Mycobacterium ulcerans toxin, mycolactone may enhance host-seeking and oviposition behaviour by Aedes aegypti (L.) (Diptera: Culicidae).

The ecological functions of many toxins continue to remain unknown for those produced by environmental pathogens. Mycobacterium ulcerans, the causative agent of the neglected tropical disease, Buruli ulcer, produces a cytotoxic macrolide, mycolactone, whose function(s) in the environment remains elusive. Through a series of dual-choice behaviour assays, they show that mycolactone may be an interkingdom cue for the yellow fever mosquito, Aedes aegypti, seeking blood-meals as well as oviposition sites. Results provide novel insight into the evolution between bacteria and potential vectors. While further studies are needed to determine if mycolactone is an actual signal rather than simply a cue, this discovery could serve as a model for determining roles for toxins produced by other environmental pathogens and provide opportunities for developing novel strategies for disease prevention. The relationship between M. ulcerans, mycolactone, and Ae. aegypti further suggests there could be an amplification effect for the spread of pathogens responsible for other diseases, such as yellow fever and dengue.

Calorie restriction and not glucagon-like peptide-1 explains the acute improvement in glucose control after gastric bypass in Type 2 diabetes.

AIMS: To compare directly the impact of glucagon-like peptide-1 secretion on glucose metabolism in individuals with Type 2 diabetes listed for Roux-en-Y gastric bypass surgery, randomized to be studied before and 7 days after undergoing Roux-en-Y gastric bypass or after following a very-low-calorie diet. METHODS: A semi-solid meal test was used to investigate glucose, insulin and glucagon-like peptide-1 response. Insulin secretion in response to intravenous glucose and arginine stimulus was measured. Hepatic and pancreatic fat content was quantified using magnetic resonance imaging. RESULTS: The decrease in fat mass was almost identical in the Roux-en-Y gastric bypass and the very-low-calorie diet groups (3.0±0.3 and 3.0±0.7kg). The early rise in plasma glucose level and in acute insulin secretion were greater after Roux-en-Y gastric bypass than after a very-low-calorie diet; however, the early rise in glucagon-like peptide-1 was disproportionately greater (sevenfold) after Roux-en-Y gastric bypass than after a very-low-calorie diet. This did not translate into a greater improvement in fasting glucose level or area under the curve for glucose. The reduction in liver fat was greater after Roux-en-Y gastric bypass (29.8±3.7 vs 18.6±4.0%) and the relationships between weight loss and reduction in liver fat differed between the Roux-en-Y gastric bypass group and the very-low-calorie diet group. CONCLUSIONS: This study shows that gastroenterostomy increases the rate of nutrient absorption, bringing about a commensurately rapid rise in insulin level; however, there was no association with the large post-meal rise in glucagon-like peptide-1, and post-meal glucose homeostasis was similar in the Roux-en-Y gastric bypass and very-low-calorie diet groups. (Clinical trials registry number: ISRCTN11969319.).

Reversal of Type 2 diabetes after bariatric surgery is determined by the degree of achieved weight loss in both short- and long-duration diabetes.

AIM: To define the impact of duration of diabetes and extent of weight loss on the reversibility of Type 2 diabetes after bariatric surgery. METHODS: Complete data were collated from 89 individuals with Type 2 diabetes mellitus undergoing any bariatric surgical procedure in a specialist bariatric centre. People with a preoperative HbA1c < 43 mmol/mol (6.1%) were excluded. Diabetes duration was defined as: short, < 4 years; medium, 4-8 years; and long, > 8 years. RESULTS: An HbA1c of <43 mmol/mol (6.1%) was achieved by 62% of patients in the short-duration group and 26% of patients in the long-duration group. Normoglycaemia was rarely achieved in the long-duration group if weight loss was < 25 kg. In the whole cohort there was a clear relationship of greater weight loss with lower HbA1c levels (Rs = -0.53; P < 0.0001). CONCLUSIONS: The study shows that the degree of achieved weight loss is the major determinant of return to normal blood glucose levels after bariatric surgery. Normoglycaemia can be achieved in long-duration Type 2 diabetes, but a greater degree of weight loss is required than for short-duration diabetes.

