Lymphatic filariasis in the Caribbean region: the opportunity for its elimination and certification.

In order to support the case for a certification of elimination of lymphatic filariasis (LF) in some Caribbean countries, we compared the prevalence of circulating Wuchereria bancrofti antigen in communities in Guyana, Suriname, and Trinidad. For the study, we assayed school children in six communities in Guyana, five communities in Suriname, and three communities in Trinidad for the prevalence of circulating W. bancrofti antigen, using a new immunochromatographic test for LF. We also assayed adults in these three countries, with a special focus on Blanchisseuse, Trinidad, where mass treatment for LF elimination had been carried out in 1981. The prevalences of W. bancrofti circulating antigen found in the school children populations ranged from 1.7% to 33.2% in Guyana and were 0.22% overall in Suriname and 0.0% in Trinidad. Among adults in two Guyana communities the prevalences were 16.7% and 32.1%. The results were all negative from 211 adults in communities in the north, center, and south of Trinidad, as well as from 29 adults in Suriname. The data suggest that contrary to reports of LF endemicity from the World Health Organization, LF may no longer be present in Trinidad and may be of very low prevalence in Suriname. Trinidad and Tobago and other Caribbean nations proven negative could seek to be awarded a certificate of LF elimination. In Suriname the small localized pocket of infected persons who may serve as a reservoir of LF infection could be tested and appropriately treated to achieve LF elimination. Such LF-positive countries as Guyana should access new international resources being made available for LF elimination efforts. An adequate certification program would help identify which countries should seek the new LF elimination resources.     

The mycobactericidal efficacy of ortho-phthalaldehyde and the comparative resistances of Mycobacterium bovis, Mycobacterium terrae, and Mycobacterium chelonae.

OBJECTIVES: To assess the mycobactericidal efficacy of an agent relatively new to disinfection, ortho-phthalaldehyde (OPA) and to compare the resistances of three Mycobacterium species. Mycobacterium bovis (strain BCG) was compared with Mycobacterium chelonae and Mycobacterium terrae to investigate the feasibility of using either of the latter two species in tuberculocidal testing. M. chelonae (a rapid grower) and M. terrae (an intermediate grower) both grow faster and are less virulent than M. bovis (a slow grower). DESIGN: The quantitative suspension protocol specified by the Environmental Protection Agency (EPA), the Tuberculocidal Activity Test Method (EPA test), was used throughout this study. Standard suspensions of all three species were prepared in a similar manner. Two suspensions of M. bovis, created in different laboratories, were used. These were tested against two concentrations of alkaline glutaraldehyde to provide reference data. Two concentrations of OPA were evaluated against all mycobacterial test suspensions. Four replicates of each organism-disinfectant combination were performed. RESULTS: Results were assessed by analysis of variance. M. terrae was significantly more resistant to 0.05% OPA than either M. bovis or M. chelonae. At 0.21% OPA, M. terrae was slightly more susceptible than one test suspension of M. bovis, but not significantly different from the other. M. chelonae was significantly less resistant than the other species at both OPA concentrations. At their respective minimum effective concentration, OPA achieved a 6-log10 reduction of M. bovis in nearly one sixth the time required by glutaraldehyde (5.5 minutes vs. 32 minutes). CONCLUSIONS: These data, along with other recent studies, lend support to the idea that M. terrae may be a suitable test organism for use in the tuberculocidal efficacy testing of disinfectants. They also confirm the relatively rapid tuberculocidal activity of OPA.     

Transmission of Mycobacterium tuberculosis depending on the age and sex of source cases.

This study estimated to what extent tuberculosis transmission in the Netherlands depends on the age and sex of source cases. DNA fingerprints of Mycobacterium tuberculosis isolates were matched to patient information in the Netherlands Tuberculosis Register for 1993-1998. Clusters were defined as groups of patients with pulmonary tuberculosis whose isolates had identical DNA fingerprints. Source cases were assigned by using two models. The first-case model assumed that the first diagnosed case was the source case. The incidence rate model estimated source case probabilities from the incidence rates of potential source cases and the time of diagnosis. DNA fingerprints of 6,102 isolates were matched to patient information on 5,080 (83%) cases, 3,479 of whom had pulmonary disease. According to both models, the number of infectious cases generated per source case was lower for female than for male source cases and decreased with increasing age of the source case. The authors concluded that transmission of tuberculosis is associated with the age and sex of source cases as well as the age of secondary cases. Increased transmission among immigrant groups in the Netherlands is largely attributable to the relatively young age of immigrant source cases.     

