Assessment of Post-Vaccination Phagocytic Activation Using Candida albicans Killing Assays.

Candida albicans is an important opportunistic fungal pathogen. It is now the fourth leading cause of nosocomial bloodstream infections and a great threat to the immuncompromised patients attributed to the disseminated candidiasis with the mortality up to 40%. Phagocytic cells are the first line of defense against Candida infections. Antibodies induced by vaccination can effectively enhance the capacities of phagocytosis and killing of neutrophils and macrophages. In this chapter, flow cytometric analysis (FACS) and killing assay by plate culture methods are introduced to evaluate the phagocytosis and killing of strains of Candida albicans opsonized with immune serum obtained from mice vaccinated with yeast and recombinant enolase.

Improved vaccines through targeted manipulation of the body's immunological risk-assessment?

Recent advances have highlighted the outstanding role of the innate immune system for instructing adaptive immunity. Translating this knowledge into successful immunotherapies like vaccines, however, has proven to be a difficult task. This essay is based on the hypothesis that immune responses are tightly scaled to the infectious threat posed by a given microbial stimulus. A meticulous immunological risk-assessment process is therefore instrumental for eliciting well-balanced responses and maintaining immune homeostasis. The immune system makes fine distinctions, for example, between live and dead bacteria, or pathogenic and non-pathogenic microorganisms. Here, I discuss recent evidence for some of the mechanisms underlying these distinctions and speculate on strategies for therapeutically targeting the immunological risk-assessment machinery.

Nasty viruses, costly plasmids, population dynamics, and the conditions for establishing and maintaining CRISPR-mediated adaptive immunity in bacteria.

Clustered, Regularly Interspaced Short Palindromic Repeats (CRISPR) abound in the genomes of almost all archaebacteria and nearly half the eubacteria sequenced. Through a genetic interference mechanism, bacteria with CRISPR regions carrying copies of the DNA of previously encountered phage and plasmids abort the replication of phage and plasmids with these sequences. Thus it would seem that protection against infecting phage and plasmids is the selection pressure responsible for establishing and maintaining CRISPR in bacterial populations. But is it? To address this question and provide a framework and hypotheses for the experimental study of the ecology and evolution of CRISPR, I use mathematical models of the population dynamics of CRISPR-encoding bacteria with lytic phage and conjugative plasmids. The results of the numerical (computer simulation) analysis of the properties of these models with parameters in the ranges estimated for Escherichia coli and its phage and conjugative plasmids indicate: (1) In the presence of lytic phage there are broad conditions where bacteria with CRISPR-mediated immunity will have an advantage in competition with non-CRISPR bacteria with otherwise higher Malthusian fitness. (2) These conditions for the existence of CRISPR are narrower when there is envelope resistance to the phage. (3) While there are situations where CRISPR-mediated immunity can provide bacteria an advantage in competition with higher Malthusian fitness bacteria bearing deleterious conjugative plasmids, the conditions for this to obtain are relatively narrow and the intensity of selection favoring CRISPR weak. The parameters of these models can be independently estimated, the assumption behind their construction validated, and the hypotheses generated from the analysis of their properties tested in experimental populations of bacteria with lytic phage and conjugative plasmids. I suggest protocols for estimating these parameters and outline the design of experiments to evaluate the validity of these models and test these hypotheses.

Proceedings of a Consensus Conference: pathogen inactivation-making decisions about new technologies.

Significant progress has been made in reducing the risk of pathogen transmission to transfusion recipients. Nonetheless, there remains a continuing risk of transmission of viruses, bacteria, protozoa, and prions to recipients. These include many of the viruses for which specific screening tests exist as well as pathogens for which testing is currently not being done, including various species of bacteria, babesiosis, variant Creutzfeld-Jacob disease, hepatitis A virus, human herpes virus 8, chikungunya virus, Chagas disease, and malaria. Pathogen inactivation (PI) technologies potentially provide an additional way to protect the blood supply from emerging agents and also provide additional protection against both known and as-yet-unidentified agents. However, the impact of PI on product quality and recipient safety remains to be determined. The purpose of this consensus conference was to bring together international experts in an effort to consider the following issues with respect to PI: implementation criteria; licensing requirements; blood service and clinical issues; risk management issues; cost-benefit impact; and research requirements. These proceedings are provided to make available to the transfusion medicine community the considerable amount of important information presented at this consensus conference.