Laparoscopic adjustable gastric band survival in a high-volume bariatric unit.

BACKGROUND: Although laparoscopic adjustable gastric bands (LAGBs) have been shown to be efficacious, their long-term usefulness has been questioned. This study examined the fate of LAGBs in a unit with over a decade of experience in their use. Patient factors related to the need for, and timing of, band removal were investigated. METHODS: A prospectively maintained database was used to identify all patients with a LAGB. Patient demographics, need for band removal and band survival were examined. Logistic regression modelling was done and Kaplan-Meier curves were calculated for band survival. RESULTS: Between 2000 and 2012, 674 bands were placed in 665 patients. Of these, 143 (21.2 per cent) were removed. There was no difference in rates of removal by sex (P = 0.910). The highest rates of removal were in patients aged less than 40 years (26.7 per cent), and those with a BMI greater than 60 kg/m2 (28.6 per cent). Earlier band removal was seen in younger patients (P = 0.002). Rates of removal increased linearly by earlier year of placement. Of bands placed 4 or more years previously, 35.0 per cent required removal. Eighty-three patients (58.0 per cent) who had a LAGB removed went on to have a further bariatric procedure (band to bypass, 66; band to sleeve, 17). CONCLUSION: Even in experienced hands LAGB does not appear to be a definitive solution. In a large number of patients there appears to be a finite 'band life', with the majority of patients requiring conversion to a further bariatric procedure.

The intrinsic transmission dynamics of tuberculosis epidemics.

In developed countries the major tuberculosis epidemics declined long before the disease became curable in the 1940s. We present a theoretical framework for assessing the intrinsic transmission dynamics of tuberculosis. We demonstrate that it takes one to several hundred years for a tuberculosis epidemic to rise, fall and reach a stable endemic level. Our results suggest that some of the decline of tuberculosis is simply due to the natural behaviour of an epidemic. Although other factors must also have contributed to the decline, these causal factors were constrained to operate within the slow response time dictated by the intrinsic dynamics.

Mycobacterium liflandii outbreak in a research colony of Xenopus (Silurana) tropicalis frogs.

A research colony of Xenopus (Silurana) tropicalis frogs presented with nodular and ulcerative skin lesions. Additional consistent gross findings included splenomegaly with multiple tan-yellow nodular foci in the spleen and liver of diseased frogs. Copious acid-fast positive bacteria were present in touch impression smears of spleen, skin, and livers of diseased frogs. Histologically, necrotizing and granulomatous dermatitis, splenitis, and hepatitis with numerous acid-fast bacilli were consistently present, indicative of systemic mycobacteriosis. Infrequently, granulomatous inflammation was noted in the lungs, pancreas, coelomic membranes, and rarely reproductive organs. Ultrastructurally, both extracellular bacilli and intracellular bacilli within macrophages were identified. Frogs in the affected room were systematically depopulated, and control measures were initiated. Cultured mycobacteria from affected organs were identified and genetically characterized as Mycobacterium liflandii by polymerase chain reaction amplification of the enoyl reductase domain and specific variable numbers of tandem repeats. In recent years, M. liflandii has had a devastating impact on research frog colonies throughout the United States. This detailed report with ultrastructural description of M. liflandii aids in further understanding of this serious disease in frogs.

Phylogeny of immunoglobulin structure and function. I. Immunoglobulins of the lemon shark.