Potential of Mycobacterium vanbaalenii as a model organism to study drug transporters of Mycobacterium tuberculosis, Mycobacterium marinum and Mycobacterium ulcerans: homology analysis of M. tuberculosis drug transporters among mycobacterial species

Drug efflux pumps have been one of the important mechanisms of drug resistance in Mycobacterium tuberculosis. There is a prerequisite to study the behavior and mechanisms of these drug efflux pumps in detail for being considered in future anti-TB drug designing. The need of a rapid grower non-pathogenic mycobacterium with significant genomic homology for such type of studies is often being felt. During microarray and Real-Time PCR analysis of drug efflux pump genes of M. tuberculosis, we found 10 genes to be over-expressed during stress induced by common anti-TB drugs. In the present study homology analysis of these genes was done in order to know its phylogenetic relationship among other bacteria/mycobacteria. It was found that amino acid sequences of 7 out of 10 genes were significantly (>40%) identical to a non-pathogenic rapid grower environmental mycobacterium, Mycobacterium vanbaalenii. The protein sequences of M. vanbaalenii share important sequence motifs with M. tuberculosis useful for drug efflux mechanism based study across species. Like Mycobacterium smegmatis, it can be used as a model organism to study drug efflux pumps of M. tuberculosis and also other pathogenic mycobacteria such as Mycobacterium ulcerans and Mycobacterium marinum.     

Large Sequence Polymorphisms of the Euro-American lineage of Mycobacterium tuberculosis: A phylogenetic reconstruction and evidence for convergent evolution in the DR locus

The Euro-American lineage of the Mycobacterium tuberculosis complex consists of 10 sublineages, each defined by a deletion of a large genomic region (RD, region of difference); by spoligotyping, that probes the polymorphism of the Direct Repeat (DR) locus, the Euro-American strains are classified into 5 lineages (T, Haarlem, LAM, S and X) and 34 sublineages, but the relationships between the RD-defined sublineages and the spoligotype groupings are largely unclear.By testing a global sample of 158 Euro-American strains, mutually exclusive deletions of RD115, RD122, RD174, RD182, RD183, RD193, RD219, RD726 or RD761 were found in 122 strains; deletion of RD724, typical of strains from Central Africa, was not found. The RD-defined sublineages, tested for katG463/gyrA95 polymorphism, belonged to Principal Genotypic Group (PGG) 2, with the exception of RD219 sublineage belonging to PGG3; the 36 strains with no deletion were of either PGG2 or 3. Based on these polymorphisms, a phylogenetic reconstruction of the Euro-American lineage, that integrates the previously reported phylogeny, is proposed.Although certain deletions were found to be associated to certain spoligotype lineages (i.e., deletion RD115 to T and LAM, RD174 to LAM, RD182 to Haarlem, RD219 to T), our analysis indicates a general lack of concordance between RD-defined sublineages and spoligotype groupings. Moreover, of the 42 spoligotypes detected among the study strains, sixteen were shared by strains belonging to different RD sublineages. IS6110-RFLP analysis of strains sharing spoligotypes confirmed a poor genetic relatedness between strains of different RD sublineages. These findings provide evidence for the occurrence of a high degree of homoplasy in the DR locus leading to convergent evolution to identical spoligotypes. The incongruence between Large Sequence Polymorphism and spoligotype polymorphism argues against the use of spoligotyping for establishing phylogenetic relationships within the Euro-American lineage.     

Genome sequence based, comparative analysis of the fluorescent amplified fragment length polymorphisms (FAFLP) of tubercle bacilli from seals provides molecular evidence for a new species within the Mycobacterium tuberculosis complex.

