Colony-stimulating factor-1 receptor protein expression is a specific marker for goldfish (Carassius auratus L.) macrophage progenitors and their differentiated cell types

Signaling through the colony-stimulating factor-1 receptor (CSF-1R) mediates the proliferation, differentiation, and activation of macrophages and their progenitors. In this study we report on the use of an anti-goldfish CSF-1R antibody to specifically recognize a population of CSF-1R positive cells from goldfish tissues. Furthermore, using our previously characterized primary kidney macrophage culture system, we show that CSF-1R positive cells include monocytes, macrophages, and their progenitor cells. Freshly isolated progenitor cells had a higher median florescent intensity ratio than those progenitor cells cultured for up to four days. The decrease in CSF-1R expression on the progenitor cells coincides with the appearance and development of monocytes and macrophages. Monocytes were consistently CSF-1R⁺ and maintained the high level of CSF-1R expression as they developed into macrophages. Like that of mammalian systems, CSF-1R is expressed on all macrophage sub-populations (progenitors, monocytes, macrophages), and CSF-1R expression increases with macrophage development in teleosts.     

Application of Two-spotted Spider Mite Tetranychus urticae for Plant-pest Interaction Studies

The two-spotted spider mite, Tetranychus urticae, is a ubiquitous polyphagous arthropod herbivore that feeds on a remarkably broad array of species, with more than 150 of economic value. It is a major pest of greenhouse crops, especially in Solanaceae and Cucurbitaceae (e.g., tomatoes, eggplants, peppers, cucumbers, zucchini) and greenhouse ornamentals (e.g., roses, chrysanthemum, carnations), annual field crops (such as maize, cotton, soybean, and sugar beet), and in perennial cultures (alfalfa, strawberries, grapes, citruses, and plums)^1,2. In addition to the extreme polyphagy that makes it an important agricultural pest, T. urticae has a tendency to develop resistance to a wide array of insecticides and acaricides that are used for its control^3-7.T. urticae is an excellent experimental organism, as it has a rapid life cycle (7 days at 27 °C) and can be easily maintained at high density in the laboratory. Methods to assay gene expression (including in situ hybridization and antibody staining) and to inactivate expression of spider mite endogenous genes using RNA interference have been developed^8-10. Recently, the whole genome sequence of T. urticae has been reported, creating an opportunity to develop this pest herbivore as a model organism with equivalent genomic resources that already exist in some of its host plants (Arabidopsis thaliana and the tomato Solanum lycopersicum)¹¹. Together, these model organisms could provide insights into molecular bases of plant-pest interactions.Here, an efficient method for quick and easy collection of a large number of adult female mites, their application on an experimental plant host, and the assessment of the plant damage due to spider mite feeding are described. The presented protocol enables fast and efficient collection of hundreds of individuals at any developmental stage (eggs, larvae, nymphs, adult males, and females) that can be used for subsequent experimental application.     

Macrophage colony-stimulating factor (CSF-1) induces pro-inflammatory gene expression and enhances antimicrobial responses of goldfish (Carassius auratus L.) macrophages

We report on the regulation of pro-inflammatory functions of goldfish macrophages and induction of gene expression by recombinant goldfish CSF-1 (rgCSF-1). Recombinant goldfish TNFα-2 (rg TNFα-2), rgIFNγ but not rgTGFβ induced time-dependent increase of CSF-1 expression in macrophages. Treatment of goldfish macrophages with rgCSF-1 increased expression of several immune genes including CXCL-8 (= IL-8), CCL-1, TNFα-1, TNFα-2, IL-1β-1, IL-1β-2, IL-12-p35, IL-12-p40, IFN, IL-10 and iNOS A and B. The rgCSF-1 treatment did not significantly alter the mRNA levels of TGFβ and NRAMP in macrophages up to 48 h post treatment. However, at 72 h post treatment, the expression of TGFβ increased whereas that of NRAMP decreased. The treatment of macrophages with rgCSF-1 enhanced their respiratory burst and nitric oxide responses that were abrogated after addition of soluble CSF-1 receptor (sCSF-1R) to cell cultures. Macrophages exhibited a concentration-dependent chemotactic response toward rgCSF-1 as well as an increase in phagocytic activity that was abrogated after addition of sCSF-1R to cell cultures. Our results indicate that in addition to being an important growth factor of goldfish macrophages, rgCSF-1 also plays a central role in the regulation of their pro-inflammatory responses.     

The induction of nitric oxide response of carp macrophages by transferrin is influenced by the allelic diversity of the molecule

The central role of transferrin (Tf) as an iron transporting protein has been extended by observations that modified versions of Tf also participate in the regulation of innate immunity. We report on the isolation of two carp Tf proteins (alleles D and G) to purity using rivanol precipitation and ion-exchange chromatography, and describe the activation of head kidney-derived carp macrophages by cleaved Tf. We demonstrate the superiority of the D-type over the G-type Tf in inducing nitric oxide (NO) and confirm previous observations that full-length Tf cannot induce NO in fish macrophages. We believe that cleaved Tf fragments should be considered to be "alarmins". We discuss the possibility that parasites such as Trypanoplasma borreli cleave Tf and use Tf fragments to their advantage by modulating the NO induction in carp macrophages.     

A reexamination of the propensities of amino acids towards a particular secondary structure: classification of amino acids based on their chemical structure

The correlation between the primary and secondary structures of proteins was analysed using a large data set from the Protein Data Bank. Clear preferences of amino acids towards certain secondary structures classify amino acids into four groups: α-helix preferrers, strand preferrers, turn and bend preferrers, and His and Cys (the latter two amino acids show no clear preference for any secondary structure). Amino acids in the same group have similar structural characteristics at their Cβ and Cγ atoms that predicts their preference for a particular secondary structure. All α-helix preferrers have neither polar heteroatoms on Cβ and Cγ atoms, nor branching or aromatic group on the Cβ atom. All strand preferrers have aromatic groups or branching groups on the Cβ atom. All turn and bend preferrers have a polar heteroatom on the Cβ or Cγ atoms or do not have a Cβ atom at all. These new rules could be helpful in making predictions about non-natural amino acids.

Survival of Giardia lamblia trophozoites after exposure to UV light

The ability of Giardia lamblia trophozoites to reproduce after exposure to different fluences of UV radiation was determined using an in vitro-cultured method. The rate of parasite reproduction following UV exposure was measured by direct enumeration of trophozoites cultured in Diamond's Trypticase Yeast extract-Iron (TYI)-S-33 medium. The results suggested that some G. lamblia trophozoites may survive or are reactivated following exposure to UV fluences up to 10 mJ cm(-2). In addition, trophozoites exposed to a UV fluence of 1 mJ cm(-2) were infectious to Mongolian gerbils. Evidence of survival or reactivation at UV fluences of 20 and 40 mJ cm(-2) was ambiguous and statistically inconclusive, while at 100 mJ cm(-2) there was no evidence of survival or reactivation. This finding may have implications for criteria used by the drinking water and wastewater treatment industry to ensure safe reduction of G. lamblia cysts by UV disinfection processes.