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Evaluation of a micromethod gallery (API Staph) for the identification of staphylococci and micrococci.

A collection of 300 well-characterised strains of staphylococci and micrococci was examined by a commercially available gallery micromethod (API Staph). The results were compared with biotyping by conventional methods. The gallery micromethod broadly agreed with the biotyping scheme used but gave an identification from the index supplied in less than 30% of the trials. Reproducibility was better after 48 h incubation than after 24 h but was poor for the tests for phosphatase and acetoin. When compared with the results of conventional tests, the tests for acetoin, phosphatase and urea were unsatisfactory.

Bacteriological Techniques↗

Biotyping of Haemophilus using API 10S--an epidemiological tool?

One hundred and ninety-nine strains of Haemophilus isolates were biotyped by Kilian's method(1) and a modified API 10S strip and the results compared. One hundred percent correlation was found between the two systems. The ONPG test proved of value in differentiating between Haemophilus influenzae and Haemophilus parainfluenzae when there was growth factor disc failure.

Adolescent↗

Identification of the Enterobacteriaceae: a comparison of the Enterotube II with the API 20E.

Two commercial methods for the identification of the Enterobacteriaceae, API 20E and Enterotube II, were compared using the results obtained with 235 cultures of fresh clinical isolates. Enterotube II was an improvement on the original Enterotube, but the availability of two differing indices, one using the Voges-Proskauer (VP) test result and one without, is criticised.

Bacteriological Techniques↗

Rapid identification of thermophilic Naegleria, including Naegleria fowleri using API ZYM system.

The suitability of the API ZYM system for identifying thermophilic Naegleria species, based on enzyme presence and activity, was investigated. Replicate testing on strains of N fowleri, N lovaniensis, and N australiensis cultured in a monoxenic and an axenic medium showed that the system could provide a rapid and reproducible means of identifying the species soon after primary isolation. No single enzyme was found specific for any one species, but considerable differences were found in the patterns of activity of acid phosphatase and leucine arylamidase. When these were compared the species could be differentiated. Use of the system in conjunction with a simple culture method is proposed as a readily available means of monitoring environmental and public bathing sites to prevent primary amoebic meningoencephalitis.

Acid Phosphatase↗

Comparison of identification of Enterobacteriaceae by API 20E and Sensititre Autoidentification System.

Of 251 isolates of the Enterobacteriaceae identified to species level by API 20E, 208 (83%) were similarily identified by the Sensititre Autoidentification System. Both systems shared a common problem in that discrimination between species of the genera Klebsiella, Enterobacter, and Serratia was poor. The eight digit biocode generated by the Sensititre system for individual isolates is not reproducible and therefore not of epidemiological value.

Bacteriological Techniques↗

Identification of Pseudomonas aeruginosa with the API-20E system.

The API-20E multitest system was found to be capable of identifying Pseudomonas aeruginosa isolates at least to the level of the Pseudomonas fluorescens group within 18 h at 35 degrees C. Additional tests, such as the acetamide reaction, growth at 42 degrees C, and the oxidative (OF) glucose test, assisted in the speciation of P. aeruginosa.

Bacteriological Techniques↗

Physical, chemical, and enzymatic studies on the major sucrase of honey bees (Apis mellifera).

