Digital Image Analysis of Internal Light Spots of Appressoria of Colletotrichum acutatum

ABSTRACT The initial penetration process of appressoria of Colletotrichum acutatum on almond leaves was studied using digital image analysis of light micrographs and scanning electron microscopy. For image analysis, a series of sequential, partially focused digital micrographs of appressoria was analyzed to generate a single, completely focused montage image with a continuous in-focus depth of field. In studies on the development of the internal light spot (ILS), we observed that 50.4% of the appressoria formed an ILS after leaves were inoculated and incubated for 12 h at 20 degrees C, and that this increased to 95.8% after 24 h. Comparative image analyses of appressoria with and without ILSs using depth relief mapping and line profile software options showed that the ILS had a depth relief that was below that of the leaf surface. Depth relief analysis in the ILS region during incubation revealed an increase in depth in this area of up to 1.8 mum in some of the appressoria. A comparative morphological study of the ILS in montage images and the penetration pore of appressoria in scanning electron micrographs showed similar shapes and dimensions of the two structures in the appressorium. Light micrographs of histological sections confirmed fungal penetration and internal vesicle formation in almond leaves within 36 h after inoculation and incubation at 20 degrees C. This study represents the first direct evidence that the ILS in appressoria corresponds to the penetration pore and the developing penetration peg using a rapid, digital image analysis technique.     

Effect of rice growth stages and silicon on sheath blight development

ABSTRACT The objective of this study was to determine the effect of silicon (Si) and rice growth stages on tissue susceptibility to sheath blight (Rhizoctonia solani Kühn) under controlled conditions. Rice plants (cv. Rio Formoso) were grown in pots containing low-Si soil amended with Si at 0, 0.48, 0.96, 1.44, and 1.92 g pot(-1) and inoculated with R. solani at the following days after emergence: 45 (four-leaf stage), 65 (eight-leaf stage), 85 (tillering), 117 (booting), and 130 (panicle exsertion). For plants inoculated with R. solani at all growth stages, Si concentration in straw increased as rate of Si increased from 0 to 1.92 g pot(-1). Concentration of calcium in the straw did not differ among plant growth stages. Although incubation period was not affected by the amount of Si added to the soil, this variable was shorter at booting and panicle exsertion stages. As the rates of Si increased in the soil, the total number of sheath blight lesions on sheaths and total area under the relative lesion extension curve decreased at all plant growth stages. The severity of sheath blight was lower at booting and panicle exsertion stages as the rates of Si increased in the soil. In general, plants grown in Si-nonamended pots and inoculated with R. solani were more vulnerable to infection at all growth stages, but especially at 45 days after emergence. Plant dry weights for inoculated plants increased as the Si rates increased from 0 to 1.92 g pot(-1). The greatest dry weight increases occurred for plants inoculated at booting and panicle exsertion stages. Si fertilization is a promising method for controlling sheath blight in areas where soil is Si deficient and when cultivars that exhibit an acceptable level of resistance to sheath blight are not available for commercial use.     

Serum concentrations of pepsinogen A in healthy dogs after food deprivation and after feeding

OBJECTIVE: To develop and validate an ELISA for measurement of serum canine pepsinogen A (cPG A) as a diagnostic marker of gastric disorders in dogs and to measure serum cPG A in healthy dogs after food deprivation and after feeding. SAMPLE POPULATION: Sera from 72 healthy dogs. PROCEDURE: A sandwich ELISA was developed and validated. The reference range for serum concentrations of cPG A was determined in 64 healthy dogs. Postprandial changes in serum concentrations of cPG A were evaluated in 8 healthy dogs. RESULTS: Assay sensitivity was 18 microg/L, and the maximum detectable concentration was 1,080 microg/L. The observed-to-expected ratio (O:E) for 3 serial dilutions of 3 serum samples ranged from 69.3 to 104.1%. The O:E for 3 serum samples spiked with 8 concentrations of cPG A ranged from 58.8 to 120.4%. Coefficients of variation for intra- and interassay variability of 3 serum samples ranged from 7.6 to 11.9% and from 10.1 to 13.1%, respectively. Mean +/- SD serum concentration of cPG A in healthy dogs was 63.8 +/- 31.0 microg/L and the reference range was 18 to 129 microg/L. Significant increases in serum concentrations of cPG A were observed between 1 and 7 hours after feeding. CONCLUSIONS AND CLINICAL RELEVANCE: The ELISA for measuring cPG A was sufficiently sensitive, linear, accurate, precise, and reproducible for clinical use. Serum concentrations of cPG A increase substantially after feeding, which should be taken into account when conducting clinical studies.     

