Age-Associated Pathology in Rhesus Macaques (Macaca mulatta).

The rhesus macaque (Macaca mulatta) is one of the most extensively used nonhuman primate models for human diseases. This article presents a literature review focusing on major organ systems and age-associated conditions in humans and primates, combined with information from the Wisconsin National Primate Research Center Electronic Health Record database to highlight and contrast age-associated lesions in geriatric rhesus macaques with younger cohorts. Rhesus macaques are excellent models for age-associated conditions, including diabetes, osteoarthritis, endometriosis, visual accommodation, hypertension, osteoporosis, and amyloidosis. Adenocarcinoma of the large intestine (ileocecocolic junction, cecum, and colon) is the most common spontaneous neoplasm in the rhesus macaque. A combination of cross-sectional and longitudinal studies is required to truly define mechanisms of maturation, aging, and the pathology of age-associated conditions in macaques and thus humans. The rhesus macaque is and will continue to be an appropriate and valuable model for investigation of the mechanisms and treatment of age-associated diseases.

Real-Time PCR Assay for Detection of Sphacelotheca reiliana Infection in Maize (Zea mays) Seedlings and Evaluation of Seed Treatment Efficacy.

Head smut of maize, caused by the fungus Sphacelotheca reiliana, is an economically important disease in all major maize-producing countries. Although seed treatments are commonly used for management purposes, evaluating these treatments for efficacy is both time consuming and inefficient. Therefore, in order to improve the capacity for evaluating seed treatment fungicides, we developed a real-time PCR-based seedling assay for S. reiliana infection. We optimized growth chamber conditions and inoculation methods to achieve infection incidence of 60 to 80% in inoculated, nontreated controls. The effects of five commercially available fungicidal seed treatment formulations on seedling infection incidence were compared by PCR analysis of root and mesocotyl tissues. Tebuconazole, fludioxonil, sedaxane, and Maxim Quattro (fludioxonil+mefenoxam+azoxystrobin+thiabendazole) reduced the incidence of infection (P < 0.05) compared with the control, but no difference was found between the azoxystrobin treatment and the control. All rates tested for both sedaxane and tebuconazole were equally effective for seeds coated with 106 teliospores∙seed-1. Sedaxane, at a rate of 0.1 mg/kernel, eliminated seedling infection if seeds were infested with a low inoculum concentration (101 teliospores∙seed-1). The assay developed here is a valuable tool not only for the detection of fungal infection at the seedling stage, but also for testing the relative efficacies of seed treatments for reducing incidence of infection.

Transgenic Virus Resistance in Crop-Wild Cucurbita pepo Does Not Prevent Vertical Transmission of Zucchini yellow mosaic virus.

Zucchini yellow mosaic virus (ZYMV) is an economically important pathogen of cucurbits that is transmitted both horizontally and vertically. Although ZYMV is seed-transmitted in Cucurbita pepo, the potential for seed transmission in virus-resistant transgenic cultivars is not known. We crossed and backcrossed a transgenic squash cultivar with wild C. pepo, and determined whether seed-to-seedling transmission of ZYMV was possible in seeds harvested from transgenic backcrossed C. pepo. We then compared these transmission rates to those of non-transgenic (backcrossed and wild) C. pepo. The overall seed-to-seedling transmission rate in ZYMV was similar to those found in previous studies (1.37%), with no significant difference between transgenic backcrossed (2.48%) and non-transgenic (1.03%) backcrossed and wild squash. Fewer transgenic backcrossed plants had symptom development (7%) in comparison with all non-transgenic plants (26%) and may be instrumental in preventing yield reduction due to ZYMV. Our study shows that ZYMV is seed transmitted in transgenic backcrossed squash, which may affect the spread of ZYMV via the movement of ZYMV-infected seeds. Deep genome sequencing of the seed-transmitted viral populations revealed that 23% of the variants found in this study were present in other vertically transmitted ZYMV populations, suggesting that these variants may be necessary for seed transmission or are distributed geographically via seeds.

Special consideration regarding the assessment and management of patients being treated with mandibular advancement oral appliance therapy for snoring and obstructive sleep apnea.