A sucrase from honey bees (Apis mellifera) which precipitates between ammonium sulfate saturations of 50 and 70% (5 mg protein per millilitre) and which makes up the major portion of the sucrases of honey bees was purified to homogeneity as shown by several criteria. A large part of the sucrase was found in the head while most of the rest was in the abdomen (a small amount was in the thorax). The enzyme precipitated between the same values of ammonium sulfate saturation as did the sucrase in honey and honey sucrase exhibited kinetics very similar to those of this enzyme. The enzyme was found to be a relatively nonspecific alpha-glucosidase and was shown to have transglucosidase activity. The production of glucose from sucrose was rectilinear when plotted by the Hofstee method at low substrate concentrations but decreased at high sucrose concentrations. The production of fructose was rectilinear throughout the concentration range used. The production of both glucose and rho-nitrophenol when rho nitrophenyl alpha-D-glucoside was the substrate was linear by the Hofstee plot. These effects were found to be due to transglucolysis and a mechanism of action is proposed. Amino acid and amino sugar analyses indicated that the sucrase was a glycoprotein. The molecular weight was found to be between 51000 and 82000 by three different methods and an so20.w value of 4.0 S was obtained. There was no evidence for subunit structure. Tests of the enzyme under various denaturation conditions did not reveal any unusual stabilities. The sucrase bound very tightly to a hydrophobic column. Iodoacetic acid decreased the activity of the sucrase but a large concentration was needed to bring about a 50% activity loss. Reducing agents caused some activity declines. Diethyl pyrocarbonate activated the enzyme.

Amino Acids↗

Effect of Apis mellifera propolis from two Brazilian regions on caries development in desalivated rats.

The purpose of the present study was to evaluate the effect of Apis mellifera propolis collected from two regions of Brazil on caries development in desalivated rats. Ethanolic extracts of propolis (EEP) were prepared from crude propolis samples collected in Minas Gerais state (MG), southeastern Brazil, and Rio Grande do Sul state (RS), southern Brazil. The flavonoid composition of EEP was analyzed by high-performance thin-layer chromatography (HPTLC) and reversed-phase high-performance liquid chromatography (HPLC). For the animal study, 30 specific pathogen-free Wistar rats were infected with Streptococcus sobrinus 6715 and surgically desalivated. The rats were randomly divided into three groups which were treated with 80% ethanol (control), EEP from MG and EEP from RS. The animals were placed in a König-Höfer programmed feeder and received 17 meals of diet 2000 daily at hourly intervals. The solutions were applied on the rat molars (25 microl on molars of each quadrant) twice a day, by using graduate syringes. After 3 weeks, the animals were killed by CO(2) asphyxiation. For microbial assessment, the left jaw was removed and sonicated in 154 mM NaCl solution. Dental caries was evaluated according to Larson's modification of Keyes' system. The HPTLC patterns and HPLC profiles demonstrated that both quality and quantity of flavonoid aglycones of EEP from MG were different compared to EEP from RS. In general, it is apparent that EEP from RS contained the highest concentrations of pinocembrin, chrysin, acacetin and galangin. The group of animals treated with EEP from RS showed the lowest smooth-surface and sulcal caries scores as well as less caries severity in smooth-surface and sulcal lesions, and these data were statistically different when compared with the control group. The group treated with EEP from MG only demonstrated a significant difference in the severity of sulcal lesions when compared to the control group. The percentage of S. sobrinus was lower in the groups treated with EEP, but did not differ statistically from the control group. The results showed that the cariostatic effect of propolis depends on its composition, and consequently the region of collection of propolis samples.

Analysis of Variance↗

Effects of Apis mellifera propolis on the activities of streptococcal glucosyltransferases in solution and adsorbed onto saliva-coated hydroxyapatite.