Usefulness of serological examinations in the analysis of Salmonella infections in pig herds

The development of the antibody concentration against lipopolysaccharide (LPS) of S. Typhimurium und S. Choleraesuis in rearing pigs during the fattening period and in breeding sows of the corresponding age was recorded. The studies revealed the following results. Antibodies of isotypes IgG1 and IgG2 revealed a more pronounced specificity against the according Salmonella serovar than IgM antibodies. The calculated "antibody percent value" based on the total amount of Salmonella antibodies is mainly determined by the IgM antibodies in sera and meat juice, respectively. In fattening pigs a significant increase of antibodies against IgM and total Ig was observed between week 3 and 10 after beginning of the rearing period. In breeding pigs this increase was detectable already earlier. In only 3 out of 10 groups an increase of IgG1 and IgG2 was also seen. The detected significant increase of total Ig and IgM in the other groups might be the result of a less intensive exposure to salmonellas or it might be due to an increase of unspecific antibodies induced by other antigens. Serological investigations represent a valuable tool to record the intensity and development in time of the Salmonella exposure in pigs farms. Examination of total Ig is an appropriate method to detect pig herds with a high level of Salmonella exposure, for detailed epidemiological studies in pig farms the examination of antibody isotypes will give more comprehensive information.     

Canine brucellosis--a case report

Brucella (B.) canis was isolated from ejaculate of a 4-year old Korthals-Griffon male dog after occurrence of epididymitis and orchitis. Despite several trials of therapy with different antibiotics relapes occurred, with B. canis being isolated from ejaculate, blood and urine samples, respectively. Bacteriological examinations were added by serological testing over a period of about 1.5 years. During the study SAT serum titre steadily dropped from 1:200 to 1:50. By CFT, B. canis antibodies were detectable at the beginning with a titre of 1:320 and to the end of the study with titres between 1:80 and 1:160.     

Effects of butyrate on apoptosis in the pig colon and its consequences for skatole formation and tissue accumulation

Evidence exists that butyrate inhibits apoptosis of colon crypt cells in vivo so that less tryptophan from cell debris is available for skatole formation by microbes in the pig colon. In this study, potato starch containing a high proportion of resistant starch was fed to test the hypothesis that increased butyrate formation will occur in the colon and contribute to reduced epithelial cell apoptosis, thus leading to reduced skatole formation and absorption. Two groups of six barrows were provided with catheters in the jugular vein and fed either a ration with pregelatinized starch (high ileal digestibility; controls) or potato starch (low ileal digestibility; PS) as the main carbohydrate. All pigs were fed 31 MJ of metabolizable energy and 381 g of crude protein per day. The controls were fed for 19 d. The PS group received the same control ration for 10 d, and then changed to the PS ration. The total feeding period of PS consisted of a 5 d adaptation period followed by another 19 d. In the continously sampled feces, pH, short chain fatty acids, and skatole were determined. Skatole was additionally measured in blood plasma that was sampled daily. After killing barrows at the end of the feeding period, fat tissue for skatole measurement and colon tissue for histological quantification of mitosis and apoptosis were obtained. Feeding potato starch led to a rapid 2.2 fold increase of fecal butyrate when compared both with the control period of the PS group and the control group (P < 0.001). PS feeding resulted in a decrease in pH from 7.3 to 5.3 (P < 0.001) and apoptosis from 2.06 cells/crypt to 0.90 cells (P < 0.01), whereas there was no change in mitosis. Consequently, skatole decreased both in feces (controls vs PS group: 120.0 vs 1.9 microg/g; P < 0.001) and in blood plasma (1.6 vs 0.2 ng/mL; P < 0.001). The mean concentration of skatole in fat tissue was 167 ng/g tissue in controls, and below the detection limit (0.8 ng/g) in the PS group (P < 0.001). It is concluded that butyrate-dependent inhibition of apoptosis in the colon due to potato starch feeding efficiently inhibits skatole production in barrows. Because of the depressed skatole levels, improved sensory quality of pork is possible.     

Effects of probiotics: alone and in a mixture of Biosacc plus Zinc Bacitracin on the caecal microflora of Japanese quail

The aim of this study was to investigate the effects of two commercially available probiotics, alone and in combination with an antibiotic, on the caecal flora of Japanese quail (Coturnix coturnix japonica) reared under unstressed conditions. Thirty-four 90-day-old Japanese quail were selected for this study. The birds were divided into four groups, two groups of nine birds and two groups of eight birds. The animals in these groups were given feed containing 0.5 kg per tonne of probiotics alone (Protexin or Biosacc), a mixture of probiotic plus antibiotic (Biosacc plus Zinc Bacitracin at 0.5 and 0.0525 ppm, respectively), and a group where no supplement was added to be used as controls. The total count of aerobic bacteria, lactobacilli, enterobacteriaceae, coliforms, enteroccoci, salmonellae, sulphite-reducing anaerobic bacteria (clostridia), and pH values in the caecal content of the birds were examined. No significant differences were detected among the four groups for pH values and bacterial number (p>0.05), except for sulphite-reducing anaerobic bacteria (p<0.001). These results suggest that the use of probiotics alone and/or a mixture of a probiotic plus antibiotic as a feed supplement does not have a major suppressing effect on the majority of bacterial groups in the caecal flora of mature, healthy Japanese quail reared in unstressed conditions.