This position paper, as developed by a Task Force of the American Academy of Craniofacial Pain on Mandibular Advancement Oral Appliance Therapy for Snoring and Obstructive Sleep Apnea, contains recommendations for dentists engaged in the management of patients with snoring and obstructive sleep apnea utilizing mandibular advancement oral appliances. The recommendations are supported by current scientific evidence, published standards and guidelines, and expert panel consensus. Snoring and obstructive sleep apnea (OSA) affects millions of people. Oral appliance therapy (OAT) is recognized as an effective therapy for many with primary snoring and mild to moderate OSA, as well as those with more severe OSA who cannot tolerate positive airway pressure (PAP) therapies. Dentists are playing a much larger role in the screening and management of patients with snoring and OSA as part of a multi-disciplinary team. It is also recognized that OAT has the potential to cause untoward side effects, including temporomandibular joint (TMJ) pain and dysfunction. The present paper highlights the need for dentists who manage patients using mandibular advancement OAT to be competent in the assessment, diagnosis and management of temporomandibular disorders (TMDs) and craniofacial pain disorders. The authors of this article are all clinically engaged in the management of patients with snoring and OSA, and reached consensus based on their review of the current evidence, published guidelines and clinical experience. It is the opinion of the authors that dentists experienced and knowledgeable in the assessment, diagnosis and management of TMD and craniofacial pain applying this knowledge to the management of patients with snoring and OSA using OAT will provide their patients with the best prognosis and most successful treatment outcomes.

Deep sequencing reveals persistence of intra- and inter-host genetic diversity in natural and greenhouse populations of zucchini yellow mosaic virus.

The genetic diversity present in populations of RNA viruses is likely to be strongly modulated by aspects of their life history, including mode of transmission. However, how transmission mode shapes patterns of intra- and inter-host genetic diversity, particularly when acting in combination with de novo mutation, population bottlenecks and the selection of advantageous mutations, is poorly understood. To address these issues, this study performed ultradeep sequencing of zucchini yellow mosaic virus in a wild gourd, Cucurbita pepo ssp. texana, under two infection conditions: aphid vectored and mechanically inoculated, achieving a mean coverage of approximately 10 ,000×. It was shown that mutations persisted during inter-host transmission events in both the aphid vectored and mechanically inoculated populations, suggesting that the vector-imposed transmission bottleneck is not as extreme as previously supposed. Similarly, mutations were found to persist within individual hosts, arguing against strong systemic bottlenecks. Strikingly, mutations were seen to go to fixation in the aphid-vectored plants, suggestive of a major fitness advantage, but remained at low frequency in the mechanically inoculated plants. Overall, this study highlights the utility of ultradeep sequencing in providing high-resolution data capable of revealing the nature of virus evolution, particularly as the full spectrum of genetic diversity within a population may not be uncovered without sequence coverage of at least 2500-fold.

Meeting report: Spontaneous lesions and diseases in wild, captive-bred, and zoo-housed nonhuman primates and in nonhuman primate species used in drug safety studies.

The combination of loss of habitat, human population encroachment, and increased demand of select nonhuman primates for biomedical research has significantly affected populations. There remains a need for knowledge and expertise in understanding background findings as related to the age, source, strain, and disease status of nonhuman primates. In particular, for safety/biomedical studies, a broader understanding and documentation of lesions would help clarify background from drug-related findings. A workshop and a minisymposium on spontaneous lesions and diseases in nonhuman primates were sponsored by the concurrent Annual Meetings of the American College of Veterinary Pathologists and the American Society for Veterinary Clinical Pathology held December 3-4, 2011, in Nashville, Tennessee. The first session had presentations from Drs Lowenstine and Montali, pathologists with extensive experience in wild and zoo populations of nonhuman primates, which was followed by presentations of 20 unique case reports of rare or newly observed spontaneous lesions in nonhuman primates (see online files for access to digital whole-slide images corresponding to each case report at http://www.scanscope.com/ACVP%20Slide%20Seminars/2011/Primate%20Pathology/view.apml). The minisymposium was composed of 5 nonhuman-primate researchers (Drs Bradley, Cline, Sasseville, Miller, Hutto) who concentrated on background and spontaneous lesions in nonhuman primates used in drug safety studies. Cynomolgus and rhesus macaques were emphasized, with some material presented on common marmosets. Congenital, acquired, inflammatory, and neoplastic changes were highlighed with a focus on clinical, macroscopic, and histopathologic findings that could confound the interpretation of drug safety studies.