Propolis, a resinous hive product collected by Apis mellifera bees, has been used for thousands of years in folk medicine. Ethanolic extracts of propolis (EEP) have been shown to inhibit the activity of a mixture of crude glucosyltransferase (Gtf) enzymes in solution. These enzymes synthesize glucans from sucrose, which are important for the formation of pathogenic dental plaque. In the present study, the effects of propolis from two different regions of Brazil on the activity of separate, purified Gtf enzymes in solution and on the surface of saliva-coated hydroxyapatite (sHA) beads were evaluated. The EEP from Minas Gerais (MG; Southeastern Brazil) and Rio Grande do Sul (RS; Southern Brazil) were tested for their ability to inhibit the enzymes GtfB (synthesis of insoluble glucan), GtfC (insoluble/soluble glucan) and GtfD (soluble glucan). The effects of propolis on Gtf from Streptococcus sanguis (soluble glucan synthesis) was also explored. The EEP from both regions effectively inhibited the activity of all Gtfs in solution (75-95%) and on the surface of sHA beads (45-95%) at concentrations between 0.75 and 3.0 mg of propolis/ml. However, the two samples of propolis showed different levels of inhibition on each of the enzymes tested. In general, EEP RS demonstrated a significantly higher inhibitory activity on GtfB and C activities (both solution and surface assays) than EEP MG at concentrations between 0.047 and 0.187 mg/ml (p<0.05). EEP MG, on the other hand, exhibited a greater inhibitory effect on the activities of surface GtfD (at 0.375, 0.75 and 1.5 mg/ml) and S. sanguis Gtf (at 1.5 and 3.0 mg/ml; p<0.05). These data indicate that EEP is a potent inhibitor of Gtf enzymes in solution and adsorbed on an experimental pellicle; however, its effect on Gtf activity is variable depending on the geographical origin of the propolis samples. There is a need to identify the active compounds of propolis.

Bacterial Proteins↗

Detoxified hymenoptera venoms: preliminary studies of in vitro cytotoxicity and antigenicity of Apis mellifera 'venomoid'.

The preparation of detoxified venom (venomoid) of Apis mellifera is described. The venomoid obtained by formaldehyde detoxification retained significant antigenicity. Radioallergosorbent test (RAST) inhibition studies indicated a loss of the affinity of the venomoid for the venom-specific human IgE antibody. In rabbits the venomoid was capable of producing precipitating antibodies which cross-reacted with the original venom. Cytotoxic activity of venomoid as measured by nonimmunological histamine release and hemolytic activity with human cells was not detectable in vitro. Further studies seem warranted for the establishment of a venomoid as a diagnostic and/or therapeutic agent in the management of the hypersensitivity reactions to venom.

Animals↗

Mechanism of action of honey bee (Apis mellifera L.) venom on different types of muscles.

1. The effect of crude honeybee (Apis mellifera) venom on the skeletal, smooth as well as cardiac muscles were studied in this investigation. 2. Perfusion of gastrocnemius-sciatic nerve preparation of frogs with 1 microgram/ml venom solution has weakened the mechanical contraction of the muscle without recovery. Blocking of nicotinic receptors with 3 micrograms/ml flaxedil before bee venom application sustained normal contraction of gastrocnemius muscle. 3. The electrical activity of duodenum rabbits was recorded before and after the application of 1 microgram/ml venom solution. The venom has depressed the amplitude of the muscle contraction after 15 min pretreatment with atropine nearly abolished the depressor effect of the venom on smooth muscle. 4. In concentrations from 0.5-2 micrograms/ml, bee venom caused decrease of heart rate of isolated perfused toad heart. This bradycardia was accompanied by elongation in the P-R interval. A gradual and progressive increase in the R-wave amplitude reflected a positive inotropism of the venom. Application of 5 micrograms/ml verapamil, a calcium channels blocking agent, abolished the noticed effect of the venom. 5. Marked electrocardiographic changes were produced within minutes of the venom application on the isolated perfused hearts, like marked injury current (elevation or depression of the S-T segment), atrioventricular conduction disturbances and sinus arrhythmias. Atropine and nicotine could decrease the toxic effect of the venom on the myocardium. 6. Results of the present work lead to the suggestion that bee venom is mediated through the peripheral cholinergic neurotransmitter system. General neurotoxicity of an inhibitory nature involving the autonomic as well as neuromuscular system are established as a result of the venom, meanwhile a direct effect on the myocardium membrane stabilization has been suggested.

Animals↗

Occurrence of intercellular bridges between follicle epithelial cells in the ovary of Apis mellifica queens.

