The potato cyst nematode Globodera pallida is a serious pest of potato crops. Nematode FMRFamide-like peptides (FLPs) are one of the most diverse neuropeptide families known, and modulate sensory and motor functions. As neuromuscular function is a well-established target for parasite control, parasitic nematode FLP signaling has significant potential in novel control strategies. In the absence of transgenic parasitic nematodes and the reported ineffectiveness of neuronal gene RNAi in Caenorhabditis elegans, nothing is known about flp function in nematode parasites. In attempts to evaluate flp function in G. pallida, we have discovered that, unlike in C. elegans, these genes are readily susceptible to RNAi. Silencing any of the five characterized G. pallida flp genes (Gp-flp-1, -6, -12, -14, or -18) incurred distinct aberrant behavioral phenotypes consistent with key roles in motor function. Further delineation of these effects revealed that double-stranded RNA exposure time (> or = 18 h) and concentration (> or = 0.1 microg/ml) were critical to the observed effects, which were reversible. G. pallida flp genes are essential to coordinated locomotory activities, do not display redundancy, and are susceptible to RNAi, paving the way for the investigation of RNAi-mediated flp gene silencing as a novel plant parasite control strategy.
Disparity for HLA-A or HLA-B antigens increases the risk of marrow graft rejection, but the relevance of HLA-C is unknown because typing methods have not been sufficiently accurate for clinical use. We designed a matched case-control study and employed DNA sequencing methods to evaluate the role of HLA-C disparity in 21 patients who experienced graft failure (cases) following transplantation with unmanipulated marrow from either HLA-A, B serologically matched, DRB1 matched (n = 14) or single locus mismatched (n = 7) unrelated donors. For each case, two patients who successfully engrafted were selected as controls based on similarity for factors known or suspected to influence engraftment. The estimated odds ratio (OR) of graft failure for an HLA-C mismatch relative to match (univariable model) was 5.2 (95% CI, 1.4, 19; P = .01). Serologically undetectable HLA-A or HLA-B allele disparity was also associated with graft failure. The association between HLA-C disparity and graft failure remained significant even after accounting for the contribution of HLA-A and/or HLA-B allele disparity (OR 4.0; 95% CI, 1.1, 15; likelihood ratio test P = .03). These results show that HLA-C functions as a transplantation antigen and that HLA-A and HLA-B allele mismatches are biologically important. Molecular-based methods for pretransplant assessment of class I compatibility should be implemented for the selection of unrelated marrow donors.
A double-blind study was carried out to evaluate the relative performance and reliability of the PCR/SSOP assay compared to conventional serological typing in identifying HLA-DR alleles. A total of 268 consecutive samples were entered into the study. In 14 (5.2%) of the cases, HLA-DR serology could not be performed due to poor cell viability, while in seven (2.6%) of the cases, PCR/SSOP typing could not be performed due to poor amplification or to contamination with exogenous DNA. Among samples that were successfully typed by both methods, serologic typing correctly identified 455/465 (97.9%) DR antigens, while PCR/SSOP correctly identified 464/465 (99.8%) DR alleles (p = 0.0117, McNemar's test). The majority of discrepancies in serologic typing resulted from a lack of discriminative alloantisera to identify DR6 or DR103. For the overall sample set (N = 268), serology provided accurate results in 244 (91.0%) cases, while PCR/SSOP provided accurate results in 260 (97.0%) cases (p = 0.0037). The results of this study demonstrate that PCR/SSOP typing for HLA-DRB1 alleles provides results that are equal to or surpass serological typing for HLA-DR antigens. In addition, the PCR/SSOP approach offers the advantages of better reagent availability, lower cost, more rapid turn-around time, and greater accuracy, all of which would warrant its use as an HLA typing method of choice.
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