A critical review of Dr. Charles S. Greene's article titled "Managing the Care of Patients with Temporomandibular Disorders: a new Guideline for Care" and a revision of the American Association for Dental Research's 1996 policy statement on temporomandibular disorders, approved by the AADR Council in March 2010, published in the Journal of the American Dental Association September 2010.

Dr. Charles Greene's article, "Managing the Care of Patients with TMDs A New Guideline for Care," and the American Association for Dental Research's (AADR) 2010 Policy Statement on Temporomandibular Disorders, published in the Journal of the American Dental Association (JADA) September 2010, are reviewed in detail. The concept that all temporomandibular disorders (TMDs) should be lumped into one policy statement for care is inappropriate. TMDs are a collection of disorders that are treated differently, and the concept that TMDs must only be managed within a biopsychosocial model of care is inappropriate. TMDs are usually a musculoskeletal orthopedic disorder, as defined by the AADR. TMD orthopedic care that is peer-reviewed and evidence-based is available and appropriate for some TMDs. Organized dentistry, including the American Dental Association, and mainstream texts on TMDs, support the use of orthopedics in the treatment of some TMDs. TMDs are not psychological or social disorders. Informed consent requires that alternative care is discussed with patients. Standard of care is a legal concept that is usually decided by a court of law and not decided by a policy statement, position paper, guidelines or parameters of care handed down by professional organizations. The 2010 AADR Policy Statement on TMD is not the standard of care in the United States. Whether a patient needs care for a TMD is not decided by a diagnostic test, but by whether the patient has significant pain, dysfunction and/or a negative change in quality of life from a TMD and they want care. Some TMDs need timely invasive and irreversible care.

Experimental Verification of Seed Transmission of Zucchini yellow mosaic virus.

Within two decades of its discovery, Zucchini yellow mosaic virus (ZYMV) achieved a global distribution. However, whether or not seed transmission occurs in this economically significant crop pathogen is controversial, and the relative impact of seed transmission on the epidemiology of ZYMV remains unclear. Using reverse transcription-polymerase chain reaction, we observed a seed transmission rate of 1.6% in Cucurbita pepo subsp. texana and show that seed-infected C. pepo plants are capable of initiating horizontal ZYMV infections, both mechanically and via an aphid vector (Myzus persicae). We also provide evidence that ZYMV-infected seeds may act as effective viral reservoirs, partially accounting for the current geographic distribution of ZYMV. Finally, the observation that ZYMV infection of C. pepo seeds results in virtually symptomless infection, coupled with our finding that an antibody test failed to detect vertically transmitted ZYMV in infected seed, highlights the urgent need to standardize current detection methods for seed infection.

Studies on congenital osteopetrosis in microphthalmic mice using organ cultures: impairment of bone resorption in response to physiologic stimulators.

The mechanism of congenital osteopetrosis in microphthalmic (mi) mice has been examined in bone organ cultures. Resorption was measured by the release of previously incorporated 45Ca in fetal long bones and newborn calvaria from mi mice and heterozygous or homozygous normal litter mates. Bones from mi mice showed a generalized resorption defect with decreased spontaneous or control resorption and failure to respond to parathyroid hormone (PTH), prostaglandin E2, 1,25 dihydroxy vitamin D3, vitamin A, or osteoclast activating factor (OAF) from human peripheral leukocytes or mouse spleen cells. Bones from heterozygotes showed a smaller response to PTH than bones from homozygous normals. Mutant bones failed to show an increase in lysosomal enzyme release in response to PTH or vitamin A, agents which increased release from bones of homozygous normals. Proline incorporation into collagenase-digestible protein was similar in cultures of normal and mutant bone and was inhibited by PTH and OAF. These results indicate that congenital osteopetrosis in mi mice is due to a generalized defect in the function and hormonal response of osteoclasts and suggests that this cell line is separate from the osteoblast cell line which shows no impairment of hormonal response.

Detection of PrPsc in blood from sheep infected with the scrapie and bovine spongiform encephalopathy agents.

