Digest Chitin Research Articles

Phage Adsorption and Cell Adherence Are Motility-dependent Characteristics of the Gliding Bacterium Cytophaga johnsonae

Summary: Mutants of the gliding bacterium Cytophaga johnsonae that are incapable of movement are called truly nonmotile (TNM) to distinguish them from other mutants that are motile but produce nonspreading colonies. All TNM mutants are pleiotropic, being nonmotile, unable to digest chitin, resistant to all phages that infect wild-type cells, nonadherent and having less hydrophobic surfaces than do wild-type cells. In this study, we tested the idea that the TNM pleiotropy is the result of blocking cell surface movement, rather than of loss or alteration of a specific cell surface component. Motility of wild-type cells was blocked by addition of chemicals, and treated cells were compared with untreated cells for their ability to adhere to spheroidal hydroxyapatite (SHA) beads and to adsorb phages, two characteristics used as an index of the TNM pleiotropy. All the chemicals tested that blocked motility also reduced phage adsorption and adherence to SHA to approximately the same extent seen with TNM mutants. The chemicals tested (carbonyl cyanide m-chlorophenylhydrazone, cyanide, azide and photoactivated eosin Y and rose bengal) were sufficiently different from each other to reduce the possibility that each chemical inhibited phage adsorption and cell adherence by similar secondary effects, independent of their effects on motility. It was also shown that the pleiotropy of TNM mutants is not caused by their inability to maintain a membrane potential. The results are compatible with the conclusion that the TNM pleiotropy is manifested whenever cell movement is stopped, whether by mutation or by use of chemical inhibitors, and they are inconsistent with the idea that all TNM mutants are pleiotropic because they all carry a lesion in the same gene that codes for expression of surface components required for all characteristics affected. The reason that stopping motility influences several seemingly unrelated properties is not known but is probably related to adaptations required for the organism to interact with its environment through a cell surface covered with a slime that is normally kept in motion by components of the machinery of gliding motility.

Characteristics of Crassostrea virginica crystalline style chitin digestion

1. 1. Fundamental chitin digestion characteristics of Crassostrea virginica crystalline style were investigated. 2. 2. Optimum temperature and pH were 34°C and 4.8. respectively. 3. 3. The colloidal regenerated chitin (0.56mol/0.5 ml: GlcNAc equivalents) was saturating under all enzyme levels encountered. 4. 4. There was no evidence of end product inhibition, even after 100 hr incubation. 5. 5. Calculated K m for the chitinase complex was 1.19mM when determined using a 30 min assay, but was only 0.70 mM when determined using a 4.6 hr assay. 6. 6. Both K m values are lower than reported for similar assays in other molluscs and for most bacteria. 7. 7. Effect of substrate preparation on the kinetics are discussed. 8. 8. Eight peaks of chitinase activity were resolved by DEAE-Fractogel ion exchange chromatography.

The growth of rainbow trout ( Salmo gairdneri) given diets containing chitin and its relationship to chitinolytic enzymes and chitin digestibility

The growth of rainbow trout fed diets containing 4, 10 and 25% chitin, over a 12-week period, was significantly depressed ( P < 0.001) when compared with controls fed diets containing 25% starch. There was no difference in growth rate between control fish and those fed diets containing 10% N-acetyl glucosamine (GlcNAc). Isotopically labelled amino sugars were shown to be oxidized when injected intraperitoneally into rainbow trout. Relatively high levels were found of chitinase activity in the stomachs and of chitobiase in the intestines. These enzyme activities were similar in all the trout, irrespective of the amount of chitin in their diets, except that chitobiase in the intestines of fish fed diets containing GlcNAc showed higher levels of activity than the controls ( P < 0.05). Chitin was not significantly digested when fed at 10 and 30% of the diet but the apparent digestibility of pre-cooked starch was 50% when fed at either 15 or 25%, on the basis of the inert indicator (Cr 2O 3) method. Chitinase and chitobiase activities were not reduced in fish fed diets containing 10% chitin and an antibiotic (Tribrissen ®) although the alimentary microflora were completely eliminated. Enzyme activities were not enhanced when live chitinolytic bacteria ( Vibrio alginolyticus) were incorporated into diets with 10% chitin; these bacteria were only recovered from the intestine. The evidence indicates an endogenous origin of chitinolytic enzymes in the trout gastro-intestinal tract. The presence of either antibiotic or bacteria in the diet had no effect on the digestibility of chitin.

