P. S. Ojiambo

Distribution and toxigenicity of Aspergillus species isolated from maize kernels from three agro-ecological zones in Nigeria

Maize samples were collected during a survey in three agro-ecological zones in Nigeria to determine the distribution and aflatoxin-producing potential of members of Aspergillus section Flavi. The three agro-ecological zones were, Derived Savannah (DS) and Southern Guinea Savannah (SGS) in the humid south and North Guinea Savannah (NGS) in the drier north. Across agro-ecological zones, Aspergillus was the most predominant fungal genera identified followed by Fusarium with mean incidences of 70 and 24%, respectively. Among Aspergillus, A. flavus was the most predominant and L-strains constituted >90% of the species identified, while the frequency of the unnamed taxon S(BG) was <3%. The incidence of atoxigenic strains of A. flavus was higher in all the districts surveyed except in the Ogbomosho and Mokwa districts in DS and SGS zones, respectively, where frequency of toxigenic strains were significantly (P<0.05) higher than that of atoxigenic strains. The highest and lowest incidence of aflatoxin positive samples was recorded in the SGS (72%) and NGS (20%), respectively. Aflatoxin contamination in grain also followed a similar trend and the highest mean levels of B-aflatoxins were detected in maize samples obtained from Bida (612 ng g(-1)) and Mokwa (169 ng g(-1)) districts, respectively, in the SGS. Similarly, the highest concentrations of G-aflatoxins were detected in samples from Akwanga district in the SGS with a mean of 193 and 60 ng g(-1), respectively. When agro-ecological zones were compared, B-aflatoxins were significantly (P<0.05) higher in SGS than in NGS, and intermediate in maize samples from the DS agro-ecological zone.

Evaluation of atoxigenic isolates of Aspergillus flavus as potential biocontrol agents for aflatoxin in maize

Aflatoxin contamination resulting from maize infection by Aspergillus flavus is both an economic and a public health concern. Therefore, strategies for controlling aflatoxin contamination in maize are being investigated. The abilities of eleven naturally occurring atoxigenic isolates in Nigeria to reduce aflatoxin contamination in maize were evaluated in grain competition experiments and in field studies during the 2005 and 2006 growing seasons. Treatments consisted of inoculation of either grains in vials or ears at mid-silking stage in field plots, with the toxigenic isolate (La3228) or atoxigenic isolate alone and co-inoculation of each atoxigenic isolate and La3328. Aflatoxin B1 + B2 concentrations were significantly (p < 0.05) lower in the co-inoculation treatments compared with the treatment in which the aflatoxin-producing isolate La3228 was inoculated alone. Relative levels of aflatoxin B1 + B2 reduction ranged from 70.1% to 99.9%. Among the atoxigenics, two isolates from Lafia, La3279 and La3303, were most effective at reducing aflatoxin B1 + B2 concentrations in both laboratory and field trials. These two isolates have potential value as agents for the biocontrol of aflatoxin contamination in maize. Because these isolates are endemic to West Africa, they are both more likely than introduced isolates to be well adapted to West African environments and to meet regulatory concerns over their use throughout that region.

Comparison of Field, Greenhouse, and Detached-Leaf Evaluations of Soybean Germplasm for Resistance to Phakopsora pachyrhizi

Fourteen soybean accessions and breeding lines were evaluated for resistance to soybean rust caused by the fungus Phakopsora pachyrhizi. Evaluations were conducted in replicated experiments in growth chambers using detached leaves and under greenhouse and field conditions. In growth-chamber experiments, inoculation of detached leaves with 1 × 106 spores/ml resulted in a significantly (P < 0.0001) higher total number of pustules and spores per unit leaf area than inoculations with lower spore concentrations. Amending agar medium with plant hormones significantly (P < 0.0001) aided retention of green leaf color in detached leaves. Leaf pieces on a medium containing kinetin at 10 mg/liter had 5% chlorosis at 18 days after plating compared with leaf pieces on media amended with all other plant hormones, which had higher levels of chlorosis. Leaf age significantly affected number of pustules (P = 0.0146) and number of spores per pustule (P = 0.0088), and 3- to 4-week-old leaves had a higher number of pustules and number of spores per pustule compared with leaves that were either 1 to 2 or 5 to 6 weeks old. In detached-leaf and greenhouse screening, plants were evaluated for days to lesion appearance, days to pustule formation, days to pustule eruption, lesion number, lesion diameter, lesion type, number of pustules, and spores per pustule in 1-cm2 leaf area. Plants also were evaluated for diseased leaf area (in greenhouse and field screening) and sporulation (in field screening) at growth stage R6. There were significant (P < 0.0001) differences among genotypes in their response to P. pachyrhizi infection in the detached-leaf, greenhouse, and field evaluations. Accessions PI 594538A, PI 417089A, and UG-5 had very low levels of disease compared with the susceptible checks and all other genotypes. Detached-leaf, greenhouse, and field results were comparable, and there were significant correlations between detached-leaf and greenhouse (absolute r = 0.79; P < 0.0001) and between detached-leaf and field resistance (absolute r = 0.83; P < 0.0001) across genotypes. The overall results show the utility of detached-leaf assay for screening soybean for rust resistance.