The role of blood in the iatrogenic transmission of transmissible spongiform encephalopathy (TSE) or prion disease has become an increasing concern since the reports of variant Creutzfeldt-Jakob disease (vCJD) transmission through blood transfusion from humans with subclinical infection. The development of highly sensitive rapid assays to screen for prion infection in blood is of high priority in order to facilitate the prevention of transmission via blood and blood products. In the present study we show that PrP(sc), a surrogate marker for TSE infection, can be detected in cells isolated from the blood from naturally and experimentally infected sheep by using a rapid ligand-based immunoassay. In sheep with clinical disease, PrP(sc) was detected in the blood of 55% of scrapie agent-infected animals (n = 80) and 71% of animals with bovine spongiform encephalopathy (n = 7). PrP(sc) was also detected several months before the onset of clinical signs in a subset of scrapie agent-infected sheep, followed from 3 months of age to clinical disease. This study confirms that PrP(sc) is associated with the cellular component of blood and can be detected in preclinical sheep by an immunoassay in the absence of in vitro or in vivo amplification.

The effects of parathyroid hormone, colchicine, and calcitonin on the ultrastructure and the activity of osteoclasts in organ culture.

The ultrastructure of osteoclasts was examined in fetal rat bones after stimulation or inhibition of resorption in culture. A central ruffled border area completely encircled by a clear zone was considered to represent the resorbing system of the cell. The proportion of ruffled border and clear zone in osteoclast cross sections was compared with changes in bone resorption as measured by the release of previously incorporated radioactive calcium ((45)Ca). In control cultures 55% of the osteoclast cross sections showed an area closely apposed to bone and this consisted mainly of clear zone; only 11% showed ruffled borders. Treatment with parathyroid hormone (PTH) increased (45)Ca release, increased the frequency of finding areas closely apposed to bone (79%), and markedly increased the frequency of the ruffled border area (64%). Colchicine given concurrently with PTH decreased the number of osteoclasts. Colchicine or calcitonin treatment after PTH stimulation decreased the proportion of ruffled border area significantly by 1 h; this was followed by a decrease in (45)Ca release. These inhibited osteoclasts resembled osteoclasts from control, unstimulated cultures, suggesting that the cells had returned to their inactive state. Colchicine-treated osteoclasts also showed a loss of microtubules and a massive accumulation of 100 A filaments, suggesting that synthesis of microtubular subunits had increased.

Complement-dependent stimulation of prostaglandin synthesis and bone resorption.

Complement-sufficient heterologous serum induced prostaglandin synthesis and resultant resorption in cultures of fetal rat long bones. Bone resorption was enhanced with unheated normal rabbit serum as compared to heated serum or serum from rabbits lacking the sixth component of complement (C6). Addition of functionally purified C6 restored resorptive activity in C6-deficient serum. Concentrations of prostaglandin E were increased in the culture media of bones incubated with complement-sufficient serum. The resorptive effects of active serum as well as the appearance of prostaglandin E in the media were inhibited by indomethacin.

Bone resorbing activity in supernatant fluid from cultured human peripheral blood leukocytes.

A new soluble mediator was found in supernatant fluid from cultures of human peripheral blood leukocytes that were stimulated by phytohemagglutinin, or by antigenic material present in human dental plaque deposits. This soluble Jactor produced bone resorption in organ cultures of fetal rat bones as measured by increased release of calcium-45, and also increased the number of active osteoclasts.

Parathyroid hormone-related protein of malignancy: active synthetic fragments.

Peptides corresponding to the amino-terminal region of the parathyroid hormone-related protein (PTHrP) of humoral hypercalcemia of malignancy were synthesized. A 34-amino acid peptide, PTHrP(1-34), was two to four times more potent than bovine or human PTH(1-34) in bioassays promoting the formation of adenosine 3',5'-monophosphate (cAMP) and plasminogen activator activity in osteogenic sarcoma cells and adenylate cyclase activity in chick kidney membranes. Like parathyroid hormone itself, in which the activity resides in the first 34 residues, PTHrP peptides of less than 30 residues from the amino terminus showed substantially reduced activity. PTHrP(1-34) had only 6% of the potency of bovine PTH(1-34) in promoting bone resorption in vitro. PTHrP(1-34) strongly promoted the excretion of cAMP and phosphorus and reduced the excretion of calcium in the isolated, perfused rat kidney consistent with the symptoms seen in malignant hypercalcemia.