Tuberculosis in seals is caused by a member of the Mycobacterium tuberculosis complex referred to as the 'seal bacillus'. Fluorescent amplified-fragment length polymorphism (FAFLP) analysis was applied to isolates from four Australian and six Argentinean seals and compared with FAFLP pattern for standard strains belonging to the M. tuberculosis complex. The FAFLP profiles derived from EcoRI/MseI restricted fragments of blind coded DNA samples differentiated the seal bacillus from other members of the M. tuberculosis complex. According to the phylogenetic analysis performed using FAFLP data, seal bacilli appear to have diverged significantly from other members of the M. tuberculosis complex. We describe the suitability of a panel of 19 highly polymorphic markers for rapid identification and comparative genomic analyses of the seal bacillus strains. It is likely that these bacilli got separated from the M. tuberculosis lineage as a result of different insertion deletion events occurring on a genome wide scale. Our analysis reveals that the seal bacillus and M. bovis are genetically related and therefore, might have originated from a common ancestor. Our data additionally support the hypothesis that seal bacillus occupies a unique taxonomic position within the M. tuberculosis complex.     

Subdivision of Mycobacterium tuberculosis into five variants for epidemiological purposes: methods and nomenclature.

Virulent strains of Mycobacterium tuberculosis isolated from humans are divisible into five variants by using four tests: oxygen requirement (aerobic or microaerophilic), nitrate reductase activity, susceptibility to pyrazinamide (60 micrograms/ml) and susceptibility to thiophene-2-carboxylic acid hydrazide (5 micrograms/ml). The five variants are referred to as Classical human, Asian human, bovine, African I and African II. The relation of these variants to previously described types is discussed. This simple division has been shown to be useful in epidemiological studies.     

DNA fngerprinting of Mycobacterium tuberculosis: lessons learned and implications for the future

DNA fingerprinting of Mycobacterium tuberculosis--a relatively new laboratory technique--offers promise as a powerful aid in the prevention and control of tuberculosis (TB). Established in 1996 by the Centers for Disease Control and Prevention (CDC), the National Tuberculosis Genotyping and Surveillance Network was a 5-year prospective, population-based study of DNA fingerprinting conducted from 1996 to 2000. The data from this study suggest multiple molecular epidemiologic and program management uses for DNA fingerprinting in TB public health practice. From these data, we also gain a clearer understanding of the overall diversity of M. tuberculosis strains as well as the presence of endemic strains in the United States. We summarize the key findings and the impact that DNA fingerprinting may have on future approaches to TB control. Although challenges and limitations to the use of DNA fingerprinting exist, the widespread implementation of the technique into routine TB prevention and control practices appears scientifically justified.     

Measuring the quality of hospital tuberculosis services: a prospective study in four Zimbabwe hospitals.

OBJECTIVE: To show how the use of a prospective approach to measuring the quality of services for a specific diagnosis can generate useful information for improving the quality of services in environments with limited information technology and data. DESIGN: Tracer approach focusing on intensive treatment of tuberculosis in hospital. The study was conducted in Zimbabwe in 1999. Local tuberculosis management guidelines were first translated into explicit quality assessment criteria and a panel of public health experts assisted in weighting different factors (structural and process) of the criteria. Factor weightings were based on both local knowledge and experience, and potential contribution of a factor to the likelihood of a positive outcome. A total of 138 patients was recruited into the study cohort at admission and followed up to discharge. An assessment of what was done to and for the patient was made for the entire hospitalization episode using explicit criteria. Comparisons were made between actual and maximum performance scores. SETTING: The study was conducted at four regional referral hospitals. The hospitals serve at least six secondary hospitals, and several public and private primary care facilities. The hospitals have a dual role as they also provide secondary care to their immediate catchment population. RESULTS: Notable quality gaps are observed between actual and maximum quality levels in all four hospitals although the size of the gap differed significantly. Variation in the quality of services between the hospitals is explained by distinguishable differences in structural and process aspects of tuberculosis management. CONCLUSIONS: It is feasible to conduct prospective quality assessment in developing countries with minimal disruption of routine activities. The study also showed that prospective exploration of health care quality for a specific diagnosis can provide insights into hospital-level quality issues. Such information is useful for monitoring and improving the quality of hospital services in general.     