Digestion of chitin by the labial glands of Acromyrmex octospinosus Reich (Hymenoptera: Formicidae)

The capability of the attine ant, Acromyrmex octospinosus, to digest chitin was studied. β-N-Acetylglucosaminidase, chitobiase, and chitinase activities of labial gland extracts were demonstrated. The biochemical characteristics (pH optimum, Michaelis constant, temperature influence) were investigated and the results are discussed in comparison with published data. The absence of microflora in the labial glands suggests the direct production of the chitinolytic system by labial glands. Its function in the digestion of the chitinous symbiotic fungus walls was examined. Effective degradation of this fungal material has been demonstrated in the infrabuccal pocket. Acromyrmex octospinosus appears to have adapted to a fungus diet by synthesis of specific enzymes. This result is discussed in relation to a previous hypothesis concerning mycophagous insects. Some hypotheses regarding the role of the chitinolytic activities are formulated.

Insect endochitinases: Glycoproteins from moulting fluid, integument and pupal haemolymph of Manduca sexta L.

Three endochitinases have been isolated from the tobacco hornworm, Manduca sexta, by ammonium sulphate fractionation and chromatographic procedures. The purified enzymes show single bands on sodium dodecyl sulphate acrylamide gel electrophoresis and have apparent mol. wts of 7.5 × 10 4 (I), 6.2 × 10 4 (II) and 5.0 × 10 4 (III). They are present in prepharate pupal integument, larval integument prior to apolysis, moulting fluid and pupal haemolymph. I and III are glycoproteins that contain glucosamine and several neutral hexoses. II was not examined for carbohydrate. The endochitinases cleave glycol chitin and chitin oligosaccharides ultimately to disaccharide and trisaccharide products. The longer substrates are preferred and no activity is expressed toward p- nitrophenyl-2-acetamido-2-deoxy-β- d-glucopyranoside , the β (1 → 4) linked dimer of N-acetylglucosamine or Micrococcus lysodeikticus cell walls. K m and k cat values for glycol chitin are 0.2 mg/ml and 2 sec −1, respectively. The endochitinases are quite stable over a wide range of pH and exhibit maximum activity around pH 6 with the oligosaccharide substrates. Antibody raised against III cross reacts with II but not with I. The endochitinases in integument and moulting fluid probably digest chitin in old cuticle prior to moulting.

Evidence for chitin synthesis in an insect cell line

AbstractCells from the continuous MRRL‐CH line derived from embryos of the tobacco hornworm synthesized chitin. Digestion of the washed pellet from [14C]‐N‐acetylglucosamine‐labeled cells by chitinase yielded a water‐soluble labeled compound. The lyophilized residue from the supernatant of the chitin digestion was analyzed by gas‐liquid chromatography as its trimethylsilyl derivative. The major component cochromatographed with derivitized chitobiose. The presence of chitobiose was confirmed by gas chro‐matography‐mass spectrometry. The synthesis of chitin by this cell line is inhibited by diflubenzuron.

Digestion of cuticle chitin during the moult of Manduca sexta (Lepidoptera: Sphingidae)

At the time of moulting, the old cuticle of Manduca sexta acquires the ability to break down its constituent chitin. The appearance of such endogenous moulting chitinase activity in vivo is not related to the occurrence of apolysis but results from exposure to moulting fluid; chitinase activity is induced in intact or deproteinized cuticle in vitro by incubation with enzymatically active moulting fluid ( Bade, 1970). Acquisition of endogenous chitinase activity by the chitin of intact cuticle in vitro requires several steps, one of which involves an enzymatic reaction which is a function of both temperature and concentration of moulting fluid. The process of rendering cuticle chitin able to serve as substrate for chitinase, can be inhibited by anti-trypsin agents and is postulated to be a novel variant of the biological activation of a biopolymer by a protease. Moulting fluid chitinase becomes attached to deproteinized chitin in vitro. Attachment occurs very rapidly, and the extent of this attachment with time is a function of temperature. Digestion of old cuticle during the moult is shown to be an orderly process which comprises a number of steps in sequence; some steps probably overlap in part.

Nutritive Value of Crab Meal for Young Ruminating Calves

Three trials, utilizing 26 male Holstein calves, were to evaluate the nutritive value of dried crab meal in ruminant feeds. In trials 1 and 2 crab meal (29% crude protein and 10% crude fiber) was substituted in amounts up to 20% for a basal ration (16% crude protein and 19% crude fiber) formulated to satisfy the National Research Council requirements for young rapidly growing animals. No declines were significant for body weight gain, feed intake, or feed efficiency in trials 1 and 2. Nitrogen digestion and retention were not altered by crab meal substitution in trial 1, but nitrogen retention was reduced by 20% crab meal substitution in trial 2. This reduction was associated with increased urine outputs. Crab meal rations contained excessive calcium, sodium, chlorine, and bromine which may have caused mineral imbalance increasing urine output. Digestibilities of dry matter, fiber, and calcium were not significantly different for basal and crab meal rations. Digestibility of chitin from the crab meal was measured as 66% but was highly variable from 26 to 87%. In trial 3 a crab meal addition was made to a protein-deficient ration (crude protein = 7.2%) resulting in a crude protein of 10.3%. Average daily gain and feed intake were higher for the supplemented ration.