Hitherto unknown intercellular bridges or fusomes between the follicle epithelial cells investing the oocytes of Apis mellifica queens have been observed both with light and electron microscopy. Usually each follicle cell has 2-3 intercellular bridges. In surfacial paraffin sections, the intercellular bridges can be seen to connect a series of follicle cells which may be branching. The intercellular bridges lie close to the egg cortex and this position is relatively constant. The width of the fusomal ring canal varies in different developmental stages. In stages 3 and 4 of oogenesis, which are the main vitellogenic stages, the intercellular bridges measure 0-5 micron, while in stages 1 and 2 they have a diameter ranging from 1-5 to 3-5 micron. In these stages the intercellular bridges are provided with numerous transverse microfilaments which disappear later. The fusomal lips are thickened and consist of electron-dense material and an additional layer of less electron-dense material both inside and outside. Ribosomes flow across the bridge. The intercellular bridges may serve to synchronize the differentiation and functional activity of the follicle epithelium during the course of oogenesis.

Animals↗

Motor pattern of the stinging response in the honeybee Apis mellifera

In the stinging response of the worker honeybee (Apis mellifera), rhythmic movements of the lancets on the stylet are produced by alternating contractions of a set of stinging muscles (a protractor, M198, and a retractor, M199) on each side during co-contraction of the frucula muscles (M197s) on both sides. In this study, stinging movements were elicited by tactile stimulation to the sternum in isolated abdomens, in intact animals and in preparations in which the connectives between the sixth and terminal abdominal ganglia were cut. There was a close relationship among the following three temporal variables of stinging motoneurone pattern: the interval between successive bursts of a stinging muscle, the duration of a burst and the time lag between the bursts of homologous stinging muscles on both sides. All of these variables increased linearly as the sting was inserted deeper into a soft object and the tension on the lancets increased. When sensory nerves from the proprioceptors (campaniform sensilla on the tapering sting shaft and hair plates at the basal cuticular plate) were cut on both sides, the relative timing of bursts of homologous stinging muscles on both sides and antagonistic stinging muscles on each side became more variable. When a proprioceptive input was removed from one side during penetration of the sting, the frequency of the bursts of stinging muscles was higher and the duration of bursts was shorter on the cut side than on the intact side; nevertheless, a sting muscle was still activated out of phase with its antagonistic muscle on the ipsilateral side and its homologous muscle on the contralateral side. These results suggest that the motor pattern driving the rhythmic movements of stinging muscles is produced by a central pattern generator consisting of a pair of oscillators located in the terminal abdominal ganglion and that the precise timing of the motor pattern in a hemiganglion is controlled mainly by proprioceptive inputs on its own side.

Journal Article↗

Mechanisms of thermal stability during flight in the honeybee apis mellifera

Thermoregulation of the thorax allows honeybees (Apis mellifera) to maintain the flight muscle temperatures necessary to meet the power requirements for flight and to remain active outside the hive across a wide range of air temperatures (Ta). To determine the heat-exchange pathways through which flying honeybees achieve thermal stability, we measured body temperatures and rates of carbon dioxide production and water vapor loss between Ta values of 21 and 45 degrees C for honeybees flying in a respirometry chamber. Body temperatures were not significantly affected by continuous flight duration in the respirometer, indicating that flying bees were at thermal equilibrium. Thorax temperatures (Tth) during flight were relatively stable, with a slope of Tth on Ta of 0.39. Metabolic heat production, calculated from rates of carbon dioxide production, decreased linearly by 43 % as Ta rose from 21 to 45 degrees C. Evaporative heat loss increased nonlinearly by over sevenfold, with evaporation rising rapidly at Ta values above 33 degrees C. At Ta values above 43 degrees C, head temperature dropped below Ta by approximately 1-2 degrees C, indicating that substantial evaporation from the head was occurring at very high Ta values. The water flux of flying honeybees was positive at Ta values below 31 degrees C, but increasingly negative at higher Ta values. At all Ta values, flying honeybees experienced a net radiative heat loss. Since the honeybees were in thermal equilibrium, convective heat loss was calculated as the amount of heat necessary to balance metabolic heat gain against evaporative and radiative heat loss. Convective heat loss decreased strongly as Ta rose because of the decrease in the elevation of body temperature above Ta rather than the variation in the convection coefficient. In conclusion, variation in metabolic heat production is the dominant mechanism of maintaining thermal stability during flight between Ta values of 21 and 33 degrees C, but variations in metabolic heat production and evaporative heat loss are equally important to the prevention of overheating during flight at Ta values between 33 and 45 degrees C.

