Probing the surface ultrastructure of Brevibacillus laterosporus using atomic force microscopy.

One of the main obstacles to studying the surface ultrastructure of microbial cells by atomic force microscopy (AFM) is determining how to immobilize live cells on the AFM substrates. Each method has its own advantages and disadvantages. The aim of this study was to characterize a new simple and inexpensive method using two types of polyethersulfone (PES) membrane filters that differ in pore size (micropore and nanopore) to immobilize live and dead Brevibacillus laterosporus for AFM imaging. B. laterosporus was easily trapped by the microporous PES membrane, facilitating the successful AFM scanning of the bacterial surface ultrastructure. In addition, B. laterosporus strongly attached to the nanoporous membranes and withstood the pulling forces exerted by the AFM tip during scanning. These methods of immobilization did not affect the cell viability. The nanostructure and roughness of the bacterial surface were also observed for live, fixed, and air-dried cells. Live and dead bacteria displayed similar morphologies at low resolution, while at high resolution, live bacteria displayed a more convoluted surface ("brain-like structure").

Observations on house fly larvae midgut ultrastructure after Brevibacillus laterosporus ingestion.

The pathological and histopathological course caused by Brevibacillus laterosporus on house fly larvae has been investigated conducting observations on insect behavior and midgut ultrastructure. After dissection and fixation, gut tissues were analyzed under transmission electron microscopy (TEM) in order to compare in vivo-treated and non-treated (control) fly specimens. Treated larvae showed extensively reduced feeding and growth rate, then became sluggish and died within 72 h. A progressive midgut epithelium deterioration was observed in treated larvae, compared to the control. Ultrastructural changes consisted of microvilli disruption, cytoplasm vacuolization and general disorganization, endoplasmic reticulum deformation, mitochondria alteration. Deterioration became progressively more dramatic until the infected cells released their content into the gut lumen. Disruption was associated also with midgut muscular sheath and connective tissue. These ultrastructural changes are similar to those widely described for other entomopathogenic bacteria, such as Bacillus thuringiensis, against different insect species. The rapid disruption of cellular fine structure supports a hypothesis based on an interaction of toxins with the epithelial cell membranes reminiscent of the specific B. thuringiensis δ-endotoxins mechanism of action on other insect targets.