scheme according to their O, H, and Vi antigens (4, 40). The O antigen contains multiple repeats of an oligosaccharide unit (O unit), which, together with lipid A and core oligosaccharides, form the lipopolysaccharide present in the outer membranes of gram-negative bacteria (7). Many of the genes required for O-antigen biosynthesis are organized in a large regulon termed the rfb gene cluster (47). rfb gene clusters have been characterized from a growing number of gram-negative bacteria; this operon is located between galF and gnd in Salmonella enterica and Escherichia coli (49). In general, rfb genes have a low GϩC content (usually less than 40%); the deviation in GϩC content from that of typical S. enterica genes (51%) suggests that rfb DNA originated in species other than S. enterica and was captured by lateral gene transfer (46, 56). Typically, three classes of genes are found in rfb clusters: (i) genes for synthesis of nucleotide sugars specific to the respective O antigen, (ii) sugar transferase genes to build the O subunit, and (iii) the O-antigen polymerase (wzy) and transport protein (wzx) genes for assembly of the O subunit into the O antigen (49).There are 46 O serogroups described in the KauffmannWhite scheme; serogroups were originally designated by alphabetic letters, and then it was necessary to continue with numbers 51 to 67. For consistency in the scheme, all serogroups were given a number designation; however, the most common serogroups (A to E) are commonly designated by ...

No abstract

The administration of an efficacious vaccine is the most effective long-term measure to control the oculogenital infections caused by Chlamydia trachomatis in humans. Chlamydia genome sequencing has identified a number of potential vaccine candidates, and the current challenge is to develop an effective delivery vehicle for induction of a high level of mucosal T and complementary B cell responses. Vibrio cholerae ghosts (VCG) are nontoxic, effective delivery vehicles with potent adjuvant properties, and are capable of inducing both T cell and Ab responses in mucosal tissues. We investigated the hypothesis that rVCG could serve as effective delivery vehicles for single or multiple subunit chlamydial vaccines to induce a high level of protective immunity. rVCG-expressing chlamydial outer membrane proteins were produced by a two-step genetic process, involving cloning of Omp genes in V. cholerae, followed by gene E-mediated lysis of the cells. The immunogenicity and vaccine efficacy of rVCG-expressing single and multiple subunits were compared. Immunologic analysis indicated that i.m. immunization of mice with either vaccine construct induced a strong mucosal and systemic specific Th1 response against the whole chlamydial organism. However, there was an immunogenic advantage associated with the multiple subunit vaccine that induced a higher frequency of Th1 cells and a relatively greater ability to confer protective immunity, compared with the single subunit construct. These results support the operational theory that the ability of a vaccine to confer protective immunity against Chlamydia is a function of the level of Th1 response elicited.

High maternal viral load increases the likelihood of perinatal transmission of HIV-1 in women without AIDS and advanced immunosuppression. HIV-1 infected pregnant women without advanced disease, shown by others to have the lowest risk of perinatal transmission, may benefit the most from efforts to identify and decrease viral load at delivery.

Background Malnutrition in advanced cancer patients is common but limited and inconclusive data exists on the effectiveness of nutrition interventions. Feasibility and acceptability of a novel family-based nutritional psychosocial intervention were established recently. The aims of this present study were to assess the feasibility of undertaking a randomised controlled trial of the latter intervention, to pilot test outcome measures and to explore preliminary outcomes. Methods Pilot randomised controlled trial recruiting advanced cancer patients and family caregivers in Australia and Hong Kong. Participants were randomised and assigned to one of two groups, either a family-centered nutritional intervention or the control group receiving usual care only. The intervention provided 2–3 h of direct dietitian contact time with patients and family members over a 4–6-week period. During the intervention, issues with nutrition impact symptoms and food or eating-related psychosocial concerns were addressed through nutrition counselling, with a focus on improving nutrition-related communication between the dyads and setting nutritional goals. Feasibility assessment included recruitment, consent rate, retention rate, and acceptability of assessment tools. Validated nutritional and quality of life self-reported measures were used to collect patient and caregiver outcome data, including the 3-day food diary, the Patient-Generated Subjective Global Assessment Short Form, the Functional Assessment Anorexia/Cachexia scale, Eating-related Distress or Enjoyment, and measures of self-efficacy, carers’ distress, anxiety and depression. Results Seventy-four patients and 54 family caregivers participated in the study. Recruitment was challenging, and for every patient agreeing to participate, 14–31 patients had to be screened. The consent rate was 44% in patients and 55% in caregivers. Only half the participants completed the trial’s final assessment. The data showed promise for some patient outcomes in the intervention group, particularly with improvements in eating-related distress (p = 0.046 in the Australian data; p = 0.07 in the Hong Kong data), eating-related enjoyment (p = 0.024, Hong Kong data) and quality of life (p = 0.045, Australian data). Energy and protein intake also increased in a clinically meaningful way. Caregiver data on eating-related distress, anxiety, depression and caregiving burden, however, showed little or no change. Conclusions Despite challenges with participant recruitment, the intervention demonstrates good potential to have positive effects on patients’ nutritional status and eating-related distress. The results of this trial warrant a larger and fully-powered trial to ascertain the effectiveness of this intervention. Trial registration The trial was registered with the Australian & New Zealand Clinical Trials Registry, registration number ACTRN12618001352291.

The qualitative NASBA RNA assay is highly specific and more sensitive than DNA PCR. Qualitative RNA assays may be useful for diagnosing and excluding perinatal HIV infection in children after the first week of life for such purposes as initiating antiretroviral therapy and other treatment, resolving parental uncertainty, determining timing of transmission, and providing endpoints for intervention trials.