Lemon sharks immunized with bovine serum albumin produced two molecular forms of antibodies detectable by passive hemagglutination of antigen-coated, tanned sheep erythrocytes. Throughout the course of immunization 2-ME-sensitive antibody was associated with a 19S immunoglobulin fraction (4-5 mg/ml serum) while late in the course of immunization antibody was found also associated with a 7S immunoglobulin fraction (7-8 mg/ml serum). No evidence for any anamnestic response was found in these animals. Naturally occurring hemagglutinins for sheep erythrocytes were found to be 2-ME-sensitive and present in the 19S immunoglobulin fraction. These immunoglobulin fractions were readily purified by DEAE-cellulose chromatography and Sephadex G-200 gel filtration. Both immunoglobulin molecules yielded equimolar amounts of H and L polypeptide chains when subjected to extensive reduction and alkylation followed by gel filtration in 5 M guanidine-HCl. Antigenically reactive H and L chains were obtained by partial reduction and alkylation followed by gel filtration in 1 M propionic acid. The 7S and 19S immunoglobulin H chains were indistinguishable by fingerprints of tryptic digests, disc electrophoretic patterns, antigenic properties, and mass (molecular weight approximately 70,000), thus suggesting these two molecules to belong to the same immunoglobulin class. The shark 19S and 7S immunoglobulin L chains were indistinguishable from each other by similar criteria and were different from the H chains. These L chains exhibited the electrophoretic heterogeneity of their mammalian counterparts. The 7S (shark immunoglobulin) molecule was shown to have a molecular weight of approximately 160,000 and to consist of 2H and 2L polypeptide chains (total mass congruent with180,000). The 19S molecule was shown to have a molecular weight of 800,000-900,000; therefore, there were probably five 7S subunits per 19S molecule, comparable to mammalian gammaM. Other reasons for considering the 7S and the 19S lemon shark molecules to belong to a class of immunoglobulins comparable to the gammaM class of mammals are that they both have high carbohydrate contents, and H chains of mass similar to micro chains. The lemon shark serum proteins with electrophoretic mobilities comparable to gamma G of mammals were not related to the immunoglobulins of this species. These proteins had no antibody activity and had no antigenic or chemical similarity to either the H chains, the L chains, or the intact immunoglobulin molecules from the lemon shark.

Phylogeny of immunoglobulin structure and function. V. Valences and association constants of teleost antibodies to a haptenic determinant.

The giant grouper, Epinephelus itaira, was shown to synthesize 16 and 6.4S antibodies specific for the dinitrophenyl determinant (DNP). Sera obtained at various intervals between 1 month and 2 yr after initial immunization contained both species of antibody; no temporal synthesis was evident. Equilibrium dialysis studies employing epsilon-dinitrophenyl-amino caproic acid were conducted with purified grouper antibodies specific for DNP. The 16S antibody preparations obtained at 1 and 2 months of immunization showed heterogeneity of hapten binding indicative of two populations of combining sites. One-half of these sites (an average of four sites per 16S molecule) exhibited an average intrinsic association constant (K(o)) of approximately 10(6)M(-1); the K(o) of the remaining four-sites was approximately 10(4)M(-1). Thus, the valence of the grouper 16S antibody molecule appears to be eight although the distribution of the high and low K(o) sites is unknown, i.e., are they each on the same or on different molecules? The 16S antibody preparations obtained after more prolonged immunization exhibited increasingly lower K(o) values; the so-called low K(o) sites were no longer detectable. These findings are in contrast to reports of rabbit IgG antibodies showing an increase in K(o) with increased time. The 6.4S antibody preparations obtained from the 1 and 2 month antisera had K(o) values of approximately 10(6)M(-1) and a valence of one. These antibodies would not precipitate with antigen. The 6.4S antibody preparations obtained at later times showed decreasing K(o) values comparable to those of the 16S antibodies from the same bleedings. Studies on the thermodynamic parameters of the hapten-antibody interaction showed the grouper 16 and 6.4S antibodies to be similar to each other. These data also showed that the enthalpy and entropy changes of grouper antibody-hapten reactions resemble those reported for rabbit IgG antibodies to this hapten. It is thus suggested that, although considerable evolution of immunoglobulin classes has occurred between fish and rabbits, the antibody combining site may have remained relatively unchanged during a large part of evolutionary time.

