Incidence of Mite-Vectored Viruses of Wheat in the Texas High Plains and Interactions With Their Host and Vector.

Mite-vectored virus diseases of wheat are common throughout the Great Plains and cause significant economic losses to growers each year. These diseases are caused by Wheat streak mosaic virus (WSMV), Triticum mosaic virus (TriMV), and Wheat mosaic virus (WMoV), all of which are transmitted by the wheat curl mite (WCM), Aceria tosichella Keifer. New wheat cultivars with tolerance or resistance to WSMV have been released recently, but their widespread cultivation and potential impact on mite-transmitted virus incidence in the Texas Panhandle was unknown. A total of 648 symptomatic wheat samples were collected from 26 counties, predominately in the Texas Panhandle, and tested by enzyme-linked immunosorbent assay (ELISA) for WSMV, TriMV, and WMoV. Samples that tested negative by ELISA were subsequently tested by real-time quantitative PCR (qPCR) for each virus. Approximately 93% of the samples tested by ELISA were positive for WSMV, 43% were positive for TriMV, and 7% were positive for WMoV. Eleven samples tested positive only for TriMV, but none were positive only for WMoV. When samples that tested negative for the different viruses by ELISA were retested by real-time qPCR, detection of each virus was significantly increased. When results of the ELISA test and qPCR were combined, 100% of the 648 samples tested positive for WSMV, approximately 94% were positive for TriMV, and 23% were positive for WMoV. This demonstrated that the incidence of TriMV in the Texas High Plains is much greater than previously reported. The fact that real-time qPCR revealed over a 2-fold increase in the incidence of TriMV and a 3-fold increase in WMoV demonstrated that the ELISA test, which is commonly used by diagnostic laboratories in the Great Plains, should not be used for studies requiring a high degree of sensitivity and accuracy in virus detection. After initial virus infection status was determined, samples that tested positive for WSMV and TriMV were further observed for WCM infestation. A total of 292 samples were inspected and a total of 101 mites were collected from 40 tillers. Individual mites and the tillers from which they were recovered were tested by real-time qPCR to determine how copy numbers of WSMV and TriMV in mites and host tissue compared, and whether the WSMV/TriMV copy number ratio in individual mites was similar to that of the host tissue from which they were collected. In all mites and tillers tested, the WSMV copy number was always higher than that of TriMV and copy numbers of both viruses were always higher in plant tissue than in mites. Although there was a significant correlation between the WSMV/TriMV copy number ratio in plant tissue and in associated mites, the correlation coefficient was very low (r = 0.31, P = 0.0248). In the majority of comparisons, the WSMV/TriMV ratio was higher in individual mites than in the tiller from which they were recovered. The reason for this increase is unknown but indicates that mites may preferentially acquire WSMV from tillers coinfected with WSMV and TriMV, a finding that could have significant implications for virus transmission and disease epidemiology.

Population Dynamics of Released Potato Psyllids and their Bacteriliferous Status in Relation to Zebra Chip Incidence in Caged Field Plots.

Potato psyllids vector 'Candidatus Liberibacter solanacearum' (Lso), the putative causal agent of potato zebra chip (ZC). Currently, sticky traps are the primary psyllid monitoring tools used by growers for making management decisions. However, the reliability of sticky traps in predicting psyllid numbers in potato fields has always been questioned. In 2013 and 2014, experiments were conducted in covered field plots at the Texas A&M AgriLife Research Station at Bushland to investigate the relationships among initial psyllid numbers, psyllids captured on sticky traps and their Lso status, and zebra chip incidence. Three densities of Lso-positive psyllids (5, 15, or 30/cage) were released under 2-week-old potato canopies with four replications in plot sizes of 7.6 to 9 m by 5 rows. Psyllids were released under the first plant in the center row and monitored weekly with a yellow sticky trap from the opposite end. Number of plants with zebra chip symptoms also was counted weekly beginning one month after infestation with psyllids. The total number of psyllids captured on sticky traps and disease incidence levels generally corresponded to the levels of psyllid density treatments (5 < 15 < 30), but the differences became more apparent toward the end of the experiments. Psyllid numbers in the different density treatments fluctuated more or less in synchrony over time, which appeared to reflect periodic emergence of new generations of psyllids. Initially, all captured psyllids tested positive for Lso. However, the proportions of psyllids testing positive declined dramatically after a few weeks, which suggested that the new generations of psyllids were devoid of Lso. Over all, less than 50% of captured psyllids tested positive for the pathogen. The decline in proportions of psyllids testing positive for Lso following successive generations has significant relevance to field situations and may partly explain why there are generally low percentages of Lso-positive psyllids under field conditions.

