Discrimination between Francisella tularensis and Francisella-like endosymbionts when screening ticks by PCR

The presence of Francisella-like endosymbionts in tick species known to transmit tularemia poses a potential diagnostic problem for laboratories that screen tick samples by PCR for Francisella tularensis. Tick samples initially considered positive for F. tularensis based on standard 16S rRNA gene PCR were found to be positive only for Francisella-like endosymbionts using a multitarget F. tularensis TaqMan assay (ISFtu2, tul4, and iglC) and 16S rRNA gene sequencing. Specificity of PCR-based diagnostics for F. tularensis should be carefully evaluated with appropriate specimen types prior to diagnostic use.     

Aquatic Francisella-like bacterium associated with mortality of intensively cultured hybrid striped bass Morone chrysops x M. saxatilis

The present study identifies an emerging disease associated with an aquatic Francisella-like bacterium that can cause mortality in hybrid striped bass Morone chrysops x M. saxatilis reared intensively in freshwater. Clinically affected fish were lethargic, had scattered haemorrhagic cutaneous lesions and diffuse gill pallor. The head kidney and spleen were markedly swollen and contained numerous interstitial granulomas; histological examination revealed small, pleomorphic Gram-negative coccobacilli within vacuolated cells. The bacterium could not be cultured from head kidney homogenates either with standard or enriched microbiological media or following inoculation of a Chinook salmon embryo (CHSE)-214 cell line. No amplification product was obtained from head kidney DNA by polymerase chain reaction (PCR) assay using Piscirickettsia salmonis-specific primers. PCR analysis of infected head kidney homogenate with primers designed for the eubacterial 16S rRNA produced a single amplicon. Phylogenetic analysis of this DNA sequence demonstrated that the sequence aligned most closely with members of the genus Francisella, identified from tilapia Oreochromis spp. in Taiwan and an aquatic Francisella species that was recently isolated from the three-line grunt Parapristipoma trilineatum in Japan. This Francisella-like disease was transmitted to naive hybrid striped bass fingerlings by intraperitoneal injection of tissue homogenates prepared from a natural outbreak. All fish developed gross and histological lesions identical to those from natural outbreaks. Intracellular Gram-negative bacteria were observed within the cytoplasm of cells (presumably macrophages) within the granulomas, but bacteria were not recovered. The 16S DNA sequence of the bacterium obtained from tissues of experimentally infected fish was identical to that obtained from the fish used as infected donor tissue.     

Amylose crystallization from concentrated aqueous solution

Maize amylose, separated from granular starch by means of an aqueous leaching process, was used to investigate spherulite formation from concentrated mixtures of starch in water. Amylose (10-20%, w/w) was found to form a spherulitic semicrystalline morphology over a wide range of cooling rates (1-250 degrees C/min), provided it was first heated to >170 degrees C. This is explained through the effect of temperature on chain conformation. A maximum quench temperature of approximately 70 degrees C was required to produce spherulitic morphology. Quench temperatures between 70 and 110 degrees C produced a gel-like morphology. This is explained on the basis of the relative kinetics of liquid-liquid phase separation vis-à-vis crystallization. The possibility of the presence of a liquid crystalline phase affecting the process of spherulite formation is discussed.