The impacts of larval density and protease inhibition on feeding in medicinal larvae of the greenbottle fly Lucilia sericata

Larval therapy, the therapeutic use of blowfly larvae to treat chronic wounds, is primarily used in debridement. There are, however, gaps in current knowledge of the optimal clinical application of the therapy and mechanisms of action in the debridement process. Using an artificial assay, two studies were undertaken to investigate these aspects of larval debridement by Lucilia sericata Meigen (Diptera: Calliphoridae); the first studied the effects of the density of larvae on tissue digestion and larval mass, and the second considered the effects on the same parameters of incorporating protease inhibitors into the feeding substrate. The total mass of tissue digested increased with larval density until saturation was observed at 5.0–7.5 larvae/cm². This range was considered optimal as lower doses resulted in the removal of less tissue and higher doses offered no additional tissue removal and appeared to exacerbate competition for feeding. In the second study, increased protease inhibitor concentration led to significant decreases in tissue digestion and larval mass, suggesting that serine proteases, particularly trypsin, may play major roles in larval digestion. Such information is important in elucidating the main constituents that make up larval digestive products and may be significant in the development of new therapies.     

Combined effects of photodynamic therapy and irrigants in disinfection of root canals

In this study, the combined effects of photodynamic therapy and irrigants in eradicating common endodontic pathogens are evaluated. Roots of 80 extracted single rooted teeth are divided into 2 groups (1) mechanical flushing; (2) antibacterial irrigation. After cleaning and shaping, they are inoculated with either (A) Streptococcus mutans or (B) Enterococcus faecalis and incubated. They are again subdivided and either only irrigated or irrigated and lased. Dentin shavings are taken from root canal walls and cultured. Statistical analysis using One‐Way ANOVA and Post‐hoc tests are done. The combination eradicated both bacteria. Antibacterial irrigants controlled S. mutans better than PDT (p = 0.041). The combination of PDT and antibacterial irrigation proposed in this study can be used in all primary cases for thorough and reliable disinfection of root canals but may be highly effective in resistant cases like endodontic failures, as E. faecalis is prevalent in such cases.

     

Denaturing gradient gel electrophoresis analysis with different primers of subgingival bacterial communities under mechanical debridement

DGGE of 16S rDNA is one of the most frequently used methods to study microbial communities. In this study, the DGGE profiles of different 16S rDNA regions of the periodontal pathogens Porphyromonas gingivalis, Fusobacterium nucleatum, and Prevotella nigrescens were investigated. The results suggested that V3-V5 and V6-V8 fragments may be suitable for community analysis of subgingival bacteria. Further analysis of subgingival samples with V3-V5 and V6-V8 regions as target fragments suggested that, in chronic periodontitis, re-colonization by periodontal bacteria with a population very similar to the baseline may occur by 6 weeks after mechanical debridement.     

The influence of territory characteristics and food supply on the breeding performance of the Red-backed Shrike (<Emphasis Type="Italic">Lanius collurio</Emphasis>) in an extensively farmed region of eastern Poland

The breeding performance of the Red-backed Shrike (Lanius collurio) in relation to the territory characteristics was investigated. The study was conducted in an area of high agricultural land use, characterised by small field size, a more mosaic habitat and low use of mineral fertilisers in comparison to Western European countries. The influence of habitat structure and composition in the territory on the date of clutch initiation, nest predation and clutch size in the Red-backed Shrike was not found, but such an influence on the number of nestlings was shown. The minimum adequate model explained 18.3% of the variation between territories in nestling’s number. The number of fledglings was positively correlated with the area of pastures, meadows, orchards and fallows within territories, and negatively correlated with length of overhead transmission lines. The number of fledglings was correlated with food abundance within territories, based on to the results from pitfall traps. There was no correlation between territory food abundance and the size of clutch. The presence of pastures, meadows and fallows, which are foraging places, thus seems to be crucial for the protection of the Red-backed Shrike.     

蛆虫清创疗法在难愈性感染创面的临床应用

蛆虫治疗感染创面已有近千年的历史,始载于《本草纲目》,具有良好的疗效。20世纪40年代以后,随着抗生素的广泛使用,蛆虫治疗感染创面的方法逐渐被遗弃。但是随着抗生素的滥用,以及细菌严重耐药性的出现,使得人们开始重新审视这种古老而又充满生机的方法。在难愈性感染创面的治疗过程中,蛆虫通过多种作用机制能够促进创面的愈合,清除大量的坏死组织,促进肉芽组织生长,并有显著的抑菌作用。蛆虫通过多种作用机制协同作用能够促进难愈性感染伤口的愈合,值得进一步的临床研究及推广。本文广泛查阅近年国内外相关文献进行回顾与综合分析。对蛆虫清创疗法在难愈性感染创面的临床应用进行综述。     

Genetic evidence supporting the existence of two diverged groups in the goby Gymnogobius castaneus

The genetic population structure of the Japanese freshwater goby Gymnogobius castaneus was investigated on the basis of analysis of gene products of 19 allozyme loci. Two diverged groups were detected, one being endemic to the Kanto region and the other extensively distributed in eastern Japan. These two groups were distinguishable from each other by a complete allelic substitution in one locus, G3PDH*. In the Kanto region, both groups were distributed in the same river basin, being distinguishable by a complete allelic substitution in four loci, G3PDH*, GPI-2*, PGDH*, and PGM*. These results suggest that these two groups showed reproductive isolation.     