Effects of Potato-Psyllid-Vectored 'Candidatus Liberibacter solanacearum' Infection on Potato Leaf and Stem Physiology.

The bacterium 'Candidatus Liberibacter solanacearum' is associated with zebra chip disease (ZC), a threat to potato production in North America and New Zealand. It is vectored by potato psyllids. Previous studies observed that 'Ca. L. solanacearum' infection causes potato tubers to undergo ZC-symptom-associated shifts in physiology, such as increased levels of amino acids, sugars, and phenolics. However, little is known about how 'Ca. L. solanacearum' infections caused by psyllid vector feeding may affect metabolism in potato foliage and stems. This study compared metabolism in potato plants fed upon by 'Ca. L. solanacearum'-positive psyllids with potato plants not exposed to psyllids. Foliar levels of asparagine, aspartic acid, glutamine, fructose, glucose, sucrose, a ferulic acid derivative, and quinic acid were lower in 'Ca. L. solanacearum'-inoculated than noninfected plants. However, foliar levels of proline, serine, four phenolic compounds, and most terpenoids were greater in 'Ca. L. solanacearum'-inoculated than noninfected plants. Upper stem levels of asparagine and aspartic acid, upper and lower stem levels of ellagitannins and most monoterpenoids, and lower stem level of sesquiterpenoids were greater in 'Ca. L. solanacearum'-inoculated than noninfected plants. These results suggest that many defense-related terpenoid compounds might increase in plants which had psyllids inoculate 'Ca. L. solanacearum'. This could impact progression and spread of ZC.

Similarities and differences in physiological responses to 'Candidatus Liberibacter solanacearum' infection among different potato cultivars.

Zebra chip disease (ZC), putatively caused by the fastidious bacterium 'Candidatus Liberibacter solanacearum', is a threat to potato growers worldwide. However, little is known about biochemical shifts in different potato genotypes in response to 'Ca. L. solanacearum' infection. To address this, 'Red La Soda', 'Russet Norkotah', and 'FL 1867' potato were infected with 'Ca. L. solanacearum' 4, 3, 2, and 1 weeks before harvest to observe variability in cultivar responses to 'Ca. L. solanacearum' infection. ZC symptoms, 'Ca. L. solanacearum' titers, and tuber biochemistry were assessed. Red La Soda tubers exhibited greater symptoms when infected for 4 weeks than Russet Norkotah or FL 1867 tubers. 'Ca. L. solanacearum' titers did not vary among cultivars. Tuber levels of amino acids, carbohydrates, and phenolics varied among cultivars but no consistent trends were observed. Individual amino acids and phenolics were greater in FL 1867 than Red La Soda, whereas others were greater in Red La Soda or Russet Norkotah than FL 1867. Most amino acids, carbohydrates, and phenolics were positively associated with infection duration and symptoms regardless of cultivar. Associations between most of the evaluated compounds and 'Ca. L. solanacearum' titer were positive in Red La Soda. However, no associations between 'Ca. L. solanacearum' quantity and compounds were observed in FL 1867 and Russet Norkotah.

Zebra Chip Disease Development in Relation to Plant Age and Time of 'Candidatus Liberibacter solanacearum' Infection.

