Selection of bacterial antagonists for the biological control of Botrytis cinerea in apple (Malus domestica) and in comparison with application of thiabendazole.

In this study fifteen strains of identified Pseudomonas fluorescens and Bacillus subtilis were investigated for biological control activity against Botrytis cinerea. P. fluorescens P-35 and B. subtilis B-16 showed the most inhibitory zone in dual culture assay against B. cinerea, In vitro. After ten days, P. fluorescens P-5 and B. subtilis B-3 showed the considerable results against B. cinerea on apple fruits and could reduce the grey mould from 100% to less than 35%. After twenty days, P. fluorescens P-5 and 8. subtilis B-16 decreased the disease from 100% to less than 65%. Also, application of thiabendazol at 1500 mg/litre was more effective and could reduce the disease from 100% to 30% and 60%, after 10 and 20 days respectively. Results indicated that there is no significant difference among the treatments (thiabendazol and bacterial strains). So, bacterial strains could not only control the disease but also be a reliable replacement instead of Thiabendazol.

Expression and secretion of the protective antigen of Bacillus anthracis in Bacillus brevis.

We used the Bacillus brevis-pNU212 system to develop a mass production system for the protective antigen (PA) of Bacillus anthracis. A moderately efficient expression-secretion system for PA was constructed by fusing the PA gene from B. anthracis with the B. brevis cell-wall protein signal-peptide encoding region of pNU212, and by introducing the recombinant plasmid, pNU212-mPA, into B. brevis 47-5Q. The clone producing PA secreted about 300 microg of recombinant PA (rPA) per ml of 5PY-erythromycin medium after 4 days incubation at 30 degrees C. The rPA was fractionated from the culture supernatant of B. brevis 47-5Q carrying pNU212-mPA using ammonium sulfate at 70% saturation followed by anion exchange chromatography on a Hitrap Q, a Hiload 16/60 Superdex 200 gel filtration column and a phenyl sepharose hydrophobic interaction column, yielding 70 mg rPA per liter of culture. The N-terminal sequence of the purified rPA was identical to that of native PA from B. anthracis. The purified rPA exhibited cytotoxicity towards J774A.1 cells when combined with lethal factor. The rPA formulated in either Rehydragel HPA or MPL-TDM-CWS adjuvant (Ribi-Trimix) elicited the expression of a large amount of anti-PA and neutralizing antibodies in guinea pigs and completely protected them against a 100 LD50 challenge with fully virulent B. anthracis spores.

Paenibacillus macerans pseudobacteremia resulting from contaminated blood culture bottles in a neonatal intensive care unit.

Paenibacillus species are gram-positive, rod-shaped, spore-forming aerobes that are abundant in nature and closely related to Bacillus. Between June 24 and June 30, 1999, 8 neonates in our neonatal intensive care unit had positive blood cultures for Paenibacillus macerans. This cluster of positive blood cultures with an unusual pathogen suggested a pseudoepidemic. Investigation revealed that the most likely etiology of the pseudobacteremia was environmental contamination of the rubber stoppers in blood culture bottles. This was confirmed by environmental sampling and simulated inoculation studies. This pseudobacteremia outbreak highlights the importance of adhering to well-established methods for blood culture collection and ongoing infection control surveillance.

Management of suspected nosocomial infection: an audit of 19 hospitalized patients with septicemia caused by Bacillus species.

From April to August of 2000, Bacillus spp. were detected in the blood culture of 29 patients in a hospital in Japan. Of these patients, 19 had clinical signs of septicemia; positive culture in the remaining 10 patients was attributed to contamination with skin flora at the site of puncture. Of the 18 strains evaluated, 15 were Bacillus cereus, 2 were Bacillus subtilis, and one was Bacillus licheniformis. The only hospital death observed was that of a patient who had no clinical signs of septicemia at the time of blood sampling. That death is now considered attributable to the underlying neoplasm. The hospital committee for prevention of nosocomial infection concluded after a critical review of the patient records that the cause of septicemia in most cases had been contaminated intravenous lines. To control the situation, the committee recommended the use of a new skin disinfectant, and medical personnel were advised to avoid infusion pauses with interruption of intravenous lines and to replace the caps for the stopcocks with new ones each time the caps were removed. These measures were rigorously observed in addition to the conventional measures for preventing catheter sepsis, and the incidence of septicemia due to the Bacillus spp. declined dramatically thereafter.