Evaluation of Soybean Germplasm for Resistance to Soybean Rust (Phakopsora pachyrhizi) in Nigeria

Soybean rust, caused by Phakopsora pachyrhizi, is one of the most important constraints to soybean production worldwide. The absence of high levels of host resistance to the pathogen has necessitated the continued search and identification of sources of resistance. In one set of experiments, 178 soybean breeding lines from the International Institute of Tropical Agriculture were rated for rust severity in the field in 2002 and 2003 at Ile-Ife, Yandev, and Ibadan, Nigeria. Thirty-six lines with disease severity ≤3 (based on a 0-to-5 scale) were selected for a second round of evaluation in 2004 at Ibadan. In the third round of evaluation under inoculated field conditions, 11 breeding lines with disease severity ≤2 were further evaluated for rust resistance at Ibadan in 2005 and 2006. The breeding lines TGx 1835-10E, TGx 1895-50F, and TGx 1903-3F consistently had the lowest level of disease severity across years and locations. In another set of experiments, 101 accessions from the United States Department of Agriculture-Agricultural Research Service and National Agriculture Research Organization (Uganda) were evaluated in the first round in 2005 under inoculated conditions in the screenhouse; 12 accessions with disease severity ≤20% leaf area infected were selected for evaluation in the second round in 2005 and 2006 under inoculated field conditions at Ibadan. Highly significant differences (P < 0.0001) in disease severity were observed among the 101 accessions during this first round of rust evaluation. Significant (P < 0.0001) differences in rust severity and sporulation also were observed among the 12 selected accessions. Accessions PI 594538A, PI 417089A, and UG-5 had significantly (P < 0.05) lower disease severity than all other selected accessions in both years of evaluation, with rust severities ranging from 0.1 to 2.4%. These results indicate that some of the breeding lines (TGx 1835-10E, TGx 1895-50F, and TGx 1903-3F) and accessions (PI 594538A, PI 417089A, and UG-5) would be useful sources of soybean rust resistance genes for incorporation into high-yielding and adapted cultivars.

Spatiotemporal Spread of Cucurbit Downy Mildew in the Eastern United States

The dynamics of cucurbit downy mildew, caused by Pseudoperonospora cubensis, in the eastern United States in 2008 and 2009 were investigated based on disease records collected in 24 states as part of the Cucurbit downy mildew ipmPIPE monitoring program. The mean season-long rate of temporal disease progress across the 2 years was 1.4 new cases per day. Although cucurbit downy mildew was detected in mid-February and early March in southern Florida, the disease progressed slowly during the spring and early summer and did not enter its exponential phase until mid-June. The median nearest-neighbor distance of spread of new disease cases was ≈110 km in both years, with ≈15% of the distances being >240 km. Considering disease epidemics on all cucurbits, the epidemic expanded at a rate of 9.2 and 10.5 km per day in 2008 and 2009, respectively. These rates of spatial spread are at the lower range of those reported for the annual spread of tobacco blue mold in the southeastern United States, a disease that is also aerially dispersed over long distances. These results suggest that regional spread of cucurbit downy mildew may be limited by opportunities for establishment in the first half of the year, when fewer cucurbit hosts are available for infection. The O-ring statistic was used to determine the spatial pattern of cucurbit downy mildew outbreaks using complete spatial randomness as the null model for hypothesis testing. Disease outbreaks in both years were spatially aggregated and the extent of spatial dependence was up to 1,000 km. Results from the spatial analysis suggests that disease outbreaks in the Great Lakes and mid-Atlantic regions may be due to the spread of P. cubensis sporangia from outbreaks of the disease near the Georgia/South Carolina/North Carolina border rather than from overwintering sites in southern Florida. Space-time point pattern analysis indicated strong (P < 0.001) evidence for a space-time interaction and a space-time risk window of ≈3 to 5 months after first disease outbreak and 300 to 600 km was detected in both years. Results of this study support the hypothesis that infection of cucurbits by P. cubensis appears to be an outcome of a contagion process, and the relative large space-time window suggests that factors occurring on a large spatial scale (≈1,000 km) facilitate the spread of cucurbit downy mildew in the eastern United States.