A 2-year field study was conducted to evaluate plant susceptibility to 'Candidatus Liberibacter solanacearum', the putative causal agent of zebra chip disease (ZC). Incubation period of ZC, the rate of symptom progress, and the rate of pathogen population growth were evaluated for individual plants infested on different weeks after emergence. In foliage, incubation period was between 21 and 28 days. The pathogen was detected within leaf tissue in 3 to 4 weeks, regardless of the time of infestation. The rates of foliar symptom progress and pathogen population growth were uniform among all infestations. Although symptoms were observed in only 1.3% of tubers from plants infested 2 weeks before harvest, 74% of these tubers tested positive for the pathogen. There was a positive correlation between symptom severity and titer in the foliage. Within tubers, however, the relationship was negative but nonsignificant. Pathogen titer reached detectable levels some time between 7 to 14 days following infestation. Although yield reduction was significant only in plants infested during early stages of their growth, chemical management of potato psyllids needs to be continued until at least a week before harvest to minimize ZC impact on the tuber quality.

Zebra chip disease and potato biochemistry: tuber physiological changes in response to 'Candidatus Liberibacter solanacearum' infection over time.

Zebra chip disease, putatively caused by the bacterium 'Candidatus Liberibacter solanacearum', is of increasing concern to potato production in Mexico, the United States, and New Zealand. However, little is known about the etiology of this disease and changes that occur within host tubers that result in its symptoms. Previous studies found that increased levels of phenolics, amino acids, defense proteins, and carbohydrates in 'Ca. L. solanacearum'-infected tubers are associated with symptoms of zebra chip. This study was conducted to quantify variations in levels of these biochemical components in relation to the time of infestation, symptom severity, and 'Ca. L. solanacearum' titer. Levels of phenolics, peroxidases, polyphenol oxidases, and reducing sugars (glucose and, to some extent, fructose) changed during infection, with higher levels occurring in tubers infected at least 5 weeks before harvest than in those infected only a week before harvest and those of controls. Compared with the apical tuber ends, greater levels of phenolics, peroxidases, and sucrose occurred at the basal (stolon attachment) end of infected tubers. With the exception of phenolics, concentrations of the evaluated compounds were not associated with 'Ca. L. solanacearum' titer. However, there were significant associations between biochemical responses and symptom severity. The lack of a linear correlation between most plant biochemical responses and 'Ca. L. solanacearum' titer suggests that shifts in metabolic profiles are independent of variations in 'Ca. L. solanacearum' levels.

Characterization of management and environmental factors associated with regional variations in potato zebra chip occurrence.

Potato zebra chip (ZC), caused by the bacterial pathogen 'Candidatus Liberibacter solanacearum', which is vectored by the potato psyllid (Bactericera cockerelli), has caused widespread damage to U.S. potato production ever since its first discovery in south Texas in 2000. To determine the influence of environmental factors and management practices on ZC occurrence, data on management and meteorological variables, field locations, and psyllid counts were collected over a 3-year period (2010 to 2012) from six locations across the central United States (south Texas to Nebraska). At these locations, ZC-symptomatic plants were counted in 26 fields from systematically established 20 m × 30 m plots around the field edges and field interiors. Mean numbers of symptomatic plants per plot were classified into two intensity classes (ZC ≤ 3 or ZC > 3) and subjected to discriminant function and logistic regression analyses to determine which factors best distinguish between the two ZC intensity classes. Of all the variables, location, planting date, and maximum temperature were found to be the most important in distinguishing between ZC intensity classes. These variables correctly classified 88.5% of the fields into either of the two ZC-intensity classes. Logistic regression analysis of the individual variables showed that location accounted for 90% of the variations, followed by planting date (86%) and maximum temperature (70%). There was a low but significant (r = -0.44983, P = 0.0211) negative correlation between counts of psyllids testing positive for pathogen and latitudinal locations, indicating a south-to-north declining trend in counts of psyllids testing positive for the pathogen. A similar declining trend also was observed in ZC occurrence (r = -0.499, P = 0.0094).