Assessing the diet of breeding bearded vultures (<Emphasis Type="Italic">Gypaetus barbatus</Emphasis>) in mid-20th century in Spain: a comparison to recent data and implications for conservation

We describe the diet of bearded vultures (Gypaetus barbatus) in southern Spain during the period from 1940 to 1950 based on the remains discovered in two historical nests and compare (using the same method) the anatomical and taxonomical bones found with recent data (2000–2001). At both times, the most important prey were medium-sized mammals (old: 73.3%, n = 30 vs recent: 71.2%, n = 73) with Ovis/Capra being the most prevalent (70% and 68.5%, respectively). Concerning the anatomical remains found, in old nests, long bones were the most frequently encountered skeletal part (30%) followed by the vertebral column (23%) and skull (23%); whereas, in recent nests, the most common remains were extremities (49.3%) followed by long bones (19.2%) and skulls (15.1%). These differences probably are due to differences in taphonomic conservation as a consequence of bone density and the removal by other scavenger species. The results suggest similar dietary habits between periods, with domestic species being the most important prey species group (old: 93.3% vs recent: 82.2%). These results emphasise the importance of the management of extensive livestock for the conservation of the bearded vulture.     

Recombinant Lucilia sericata chymotrypsin in a topical hydrogel formulation degrades human wound eschar ex vivo

Larval biotherapy is a debridement tool used in wound management. The mechanism of action involves degradation of eschar by serine proteases including chymotrypsin within the alimentary fluids of first instar Lucilia sericata. With the rationale of obviating some limitations of biotherapy, including cost, complexity of use, and patient reticence, the present study describes a mobile hydrogel formulation containing freeze-dried recombinant L. sericata chymotrypsin designed for topical application. Neither freeze-drying nor formulation into the hydrogel significantly attenuated the measured activity of released enzyme compared to fresh-frozen enzyme in aqueous solution. Gel electrophoresis confirmed qualitatively that the chymotrypsin/hydrogel formulation both with and without supplementary urea at 10% (w) /(v) degraded human chronic wound eschar ex vivo. Mindful that the hallmark of intractability of chronic wounds is aberrant biochemistry, the pH activity profile for the enzyme/hydrogel formulation was compared with exudate pH in chronic wounds of mixed aetiology in a cohort of 48 hospital in-patients. Five patients' wounds were acidic, however, the remainder were predominantly alkaline and coincided with the pH optimum for the insect enzyme. Thus, a recombinant L. sericata chymotrypsin and hydrogel formulation could represent a pragmatic alternative to larval therapy for the management of chronic wounds.     

Wound debridement potential of glycosidases of the wound-healing maggot, Lucilia sericata

The wound-healing maggot, Lucilia sericata Meigen (Diptera: Calliphoridae), degrades extracellular matrix components by releasing enzymes. The purpose of this study was to investigate the glycosylation profiles of wound slough/eschar from chronic venous leg ulcers and the complementary presence of glycosidase activities in first-instar excretions/secretions (ES1) and to define their specificities. The predominant carbohydrate moieties present in wound slough/eschar were determined by probing one-dimensional Western blots with conjugated lectins of known specificities. The presence of specific glycosidase activities in ES1 was determined using chromogenic and fluorogenic substrates. The removal of carbohydrate moieties from slough/eschar proteins by glycosidases in ES1 was determined by two-dimensional electrophoresis and Emerald 300 glycoprotein staining. α-d-glucosyl, α-d-mannosyl and N-acetlyglucosamine residues were detected on slough/eschar-derived proteins. Furthermore, it was demonstrated that the treatment of slough/eschar with ES1 significantly reduced uptake of the carbohydrate-specific stain. Subsequently, α-d-glucosidase, α-d-mannosidase and N-acetylglucosaminidase activities were identified in ES1. Specific chromogenic and fluorogenic substrates and gel filtration chromatography showed that these activities result from distinct enzymes. These activities were mirrored in the removal of α-d-glucosyl, α-d-mannosyl and N-acetylglucosamine residues from proteins of slough/eschar from maggot-treated wounds. These data suggest that maggot glycosidases remove sugars from slough/eschar proteins. This may contribute to debridement, which is ultimately accomplished by a suite of biochemically distinct enzymes present in ES1.