Resurgence of Pseudoperonospora cubensis: The Causal Agent of Cucurbit Downy Mildew

The downy mildew pathogen, Pseudoperonospora cubensis, which infects plant species in the family Cucurbitaceae, has undergone major changes during the last decade. Disease severity and epidemics are far more destructive than previously reported, and new genotypes, races, pathotypes, and mating types of the pathogen have been discovered in populations from around the globe as a result of the resurgence of the disease. Consequently, disease control through host plant resistance and fungicide applications has become more complex. This resurgence of P. cubensis offers challenges to scientists in many research areas including pathogen biology, epidemiology and dispersal, population structure and population genetics, host preference, host-pathogen interactions and gene expression, genetic host plant resistance, inheritance of host and fungicide resistance, and chemical disease control. This review serves to summarize the current status of this major pathogen and to guide future management and research efforts within this pathosystem.

Pathogenic Variation of Phakopsora pachyrhizi Infecting Soybean in Nigeria

Soybean rust, caused by Phakopsora pachyrhizi, is an important disease in Nigeria and many other soybean-producing countries worldwide. To determine the geographical distribution of soybean rust in Nigeria, soybean fields were surveyed in the Derived Savanna (DS), Northern Guinea Savanna (NGS), and Southern Guinea Savanna (SGS) agroecological zones in Nigeria between 2004 and 2006. Disease severity in each zone was determined and analyzed using geostatistics. Prevalence of infected fields and disease severity in surveyed fields were significantly (P < 0.05) different between geographical zones with both variables being higher in the DS zone than in either NGS or SGS zones. Geostatistical analysis indicated that the spatial influence of disease severity at one location on severity at other locations was between 75 and 120 km. An exponential model best described the relationship between semivariance and lag distance when rust severity was high. Spatial interpolation of rust severity showed that locations in the DS zone were more conducive for the rust epidemic compared to areas in the NGS zone. In the 2005 survey, 116 purified isolates were established in culture on detached soybean leaves. To establish the nature of pathogenic variation in P. pachyrhizi, a set of four soybean accessions with Rpp(1), Rpp(2), Rpp(3), and Rpp(4) resistance genes, two highly resistant and two highly susceptible genotypes were inoculated with single uredinial isolates. Principal component analysis on the number of uredinia per square centimeter of leaf tissue for 116 isolates indicated that an adequate summary of pathogenic variation was obtained using only four genotypes. Of these four, PI 459025B (with Rpp(4) gene) and TGx 1485-1D had the lowest and highest number of uredinia per square centimeter, respectively. Based on cluster analysis of the number of uredinia per square centimeter, seven pathotype clusters were determined. Isolates in cluster III were the most virulent, while those in cluster IV were the least virulent. Shannons index (H) revealed a more diverse pathogen population in the DS zone (H = 1.21) compared to the rust population in SGS and NGS with H values of 1.08 and 0.91, respectively. This work will be useful in breeding and management of soybean rust by facilitating identification of resistant genotypes and targeting cultivars with specific resistance to match prevailing P. pachyrhizi pathotypes in a given geographical zone.

A Quantitative Review of Fungicide Efficacy for Managing Downy Mildew in Cucurbits

