Determination and quantification of Schistosoma mansoni cercarial emergence from Biomphalaria glabrata snails.

Living and fixed samples of Schistosoma mansoni -infected Biomphalaria glabrata snails were used to determine the relative contributions of different snail tissues to cercarial emergence (shedding). Three methods of observations were employed: (1) direct microscopical observations of shedding snails; (2) microscopic analysis of 5 µm serial sections (H&E stained) of actively shedding snails; and (3) scanning electron microscopic (SEM) observations of snails that were fixed while actively shedding. For this investigation, there were advantages and disadvantages to using each method. We confirmed the results of others that there were 3 tissues of the snail that contributed most prominently to cercarial release (mantle collar, pseudobranch, and headfoot). Based on histological analysis of cercarial accumulations in presumed shedding sites in these 3 tissues, 57% of the cercariae could be seen in the mantle collar, 30.6% in the pseudobranch, and 12.5% in the headfoot. Other anterior structures were involved to a much lesser extent. SEM observations clearly showed cercariae emerging either body first, tail first, or likely emerging en masse from blebs, especially from the mantle collar. These studies provide a more quantitative appraisal of the role the different anterior snail tissues play in cercarial emergence.

Induction of the Yersinia pestis PhoP-PhoQ regulatory system in the flea and its role in producing a transmissible infection.

Transmission of Yersinia pestis is greatly enhanced after it forms a bacterial biofilm in the foregut of the flea vector that interferes with normal blood feeding. Here we report that the ability to produce a normal foregut-blocking infection depends on induction of the Y. pestis PhoP-PhoQ two-component regulatory system in the flea. Y. pestis phoP-negative mutants achieved normal infection rates and bacterial loads in the flea midgut but produced a less cohesive biofilm both in vitro and in the flea and had a greatly reduced ability to localize to and block the flea foregut. Thus, not only is the PhoP-PhoQ system induced in the flea gut environment, but also this induction is required to produce a normal transmissible infection. The altered biofilm phenotype in the flea was not due to lack of PhoPQ-dependent or PmrAB-dependent addition of aminoarabinose to the Y. pestis lipid A, because an aminoarabinose-deficient mutant that is highly sensitive to cationic antimicrobial peptides had a normal phenotype in the flea digestive tract. In addition to enhancing transmissibility, induction of the PhoP-PhoQ system in the arthropod vector prior to transmission may preadapt Y. pestis to resist the initial encounter with the mammalian innate immune response.

A novel in vitro cultivation system to produce and isolate soluble factors released from hyphae of arbuscular mycorrhizal fungi.

The Petri plate dual in vitro culture system for arbuscular mycorrhizal fungi (AMF) is ill-suited for the production and isolation of extraradically released soluble factors, thereby preventing the further characterization of such compounds. To overcome this technical bottleneck, we here describe a novel cultivation system using the standard AMF Glomus intraradices-carrot dual culture as a model. This new system is capable of producing soluble factors in copious amounts without any interference from host roots. The greatest advantages of this culture method include ease of handling and reusability of the culture vessel, thus making it a very cost effective system.