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.     

Anthropogenic influences on potential fire spread in a pyrogenic ecosystem of Florida, USA

Fire has historically been an important ecological factor maintaining southeastern U.S. vegetation. Humans have altered natural fire regimes by fragmenting fuels, introducing exotic species, and suppressing fires. Little is known about how these alterations specifically affect spatial fire extent and pattern. We applied historic (1920 and 1943) and current (1990) GIS fuels maps and the FARSITE fire spread model to quantify the differences between historic and current fire spread distributions. We held all fire modeling variables (wind speed and direction, cloud cover, precipitation, humidity, air temperature, fuel moistures, ignition source and location) constant with exception of the fuel models representing different time periods. Model simulations suggest that fires during the early 1900's burned freely across the landscape, while current fires are much smaller, restricted by anthropogenic influences. Fire extent declined linearly with patch density, and there was a quadratic relationship between fire extent and percent landscape covered by anthropogenic features. We found that as little as 10 percent anthropogenic landcover caused a 50 percent decline in fire extent. Most landscapes (conservation or non-conservation areas) are now influenced by anthropogenic features which disrupt spatial fire behavior disproportionately to their actual size. These results suggest that land managers using fire to restore or maintain natural ecosystem function in pyrogenic systems will have to compensate for anthropogenic influences in their burn planning. This revised version was published online in May 2005 with corrections to the Cover Date. This revised version was published online in July 2006 with corrections to the Cover Date.     

Evaluation of soybean germplasm conserved in NIAS genebank and development of mini core collections

Genetic variation and population structure among 1603 soybean accessions, consisted of 832 Japanese landraces, 109 old and 57 recent Japanese varieties, 341 landrace from 16 Asian countries and 264 wild soybean accessions, were characterized using 191 SNP markers. Although gene diversity of Japanese soybean germplasm was slight lower than that of exotic soybean germplasm, population differentiation and clustering analyses indicated clear genetic differentiation among Japanese cultivated soybeans, exotic cultivated soybeans and wild soybeans. Nine hundred ninety eight Japanese accessions were separated to a certain extent into groups corresponding to their agro-morphologic characteristics such as photosensitivity and seed characteristics rather than their geographical origin. Based on the assessment of the SNP markers and several agro-morphologic traits, accessions that retain gene diversity of the whole collection were selected to develop several soybean sets of different sizes using an heuristic approach; a minimum of 12 accessions can represent the observed gene diversity; a mini-core collection of 96 accession can represent a major proportion of both geographic origin and agro-morphologic trait variation. These selected sets of germplasm will provide an effective platform for enhancing soybean diversity studies and assist in finding novel traits for crop improvement.     

Non-traditional vectors for paralytic shellfish poisoning

Paralytic shellfish poisoning (PSP), due to saxitoxin and related compounds, typically results from the consumption of filter-feeding molluscan shellfish that concentrate toxins from marine dinoflagellates. In addition to these microalgal sources, saxitoxin and related compounds, referred to in this review as STXs, are also produced in freshwater cyanobacteria and have been associated with calcareous red macroalgae. STXs are transferred and bioaccumulate throughout aquatic food webs, and can be vectored to terrestrial biota, including humans. Fisheries closures and human intoxications due to STXs have been documented in several non-traditional (i.e. non-filter-feeding) vectors. These include, but are not limited to, marine gastropods, both carnivorous and grazing, crustacea, and fish that acquire STXs through toxin transfer. Often due to spatial, temporal, or a species disconnection from the primary source of STXs (bloom forming dinoflagellates), monitoring and management of such non-traditional PSP vectors has been challenging. A brief literature review is provided for filter feeding (traditional) and non-filter feeding (non-traditional) vectors of STXs with specific reference to human effects. We include several case studies pertaining to management actions to prevent PSP, as well as food poisoning incidents from STX(s) accumulation in non-traditional PSP vectors.     

Modelling the influence of site and weed competition on juvenile modulus of elasticity in Pinus radiata across broad environmental gradients

Data from a nationwide set of Pinus radiata D. Don plots established at a range of conventional stand densities were analysed at age 6 to (i) determine how environment and competition from weeds influence dynamic modulus of elasticity (E) of the lower stem base, (ii) develop a predictive multiple regression model of E for basal stemwood and (iii) identify significant direct and indirect environmental influences (through stem slenderness) on E using path analysis.Site had a highly significant (P < 0.001) influence on E, which exhibited a 3-fold range from 1.6 to 5.3 GPa, across 30 sites. When compared to the weed-free controls, weed competition had a significant (P < 0.0001) and substantial effect on E, increasing values by on average 16% (2.76 GPa vs. 2.38 GPa).The positive linear relationship between stem slenderness (determined as tree height/ground-line tree diameter) and E was by far the strongest relationship (R² = 0.71; P < 0.001) among the 20 variables that were significantly related to E. A multiple regression model that included stem slenderness, mean minimum air temperature in mid-autumn, T_min, as positive linear relationships and net nitrogen (N) mineralisation in a negative linear form accounted for 86% of the variance in E. A cross-validation indicated that this model was stable and unbiased, with the validation accounting for 82% of the variance in E. The final path analysis model included T_min, net N mineralisation, below canopy solar radiation and stem slenderness as significant (P < 0.05) direct influences on E. Below canopy radiation, maximum air temperature during mid-summer, soil total phosphorus and carbon:nitrogen ratio were indirectly associated with E through their significant (P < 0.05) direct relationship with stem slenderness.These results provide considerable insight into how environment regulates E of juvenile P. radiata. Low fertility sites that have warm air temperatures and either a high canopy leaf area index, or high levels of woody weed competition, are most likely to produce trees with high stem slenderness and high E. Conversely, sites that are cool over summer and autumn and high in fertility, with low levels of intra- or inter-specific competition for light are likely to produce trees with low stem slenderness and low E.     