A meta-analysis of the effect of fungicides on cucurbit downy mildew was conducted using data previously published in Fungicide & Nematicide Tests and Plant Disease Management Reports from 2000 to 2008. Standardized mean effect size (the difference in disease intensity expressed in standard deviation units between the fungicide treatment and its corresponding untreated control) was calculated for each of the 105 field studies evaluating the effects of fungicides on cucurbit downy mildew. Fixed- and random-effects meta-analyses were performed on the log-transformed standardized mean effect sizes to estimate the overall mean effect size (L+), and to determine the variability in the effect size among studies. Fungicides led to a significant (P<0.001) reduction in disease with an L+ value of 1.198, indicating that, overall, fungicides were highly effective against cucurbit downy mildew. Fixed- and random-effects meta-analyses were then conducted to determine the effects of moderator variables on L+. The fixed-effects model resulted in narrower 95% confidence intervals and generally lower estimates of moderator subgroup mean effect size (L) than the random-effects models. Fungicide efficacy was significantly (P<0.001) greater in cucumber, with L being 26.5% higher in cucumber than in other cucurbits. Year of study, number of sprays, and disease pressure in the control significantly (P<0.001) affected L. Fungicide efficacy was significantly lower during the 2004-05 season than prior to or after the 2004-05 season. Studies in which disease pressure was moderate had a significantly higher L than studies with either low or high disease pressure. Fungicide efficacy was ≥22% in studies that received 5 to 6 sprays than in studies that received 1 to 4 or >6 sprays. Fungicide products led to a significant (P<0.001) reduction in disease, although there was substantial between-study variability. The pyridinylmethyl-benzamide group of fungicides (fluopicolide) was the most effective, followed by the carbamate (propamocarb) and quinone inside inhibitors (cyazofamid) group of fungicides, while the carboxylic acid amide group (mandipropamid and dimethomorph) was the least effective. A combination of protectant and systemic fungicides resulted in a proportional increase in L compared with sole application of either protectant or systemic fungicides.

Resurgence of Cucurbit Downy Mildew in the United States: A Watershed Event for Research and Extension

In 2004, an outbreak of cucurbit downy mildew (CDM) caused by the oomycete Pseudoperonospora cubensis (Berk. & M. A. Curtis) Rostovzev resulted in an epidemic that stunned the cucumber (Cucumis sativus L.) industry in the eastern United States. The disease affects all major cucurbit crops, including cucumber, muskmelon, squashes, and watermelon. Although the 2004 epidemic began in North Carolina, the cucumber crop from Florida to the northern growing regions in the United States was devastated, resulting in complete crop loss in several areas. Many cucumber fields were abandoned prior to harvest. The rapid spread of the disease coupled with the failure of fungicide control programs surprised growers, crop consultants, and extension specialists. The epidemic raised several fundamental questions about the potential causes for the resurgence of the disease. Some of these questions revolved around whether the epidemic would recur in subsequent years and the possible roles that changes in the host, pathogen, and environment may have played in the epidemic.

Quantitative models for germination and infection of Pseudoperonospora cubensis in response to temperature and duration of leaf wetness.

The influence of temperature and leaf wetness duration on germination of sporangia and infection of cantaloupe leaves by Pseudoperonospora cubensis was examined in three independent controlled-environment experiments by inoculating plants with a spore suspension and exposing them to a range of leaf wetness durations (2 to 24 h) at six fixed temperatures (5 to 30 degrees C). Germination of sporangia was assessed at the end of each wetness period and infection was evaluated from assessments of disease severity 5 days after inoculation. Three response surface models based on modified forms of the Weibull function were evaluated for their ability to describe germination of sporangia and infection in response to temperature and leaf wetness duration. The models estimated 15.7 to 17.3 and 19.5 to 21.7 degrees C as the optimum temperature (t) range for germination and infection, respectively, with little germination or infection at 5 or 30 degrees C. For wetness periods of 4 to 8 h, a distinct optimum for infection was observed at t = 20 degrees C but broader optimum curves resulted from wetness periods >8 h. Model 1 of the form f(w,t) = f(t) x (1 - exp{-[B x w](D)}) resulted in smaller asymptotic standard errors and yielded higher correlations between observed and predicted germination and infection data than either model 2 of the form f(w,t) = A{1 - exp[- f(t) x (w - C)](D)} or model 3 of the form f(w,t) = [1 - exp{-(B x w)(2)}]/cosh[(t - F)G/2]. Models 1 and 2 had nonsignificant lack-of-fit test statistics for both germination and infection data, whereas a lack-of-fit test was significant for model 3. The models accounted for approximately 87% (model 3) to 98% (model 1) of the total variation in the germination and infection data. In the validation of the models using data generated with a different isolate of P. cubensis, slopes of the regression line between observed and predicted germination and infection data were not significantly different (P > 0.2487) and correlation coefficients between observed and predicted values were high (r(2) > 0.81). Models 1 and 2 were used to construct risk threshold charts that can be used to estimate the potential risk for infection based on observed or forecasted temperature and leaf wetness duration.