Evolution of policies on human resources for health: opportunities and constraints in four post-conflict and post-crisis settings

Background

Few studies look at policy making in the health sector in the aftermath of a conflict or crisis and even fewer specifically focus on Human Resources for Health, which is a critical domain for health sector performance. The main objective of the article is to shed light on the patterns and drivers of post-conflict policy-making. In particular, we explore whether the post -conflict period offers increased chances for the opening of ‘windows for opportunity’ for change and reform and the potential to reset health systems.

Methods

This article uses a comparative policy analysis framework. It is based on qualitative data, collected using three main tools - stakeholder mapping, key informant interviews and document reviews - in Uganda, Sierra Leone, Cambodia and Zimbabwe.

Results

We found that HRH challenges were widely shared across the four cases in the post-conflict period but that the policy trajectories were different – driven by the nature of the conflicts but also the wider context. Our findings suggest that there is no formula for whether or when a ‘window of opportunity’ will arise which allows health systems to be reset. Problems are well understood in all four cases but core issues – such as adequate pay, effective distribution and HRH management – are to a greater or lesser degree unresolved. These problems are not confined to post-conflict settings, but underlying challenges to addressing them – including fiscal space, political consensus, willingness to pursue public objectives over private, and personal and institutional capacity to manage technical solutions – are liable to be even more acute in these settings. The role of the MoH emerged as weaker than expected, while the shift from donor dependence was clearly not linear and can take a considerable time.

Conclusions

Windows of opportunity for change and reform can occur but are by no means guaranteed by a crisis – rather they depend on a constellation of leadership, financing, and capacity. Recognition of urgency is certainly a facilitator but not sufficient alone. Post-conflict environments face particularly severe challenges to evidence-based policy making and policy implementation, which also constrain their ability to effectively use the windows which are presented.

     

WHO cooperative studies on the phage-typing of mycobacteria. 1. Phage lysis of Czechoslovak and Italian strains of Mycobacterium tuberculosis

Although phage-typing of some pathogenic microbes—e.g., Escherichia coli and the genera Salmonella, Staphylococcus, and Streptomyces—is in routine use, little progress has yet been made in the phage-typing of mycobacteria, chiefly because mycobacteriophages possess a polyvalence that prevents the reliable identification of individual species of mycobacterium. Furthermore, laboratories cannot compare their results because they employ different methods and culture media and also because the phages used cannot be accurately identified. It has been shown that internationally reproducible results can be obtained for M. tuberculosis by selecting suitable mycobacteriophages and sensitive culture media, and by using a standardized technique of phage-typing. Under these conditions, phage lysis may be utilized to classify strains of M. tuberculosis.     

Phylogenetic analysis of the promoter region of the CD40L gene in primates and other mammals.

CD40L is a type II membrane protein comprised of 261 amino acids. CD40L plays a crucial role in the immune system where it is primarily expressed on activated T cells and triggers immunoglobulin class switching. The genetic disease X-linked hypergammaglobulinemia (HIGM1, XHIGM or XHIM) is caused by mutations in the CD40L gene. Individuals with HIGM1 are susceptible to recurrent infections to pathogens and a relationship has been shown to exist with malaria [Sabeti, P., Usen, S., Farhadian, S., Jallow, M., Doherty, T., Newport, M., Pinder, M., Ward, R., Kwiatkowski, D., 2002a. CD40L association with protection from severe malaria. Genes Immun. 3, 286-291]. In this paper, we phylogenetically examine the promoter region of CD40L in primates and other mammals via phylogenetic shadowing. This analysis revealed several regions of the CD40L promoter that were highly constrained and thereby inferred to be functional. These constrained regions confirmed many known regulatory sites. In addition, a novel, highly constrained upstream region was also identified which had an NF-AT recognition motif. These analyses also showed that the different mammal groups do not share an exactly similar set of promoter binding sites and taxon-specific promoters have evolved.     

A bacteriological survey of tuberculosis due to the human tubercle bacillus (Mycobacterium tuberculosis) in south-east England: 1984-91.

