Development of specific PCR primers for identification and detection of <Emphasis Type="Italic">Phytophthora capsici</Emphasis> Leon

A PCR-based method was developed for the identification and detection of Phytophthora capsici in pepper plants. Three PCR primers (CAPFW, CAPRV1 and CAPRV2) specific for P. capsiciwere designed based on the sequence of its internal transcribed spacer regions. CAPFW/CAPRV1 amplify a 452 bp product from P. capsici DNA whereas CAPFW/CAPRV2 a 595 bp fragment; neither set amplifies DNA from pepper or several fungi pathogenic to pepper. In conventional (single-round) PCR, the limit of detection was 5 pg DNA for both primer sets, whereas in nested PCR the detection limit for both was of 0.5 fg. However, when the dilution series of target DNA were spiked with plant DNA, amplification declined two-fold in both conventional and nested PCR. The CAPFW/CAPRV2 set in conventional PCR was used to detect P. capsici DNA in inoculated plants. Detection occurred as soon as 8h post-inoculation in stem samples from infected but still symptomless plants. The method was also tested to detect fungal DNA in infected soils.     

Seroprevalence and risk factors for Neospora caninum in sheep in the state Minas Gerais, southeastern Brazil

The aim of this study was to determine the prevalence and risk factors associated with infection due to Neospora caninum in serum samples from 488 sheep originating from 63 farms in 63 municipalities distributed across eight of the twelve mesoregions of the state Minas Gerais, southeastern Brazil. For detection of N. caninum the sheep serum samples were subjected to the indirect fluorescence antibody test (IFAT ≥ 50). To identify the risk factors associated with infection due to N. caninum a questionnaire was filled out for each herd by interviewing, the individual responsible for the herd, demanding information on the general characteristics of the property. Sixty-four sheep sera (13.1%; 95% CI=10.3-16.4) presented IgG-specific anti-N. caninum antibodies with the following titers: 50 (49; 76.6%), 100 (7; 10.9%), 200 (4; 6.2%), 400 (3; 4.7%) and 800 (1; 1.6%). The prevalence of infected sheep per mesoregion ranged from 0 to 28.1%. Out of the 63 farms sampled, 31 (49.2%; 95% CI=36.4-62.1) presented at least one seropositive sheep. No significant association was found between the presence of anti-N. caninum antibodies and the risk factors evaluated on the farms, except for the mesoregion variable (p=0.004; OR=0.429; CI95%=0.182-1.008). These results indicate that there is a need for additional research to define the epidemiological importance of this parasite as a cause of reproductive problems in sheep herds in Minas Gerais.     

Symbiotic N<Subscript>2</Subscript> fixation and nitrate utilisation in irrigated lucerne (<Emphasis Type="Italic">Medicago sativa</Emphasis>) systems

Symbiotic N₂ fixation by lucerne (Medicago sativa) has capacity to provide significant inputs of N to agro-ecosystems, and the species has also been shown to scavenge soil mineral N and thus act as a sink for excess reactive N. The balance between these two N cycle processes was investigated in an extensive irrigated lucerne growing region where nitrate contamination of groundwater has been reported. We sampled 18 permanent pure lucerne stands under irrigation for standing dry matter, total shoot N, and N₂ fixation using ¹⁵N natural abundance along with activity of the inducible enzyme nitrate reductase as indicators of use of soil NO₃⁻ by lucerne. On average 65% of lucerne N was obtained from symbiotic N₂ fixation. Converting standing dry matter estimates to annual N₂ fixation amounts we calculated average N₂ fixation of 311 kg N/ha, including N in roots and nodules. Uptake of N from soil by lucerne was calculated to be 181 kg N/ha/year. We were not able to identify the source of this soil mineral N, although nitrate reductase activity of lucerne was higher than that of non-N₂ fixing species examined.     

Effect of ammonia and nitrite on vigour,survival rate,moulting rate of the blue swimming crab <Emphasis Type="Italic">Portunus pelagicus</Emphasis> zoea

The effects of ammonia and nitrite on vigour, survival rate, moulting rate of zoea of blue swimming crab, Portunus pelagicus, were studied. A total of five nitrite-N treatments (26.67, 53.34, 106.68, 213.36, 426.72 mg/l) and a control (no nitrite-N added) were set up for the acute nitrite-N toxicity experiment; a total of five ammonia-N treatments (8.43, 16.86, 33.72, 67.44, 134.88 mg/l) and a control (no ammonia-N added) were set up for the acute ammonia-N toxicity experiment. The results showed that the vigour, survival rate and moulting rate of zoea of the blue swimming crabs exposed to over 53.34 mg/l were significantly different (P < 0.05) from the control group. The zoea LC₅₀ values (mg/l) of nitrite-N were 179.47, 76.56, 66.70, 37.49, 25.01, 25.35, 25.34 mg/l for 24, 36, 48, 60, 72, 84, 96 h, respectively. The vigour, survival rate and moulting rate of zoea of the blue swimming crabs exposed to over 16.86 mg/l were significantly different (P < 0.05) from the control group. The zoea LC₅₀ values (mg/l) of ammonia-N were 51.04, 39.62, 38.72, 24.43, 16.90, 13.42, 11.16 mg/l for 24, 36, 48, 60, 72, 84, 96 h, respectively. The zoeae are highly sensitive to ammonia and nitrite, and the toxicity of ammonia and nitrite on Portunus pelagicus decrease with development of this crab.