Detection of Colletotrichum coccodes from soil and potato tubers by conventional and quantitative real-time PCR

Colletotrichum coccodes is the causal agent of the potato blemish disease black dot. Two PCR primer sets were designed to sequences of the ribosomal internal transcribed spacer (ITS1 and ITS2) regions for use in a nested PCR. The genus-specific outer primers (Cc1F1/Cc2R1) were designed to regions common to Colletotrichum spp., and the species-specific nested primers (Cc1NF1/Cc2NR1) were designed to sequences unique to C . coccodes . The primer sets amplified single products of 447 bp (Cc1F1/Cc2R1) and 349 bp (Cc1NF1/Cc2NR1) with DNA extracted from 33 European and North American isolates of C. coccodes. The specificity of primers Cc1NF1/Cc2NR1 was confirmed by the absence of amplified product with DNA of other species representing the six phylogenetic groups of the genus Colletotrichum and 46 other eukaryotic and prokaryotic plant pathogenic species. A rapid procedure for the direct extraction of DNA from soil and potato tubers was used to verify the PCR assay for detecting C. coccodes in environmental samples. The limit of sensitivity of PCR for the specific detection of C. coccodes when inoculum was added to soils was 3·0 spores per g, or the equivalent of 0·06 microsclerotia per g soil, the lowest level of inoculum tested. Colletotrichum coccodes was also detected by PCR in naturally infested soil and from both potato peel and peel extract from infected and apparently healthy tubers. Specific primers and a TaqMan fluorogenic probe were designed to perform quantitative real-time (TaqMan) PCR to obtain the same levels of sensitivity for detection of C. coccodes in soil and tubers during a first-round PCR as with conventional nested PCR and gel electrophoresis. This rapid and quantitative PCR diagnostic assay allows an accurate estimation of tuber and soil contamination by C. coccodes .     

Assessment of the socio-economic impacts associated with the arrival of apple snail (Pomacea canaliculata) in Mwea irrigation scheme,Kenya

Background

In Kenya, rice (Oryza sativa L.) is mainly produced under irrigation by small-scale farmers. Mwea irrigation scheme (MIS) in Kirinyaga County accounts for 80–88% of rice production. Here, rice is the main source of livelihood and a source of revenue generation for the county. However, a recently established invasive freshwater snail, Pomacea canaliculata (Lamarck) (family: Ampullariidae), a species of apple snail, presents a serious threat to rice production.

Results

Household surveys, focus group discussions and key informant interviews highlight apple snail as a serious problem in MIS. Households that observed at least a moderate level of infestation (>20% of cultivated area) experienced significant reductions in rice yield (~14%) and net rice income (~60%). Farmers reported increased use of chemical pesticides for management of apple snail. In addition, the cost of hired labor for physical removal of egg masses and snails is resulting in substantial negative effects on net income. Farmer age, area of land owned, responsibility for decision-making, receipt of extension advice, training, and membership of a farmer organization, were all statistically significant variables to explain farmers awareness of the need for area-wide apple snail management.

Conclusion

Strategies to limit the spread of apple snail are urgently needed. A Multi-Institutional Technical Team (MITT) has been established to spearhead management efforts and consolidate advice to farmers on how to manage apple snail. However, without action to mitigate spread, the consequences could be disastrous for rice production and food security in Kenya, and for other rice growing regions across Africa. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Diagnostic and public health investigation of Mycobacterium tuberculosis infection in a dog in Ontario,Canada

A 4-y-old, female mixed-breed dog was presented to the Ontario Veterinary College for further evaluation of multiple pulmonary and hepatic masses, intrathoracic lymphadenitis, and recent development of a pyogranulomatous pleural effusion. Along with other comprehensive tests, a thoracic lymph node biopsy was performed, and Mycobacterium tuberculosis complex infection was confirmed by real-time PCR. The dog’s condition declined post-operatively, and euthanasia was elected. Postmortem examination confirmed severe granulomatous pneumonia, hepatitis, intrathoracic and intraabdominal lymphadenitis, omentitis, and nephritis. Line-probe assays performed on samples collected postmortem confirmed the species as M. tuberculosis. 24-loci MIRU-VNTR genotyping, spoligotyping, and whole-genome sequencing revealed relations to known human isolates, but no epidemiologic link to these cases was investigated. Given the concern for potential human exposure during this animal’s disease course, a public health investigation was initiated; 45 individuals were tested for M. tuberculosis exposure, and no subsequent human infections related to this animal were identified. Our case highlights the need for more readily available, minimally invasive testing for the diagnosis of canine mycobacteriosis, and highlights the ability of canid species to act as potential contributors to the epidemiology of M. tuberculosis infections.