Transgenic mice lacking class I major histocompatibility complex-restricted T cells have delayed viral clearance and increased mortality after influenza virus challenge.

To investigate the role of CD8+ T lymphocytes in recovery from influenza pneumonia, we used transgenic mice either homozygous (-/-) or heterozygous (+/-) for beta 2-microglobulin (beta 2-M) gene disruption. These mice lack major histocompatibility complex-restricted class I (CD8+) T cells. We found that after challenge with a nonlethal influenza virus, the beta 2-M (-/-) mice had significantly delayed pulmonary viral clearance. Furthermore, after challenge with a more virulent influenza virus, the beta 2-M (-/-) mice had a significantly higher mortality rate than did control mice. Thus, CD8+ T cells are important in recovery from virulent influenza infections, but other host defense mechanisms can clear the respiratory tract of more benign infections.

Experimental IgA nephropathy.

An animal model for IgA immune complex nephritis was developed. IgA immune complexes formed in vitro with an IgA anti-dinitrophenyl (DNP) derived from MOPC-315 plasmacytoma, and dinitrophenylated bovine serum albumin (DNP-BSA) produced mild focal glomerulonephritis in mice. Similar, but more severe pathological changes were produced with complexes formed in vivo either in normal mice or MOPC-315 tumor-bearing mice. In contrast to the focal nature of the PAS-positive glomerular lesions observed by light microscopy, immunofluorescent examination revealed IgA deposits in all glomeruli. This discrepancy between immunofluorescent and histopathologic findings as well as the distribution of the immune complexes within the affected glomeruli, are some of the features which bear resemblance between this experimental model and human IgA nephropathy. Fixation of complements by DNP-BSA-IgA immune complexes, formed in vitro or in vivo, was shown to occur in the glomeruli of mice with IgA immune complex nephropathy. The pattern of C3 glomerular deposits was similar to that of IgA. However, complement proved to be nonessential for complex deposition. This conclusion is based on the observation that decomplemented mice, although showing no deposition of C3 in their glomerulus, developed glomerular immunohistological changes similar to those observed in experimental mice that were not decomplemented. Polymeric IgA was observed to be critical for renal deposition of complexes and induction of nephritic histological changes. In contrast, monomeric IgA immune complexes failed to produce glomerular deposits. This finding raises the possibility that secretory IgA, which is predominantly polymeric, may play a role in human IgA-associated glomerulonephritis.

Management of anaphylaxis in primary care: Canadian expert consensus recommendations.

BACKGROUND: Anaphylaxis is often managed inadequately. We used findings from a systematic review of gaps in anaphylaxis management to develop evidence-based recommendations for gaps rated as clinically important by a panel of Canadian allergy experts. METHODS: The nominal group technique (NGT) consensus methodology was used to develop evidence-based recommendations for the management of anaphylaxis in primary care. Physician-specific gaps from our systematic review were prioritized by consensus meeting participants in two rounds, which involved the rating, discussion, and re-rating of gaps. Using current anaphylaxis guidelines, recommendations were then developed for each category of gaps that were identified by the panel as clinically important. RESULTS: Thirty unique physician gaps from the systematic review were categorized according to gaps of knowledge and anaphylaxis practice behaviors. The panel rated diagnosis of anaphylaxis, and when and how to use epinephrine auto-injectors as clinically important knowledge gaps; and rated infrequent or delayed epinephrine administration, low rate of auto-injector prescription, and infrequent or no referrals to allergy specialists after a reaction as important practice behavior gaps. Evidence from four guidelines was used to support the consensus recommendation statements for three resulting categories of gap themes: anaphylaxis management, epinephrine use, and follow-up care. CONCLUSION: We used an NGT consensus methodology to develop an educational resource for primary care physicians and allergists to better understand how to manage patients with anaphylaxis. Next steps include testing our findings against observed data in primary care settings and to develop other strategies or tools to overcome gaps in anaphylaxis management.