Ecology of Vibrio parahaemolyticus in Chesapeake Bay.

A study of the ecology of Vibrio parahaemolyticus and related vibrios in the Rhode River area of Chesapeake Bay was carried out over the period December 1970 through August 1971. The incidence of V. parahaemolyticus and related vibrios was found to be correlated with water temperature. The vibrios could not be detected in the water column during the winter months, although they were present in sediment. From late spring to early summer, when water temperatures were 14 +/- 1 C, vibrios over-wintering in sediment were released from the bottom communities and attached to zooplankton, proliferating as the temperature rose. The number of vibrios in and on plankton was reflected in the water column bacterial population densities at water temperatures of ca. 19 C. Thus, temperature of the water column in the range of 14 to 19 C was found to be critical in the annual cycle of the vibrios. Interaction between sediment, water, and zooplankton was found to be essential in the natural estuarine ecosystem. Bacterial counts of zooplankton were found to be temperature dependent. The bacterial population associated with zooplankton was found to be predominantly on external surfaces and was specific, differing from that of the sediment. Vibrio spp. and related organisms comprised the total bacterial population associated with zooplankton in summer months. The ecological role of Vibrio spp., including V. parahaemolyticus, was found to be significant, with respect to their property of chitin digestion and in relation to the population dynamics of zooplankton in Chesapeake Bay.

Decomposition of Chitin by Chytriomyces Species

Various fungi were examined for evidence of ability to digest chitin. A phycomycete, Chytriomyces, produced obvious clear zones when grown on chitin agar and was selected for more detailed study. C...

Soil Oribatei VI. Protoribates lophotrichus (Acarina: Haplozetidae) and its Associations with Microorganisms1

This soil nrihatid feeds specifically upon decaying parenchymatous leaf tissue rich in living microorganisms and derives most of its sustenance from the latter. Its eggs are deposited in such tissue and the emergent larvae consume either the same foodstuff or the excrement of the adults. Approximately 5 months are required for the development of the adult from the egg stage. Microorganisms isolated from adults after a 10-day fast resemble the Brevibacteriaceae and Achromobacteriaceae in characteristics and do not participate in the digestion of cellulose, chitin, or lignin. They may aid digestion in some undetermined way but more probably serve as food.

An investigation of chitinase activity in cockroach and termite extracts

The conditions for an unequivoval demonstration of chitinase activity in insect extracts have been investigated. The breakdown of undenatured chitin by cockroach preparations (blood, digestive juice, cuticle brei, extract of cast skins, saliva) and by an extract of whole termites has been demonstrated. The pH optimum for cockroach digestive chitinase lies between 5·4 and 6·0 and the products of digestion of chitin with this preparation were N-acetylglucosamine (75 per cent), probably its dimer and trimer, and a trace of glucosamine. Only N-acetylglucosamine and a trace of glucosamine were produced by the action of other cockroach preparations. Deacetylase activity could not be detected in the cockroach preparations.

Inability of Varied-Carpet Beetle Larvæ (Anthrenus verbasci L.) to digest Chitin

A FEW animals are known to possess in their digestive juices a chitinase enabling them to digest chitin. The occurrence of a chitinase in the gut of the snail (Helix pomatia L.) was demonstrated long ago1. A similar enzyme is present in the intestinal tract of different earthworms2, eelworms3, snails and slugs4, and in the terrestrial isopod Porcellio scaber Latr.5. Among the snails, and probably also other terrestrial pulmonate gastropods, the intestinal chitinase is not produced by the glands of the animal, but appears to be of microbial origin4.

Vertical Distribution of Bacteria in Marine Sediments

Off the coast of Southern California bacteria are more abundant in marine sediments than in the overlying water, there being thousands to millions per gram of sediment and only a few hundred or less per milliliter of water. The distribution of bacteria in the sediment is more or less independent of the depth of the overlying water, the temperature of the ocean floor, and the distance from mainland. The bacterial population seems to be influenced more greatly by the organic content of the sediments than by other factors. Vertical sections from cores 40 to 75 cm. in length show that the number of bacteria in the sediments decreases greatly with depth in the upper 5 cm. of the core and more slowly beyond. In the surface slime, aerobes are 5 to 100 times more plentiful than a aerobes, but at depths exceeding 15 centimeters about as many anaerobes as aerobes are present. Both anaerobes and aerobes have been demonstrated at all depths analyzed. The oxidation-reduction potential (intensity factor expressed as Eh) increases with core depth, although the oxygen-absorbing capacity decreases with depth. Both properties are ascribed primarily to bacterial activity. Bacteria recovered from marine sediments exhibit many biochemical processes such as ammonification, proteolysis, nitrate reduction, nitrification, urea fermentation, chitin digestion, methane production, fat hydrolysis, cellulose decomposition, glucose, xylose, and arabinose fermentation and starch hydrolysis.