Photorhabdus sp. strain Az29 is symbiotic with an Azorean nematode of the genus Heterorhabditis in a complex that is highly virulent to insects even at low temperatures. The virulence of the bacteria is mainly attributed to toxins and bacterial enzymes secreted during parasitism. The bacteria secrete proteases during growth, with a peak at the end of the exponential growth phase. Protease secretion was higher in cultures growing at lower temperatures. At 10°C the activity was highest and remained constant for over 7 days, whereas at 23 and 28°C it showed a steady decrease. Two proteases, PrtA and PrtS, that are produced in the growth medium were purified by liquid chromatography. PrtA was inhibited by 1,10-phenantroline and by EDTA and had a molecular mass of 56 kDa and an optimal activity at pH 9 and 50°C. Sequences of three peptides of PrtA showed strong homologies with alkaline metalloproteases from Photorhabdus temperata K122 and Photorhabdus luminescens W14. Peptide PrtA-36 contained the residues characteristic of metzincins, known to be involved in bacterial virulence. In vitro, PrtA inhibited antibacterial factors of inoculated Lepidoptera and of cecropins A and B. PrtS had a molecular mass of 38 kDa and was inhibited by 1,10-phenanthroline but not by EDTA. Its activity ranged between 10 and 80°C and was optimal at pH 7 and 50°C. PrtS also destroyed insect antibacterial factors. Three fragments of PrtS showed homology with a putative metalloprotease of P. luminescens TTO1. Polyclonal antibody raised against PrtA did not recognize PrtS, showing they are distinct molecules.Photorhabdus spp. are non-free-living Enterobacteriaceae (6, 19). The entomopathogen species of Photorhabdus have a symbiotic relation with nematodes of the genus Heterorhabditis for transport with their infective juveniles (IJs). The IJs actively seek for an insect host, penetrating through its natural openings and cuticle. Inside the insect hemocoel, the bacteria are released and actively multiply avoiding the host defenses and causing an acute disease condition that is followed by insect death within 48 h. The bacteria also create the nutritional conditions and protective environment for the development of its nematode symbiont (2, 21).Bacteria of the Photorhabdus genus produce toxins and other potentially virulent factors (16). The characterized toxins are organized in pathogenic islands (53) and include the Tc complex, which is responsible for oral toxicity (9, 52), and the Mcf toxin, which causes loss of insect body turgor, followed by death (14). Among other potential virulence factors, there is a complex set of extracellular enzymes, including proteases, lipases, lecithinases, chitinases, and phosphatases (5,12,20). Proteases represent an important part of these enzymes, although their role on the virulence process is yet unclear. Until now, all extracellular proteases purified and characterized from Photorhabdus were classified as metalloproteases, but in reports on proteases from different strains it has been suggested that ther...
The influence of extreme environments of volcanic origin over vertebrates and the cellular responses that these may give are almost unknown. The main objectives were to evaluate the exposure of mice to metals in the interior of houses of a small village settled inside a volcanic crater (Furnas, Azores), and the levels of apoptosis and metallothionein in the organs (lung, liver, and kidney) of those animals. Adult mice (Mus musculus) were captured in two areas, one with volcanic activity and the other without it over the last three centuries. In the excised organs, analysis of metals (Al, Cd, Pb, Zn), TUNEL assay for apoptosis, and immunohistochemistry for metallothionein were undertook. Mice from the area with volcanic activity presented higher levels of apoptosis and metallothionein than those from the area without volcanic activity. Such results were in agreement with the differences in metal burdens of the three organs, and interestingly these concentrations were similar to or higher than others found in heavily polluted areas outside the Azores. Thus, there may be a high risk of harmful effects for organisms, including humans, inhabiting areas with volcanism, where hazardous gases and metals in the air are very common during the entire day or even all year round.
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