Evaluating the use of educational videos to support the tuberculosis care cascade in remote Madagascar.

SETTING: Access to information about tuberculosis (TB) is vital to ensure timely diagnosis, treatment, and control among vulnerable communities. Improved approaches for distributing health education materials to remote populations are needed.OBJECTIVE: To evaluate the impact of two comprehensive video training curricula in improving patient, community member, and community health worker knowledge of TB in a remote area of Madagascar.DESIGN: A pre-test/post-test design was used to measure knowledge acquisition. Educational videos were short, culturally appropriate films presented at critical moments in the TB cascade of care.RESULTS: Of the total 146 participants, 86 (58.9%) improved their score on the post-test, 50 (34.2%) obtained the same score, and 10 (6.8%) received a worse score. A statistically significant difference was observed between the pre- and post-test scores, wherein scores increased by a median of 10.0% (interquartile range 0.0-20.0) after viewing the videos (P < 0.001). There was a significant difference between the number of correct answers on the pre-test and the number of correct answers on the post-test (P < 0.001).CONCLUSION: Educational videos were found to significantly improve TB knowledge among a low-literacy, remote population in Madagascar. Our findings suggest educational videos could be a powerful, low-cost, and sustainable tool to improve access to TB education materials globally.

Colostral immunoglobulin-A: synthesis in vitro of T-chain by rabbit mammary gland.

When minced mammary tissue from lactating rabbits was incubated in vitro with C(14)-labeled lysine and isoleucine, it incorporated radio-activity into colostral immunoglobulin A. The only portion of this colostral molecule with significant labeling was T-chain, with little or no labeling of light or heavy chains. It was thus demonstrated that T-chains are synthesized by mammary gland. Because the remainder of the molecule was derived from unlabeled material, in vivo it was probably derived from serum.

Electrophoretic heterogeneity of polypeptide chains of specific antibodies.

Heavy and light polypeptide chains isolated from different specific antibodies to haptens and from (gamma)G-immunoglobulin of normal rabbits have been resolved into distinct, multiple components by disc electrophoresis in polyacrylamide gels in the presence of urea. In spite of the resolution of these chains into multiple bands, different specific antibodies and normal rabbit (gamma)G-immunoglobulin were indistinguishable from each other by this method.

Electrophoretic heterogeneity of the polypeptide chains of human G-myeloma proteins.

The light and heavy polypeptide chains derived from human Gmyeloma proteins are electrophoretically heterogeneous as judged by disc electrophoresis of the polypeptide chains in urea-acrylamide gels. Individual myeloma proteins contained as many as eight light-chain and nine heavy-chain components.

Control strategies for tuberculosis epidemics: new models for old problems.

Tuberculosis, although preventable and curable, causes more adult deaths than any other infectious disease. A theoretical framework for designing effective control strategies is developed and used to determine treatment levels for eradication, to assess the effects of noneradicating control, and to examine the global goals of the World Health Organization. The theory is extended to assess how suboptimal control programs contribute to the evolution of drug resistance. A new evaluation criterion is defined and used to suggest how control strategies can be improved. In order to control tuberculosis, treatment failure rates must be lower in developing countries than in developed countries.

Comparative genomics of BCG vaccines by whole-genome DNA microarray.

Bacille Calmette-Guérin (BCG) vaccines are live attenuated strains of Mycobacterium bovis administered to prevent tuberculosis. To better understand the differences between M. tuberculosis, M. bovis, and the various BCG daughter strains, their genomic compositions were studied by performing comparative hybridization experiments on a DNA microarray. Regions deleted from BCG vaccines relative to the virulent M. tuberculosis H37Rv reference strain were confirmed by sequencing across the missing segment of the H37Rv genome. Eleven regions (encompassing 91 open reading frames) of H37Rv were found that were absent from one or more virulent strains of M. bovis. Five additional regions representing 38 open reading frames were present in M. bovis but absent from some or all BCG strains; this is evidence for the ongoing evolution of BCG strains since their original derivation. A precise understanding of the genetic differences between closely related Mycobacteria suggests rational approaches to the design of improved diagnostics and vaccines.