Thermal inactivation of Bacillus cereus spores formed at different temperatures.

The effects of the sporulation temperature in the range 20-45 degrees C on the D and z values of three isolates of Bacillus cereus (ATCC 4342, 7004 and 9818) were investigated. The strains were found to differ in their response. Higher D100 values (around 10-fold) were obtained with isolates 4342 and 9818 when the sporulation temperature increased from 20 to 45 degrees C. With isolate 7004 (the least heat resistant of the three strains), however, the most thermal tolerant spores were obtained at 35 degrees C. The z values were not significantly modified (P > 0.05) by the sporulation temperature. Mean z values of 7.46+/-0.22 degrees C for isolate 4342, 7.80+/-0.40 degrees C for 7004 and 8.09+/-0.33 degrees C for 9818 were obtained.

Chlorhexidine gluconate versus chloroxylenol for preoperative skin preparation in dogs.

The efficacy of 3% chloroxylenol (PCMX) or 4% chlorhexidine gluconate (CG) for preoperative skin preparation was assessed in 100 dogs undergoing clean or clean-contaminated surgical procedures. Replication Organism Detection and Counting (RODAC) plates were used to quantify skin bacteria colony forming units (CFU) at the operative site before and after skin preparation and immediately postoperatively. Reduction of CFU after skin preparation and immediately postoperatively was significant for each agent. However, CFU levels were significantly lower in the CG group than in the PCMX group after surgical preparation, regardless of initial CFU numbers. No significant difference in CFU counts was observed between antiseptic groups postoperatively. Within-group comparisons showed PCMX to be significantly less efficacious when the prescrub CFU number was greater than 1,000. Bacterial reduction was similar in the CG group regardless of prescrub CFU levels. The number of negative cultures after skin preparation was significantly greater with CG than with PCMX. Chlorhexidine gluconate also had fewer cultures with heavy bacterial growth (> 5 CFUs) after surgical preparation. There was no significant difference between antiseptics in the number of negative cultures or cultures with more than 5 CFUs immediately after surgery. The number of skin reactions and postoperative wound infections that occurred with each technique were similar. Three percent PCMX, as used in this study, was less effective than 4% CG in its immediate antimicrobial activity, however, this difference was not associated with an increased wound infection rate.

Laboratory and field studies on the effects of the antibiotic tylosin on honey bee Apis mellifera L. (Hymenoptera: Apidae) development and prevention of American foulbrood disease.

Laboratory and field studies were conducted to determine the effectiveness of the antibiotic tylosin in preventing and controlling infections of American foulbrood disease (AFB) of honey bees. Studies conducted on immature worker bees maintained in the laboratory revealed that honey bee larvae could tolerate quite a range of doses of antibiotic in their diet. Intermediate doses of tylosin protected very young larvae from becoming infected by Bacillus larvae at a concentration of 1.5 x 10(8) spores/ml of diet. Antibiotic treatment had no measurable effects on larval or pupal developmental rates until the dose reached a lethal level. Bees in field colonies readily consumed tylosin in powered sugar, up to a level of 800 mg/7 g sugar. No negative colony effects were noted at any dosage rates. Protection against infection by American foulbrood was compared to results obtained with 200 mg Terramycin, the standard dose of the only substance currently registered for foulbrood control. Both 200 mg Terramycin and 100 mg tylosin protected the colonies for up to 3 weeks. A 200-mg dose of tylosin protected the colony for an additional week. Doses of 100 mg or more of tylosin were adequate to eliminate signs of AFB infection in overtly diseased colonies.