Evidence Supporting a Role for Mammalian Chitinases in Efficacy of Caspofungin against Experimental Aspergillosis in Immunocompromised Rats

Objectives:Caspofungin, currently used as salvage therapy for invasive pulmonary aspergillosis (IPA), strangely only causes morphological changes in fungal growth in vitro but does not inhibit the growth. In vivo it has good efficacy. Therefore the question arises how this in vivo activity is reached. Caspofungin is known to increase the amount of chitin in the fungal cell wall. Mammals produce two chitinases, chitotriosidase and AMCase, which can hydrolyse chitin. We hypothesized that the mammalian chitinases play a role in the in vivo efficacy of caspofungin.Methods:In order to determine the role of chitotriosidase and AMCase in IPA, both chitinases were measured in rats which did or did not receive caspofungin treatment. In order to understand the role of each chitinase in the breakdown of the caspofungin-exposed cells, we also exposed caspofungin treated fungi to recombinant enzymes in vitro.Results:IPA in immunocompromised rats caused a dramatic increase in chitinase activity. This increase in chitinase activity was still noted when rats were treated with caspofungin. In vitro, it was demonstrated that the action of both chitinases were needed to lyse the f

High prevalence of chitotriosidase deficiency in Peruvian Amerindians exposed to chitin-bearing food and enteroparasites

The human genome encodes a gene for an enzymatically active chitinase (CHIT1) located in a single copy on Chromosome 1, which is highly expressed by activated macrophages and in other cells of the innate immune response. Several dysfunctional mutations are known in CHIT1, including a 24-bp duplication in Exon 10 causing catalytic deficiency. This duplication is a common variant conserved in many human populations, except in West and South Africans. Thus it has been proposed that human migration out of Africa and the consequent reduction of exposure to chitin from environmental factors may have enabled the conservation of dysfunctional mutations in human chitinases. Our data obtained from 85 indigenous Amerindians from Peru, representative of populations characterized by high prevalence of chitin-bearing enteroparasites and intense entomophagy, reveal a very high frequency of the 24-bp duplication (47.06%), and of other single nucleotide polymorphisms which are known to partially affect enzymatic activity (G102S: 42.7% and A442G/V: 25.5%). Our finding is in line with a founder effect, but appears to confute our previous hypothesis of a protective role against parasite infection and sustains the discussion on the redundancy of chitinolytic function

Insights into the diagnosis, vaccines, and control of Taenia solium, a zoonotic, neglected parasite

Taenia solium taeniasis/cysticercosis (TSTC) is a foodborne, zoonotic neglected tropical disease affecting predominately low- and middle-income countries. Humans are definitive hosts for T. solium, whereas pigs act as intermediate hosts. Taeniasis, i.e. intestinal infection with adult T. solium in the human host, occurs through ingestion of undercooked pork infected with the larval stage (porcine cysticercosis, PCC). Human cysticercosis occurs after humans ingest T. solium eggs, acting as accidental intermediate hosts. Migration of cysticerci to the human brain results in neurocysticercosis (NCC), manifesting in a variety of clinical symptoms, most notably epilepsy. NCC is the leading cause of acquired epilepsy cases in endemic areas. PCC results in reduced pork value because of condemnation or the risk of condemnation of the meat. Available serological diagnostic tests for porcine and human cysticercosis are characterized by low sensitivity and are not cost-effective. An effective vaccine for T. solium cysticercosis in pigs has been developed, although it is not yet commercially available in all endemic countries, and still no vaccine is available for use in humans. This primer highlights the recent development in the field of diagnostic tests and vaccine production and explores possible strategies for future control and eradication of T. solium. In the absence of highly specific diagnostic tests and human vaccines, treatment of infected pigs and tapeworm carriers and prevention of disease transmission remain the principal means to interrupt the zoonotic cycle of T. solium in endemic countries

Multiple protein extract microarray for profiling human food-specific immunoglobulins A, M, G and E

Existing food immunoglobulin (Ig) tests require large volumes of serum, are limited to one immunoglobulin class, are not amenable to high throughput analysis and only give a limited picture of the immunological response to food antigens. Conversely a new generation of Component Resolved Diagnostic systems using pure proteins is highly specific and totally dependent on the availability of the protein in its recombinant or natural origin form. Here we demonstrate a proof-of-concept of a microarray test based on protein extracts of food components. Our approach relies on innovations on three different fronts: the novelty of using arrayed food samples sequentially extracted with detergent and chaotropic agents, the ability to measure four different Ig classes simultaneously and the ability to analyse the generated data via a suitable bioinformatics/statistical analysis interface. This approach combines high numerical power of microarrays with automation, high throughput analysis and enables detailed investigation of the Ig profiles to food antigens. The prototype shown contains extracts of approximately 350 food ingredients that cover most of the food products found in the UK. Here we showed that the use of a sequential extraction technique to solubilise and then denature food samples has its benefits in the assessment of variations in antigenicity when tested with human sera. A patient dependent degree of class specificity was observed with human sera (IgG specificity correlates well with IgA > IgM >>>>> IgE). Besides generating a simultaneous profile for IgA, IgM, IgG and IgE the array system has shown good discrimination between challenge responders in atopic and non-atopic individuals. Poly- and mono-specific IgE responders were easily identified. The mathematical modelling of specific IgE content showed good correlations when compared with established IgE antibody testing assay (UniCAP). Although in its proof-of-principle stages, the immune profiling technique described here has the potential to provide unique insights into exposure/sensitization and establish relationships between specific immunoglobulin classes and subclasses against food protein antigens. In further developments, the immune profiling technique could also be extended to other related areas such as parasite and bacterial gut infection. Full analyses of large longitudinal and retrospective clinical trials are on going to determine the positive and negative predictive values of the technique

Prediction of tolerance in children with IgE mediated cow's milk allergy by microarray profiling and chemometric approach

The sera of a retrospective cohort (n=41) composed of children with well characterized cow's milk allergy collected from multiple visits were analyzed using a protein microarray system measuring four classes of immunoglobulins. The frequency of the visits, age and gender distribution reflected real situation faced by the clinicians at a pediatric reference center for food allergy in São Paulo, Brazil. The profiling array results have shown that total IgG and IgA share similar specificity whilst IgM and in particular IgE are distantly related. The correlation of specificity of IgE and IgA is variable amongst the patients and this relationship cannot be used to predict atopy or the onset of tolerance to milk. The array profiling technique has corroborated the clinical selection criteria for this cohort albeit it clearly suggested that 4 out of the 41 patients might have allergies other than milk origin. There was also a good correlation between the array data and ImmunoCAP results, casein in particular. By using qualitative and quantitative multivariate analysis routines it was possible to produce validated statistical models to predict with reasonable accuracy the onset of tolerance to milk proteins. If expanded to larger study groups, the array profiling in combination with the multivariate techniques show potential to improve the prognostic of milk allergic patients

Parasitic nematode modulation of allergic disease

Incidence of allergic diseases such as asthma has increased at an alarming rate in Western countries in the past few decades. However, in parts of the world in which parasitic nematode infections are highly prevalent, allergy remains uncommon. Hence, it has been postulated that nematodes offer humans protection against this type of disease. This article reviews the evidence to support this idea, considering data from human studies and results from investigations into the protective effects of nematodes in animal models of allergic disease. The evidence strongly favors a protective role for nematodes; thus, the search is on to find the molecules involved, with a view toward using them for therapeutic purposes. The article also describes the nature and mode of action of recently characterized nematode-derived molecules with antiallergic properties and highlights their therapeutic efficacy in allergy models