Mycolactone: a polyketide toxin from Mycobacterium ulcerans required for virulence.

Mycobacterium ulcerans is the causative agent of Buruli ulcer, a severe human skin disease that occurs primarily in Africa and Australia. Infection with M. ulcerans results in persistent severe necrosis without an acute inflammatory response. The presence of histopathological changes distant from the site of infection suggested that pathogenesis might be toxin mediated. A polyketide-derived macrolide designated mycolactone was isolated that causes cytopathicity and cell cycle arrest in cultured L929 murine fibroblasts. Intradermal inoculation of purified toxin into guinea pigs produced a lesion similar to that of Buruli ulcer in humans. This toxin may represent one of a family of virulence factors associated with pathology in mycobacterial diseases such as leprosy and tuberculosis.

Persistent association of Mycobacterium ulcerans with West African predaceous insects of the family belostomatidae.

A number of studies have suggested that Mycobacterium ulcerans, the etiological agent of Buruli ulcer, may be transmitted to humans by insect bites. M. ulcerans has been isolated from a predaceous aquatic insect, and PCR detection of M. ulcerans DNA in aquatic environments suggests that the organism is widely distributed within many invertebrate taxa and functional feeding groups. Thus, M. ulcerans may be concentrated through different trophic links. However, the specific environmental niche of M. ulcerans and route of transmission to humans remain a mystery. In this study, a biologically relevant infection model in which M. ulcerans-infected mosquito larvae were fed to a species of predaceous hemiptera (African Belostomatidae) was used to demonstrate the persistent colonization of M. ulcerans and subsequent transmission of bacteria to naïve prey. The association of M. ulcerans with specific anatomical compartments showed that M. ulcerans accumulates preferentially on the exoskeleton. In contrast, few organisms were found in dissected guts or salivary glands. No difference was found between the ability of wild-type M. ulcerans and an M. ulcerans isogenic mycolactone-negative mutant to colonize belostomatids. These data show that African belostomatids can successfully be colonized by M. ulcerans and support the trophic transfer of M. ulcerans within the environment.

Intra-gastric balloon therapy and weight reduction.

BACKGROUND: Intra-gastric (TBT) can achieve pre-operative weight loss in obese patients, and may make surgery safer and easier. We report our experience in weight reduction and outcome in morbidly obese patients. METHODS: Data were prospectively collected in 50 consecutive morbidly obese patients undergoing intra-gastric balloon therapy. All balloons were inserted and removed by one consultant operator and injected with 500-600 mls of methylene blue-stained saline. Patients were admitted after balloon insertion to control nausea and reflux symptoms. Analysis was performed on post-insertion data collated from the hospital obesity database and patient note review. RESULTS: The majority of patients (70%) were female. Prior to IBT, mean patient weight was 149.9 +/- 36.8 kg (male 184.7 +/- 38.1 kg, female 135 +/- 26.5 kg). At six months follow-up, mean weight of patients reduced to 133.5 +/- 34.3kg (male 166.8 +/- 33.3 kg, female 119.3 +/- 25.9 kg). The overall percentage of excess weight loss was 22% (male 17%, female 25%, p=NS). Mean patient body mass index (BMI) at balloon insertion was 52.8 (+/-8.2) kg/m2 (male 56.8, female 51.0), mean patient BMI reduced to 47 (+/-8.1) (males 51.4, females 45.1) at six months. Average BMI loss was 5.8 (male 5.4, female 6.0). CONCLUSION: IBT is an effective method of weight reduction in morbid obesity with an average excess weight loss of 22% in this series. The benefit of pre-operative weight loss in this patient group is likely to be significant, but has yet to be measured.