Transmission efficiency of 'Candidatus Liberibacter solanacearum' and potato zebra chip disease progress in relation to pathogen titer, vector numbers, and feeding sites.

ABSTRACT With diseases caused by vector-borne plant pathogens, acquisition and inoculation are two primary stages of the transmission, which can determine vector efficiency in spreading the pathogen. The present study was initiated to quantify acquisition and inoculation successes of 'Candidatus Liberibacter solanacearum', the etiological agent of zebra chip disease of potato, by its psyllid vector, Bactericera cockerelli (Hemiptera: Triozidae). Acquisition success was evaluated in relation to feeding site on the host plant as well as the acquisition access period. Inoculation success was evaluated in relation to vector number (1 and 4) on the plants. Acquisition success was influenced by the feeding site on the plant. The highest acquisition success occurred when insects had access to the whole plant. The results of the inoculation study indicated that the rate of successfully inoculated plants increased with the vector number. Plants inoculated with multiple psyllids had higher bacterial titer at the point of inoculation. Although disease incubation period was significantly shorter in plants inoculated with multiple psyllids, this effect was heterogeneous across experimental blocks, and was independent of pathogen quantity detected in the leaflets 3 days postinoculation. Disease progress was not affected by bacterial quantity injected or psyllid numbers.

Assessments of the Edge Effect in Intensity of Potato Zebra Chip Disease.

Zebra chip is a newly emerging potato disease which imparts dark colorations on fried chips, rendering them unmarketable. The disease is associated with the phloem-limited proteobacterium 'Candidatus Liberibacter solancearum', vectored by the potato psyllid Bactericera cockerelli. First reported from Mexico in the mid-1990s, the disease was observed for the first time in Texas in 2000 and is now prevalent in several potato-producing regions of the United States. In this study, we were interested in investigating whether there are edge effects in zebra chip intensity that can be assessed as a "foot print" of the associated insect vector. In 2009, we conducted studies in three fields in the Texas Panhandle in paired plots of 10 by 20 m around the field edges and 100 m infield in which symptomatic plants were counted just before harvest. The number of plot pairs (edge and infield) ranged from 15 to 18 depending on the size of the fields. In a separate study, temporal disease progress was assessed in two fields around the edges of the center-pivot circle in approximately 10-by-450-m areas. In 2010, the paired plot studies were repeated in 10 potato fields in Texas, Kansas, and Nebraska. Zebra chip intensity data from the paired-plot studies for both years were analyzed using the Wilcoxon's signed-rank test, a nonparametric equivalent of the classical (parametric) paired t test. In the 2009 study in all three fields, the edge plots had significantly greater zebra chip intensity than the infield plots (P < 0.05). Edge plots in the 2010 study also had greater zebra chip intensity in all fields and the differences were significant in the majority of fields (P < 0.05). In the diseases progress study in both fields, weekly zebra chip intensity on the edges reached its maximum after the third week of its first detection, and the disease progress curves were best fitted with the second-degree polynomial (quadratic) for both fields. The 2-year study clearly demonstrated that zebra chip intensity in potato fields was greater on the edges than in the infields. This finding has significant implications for psyllid management because greater emphasis in psyllid control strategy can be directed toward the edges for better results.

Morphometrical studies of erythrophagocytosis in the spleen.

The elimination of altered RBC was investigated morphometrically in the rat spleen by determination of the percentage of erythrophagosomes in macrophages. PHZ and diamide treated RBC led to a time dependent increase of the percentage of erythrophagosomes in comparison to normal spleens. The deformability of the target RBC was lowered as revealed by measurements with glass micropipettes. RBC-microvesicles and diamide treated RBC are primarily loaded with IgG and undergo a rapid elimination. PHZ-RBC and cells with a prolonged stay in the spleen are sequestrated by secondary mechanisms. Details of primary and secondary elimination are discussed.