Journal Article↗

Antennal malformations in light ocelli drones of Apis mellifera (Hymenoptera, Apidae).

Malformed antennae of Apis mellifera light ocelli drones were drawn, dissected and mounted permanently on slides containing Canada balsam, in order to count the olfactory discs present in each segment, in comparison with the number of those structures in normal antennae of their brothers. Some drones presented morphological abnormalities in a single segment of the right or left antenna, but others had two or more malformed segments in a same antenna. Drones with malformations in both antennae were also observed. The 4th and 5th flagellum segments were the most frequently affected. In a low number of cases the frequency of olfactory discs in malformed segments did not differ from that one recorded for normal segments. However, in most cases studied, the antennal malformations brought about a significant reduction in the number of olfactory discs from malformed segments.

Animals↗

Morphometrics and adaptatives aspects in africanized honeybees (Apis mellifera).

The introduction of the African bees (Apis mellifera scutelata) in Brazil and their expansion in the American Continent created the opportunity to study the process of species adaptation in a new environment. In that process, within intra-specific variability, normalising selection can favour individuals that present a better adaptative morphology and they will constitute the most frequent type found in the population. To test that hypothesis morphometric analyses in samples of colonies of africanized bees and in samples of the populations were performed. The development of the colonies was also evaluated in terms of the amount of their brood, honey and pollen. Analysis of the data indicates that more developed colonies are formed by individuals closer to the population average with concerning morphological traits.

Journal Article↗

Effective fall treatment of Varroa jacobsoni (Acari: Varroidae) with a new formulation of formic acid in colonies of Apis mellifera (Hymenoptera: Apidae) in the northeastern United States.

New formulations of formic acid and thymol, both individually and in combination with various essential oils, were compared with Apistan to determine their efficacy as fall treatments for control of Varroa jacobsoni (Oudemans), a parasitic mite of the honey bee, Apis mellifera L. Percent mite mortality in colonies treated with 300 ml of 65% formic acid averaged 94.2 +/- 1.41% (least square means +/- SE, n = 24), equivalent to those receiving four, 10% strips of Apistan (92.6 +/- 1.79%, n = 6). Treatment with thymol (n = 24) resulted in an average mite mortality of 75.4 +/- 5.79%, significantly less than that attained with Apistan or formic acid. The addition of essential oils did not affect treatment efficacy of either formic acid or thymol. The ratio of the coefficients of variation for percentage mortality for the formic acid (CVFA) and Apistan (CVA) groups was CVFA/CVA = 0.66. This indicates that the formic acid treatment was as consistent as the Apistan treatment. Thymol treatments did not provide as consistent results as Apistan or formic acid. Coefficient variation ratios for percentage mortality for the thymol group (CVT) with the Apistan and formic acid groups were CVT/CVA = 4.47 and CVT/CVFA = 6.76, respectively. In a second experiment, colonies received a 4-wk fall treatment of either 300 ml of 65% formic acid (n = 24) or four, 10% strips of Apistan (n = 6). The next spring, mite levels in the formic acid group (554.3 +/- 150.20 mites) were similar to those in the Apistan treatment group (571.3 +/- 145.05 mites) (P = 0.93). Additionally, the quantities of bees, brood, pollen, and nectar/honey in the two treatment groups were not significantly different (P > or = 0.50 each variable). These results suggest that formic acid is an effective alternative to Apistan as a fall treatment for varroa mites in temperate climates.

Animals↗