The occurrence and nature of bacteriologically confirmed tuberculosis due to Mycobacterium tuberculosis in South-East England in the period 1984-91 is reported and compared with the results of a study for 1977-83. Registered new cases reached a low of 1028 in 1988 but increased to 1252 in 1991. This appeared to be due to a halt in the previous decline in new cases of European patients, a small increase in the number of Indian subcontinent (ISC) patients and an increase in patients from Africa. A total of 122 patients, mostly of European ethnic origin, were known to be HIV positive. As in the 1977-83 study, disease in the ISC group affected younger patients than in the European group, tubercle bacilli were more frequently isolated from a non-pulmonary site in the ISC group (45%) than in the European group (19%) and there was a higher incidence of the South Indian variant of M. tuberculosis in the former group (17%) than in the latter (9%). The overall incidence of drug resistance has not altered significantly since the 1977-83 study but 46 strains resistant to 3 or more drugs were isolated from 4099 ISC patients, compared with 3 of 4594 strains from European patients. Six of the 122 isolates from HIV positive patients were drug-resistant but none was multi-drug resistant. The slight rise in registered bacteriologically proven cases of tuberculosis, the presence of multi-drug resistant strains, the occurrence of HIV-related tuberculosis and reports of the emergence of multi-drug-resistant HIV-related tuberculosis in other countries strongly indicate the need for continued careful surveillance.     

First-in-human trial of the post-exposure tuberculosis vaccine H56:IC31 in Mycobacterium tuberculosis infected and non-infected healthy adults

BackgroundH56:IC31 is a candidate tuberculosis vaccine comprising a fusion protein of Ag85B, ESAT-6 and Rv2660c, formulated in IC31 adjuvant. This first-in-human, open label phase I trial assessed the safety and immunogenicity of H56:IC31 in healthy adults without or with Mycobacterium tuberculosis (M.tb) infection.MethodsLow dose (15 μg H56 protein in 500 nmol IC31) or high dose (50 μg H56, 500 nmol IC31) vaccine was administered intramuscularly thrice, at 56-day intervals. Antigen-specific T cell responses were measured by intracellular cytokine staining and antibody responses by ELISA.ResultsOne hundred and twenty-six subjects were screened and 25 enrolled and vaccinated. No serious adverse events were reported. Nine subjects (36%) presented with transient cardiovascular adverse events. The H56:IC31 vaccine induced antigen-specific IgG responses and Th1 cytokine-expressing CD4⁺ T cells. M.tb-infected vaccinees had higher frequencies of H56-induced CD4⁺ T cells than uninfected vaccinees. Low dose vaccination induced more polyfunctional (IFN-γ⁺TNF-α⁺IL-2⁺) and higher frequencies of H56-specific CD4⁺ T cells compared with high dose vaccination. A striking increase in IFN-γ-only-expressing CD4⁺ T cells, displaying a CD45RA⁻CCR7⁻ effector memory phenotype, emerged after the second high-dose vaccination in M.tb-infected vaccinees. TNF-α⁺IL-2⁺ H56-specific memory CD4⁺ T cells were detected mostly after low-dose H56 vaccination in M.tb-infected vaccinees, and predominantly expressed a CD45RA⁻CCR7⁺ central memory phenotype. Our results support further clinical testing of H56:IC31.     

Update on the virulence factors of the obligate pathogen Mycobacterium tuberculosis and related tuberculosis-causing mycobacteria

Over the long course of evolution from a probable environmental reservoir, the pathogen that we know today as Mycobacterium tuberculosis has become fully capable of adapting to the life inside host cells by evading and modifying their responses to infection. Factors contributing to the success of this pathogen are numerous and thanks to a large body of work accumulated over the past decades, we are closer to understanding the remarkable complexity of tuberculosis pathogenesis. The unique type VII secretion systems and various complex lipids of the cell envelope have emerged as some of the most important and most studied factors in this regard. This review attempts to summarize recent findings on these and other virulence factors, while discussing their evolution in different closely related tuberculosis-causing bacteria as well, with the aim of exploring the processes which led M. tuberculosis to becoming one of the deadliest infections agents.